dizocilpine-maleate and Brain-Ischemia

Unprecedented community containment measures were taken following the recent outbreak of COVID-19 in Italy. The aim of the study was to explore the self-reported future compliance of citizens with such measures and its relationship with potentially impactful psychological variables.. An online survey was completed by 931 people (18-76 years) distributed across the Italian territory. In addition to demographics, five dimensions were measured: self-reported compliance with containment measures over time (today, at 7, 14, 30, 60, 90, and 180 days from now) at three hypothetical risk levels (10, 50, 90% of likelihood of contracting the COVID-19), perceived risk, generalized anxiety, intolerance of uncertainty, and relevance of several psychological needs whose satisfaction is currently precluded.. The duration of containment measures plays a crucial role in tackling the spread of the disease as people will be less compliant over time. Psychological needs of citizens impacting on the compliance should be taken into account when planning an easing of the lockdown, along with interventions for protecting vulnerable groups from mental distress.. La apendicitis aguda (AA) es la urgencia quirúrgica abdominal más frecuente. No encontramos estudios específicos que evalúen el impacto de la pandemia causada por el coronavirus 2 (SARS-Cov-2) sobre la AA y su tratamiento quirúrgico. Analizamos la influencia de esta nueva patología sobre la AA.. Estudio observacional retrospectivo en pacientes intervenidos por AA desde enero hasta abril de 2020. Fueron clasificados según el momento de la apendicectomía, antes de la declaración del estado de alarma (Pre-COVID19) y después de la declaración del estado de alarma (Post-COVID19) en España. Se evaluaron variables demográficas, duración de la sintomatología, tipo de apendicitis, tiempo quirúrgico, estancia hospitalaria y complicaciones postoperatorias.. La pandemia por SARS-Cov-2 influye en el momento de diagnóstico de la apendicitis, así como en su grado de evolución y estancia hospitalaria. La peritonitis fue lo más frecuentemente observado. Una sospecha y orientación clínica más temprana, es necesaria para evitar un manejo inadecuado de este trastorno quirúrgico común.. The primary outcome is improvement in PaO. Findings will provide timely information on the safety, efficacy, and optimal dosing of t-PA to treat moderate/severe COVID-19-induced ARDS, which can be rapidly adapted to a phase III trial (NCT04357730; FDA IND 149634).. None.. The gut barrier is crucial in cirrhosis in preventing infection-causing bacteria that normally live in the gut from accessing the liver and other organs via the bloodstream. Herein, we characterised gut inflammation by measuring different markers in stool samples from patients at different stages of cirrhosis and comparing this to healthy people. These markers, when compared with equivalent markers usually measured in blood, were found to be very different in pattern and absolute levels, suggesting that there is significant gut inflammation in cirrhosis related to different immune system pathways to that seen outside of the gut. This provides new insights into gut-specific immune disturbances that predispose to complications of cirrhosis, and emphasises that a better understanding of the gut-liver axis is necessary to develop better targeted therapies.. La surveillance de l’intervalle QT a suscité beaucoup d’intérêt durant la pandémie de la COVID-19 en raison de l’utilisation de médicaments prolongeant l’intervalle QT et les préoccupations quant à la transmission virale par les électrocardiogrammes (ECG) en série. Nous avons posé l’hypothèse que la surveillance en continu de l’intervalle QT par télémétrie était associée à une meilleure détection des épisodes de prolongation de l’intervalle QT.. Nous avons introduit la télémétrie cardiaque en continu (TCC) à l’aide d’un algorithme de surveillance automatisée de l’intervalle QT dans nos unités de COVID-19. Les mesures automatisées quotidiennes de l’intervalle QT corrigé (auto-QTc) en fonction de la fréquence cardiaque maximale ont été enregistrées. Nous avons comparé la proportion des épisodes de prolongation marquée de l’intervalle QTc (QTc long), définie par un intervalle QTc ≥ 500 ms, chez les patients montrant une suspicion de COVID-19 ou ayant la COVID-19 qui avaient été admis avant et après la mise en place de la TCC (groupe témoin. La surveillance en continu de l’intervalle QT est supérieure à la norme de soins dans la détection des épisodes de QTc long et exige peu d’ECG. La réponse clinique aux épisodes de QTc long est sous-optimale.. Exposure to a model wildfire air pollution source modifies cardiovascular responses to HC challenge, suggesting air pollution sensitizes the body to systemic triggers.. Though the majority of HIV-infected adults who were on HAART had shown viral suppression, the rate of suppression was sub-optimal according to the UNAIDS 90-90-90 target to help end the AIDS pandemic by 2020. Nonetheless, the rate of immunological recovery in the study cohort was low. Hence, early initiation of HAART should be strengthened to achieve good virological suppression and immunological recovery.. Dust in Egyptian laying hen houses contains high concentrations of microorganisms and endotoxins, which might impair the health of birds and farmers when inhaled. Furthermore, laying hens in Egypt seem to be a reservoir for ESBL-producing Enterobacteriaceae. Thus, farmers are at risk of exposure to ESBL-producing bacteria, and colonized hens might transmit these bacteria into the food chain.. The lack of significant differences in the absolute changes and relative ratios of injury and repair biomarkers by contrast-associated AKI status suggests that the majority of mild contrast-associated AKI cases may be driven by hemodynamic changes at the kidney.. Most comparisons for different outcomes are based on very few studies, mostly low-powered, with an overall low CoE. Thus, the available evidence is considered insufficient to either support or refute CH effectiveness or to recommend one ICM over another. Therefore, further well-designed, larger RCTs are required.. PROSPERO database Identifier: CRD42016041953.. Untouched root canal at cross-section perimeter, the Hero 642 system showed 41.44% ± 5.62% and Reciproc R40 58.67% ± 12.39% without contact with instruments. Regarding the untouched area, Hero 642 system showed 22.78% ± 6.42% and Reciproc R40 34.35% ± 8.52%. Neither instrument achieved complete cross-sectional root canal debridement. Hero 642 system rotary taper 0.02 instruments achieved significant greater wall contact perimeter and area compared to reciprocate the Reciproc R40 taper 0.06 instrument.. Hero 642 achieved higher wall contact perimeter and area but, regardless of instrument size and taper, vital pulp during. The functional properties of the main mechanisms involved in the control of muscle Ca. This study showed that the anti-inflammatory effect of the iron-responsive product DHA in arthritis can be monitored by an iron-like radioactive tracer (. Attenuated vascular reactivity during pregnancy suggests that the systemic vasodilatory state partially depletes nitric oxide bioavailability. Preliminary data support the potential for MRI to identify vascular dysfunction in vivo that underlies PE. Level of Evidence 2 Technical Efficacy Stage 1 J. MAGN. RESON. IMAGING 2021;53:447-455.. La evaluación de riesgo es importante para predecir los resultados postoperatorios en pacientes con cáncer gastroesofágico. Este estudio de cohortes tuvo como objetivo evaluar los cambios en la composición corporal durante la quimioterapia neoadyuvante e investigar su asociación con complicaciones postoperatorias. MÉTODOS: Los pacientes consecutivos con cáncer gastroesofágico sometidos a quimioterapia neoadyuvante y cirugía con intención curativa entre 2016 y 2019, identificados a partir de una base de datos específica, se incluyeron en el estudio. Se utilizaron las imágenes de tomografía computarizada, antes y después de la quimioterapia neoadyuvante, para evaluar el índice de masa muscular esquelética, la sarcopenia y el índice de grasa visceral y subcutánea.. In this in vitro premature infant lung model, HF oscillation of BCPAP was associated with improved CO. Our results showed that HPC significantly promotes neurogenesis after MCAO and ameliorates neuronal injury.. Inflammatory markers are highly related to signs of systemic hypoperfusion in CS. Moreover, high PCT and IL-6 levels are associated with poor prognosis.. These findings indicate that Tetrapleura tetraptera fruit has a protective potential against stroke through modulation of redox and electrolyte imbalances, and attenuation of neurotransmitter dysregulation and other neurochemical dysfunctions. Tetrapleura tetraptera fruit could be a promising source for the discovery of bioactives for stroke therapy.

Topics: 3T3-L1 Cells; A Kinase Anchor Proteins; Acetates; Achilles Tendon; Acute Kidney Injury; Acute Pain; Acyclic Monoterpenes; Adenine Nucleotides; Adhesins, Escherichia coli; Adipocytes; Adipocytes, Brown; Adipogenesis; Administration, Inhalation; Administration, Oral; Adrenal Cortex Hormones; Adsorption; Adult; Aeromonas hydrophila; Africa; Aged; Aged, 80 and over; Agrobacterium tumefaciens; Air; Air Pollutants; Air Pollution; Air Pollution, Indoor; Algorithms; Alkaloids; Alkynes; Allosteric Regulation; Amines; Amino Acid Sequence; Amino Acids; Amino Acids, Branched-Chain; Aminoisobutyric Acids; Aminopyridines; Amyotrophic Lateral Sclerosis; Anaerobic Threshold; Angiography; Angiotensin II Type 1 Receptor Blockers; Angiotensin Receptor Antagonists; Angiotensin-Converting Enzyme Inhibitors; Animal Distribution; Animal Feed; Animal Nutritional Physiological Phenomena; Animals; Ankle Joint; Anti-Bacterial Agents; Anti-HIV Agents; Anti-Inflammatory Agents; Antibodies, Bacterial; Antifungal Agents; Antimalarials; Antineoplastic Agents; Antineoplastic Agents, Phytogenic; Antioxidants; Antiretroviral Therapy, Highly Active; Antiviral Agents; Aotidae; Apelin; Apoptosis; Arabidopsis Proteins; Argentina; Arginine; Artemisinins; Arthritis, Experimental; Arthritis, Rheumatoid; Arthroscopy; Aspergillus; Aspergillus niger; Asteraceae; Asthma; ATP Binding Cassette Transporter, Subfamily B, Member 1; ATP Binding Cassette Transporter, Subfamily G, Member 2; Auditory Cortex; Autoantibodies; Autophagy; Bacteria; Bacterial Infections; Bacterial Proteins; Bacterial Typing Techniques; Base Composition; Base Sequence; Basketball; Beclin-1; Benzhydryl Compounds; Benzimidazoles; Benzo(a)pyrene; Benzofurans; Benzoxazines; Bereavement; beta Catenin; beta-Lactamase Inhibitors; beta-Lactamases; beta-Lactams; Betacoronavirus; Betaine; Binding Sites; Biofilms; Biological Assay; Biological Availability; Biological Evolution; Biomarkers; Biomechanical Phenomena; Biopolymers; Biopsy; Bismuth; Blood Glucose; Blood Platelets; Blood Pressure; Body Composition; Body Weight; Bone Marrow; Bone Marrow Cells; Bone Regeneration; Boron; Botrytis; Brain Ischemia; Brain Neoplasms; Brain-Derived Neurotrophic Factor; Brazil; Breast Neoplasms; Breath Tests; Bronchoalveolar Lavage Fluid; Burkholderia; C-Reactive Protein; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Calcification, Physiologic; Calcium; Calcium Signaling; Calorimetry, Differential Scanning; Cameroon; Camptothecin; Candida; Candida albicans; Capillaries; Carbapenem-Resistant Enterobacteriaceae; Carbapenems; Carbohydrate Conformation; Carbon; Carbon Dioxide; Carbon Isotopes; Carcinoma, Ovarian Epithelial; Cardiac Output; Cardiomyopathy, Hypertrophic; Cardiotonic Agents; Cardiovascular Diseases; Caregivers; Carps; Case-Control Studies; Catalase; Catalysis; Cats; CD4 Lymphocyte Count; Cell Culture Techniques; Cell Differentiation; Cell Line, Tumor; Cell Membrane; Cell Movement; Cell Proliferation; Cell Survival; Cells, Cultured; Cellulose; Centrosome; Ceratopogonidae; Chickens; Child; China; Cholera Toxin; Choline; Cholinesterases; Chromatography, High Pressure Liquid; Chromatography, Liquid; Chromatography, Micellar Electrokinetic Capillary; Chromatography, Reverse-Phase; Chronic Disease; Cinnamates; Cities; Citrates; Climate Change; Clinical Trials, Phase III as Topic; Coal; Coal Mining; Cohort Studies; Coinfection; Colchicine; Colony Count, Microbial; Colorectal Neoplasms; Coloring Agents; Common Cold; Complement Factor H; Computational Biology; Computer Simulation; Continuous Positive Airway Pressure; Contrast Media; Coordination Complexes; Coronary Artery Bypass; Coronavirus 3C Proteases; Coronavirus Infections; Coronavirus Protease Inhibitors; Corynebacterium glutamicum; Cosmetics; COVID-19; Creatinine; Cross-Sectional Studies; Crotonates; Crystallography, X-Ray; Cues; Culicidae; Culture Media; Curcuma; Cyclopentanes; Cyclopropanes; Cymbopogon; Cystine; Cytochrome P-450 CYP2B6; Cytochrome P-450 CYP2C19; Cytochrome P-450 CYP2C19 Inhibitors; 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Neurodegeneration induced by excitotoxicity is a common feature in various neurological disorders. This pathological condition is caused by prolonged stimulation of glutamate receptor subtypes, followed by both intracellular Ca2+ overload and activation of specific genes, resulting in synthesis of enzymes involved in cell stress response. Using experimental in vitro models of excitotoxicity, we demonstrated that glutamate exposure up-regulated tissue transglutaminase in primary cultures of both cerebellar granule cells and astrocytes. These changes were consequent to receptor-mediated Ca2+ influx, as demonstrated by the inhibition with selective antagonists, MK-801 and GYKI 52466. Early increases in different transglutaminase isoforms were also observed in global cerebral ischemia, which closely resembles neuronal damage caused by NMDA receptor activation. These findings agree with a postulated role for transglutaminases in molecular mechanisms of several neurodegenerative diseases. Indeed, increased cross-linking reactions could be of pathologic relevance, as part of biochemical changes observed in neurological disorders.

Topics: Animals; Benzodiazepines; Brain Ischemia; Calcium; Cells, Cultured; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Humans; Nerve Degeneration; Neurotoxins; Receptors, N-Methyl-D-Aspartate; Transglutaminases

Magnesium ion blocks the ion channel of the NMDA receptor at a stable condition. The ion channel competes with the binding site of the noncompetitive antagonists phencyclidine (PCP) and MK-801, which prevent a brain impairment due to the ischemia and so on. The binding ability of these antagonists is strong, an exchange with the magnesium ion is not easy, then the side effect of the schizophrenia-like behavior is caused. Recently, memantine can be used as a therapeutic drug of the moderate-to-severe Alzheimer's disease. Memantine is the noncompetitive antagonist, too, then those development details and a difference from MK-801 were explained.

Topics: Alzheimer Disease; Binding Sites; Brain Ischemia; Dizocilpine Maleate; Humans; Magnesium; Memantine; Phencyclidine; Receptors, N-Methyl-D-Aspartate; Schizophrenia

Cell-permeant Ca2+ chelators such as 1,2-bis-(2-aminophenoxy)ethane- N,N,N',N'-tetraacetic acid acetoxymethyl ester (BAPTA-AM) have been reported to protect neurons in experimental focal cerebral ischemia. However, their in vivo actions are uncertain, and their protective efficacy is proven only in brief cerebral ischemia paradigms. Here we examine their mechanism of action in vitro and duration of efficacy in vivo. Electrophysiological studies were made in CA1 neurons in rat hippocampal slices. When superfused with BAPTA-AM (30-50 microM), CA1 somatic field potential recordings showed attenuation of the population spike amplitude, and intracellular recordings showed reduced excitatory postsynaptic potentials, indicating inhibition of excitatory synaptic transmission. Also, Ca(2+)-dependent accommodation and post-spike-train hyperpolarizations were reduced, indicating Ca2+ chelation hear the internal cell membrane surface. To determine whether Ca2+ chelators reduce the size of cerebral infarction rather than simply delaying its evolution, we studied the effects of BAPTA-AM treatment on infarction size 24 h after permanent middle cerebral artery occlusion. Fischer rats (n = 8 per group) were pretreated with saline, BAPTA-AM (20 mg/kg), or MK-801 (0.5 mg/kg). Infarction volumes in animals treated with BAPTA-AM were reduced by 50.5% compared with controls (p = 0.018), whereas animals treated with MK-801 experienced a statistically insignificant infarct volume reduction (26%; p = 0.27). These data show a persistence of neuroprotection by the Ca2+ chelator at 24 h and indicate that it may act by attenuating synaptic transmission and subplasma membrane Ca2+ excess.

Topics: Animals; Brain; Brain Ischemia; Calcium; Cell Membrane Permeability; Cerebral Infarction; Chelating Agents; Dizocilpine Maleate; Egtazic Acid; Electrophysiology; In Vitro Techniques; Male; Neurons; Neuroprotective Agents; Rats; Rats, Wistar; Synaptic Transmission; Time Factors

Topics: Animals; Brain Ischemia; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Humans; Pipecolic Acids; Piperidines; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate

EAA neurotransmitters participate in a variety of physiologic processes during central nervous system development. EAAs function as neurotransmitters but also regulate development of neuronal cytoarchitecture and neuronal connectivity. EAAs play a major role in several forms of activity-dependent synaptic plasticity including learning and memory, and stabilization and elimination of synaptic connections during development. Similar molecular mechanisms may underlie plastic changes during development and neuronal destruction by overaction of EAA receptors. A critical level of EAA neurotransmitter activity is required for normal development. However, there appears to be a continuum of physiologic reactions to EAA receptor activation: underactivation can retard or disrupt normal development, whereas overactivation can lead to neuronal injury and destruction (figure 8). The susceptibility of the brain to excitotoxicity is dramatically altered during postnatal development. As a result of these changes, the contribution of NMDA receptor activation in excitotoxic brain injury may be greater during early periods of postnatal development, whereas non-NMDA receptors may make a greater contribution to excitotoxic injury in the adult brain.

Topics: Animals; Brain; Brain Ischemia; Dizocilpine Maleate; Humans; Kainic Acid; N-Methylaspartate; Quisqualic Acid; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate

The hsp70 gene is induced by denatured protein in injured cells and is an extremely sensitive and reliable marker of cells injured by ischemia, seizures, and toxins. Normal brains have little detectable hsp70 mRNA or HSP70 protein. After status epilepticus produced by systemic injections of kainic acid, however, HSP70 protein is induced in neurons but not glia in brain regions known to be injured by kainic acid. Global and focal ischemia also induce the hsp70 gene in brain. The induction of HSP70 protein in hippocampus following increasing durations of global ischemia correlates with the regional and cellular vulnerability to ischemia: CA1 neurons express HSP70 after the briefest periods of ischemia followed by CA4, CA3, dentate granule neurons, glia, and lastly, endothelial cells. Moreover, as the severity of ischemia worsens, a transcriptional and/or translational blockade of the hsp70 gene occurs in the same order so that moderate degrees of ischemia induce HSP70 in CA3 neurons and dentate granule neurons but not necrotic CA1 neurons, and severe ischemia induces HSP70 in capillary endothelial cells of hippocampus but not in any infarcted neurons or glia throughout the hippocampus. Brief periods of focal ischemia induce HSP70 primarily in neurons, suggesting that even focal ischemia can produce selective neuronal injury without infarction. In some instances, HSP70 immunoreactive astrocytes surround the HSP70 immunostained neurons. Focal ischemia that produces infarction induces HSP70 primarily in endothelial cells of cerebral blood vessels in the regions of infarction and in neurons and astrocytes on the perimeter or the penumbral area of infarction.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Brain Ischemia; Dizocilpine Maleate; Gene Expression Regulation; Heat-Shock Proteins; Humans; In Situ Hybridization; Ketamine; Phencyclidine; RNA, Messenger; Stress, Physiological

Delayed selective nerve cell death as seen in the hippocampal CA1 area of gerbils after transient forebrain ischemia goes along with neuronal hyperactivity and an early demonstrable accumulation of calcium in circumscribed groups of nerve cells. Application of NMDA receptor-blockers such as MK 801 prevents neuronal damage. This suggests the involvement of NMDA receptors which are operated by glutamate and known to mediate a special Ca2+ influx required also for establishing sustained enhancement of synaptic efficacy. Thus, the excessive postischemic accumulation of calcium, thought to be instrumental in the generation of nerve cell death, seems to result from turning on a dangerous, but primarily physiological mechanism which ran out of control. We studied the endogenous control mechanisms by which the firing pattern of nerve cells and the initiation of NMDA receptor-mediated neuronal Ca2+ influx are controlled focusing in particular on the role of adenosine. This nucleoside is released from nerve- and glial cells in larger amounts after ischemia. It counteracts at increased extracellular concentration the generation of burst discharges, an effect which is ascribed to a modulation of the dendritic membrane properties. Removal of a possible action of endogenous adenosine by receptor antagonists such as theophylline was found to enhance postischemic nerve cell death. This together with other reported experimental evidences points to a protective action of this nucleoside in the brain. The presumed mechanisms by which this effect is achieved were studied in a rat hippocampal slice using ion selective electrodes.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenosine; Animals; Brain Damage, Chronic; Brain Ischemia; Calcium; Culture Techniques; Dizocilpine Maleate; Epilepsy, Post-Traumatic; Gerbillinae; Hippocampus; Receptors, N-Methyl-D-Aspartate; Synaptic Transmission

Topics: Animals; Benzazepines; Body Temperature; Brain; Brain Injuries; Brain Ischemia; Calcium Channel Blockers; Dextromethorphan; Dizocilpine Maleate; Drug Therapy, Combination; Humans; Ischemic Attack, Transient; N-Methylaspartate

Excitotoxic mechanisms due to overactivity of the amino acid neurotransmitters glutamate and aspartate maybe responsible for brain damage after injury. In this review we examine ischaemia and shear injury, which are relevant to human head injury. The opportunities for treatment using glutamate antagonist drugs are discussed.

Topics: Animals; Brain Damage, Chronic; Brain Injuries; Brain Ischemia; Central Nervous System Agents; Dizocilpine Maleate; Drug Evaluation, Preclinical; Excitatory Amino Acid Antagonists; Glutamates; Glutamic Acid; Humans; Receptors, N-Methyl-D-Aspartate; Retrograde Degeneration

Ischemic damage, chiefly of the focal type, and axonal disruption (diffuse axonal injury) are the major factors causing brain damage after human head injury. About one third of this damage may be delayed hours or days after the injury. Evidence from four animal models, each relevant to different aspects of human head injury, has shown that excitatory amino acid-induced changes are responsible for a proportion of the posttraumatic sequelae and that these effects can be blocked by EAA antagonists. This evidence is reviewed, and the implications for the conduct of human trials with EAA antagonists are discussed.

Topics: Animals; Brain Damage, Chronic; Brain Injuries; Brain Ischemia; Central Nervous System Depressants; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Humans; Ischemic Attack, Transient; Spinal Cord Injuries

Unprecedented community containment measures were taken following the recent outbreak of COVID-19 in Italy. The aim of the study was to explore the self-reported future compliance of citizens with such measures and its relationship with potentially impactful psychological variables.. An online survey was completed by 931 people (18-76 years) distributed across the Italian territory. In addition to demographics, five dimensions were measured: self-reported compliance with containment measures over time (today, at 7, 14, 30, 60, 90, and 180 days from now) at three hypothetical risk levels (10, 50, 90% of likelihood of contracting the COVID-19), perceived risk, generalized anxiety, intolerance of uncertainty, and relevance of several psychological needs whose satisfaction is currently precluded.. The duration of containment measures plays a crucial role in tackling the spread of the disease as people will be less compliant over time. Psychological needs of citizens impacting on the compliance should be taken into account when planning an easing of the lockdown, along with interventions for protecting vulnerable groups from mental distress.. La apendicitis aguda (AA) es la urgencia quirúrgica abdominal más frecuente. No encontramos estudios específicos que evalúen el impacto de la pandemia causada por el coronavirus 2 (SARS-Cov-2) sobre la AA y su tratamiento quirúrgico. Analizamos la influencia de esta nueva patología sobre la AA.. Estudio observacional retrospectivo en pacientes intervenidos por AA desde enero hasta abril de 2020. Fueron clasificados según el momento de la apendicectomía, antes de la declaración del estado de alarma (Pre-COVID19) y después de la declaración del estado de alarma (Post-COVID19) en España. Se evaluaron variables demográficas, duración de la sintomatología, tipo de apendicitis, tiempo quirúrgico, estancia hospitalaria y complicaciones postoperatorias.. La pandemia por SARS-Cov-2 influye en el momento de diagnóstico de la apendicitis, así como en su grado de evolución y estancia hospitalaria. La peritonitis fue lo más frecuentemente observado. Una sospecha y orientación clínica más temprana, es necesaria para evitar un manejo inadecuado de este trastorno quirúrgico común.. The primary outcome is improvement in PaO. Findings will provide timely information on the safety, efficacy, and optimal dosing of t-PA to treat moderate/severe COVID-19-induced ARDS, which can be rapidly adapted to a phase III trial (NCT04357730; FDA IND 149634).. None.. The gut barrier is crucial in cirrhosis in preventing infection-causing bacteria that normally live in the gut from accessing the liver and other organs via the bloodstream. Herein, we characterised gut inflammation by measuring different markers in stool samples from patients at different stages of cirrhosis and comparing this to healthy people. These markers, when compared with equivalent markers usually measured in blood, were found to be very different in pattern and absolute levels, suggesting that there is significant gut inflammation in cirrhosis related to different immune system pathways to that seen outside of the gut. This provides new insights into gut-specific immune disturbances that predispose to complications of cirrhosis, and emphasises that a better understanding of the gut-liver axis is necessary to develop better targeted therapies.. La surveillance de l’intervalle QT a suscité beaucoup d’intérêt durant la pandémie de la COVID-19 en raison de l’utilisation de médicaments prolongeant l’intervalle QT et les préoccupations quant à la transmission virale par les électrocardiogrammes (ECG) en série. Nous avons posé l’hypothèse que la surveillance en continu de l’intervalle QT par télémétrie était associée à une meilleure détection des épisodes de prolongation de l’intervalle QT.. Nous avons introduit la télémétrie cardiaque en continu (TCC) à l’aide d’un algorithme de surveillance automatisée de l’intervalle QT dans nos unités de COVID-19. Les mesures automatisées quotidiennes de l’intervalle QT corrigé (auto-QTc) en fonction de la fréquence cardiaque maximale ont été enregistrées. Nous avons comparé la proportion des épisodes de prolongation marquée de l’intervalle QTc (QTc long), définie par un intervalle QTc ≥ 500 ms, chez les patients montrant une suspicion de COVID-19 ou ayant la COVID-19 qui avaient été admis avant et après la mise en place de la TCC (groupe témoin. La surveillance en continu de l’intervalle QT est supérieure à la norme de soins dans la détection des épisodes de QTc long et exige peu d’ECG. La réponse clinique aux épisodes de QTc long est sous-optimale.. Exposure to a model wildfire air pollution source modifies cardiovascular responses to HC challenge, suggesting air pollution sensitizes the body to systemic triggers.. Though the majority of HIV-infected adults who were on HAART had shown viral suppression, the rate of suppression was sub-optimal according to the UNAIDS 90-90-90 target to help end the AIDS pandemic by 2020. Nonetheless, the rate of immunological recovery in the study cohort was low. Hence, early initiation of HAART should be strengthened to achieve good virological suppression and immunological recovery.. Dust in Egyptian laying hen houses contains high concentrations of microorganisms and endotoxins, which might impair the health of birds and farmers when inhaled. Furthermore, laying hens in Egypt seem to be a reservoir for ESBL-producing Enterobacteriaceae. Thus, farmers are at risk of exposure to ESBL-producing bacteria, and colonized hens might transmit these bacteria into the food chain.. The lack of significant differences in the absolute changes and relative ratios of injury and repair biomarkers by contrast-associated AKI status suggests that the majority of mild contrast-associated AKI cases may be driven by hemodynamic changes at the kidney.. Most comparisons for different outcomes are based on very few studies, mostly low-powered, with an overall low CoE. Thus, the available evidence is considered insufficient to either support or refute CH effectiveness or to recommend one ICM over another. Therefore, further well-designed, larger RCTs are required.. PROSPERO database Identifier: CRD42016041953.. Untouched root canal at cross-section perimeter, the Hero 642 system showed 41.44% ± 5.62% and Reciproc R40 58.67% ± 12.39% without contact with instruments. Regarding the untouched area, Hero 642 system showed 22.78% ± 6.42% and Reciproc R40 34.35% ± 8.52%. Neither instrument achieved complete cross-sectional root canal debridement. Hero 642 system rotary taper 0.02 instruments achieved significant greater wall contact perimeter and area compared to reciprocate the Reciproc R40 taper 0.06 instrument.. Hero 642 achieved higher wall contact perimeter and area but, regardless of instrument size and taper, vital pulp during. The functional properties of the main mechanisms involved in the control of muscle Ca. This study showed that the anti-inflammatory effect of the iron-responsive product DHA in arthritis can be monitored by an iron-like radioactive tracer (. Attenuated vascular reactivity during pregnancy suggests that the systemic vasodilatory state partially depletes nitric oxide bioavailability. Preliminary data support the potential for MRI to identify vascular dysfunction in vivo that underlies PE. Level of Evidence 2 Technical Efficacy Stage 1 J. MAGN. RESON. IMAGING 2021;53:447-455.. La evaluación de riesgo es importante para predecir los resultados postoperatorios en pacientes con cáncer gastroesofágico. Este estudio de cohortes tuvo como objetivo evaluar los cambios en la composición corporal durante la quimioterapia neoadyuvante e investigar su asociación con complicaciones postoperatorias. MÉTODOS: Los pacientes consecutivos con cáncer gastroesofágico sometidos a quimioterapia neoadyuvante y cirugía con intención curativa entre 2016 y 2019, identificados a partir de una base de datos específica, se incluyeron en el estudio. Se utilizaron las imágenes de tomografía computarizada, antes y después de la quimioterapia neoadyuvante, para evaluar el índice de masa muscular esquelética, la sarcopenia y el índice de grasa visceral y subcutánea.. In this in vitro premature infant lung model, HF oscillation of BCPAP was associated with improved CO. Our results showed that HPC significantly promotes neurogenesis after MCAO and ameliorates neuronal injury.. Inflammatory markers are highly related to signs of systemic hypoperfusion in CS. Moreover, high PCT and IL-6 levels are associated with poor prognosis.. These findings indicate that Tetrapleura tetraptera fruit has a protective potential against stroke through modulation of redox and electrolyte imbalances, and attenuation of neurotransmitter dysregulation and other neurochemical dysfunctions. Tetrapleura tetraptera fruit could be a promising source for the discovery of bioactives for stroke therapy.

Topics: 3T3-L1 Cells; A Kinase Anchor Proteins; Acetates; Achilles Tendon; Acute Kidney Injury; Acute Pain; Acyclic Monoterpenes; Adenine Nucleotides; Adhesins, Escherichia coli; Adipocytes; Adipocytes, Brown; Adipogenesis; Administration, Inhalation; Administration, Oral; Adrenal Cortex Hormones; Adsorption; Adult; Aeromonas hydrophila; Africa; Aged; Aged, 80 and over; Agrobacterium tumefaciens; Air; Air Pollutants; Air Pollution; Air Pollution, Indoor; Algorithms; Alkaloids; Alkynes; Allosteric Regulation; Amines; Amino Acid Sequence; Amino Acids; Amino Acids, Branched-Chain; Aminoisobutyric Acids; Aminopyridines; Amyotrophic Lateral Sclerosis; Anaerobic Threshold; Angiography; Angiotensin II Type 1 Receptor Blockers; Angiotensin Receptor Antagonists; Angiotensin-Converting Enzyme Inhibitors; Animal Distribution; Animal Feed; Animal Nutritional Physiological Phenomena; Animals; Ankle Joint; Anti-Bacterial Agents; Anti-HIV Agents; Anti-Inflammatory Agents; Antibodies, Bacterial; Antifungal Agents; Antimalarials; Antineoplastic Agents; Antineoplastic Agents, Phytogenic; Antioxidants; Antiretroviral Therapy, Highly Active; Antiviral Agents; Aotidae; Apelin; Apoptosis; Arabidopsis Proteins; Argentina; Arginine; Artemisinins; Arthritis, Experimental; Arthritis, Rheumatoid; Arthroscopy; Aspergillus; Aspergillus niger; Asteraceae; Asthma; ATP Binding Cassette Transporter, Subfamily B, Member 1; ATP Binding Cassette Transporter, Subfamily G, Member 2; Auditory Cortex; Autoantibodies; Autophagy; Bacteria; Bacterial Infections; Bacterial Proteins; Bacterial Typing Techniques; Base Composition; Base Sequence; Basketball; Beclin-1; Benzhydryl Compounds; Benzimidazoles; Benzo(a)pyrene; Benzofurans; Benzoxazines; Bereavement; beta Catenin; beta-Lactamase Inhibitors; beta-Lactamases; beta-Lactams; Betacoronavirus; Betaine; Binding Sites; Biofilms; Biological Assay; Biological Availability; Biological Evolution; Biomarkers; Biomechanical Phenomena; Biopolymers; Biopsy; Bismuth; Blood Glucose; Blood Platelets; Blood Pressure; Body Composition; Body Weight; Bone Marrow; Bone Marrow Cells; Bone Regeneration; Boron; Botrytis; Brain Ischemia; Brain Neoplasms; Brain-Derived Neurotrophic Factor; Brazil; Breast Neoplasms; Breath Tests; Bronchoalveolar Lavage Fluid; Burkholderia; C-Reactive Protein; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Calcification, Physiologic; Calcium; Calcium Signaling; Calorimetry, Differential Scanning; Cameroon; Camptothecin; Candida; Candida albicans; Capillaries; Carbapenem-Resistant Enterobacteriaceae; Carbapenems; Carbohydrate Conformation; Carbon; Carbon Dioxide; Carbon Isotopes; Carcinoma, Ovarian Epithelial; Cardiac Output; Cardiomyopathy, Hypertrophic; Cardiotonic Agents; Cardiovascular Diseases; Caregivers; Carps; Case-Control Studies; Catalase; Catalysis; Cats; CD4 Lymphocyte Count; Cell Culture Techniques; Cell Differentiation; Cell Line, Tumor; Cell Membrane; Cell Movement; Cell Proliferation; Cell Survival; Cells, Cultured; Cellulose; Centrosome; Ceratopogonidae; Chickens; Child; China; Cholera Toxin; Choline; Cholinesterases; Chromatography, High Pressure Liquid; Chromatography, Liquid; Chromatography, Micellar Electrokinetic Capillary; Chromatography, Reverse-Phase; Chronic Disease; Cinnamates; Cities; Citrates; Climate Change; Clinical Trials, Phase III as Topic; Coal; Coal Mining; Cohort Studies; Coinfection; Colchicine; Colony Count, Microbial; Colorectal Neoplasms; Coloring Agents; Common Cold; Complement Factor H; Computational Biology; Computer Simulation; Continuous Positive Airway Pressure; Contrast Media; Coordination Complexes; Coronary Artery Bypass; Coronavirus 3C Proteases; Coronavirus Infections; Coronavirus Protease Inhibitors; Corynebacterium glutamicum; Cosmetics; COVID-19; Creatinine; Cross-Sectional Studies; Crotonates; Crystallography, X-Ray; Cues; Culicidae; Culture Media; Curcuma; Cyclopentanes; Cyclopropanes; Cymbopogon; Cystine; Cytochrome P-450 CYP2B6; Cytochrome P-450 CYP2C19; Cytochrome P-450 CYP2C19 Inhibitors; Cytokines; Databases, Genetic; Death; Dendritic Cells; Density Functional Theory; Depsides; Diabetes Mellitus, Type 2; Diamond; Diarylheptanoids; Dibenzofurans; Dibenzofurans, Polychlorinated; Diclofenac; Diet; Dietary Carbohydrates; Dietary Supplements; Diffusion Magnetic Resonance Imaging; Dioxins; Diphenylamine; Disease Outbreaks; Disease Susceptibility; Disulfides; Dithiothreitol; Dizocilpine Maleate; DNA Methylation; DNA-Binding Proteins; DNA, Bacterial; Dogs; Dose-Response Relationship, Drug; Double-Blind Method; Doublecortin Protein; Drosophila melanogaster; Droughts; Drug Carriers; Drug Combinations; Drug Delivery Systems; Drug Liberation; Drug Resistance; Drug Resistance, Bacterial; Drug Resistance, Neoplasm; Drug Screening Assays, Antitumor; Dust; Dynactin Complex; Dysferlin; Echo-Planar Imaging; Echocardiography; Edaravone; Egypt; Elasticity; Electrodes; Electrolytes; Emodin; Emtricitabine; Endometriosis; Endothelium, Vascular; Endotoxins; Energy Metabolism; Energy Transfer; Enterobacteriaceae; Enterococcus faecalis; Enterotoxigenic Escherichia coli; Environmental Monitoring; Enzyme Inhibitors; Epidemiologic Factors; Epigenesis, Genetic; Erythrocytes; Escherichia coli; Escherichia coli Infections; Escherichia coli Vaccines; Esophageal Neoplasms; Esophagectomy; Esophagogastric Junction; Esterases; Esterification; Ethanol; Ethiopia; Ethnicity; Eucalyptus; Evidence-Based Practice; Exercise; Exercise Tolerance; Extracorporeal Membrane Oxygenation; Family; Fatty Acids; Feedback; Female; Ferric Compounds; Fibrin Fibrinogen Degradation Products; Filtration; Fish Diseases; Flavonoids; Flavonols; Fluorodeoxyglucose F18; Follow-Up Studies; Food Microbiology; Food Preservation; Forests; Fossils; Free Radical Scavengers; Freund's Adjuvant; Fruit; Fungi; Gallium; Gender Identity; Gene Expression Regulation; Gene Expression Regulation, Neoplastic; Gene Expression Regulation, Plant; Gene Knockdown Techniques; Genes, Bacterial; Genes, Plant; Genetic Predisposition to Disease; Genitalia; Genotype; Glomerulonephritis, IGA; Glottis; Glucocorticoids; Glucose; Glucuronides; Glutathione Transferase; Glycogen Synthase Kinase 3 beta; Gram-Negative Bacterial Infections; Gram-Positive Bacterial Infections; Grassland; Guinea Pigs; Half-Life; Head Kidney; Heart Atria; Heart Rate; Heart Septum; HEK293 Cells; Hematopoietic Stem Cells; Hemodynamics; Hep G2 Cells; Hepacivirus; Hepatitis C; Hepatitis C, Chronic; Hepatocytes; Hesperidin; High-Frequency Ventilation; High-Temperature Requirement A Serine Peptidase 1; Hippocampus; Hirudins; History, 20th Century; History, 21st Century; HIV Infections; Homeostasis; Hominidae; Housing, Animal; Humans; Hydrocarbons, Brominated; Hydrogen Bonding; Hydrogen Peroxide; Hydrogen-Ion Concentration; Hydroxybutyrates; Hydroxyl Radical; Hypertension; Hypothyroidism; Image Interpretation, Computer-Assisted; Immunoconjugates; Immunogenic Cell Death; Indoles; Infant, Newborn; Infant, Premature; Infarction, Middle Cerebral Artery; Inflammation; Inflammation Mediators; Infrared Rays; Inhibitory Concentration 50; Injections, Intravenous; Interferon-gamma; Interleukin-23; Interleukin-4; Interleukin-6; Intermediate Filaments; Intermittent Claudication; Intestine, Small; Iridoid Glucosides; Iridoids; Iron; Isomerism; Isotope Labeling; Isoxazoles; Itraconazole; Kelch-Like ECH-Associated Protein 1; Ketoprofen; Kidney Failure, Chronic; Kinetics; Klebsiella pneumoniae; Lactams, Macrocyclic; Lactobacillus; Lactulose; Lakes; Lamivudine; Laparoscopy; Laparotomy; Laryngoscopy; Leucine; Limit of Detection; Linear Models; Lipid A; Lipopolysaccharides; Listeria monocytogenes; Liver; Liver Cirrhosis; Logistic Models; Longitudinal Studies; Losartan; Low Back Pain; Lung; Lupinus; Lupus Erythematosus, Systemic; Machine Learning; Macular Degeneration; Madin Darby Canine Kidney Cells; Magnetic Phenomena; Magnetic Resonance Imaging; Magnetic Resonance Spectroscopy; Magnetics; Malaria, Falciparum; Male; Mannans; MAP Kinase Signaling System; Mass Spectrometry; Melatonin; Membrane Glycoproteins; Membrane Proteins; Meniscectomy; Menisci, Tibial; Mephenytoin; Mesenchymal Stem Cells; Metal Nanoparticles; Metal-Organic Frameworks; Methionine; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Nude; Mice, Obese; Mice, Transgenic; Microbial Sensitivity Tests; Microcirculation; MicroRNAs; Microscopy, Video; Microtubules; Microvascular Density; Microwaves; Middle Aged; Minimally Invasive Surgical Procedures; Models, Animal; Models, Biological; Models, Molecular; Models, Theoretical; Molecular Docking Simulation; Molecular Structure; Molecular Weight; Morus; Mouth Floor; Multicenter Studies as Topic; Multiple Sclerosis; Multiple Sclerosis, Relapsing-Remitting; Muscle, Skeletal; Myocardial Ischemia; Myocardium; NAD; NADP; Nanocomposites; Nanoparticles; Naphthols; Nasal Lavage Fluid; Nasal Mucosa; Neisseria meningitidis; Neoadjuvant Therapy; Neoplasm Invasiveness; Neoplasm Recurrence, Local; Neoplasms, Experimental; Neural Stem Cells; Neuroblastoma; Neurofilament Proteins; Neurogenesis; Neurons; New York; NF-E2-Related Factor 2; NF-kappa B; Nicotine; Nitriles; Nitrogen; Nitrogen Fixation; North America; Observer Variation; Occupational Exposure; Ochrobactrum; Oils, Volatile; Olea; Oligosaccharides; Omeprazole; Open Field Test; Optimism; Oregon; Oryzias; Osmolar Concentration; Osteoarthritis; Osteoblasts; Osteogenesis; Ovarian Neoplasms; Ovariectomy; Oxadiazoles; Oxidation-Reduction; Oxidative Stress; Oxygen; Ozone; p38 Mitogen-Activated Protein Kinases; Pakistan; Pandemics; Particle Size; Particulate Matter; Patient-Centered Care; Pelargonium; Peptides; Perception; Peripheral Arterial Disease; Peroxides; Pets; Pharmaceutical Preparations; Pharmacogenetics; Phenobarbital; Phenols; Phenotype; Phosphates; Phosphatidylethanolamines; Phosphines; Phospholipids; Phosphorus; Phosphorylation; Photoacoustic Techniques; Photochemotherapy; Photosensitizing Agents; Phylogeny; Phytoestrogens; Pilot Projects; Plant Components, Aerial; Plant Extracts; Plant Immunity; Plant Leaves; Plant Oils; Plants, Medicinal; Plasmodium berghei; Plasmodium falciparum; Platelet Activation; Platelet Function Tests; Pneumonia, Viral; Poaceae; Pogostemon; Poloxamer; Poly I; Poly(ADP-ribose) Polymerase Inhibitors; Polychlorinated Biphenyls; Polychlorinated Dibenzodioxins; Polycyclic Compounds; Polyethylene Glycols; Polylysine; Polymorphism, Genetic; Polymorphism, Single Nucleotide; Population Dynamics; Portasystemic Shunt, Transjugular Intrahepatic; Positron Emission Tomography Computed Tomography; Postoperative Complications; Postprandial Period; Potassium Cyanide; Predictive Value of Tests; Prefrontal Cortex; Pregnancy; Prepulse Inhibition; Prevalence; Procalcitonin; Prodrugs; Prognosis; Progression-Free Survival; Proline; Proof of Concept Study; Prospective Studies; Protein Binding; Protein Conformation; Protein Domains; Protein Folding; Protein Multimerization; Protein Sorting Signals; Protein Structure, Secondary; Proton Pump Inhibitors; Protozoan Proteins; Psychometrics; Pulse Wave Analysis; Pyridines; Pyrrolidines; Quality of Life; Quantum Dots; Quinoxalines; Quorum Sensing; Radiopharmaceuticals; Rain; Random Allocation; Randomized Controlled Trials as Topic; Rats; Rats, Sprague-Dawley; Rats, Wistar; RAW 264.7 Cells; Reactive Oxygen Species; Receptor, Angiotensin, Type 1; Receptor, PAR-1; Receptors, CXCR4; Receptors, Estrogen; Receptors, Glucocorticoid; Receptors, Interleukin-1; Receptors, Interleukin-17; Receptors, Notch; Recombinant Fusion Proteins; Recombinant Proteins; Reducing Agents; Reflex, Startle; Regional Blood Flow; Regression Analysis; Reperfusion Injury; Reproducibility of Results; Republic of Korea; Respiratory Tract Diseases; Retrospective Studies; Reverse Transcriptase Inhibitors; Rhinitis, Allergic; Risk Assessment; Risk Factors; Rituximab; RNA, Messenger; RNA, Ribosomal, 16S; ROC Curve; Rosmarinic Acid; Running; Ruthenium; Rutin; Sarcolemma; Sarcoma; Sarcopenia; Sarcoplasmic Reticulum; SARS-CoV-2; Scavenger Receptors, Class A; Schools; Seasons; Seeds; Sequence Analysis, DNA; Severity of Illness Index; Sex Factors; Shock, Cardiogenic; Short Chain Dehydrogenase-Reductases; Signal Transduction; Silver; Singlet Oxygen; Sinusitis; Skin; Skin Absorption; Small Molecule Libraries; Smoke; Socioeconomic Factors; Soil; Soil Microbiology; Solid Phase Extraction; Solubility; Solvents; Spain; Spectrometry, Mass, Electrospray Ionization; Spectroscopy, Fourier Transform Infrared; Speech; Speech Perception; Spindle Poles; Spleen; Sporothrix; Staphylococcal Infections; Staphylococcus aureus; Stereoisomerism; Stomach Neoplasms; Stress, Physiological; Stroke Volume; Structure-Activity Relationship; Substrate Specificity; Sulfonamides; Surface Properties; Surface-Active Agents; Surveys and Questionnaires; Survival Rate; T-Lymphocytes, Cytotoxic; Tandem Mass Spectrometry; Temperature; Tenofovir; Terpenes; Tetracycline; Tetrapleura; Textiles; Thermodynamics; Thiobarbituric Acid Reactive Substances; Thrombin; Thyroid Hormones; Thyroid Neoplasms; Tibial Meniscus Injuries; Time Factors; Tissue Distribution; Titanium; Toluidines; Tomography, X-Ray Computed; Tooth; Tramadol; Transcription Factor AP-1; Transcription, Genetic; Transfection; Transgender Persons; Translations; Treatment Outcome; Triglycerides; Ubiquinone; Ubiquitin-Specific Proteases; United Kingdom; United States; Up-Regulation; Vascular Stiffness; Veins; Ventricular Remodeling; Viral Load; Virulence Factors; Virus Replication; Vitis; Voice; Voice Quality; Wastewater; Water; Water Pollutants, Chemical; Water-Electrolyte Balance; Weather; Wildfires; Wnt Signaling Pathway; Wound Healing; X-Ray Diffraction; Xenograft Model Antitumor Assays; Young Adult; Zoogloea

We previously generated an ischemic stroke in a zebrafish model using N

Topics: Animals; Brain; Brain Ischemia; Cell Death; Disease Models, Animal; Dizocilpine Maleate; Drug Evaluation, Preclinical; Edaravone; Free Radical Scavengers; Gases; Hypoxia; Larva; Neurons; Nitrogen; Receptors, N-Methyl-D-Aspartate; Zebrafish

Ischemic stroke accounts for 70-80% of stroke cases worldwide and survivors are frequently left with compromising sensorimotor deficits localized to one or more body regions. Most animal models of stroke involve transient or permanent occlusion of one or more major vessels such as the middle cerebral artery and are characterized by widespread damage to cortical and subcortical structures that result in deficits that can confound studies of neuroprotection and neurorehabilitation. Localized microinjections of the vasoconstricting peptide endothelin-1 (ET-1) into specific brain regions are becoming increasingly popular for such studies, but the pharmacology of endothelin-induced ischemic damage is poorly understood. To test the hypothesis that NMDA receptors, and particularly those containing the NR2B subunit, are involved in ET-1-mediated excitotoxicity and functional impairment, male CD1 rats (N = 32) were pre-treated with either the non-competitive NMDA antagonist MK-801 or the NR2B-selective antagonist Ro25-6981 (or vehicle) prior to unilateral microinjections of endothelin-1 into the somatosensory cortex and striatum. Rats were then tested using 4 established tests of sensory and/or motor function over 14 days. Lesion volumes were quantified post-mortem using standard histology and image analysis. Results confirmed reproducible lesions and significant deficits in all tests in vehicle-treated rats that were significantly reduced in both drug groups but were not different between drugs, providing evidence that endothelin-induced ischemic damage is mediated almost exclusively by NR2B-containing NMDA receptors.

Topics: Animals; Brain Ischemia; Dizocilpine Maleate; Endothelin-1; Excitatory Amino Acid Antagonists; Forelimb; Male; Microinjections; Phenols; Piperidines; Psychomotor Performance; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

S-nitrosylation, the nitric oxide-derived post-translational modification of proteins, plays critical roles in various physiological and pathological functions. In this present study, a rat model of cerebral ischemia and reperfusion by four-vessel occlusion was generated to assess MKK4 S-nitrosylation. Immunoprecipitation and immunoblotting were performed to evaluate MKK4 S-nitrosylation and phosphorylation. Neuronal loss was observed using histological detection. These results indicated that endogenous NO promoted the S-nitrosylation of MKK4. However, application of the exogenous NO donor S-nitrosoglutathione (GNSO), an inhibitor of the neuronal nitric oxide synthase 7-nitroindazole (7-NI), and the N-methyl-d-aspartate receptor (NMDAR) antagonist MK801 diminished I/R-induced S-nitrosylation and phosphorylation. These compounds also markedly decreased cerebral I/R-induced degeneration and death of neurons in hippocampal CA1 region in rats. Taken together, we demonstrated for the first time, that cerebral ischemia/reperfusion can induce S-nitrosylation of MKK4. We also found that inhibiting S-nitrosylation and activation of MKK4 resulted in marked decreases in neuronal degeneration and apoptosis, potentially via NMDAR-mediated mechanisms. These findings may lead to a new field of inquiry to investigate the underlying pathogenesis of stoke and the development of novel treatment strategies.

Topics: Animals; Brain Ischemia; CA1 Region, Hippocampal; Caspase 3; Disease Models, Animal; Dizocilpine Maleate; Indazoles; Male; MAP Kinase Kinase 4; Neurons; Phosphorylation; Rats; Rats, Sprague-Dawley; Reperfusion; S-Nitrosoglutathione; Time Factors

Cortical spreading depolarization is a metabolically costly phenomenon that affects the brain in both health and disease. Following severe stroke, subarachnoid hemorrhage, or traumatic brain injury, cortical spreading depolarization exacerbates tissue damage and enlarges infarct volumes. It is not known, however, whether spreading depolarization also occurs in the retina in vivo. We report now that spreading depolarization episodes are generated in the in vivo rat retina following retinal vessel occlusion produced by photothrombosis. The properties of retinal spreading depolarization are similar to those of cortical spreading depolarization. Retinal spreading depolarization waves propagate at a velocity of 3.0 ± 0.1 mm/min and are associated with a negative shift in direct current potential, a transient cessation of neuronal spiking, arteriole constriction, and a decrease in tissue O2 tension. The frequency of retinal spreading depolarization generation in vivo is reduced by administration of the NMDA antagonist MK-801 and the 5-HT(1D) agonist sumatriptan. Branch retinal vein occlusion is a leading cause of vision loss from vascular disease. Our results suggest that retinal spreading depolarization could contribute to retinal damage in acute retinal ischemia and demonstrate that pharmacological agents can reduce retinal spreading depolarization frequency after retinal vessel occlusion. Blocking retinal spreading depolarization generation may represent a therapeutic strategy for preserving vision in branch retinal vein occlusion patients.

Topics: Animals; Brain Ischemia; Cortical Spreading Depression; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Rats; Retina; Retinal Artery Occlusion; Serotonin 5-HT1 Receptor Agonists; Sumatriptan

Experiments on nonlinear rats subjected to global transient cerebral ischemia revealed the ability of glutamic acid to improve cerebral circulation. Consequently, the excitatory amino acid can produce adverse (neurotoxic) and positive (anti-ischemic) effects in cerebral ischemia. The cerebrovascular effect of glutamic acid in cerebral ischemia is attenuated on the background action of the MNDA receptor blocker MK-801 (0.5 mg/kg intravenously) and eliminated by bicuculline. When glutamic acid is combined with the non-competitive MNDA receptor antagonist MK-801, neither one nor another drug shows its vasodilator effect. The results are indicative of the interaction between excitatory and inhibitory systems on the level of cerebral vessels and once again confirm our previous conclusion about the decisive role of GABA(A) receptors in brain vessels in the implementation of anti-ischemic activity of endogenous compounds (melatonin) and well-known pharmacological substances (mexidol, afobazole), and new chemical compounds based on GABA-containing lipid derivatives.

Topics: Animals; Animals, Outbred Strains; Bicuculline; Brain Ischemia; Carotid Artery, Common; Cerebrovascular Circulation; Coronary Occlusion; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; GABA-A Receptor Antagonists; Glutamic Acid; Male; Neuroprotective Agents; Rats; Receptors, GABA-A; Receptors, N-Methyl-D-Aspartate

Glutamate-mediated excitotoxicity is a major cause of ischemic brain damage. MK-801 confers neuroprotection by attenuating the activation of the N-methyl-d-aspartate (NMDA) receptor, but it failed in clinical use due to the short therapeutic window. Here we aim to investigate the effects of maslinic acid, a natural product from Olea europaea, on the therapeutic time window and dose range for the neuroprotection of MK-801. Rats were administered with maslinic acid intracerebroventricularly and cerebral ischemia was induced by middle cerebral artery occlusion (MCAO) followed by reperfusion. MK-801 was administered at 1 h, 2 h, 3 h and 4 h after ischemia, respectively. The cerebral infarct volume was determined by 2,3,5-Triphenyltetrazolium chloride (TTC) staining, neuronal damage was assessed by Haematoxylin Eosin (H&E) staining, and the expression of glial glutamate transporters and glial fibrillary acidic protein (GFAP) was evaluated by immunohistochemistry and Western blot post-ischemia. Results showed that the presence of maslinic acid extended the therapeutic time window for MK-801 from 1 h to 3 h. Co-treatment of maslinic acid and MK-801 at a subthreshold dosage obviously induced neuroprotection after ischemia. The combination of these two compounds improved the outcome in ischemic rats. Moreover, maslinic acid treatment promoted the expression of GLT-1 and GFAP post-ischemia. These data suggest that the synergistic effect of maslinic acid on neurological protection might be associated with the improvement of glial function, especially with the increased expression of GLT-1. The combination therapy of maslinic acid and MK-801 may prove to be a potential strategy for treating acute ischemic stroke.

Topics: Animals; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Drug Synergism; Drug Therapy, Combination; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Glutamate Plasma Membrane Transport Proteins; Neuroprotective Agents; Olea; Rats; Triterpenes

Ischemic postconditioning (Post C), which involves administration of a brief ischemia after the initial ischemic event, has been demonstrated to be strongly neuroprotective against global cerebral ischemia (GCI) and to improve cognitive outcome. To enhance understanding of the underlying mechanisms, the current study examined the role of NMDA receptors in mediating the beneficial effects of Post C (3 min ischemia) administered 2 days after GCI in adult male rats. The results revealed that Post C was strongly neuroprotective against GCI, and that this effect was blocked by administration of the NMDA receptor antagonist MK-801. Further work revealed that the NR2A-type NMDA receptors mediate the Post C beneficial effects as administration of a NR2A-preferring antagonist (NVP-A) blocked Post C neuroprotection and cognitive enhancement, while administration of a NR2B-preferring antagonist (Ro25) was without effect. Post C significantly up-regulated NR2A levels and phosphorylation of NR2A in the hippocampal CA1 region after Post C. Post C also increased Ca(2+) influx and activation/phosphorylation of CamKIIα at Thr(286), effects that were NR2A mediated as they were blocked by NVP-A. Phosphorylation of ERK and CREB was also increased by Post C, as were two downstream CREB-dependent prosurvival factors, brain derived neurotropic factor (BDNF) and Bcl2, effects that were blocked by the NR2A antagonist, NVP-A. Taken as a whole, the current study provides evidence that NR2A-activation and downstream prosurvival signaling is a critical mediator of Post C-induced neuroprotection and cognitive enhancement following GCI.

Topics: Analysis of Variance; Animals; Brain Ischemia; Calcium; CREB-Binding Protein; Dizocilpine Maleate; Hippocampus; Immunoprecipitation; Male; Maze Learning; Neuroprotective Agents; Phenols; Piperidines; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Signal Transduction; Time Factors

Motor deficits are present in cardiac arrest survivors and injury to cerebellar Purkinje cells (PCs) likely contribute to impairments in motor coordination and post-hypoxic myoclonus. N-Methyl-D-aspartic acid (NMDA) receptor-mediated excitotoxicity is a well-established mechanism of cell death in several brain regions, but the role of NMDA receptors in PC injury remains understudied. Emerging data in cortical and hippocampal neurons indicate that the GluN2A-containing NMDA receptors signal to improve cell survival and GluN2B-containing receptors contribute to neuronal injury. This study compared neuronal injury in the hippocampal CA1 region to that in PCs and investigated the role of NMDA receptors in PC injury in our mouse model of cardiac arrest and cardiopulmonary resuscitation (CA/CPR). Analysis of cell density demonstrated a 24% loss of PCs within 24 h after 8 min CA/CPR and injury stabilized to 33% by 7 days. The subunit promiscuous NMDA receptor antagonist MK-801 protected both CA1 neurons and PCs from ischemic injury following CA/CPR, demonstrating a role for NMDA receptor activation in injury to both brain regions. In contrast, the GluN2B antagonist, Co 101244, had no effect on PC loss while protecting against injury in the CA1 region. These data indicate that ischemic injury to cerebellar PCs progresses via different cell death mechanisms compared to hippocampal CA1 neurons.

Topics: Animals; Brain Ischemia; CA1 Region, Hippocampal; Calbindins; Cardiopulmonary Resuscitation; Cell Death; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Heart Arrest; Male; Mice, Inbred C57BL; Nerve Tissue Proteins; Neurons; Neuroprotective Agents; Piperidines; Purkinje Cells; Receptors, N-Methyl-D-Aspartate; Tissue Culture Techniques

The present study aims to investigate the effect of noscapine (0.5-2.5 μM), an alkaloid from the opium poppy, on primary murine fetal cortical neurons exposed to oxygen-glucose deprivation (OGD), an in vitro model of ischemia.. Cells were transferred to glucose-free DMEM and were exposed to hypoxia in a small anaerobic chamber. Cell viability and nitric oxide production were evaluated by MTT assay and the Griess method, respectively.. The neurotoxicities produced by all three hypoxia durations tested were significantly inhibited by 0.5 μM noscapine. Increasing noscapine concentration up to 2.5 μM produced a concentration-dependent inhibition of neurotoxicity. Pretreatment of cells with MK-801 (10 μM), a non-competitive NMDA antagonist, and nimodipine (10nM), an L-type Ca(2+) channel blockers, increased cell viability after 30 min OGD, while the application of NBQX (30 μM), a selective AMPA-kainate receptor antagonist partially attenuated cell injury. Subsequently, cells treated with noscapine in the presence of thapsigargin (1 μM), an inhibitor of endoplasmic reticulum Ca(2+) ATPases. After 60 min OGD, noscapine could inhibit the cell damage induced by thapsigargin. However, noscapine could not reduce cell damage induced by 240 min OGD in the presence of thapsigargin. Noscapine attenuated nitric oxide (NO) production in cortical neurons after 30 min OGD.. We concluded that noscapine had a neuroprotective effect, which could be due to its interference with multiple targets in the excitotoxicity process. These effects could be mediated partially by a decrease in NO production and the modulation of intracellular calcium levels.

Topics: Animals; Brain Ischemia; Cell Survival; Dizocilpine Maleate; Dose-Response Relationship, Drug; Glucose; Hypoxia; Mice; Neurons; Neuroprotective Agents; NG-Nitroarginine Methyl Ester; Nimodipine; Nitric Oxide; Noscapine; Primary Cell Culture; Quinoxalines; Thapsigargin

Ischemic stroke is the most frequent cause of persistent neurological disability in Western societies. New treatment strategies are required and effective in vivo models are crucial to their development.. The current study establishes a novel in vivo rat model of focal striatal ischemia using the vasoconstrictive agent N5-(1-iminoethyl)-L-ornithine (L-NIO). Adult male Sprague Dawley rats received a unilateral intrastriatal infusion of L-NIO in combination with jugular vein occlusion.. L-NIO infusion was associated with zero mortality, low surgical complexity and a reproducible infarct, providing advantages over established models of focal ischemia. The mean infarct volume of 8.5±5.3% of the volume of the contralateral striatum resulted in blood-brain barrier dysfunction, neuronal hypoxia and ongoing neurodegeneration. Further characteristics of ischemic stroke were exhibited, including robust microglia/macrophage and astroglial responses lasting at least 35 days post-ischemia, in addition to chronic motor function impairment.. When compared to other models such as the MCAo models, the consistency in regions affected, high success rate, zero mortality, reduced surgical complexity and minimal welfare requirements of the L-NIO model make it ideal for initial high-throughput investigations into preclinical efficacy and proof of principle studies of acute ischemic stroke interventions.. We propose that the L-NIO rat model of focal striatal ischemia does not replace the use of other ischemic stroke models. Rather it provides a new, complementary tool for initial preclinical investigations into the treatment of ischemic stroke.

Topics: Animals; Blood-Brain Barrier; Brain Infarction; Brain Ischemia; Corpus Striatum; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Enzyme Inhibitors; Gene Expression Regulation; Jugular Veins; Male; Movement Disorders; Nerve Degeneration; Nerve Tissue Proteins; Neurons; Neuroprotective Agents; Ornithine; Rats; Rats, Sprague-Dawley; Time Factors

Cerebral ischemia in the territory of the middle cerebral artery (MCA) can induce delayed neuronal cell death in the ipsilateral substantia nigra (SN) remote from the primary ischemic lesion. This exofocal postischemic neuronal degeneration (EPND) may worsen stroke outcomes. However, the mechanisms leading to EPND are poorly understood. Here, we studied the time course of EPND via sequential magnetic resonance imaging (MRI) and immunohistochemistry for up to 28 days after 30 minutes' occlusion of the MCA (MCAo) and reperfusion in the mouse. Furthermore, the effects of delayed treatment with FK506 and MK-801 on the development of EPND were investigated. Secondary neuronal degeneration in the SN occurred within the first week after MCAo and was characterized by a marked neuronal cell loss on histology. Sequential neuroimaging examinations revealed transient MRI changes, which were detectable as early as day 4 after MCAo and thus heralding histologic evidence of EPND. Treatment with MK-801, an established anti-excitotoxic agent, conferred protection against EPND even when initiated days after the initial ischemic event, which was not evident with FK506. Our findings define a secondary time window for delayed neuroprotection after stroke, which may provide a promising target for the development of novel therapies.

Topics: Animals; Brain Ischemia; Cell Count; Dizocilpine Maleate; Immunohistochemistry; Immunosuppressive Agents; Infarction, Middle Cerebral Artery; Magnetic Resonance Imaging; Male; Mice; Mice, 129 Strain; Nerve Degeneration; Neuroprotective Agents; Substantia Nigra; Tacrolimus

Cerebral ischemia-reperfusion is associated with NMDA receptor-mediated calcium influx which activates neuronal nitric oxide synthase (nNOS) and consequently induces NO production. NO S-nitrosylates cellular protein and aggravates neuronal injury. Receptor-interacting protein 3 (RIP3) is a sensor molecule regulating cell apoptosis and necrosis. However, the roles of RIP3 in cerebral ischemic injury remain elusive. In this study, we reported that RIP3 could be S-nitrosylated by the exogenous NO donor GSNO in HEK293 cells and the Cys(119) residue was the key nitrosylation site. In addition, we found that cerebral ischemia induced RIP3 S-nitrosylation at different time points of reperfusion, which was coupling with RIP3 phosphorylation (which is associated with its activation) and its interaction with receptor-interacting protein 1 (RIP1), and this process facilitated cerebral ischemic injury. Treatment with NMDA receptor antagonist MK801, or nNOS inhibitor 7NI, diminished RIP3 S-nitrosylation and reduced neuronal damage. Taken together, these data demonstrated that NMDAR-dependent RIP3 S-nitrosylation induced by ischemia facilitated its activation in the early stages of ischemia, blocking this process could reduce the ischemia neuronal injury.

Topics: Aldehyde Oxidoreductases; Animals; Apoptosis; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glucose; HEK293 Cells; Humans; Hypoxia; Male; Neurons; Nitric Oxide Synthase Type I; Phosphorylation; Protein Serine-Threonine Kinases; Rats; Rats, Sprague-Dawley; Receptor-Interacting Protein Serine-Threonine Kinases; Signal Transduction; Time Factors

We explored the relationship between MK-801 concentration and neural stem cell proliferation in rats with focal cerebral ischemia-reperfusion (FCIR). A total of 60 male Sprague Dawley rats were randomized into control (six rats), sham-operation (six rats), operation (12 rats), and MK-801 groups. The MK-801 group comprised 36 rats that were subjected to different doses of MK-801 (0.2, 0.4, 0.6, 0.8, 1.0, and 1.2 mg/kg). Suture occlusion was used to establish an ischemia reperfusion model of middle cerebral artery occlusion (MCAO); 30 min before establishing the FCIR model, the MK-801 group rats were intraperitoneally injected with different doses of MK-801, while the sham-operation and control groups were injected with normal saline. Seven days after model establishment, bromodeoxyuridine-positive cerebral cortex cells adjacent to the focus of infarction were labeled for immunohistochemistry. MK-801 at a concentration of 0.4 mg/kg prevented endogenous neural stem cell proliferation, and this inhibitory effect was strengthened with increasing MK-801 concentration, especially at concentrations greater than 0.8 mg/kg. MK-801 inhibits endogenous neural stem cell proliferation in rats with FCIR, and the inhibitory effect is strengthened with increasing MK-801 concentration.

Topics: Animals; Brain Ischemia; Cell Proliferation; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Male; Neural Stem Cells; Rats; Rats, Sprague-Dawley; Reperfusion Injury

Global cerebral ischemia induces selective acute neuronal injury of the CA1 region of the hippocampus. The type of cell death that ensues may include different programmed cell death mechanisms namely apoptosis and necroptosis, a recently described type of programmed necrosis. We investigated whether necroptosis contributes to hippocampal neuronal death following oxygen-glucose deprivation (OGD), an in vitro model of global ischemia. We observed that OGD induced a death receptor (DR)-dependent component of necroptotic cell death in primary cultures of hippocampal neurons. Additionally, we found that this ischemic challenge upregulated the receptor-interacting protein kinase 3 (RIP3) mRNA and protein levels, with a concomitant increase of the RIP1 protein. Together, these two related proteins form the necrosome, the complex responsible for induction of necroptotic cell death. Interestingly, we found that caspase-8 mRNA, a known negative regulator of necroptosis, was transiently decreased following OGD. Importantly, we observed that the OGD-induced increase in the RIP3 protein was paralleled in an in vivo model of transient global cerebral ischemia, specifically in the CA1 area of the hippocampus. Moreover, we show that the induction of endogenous RIP3 protein levels influenced neuronal toxicity since we found that RIP3 knock-down (KD) abrogated the component of OGD-induced necrotic neuronal death while RIP3 overexpression exacerbated neuronal death following OGD. Overexpression of RIP1 also had deleterious effects following the OGD challenge. Taken together, our results highlight that cerebral ischemia activates transcriptional changes that lead to an increase in the endogenous RIP3 protein level which might contribute to the formation of the necrosome complex and to the subsequent component of necroptotic neuronal death that follows ischemic injury.

Topics: Animals; Antibodies; Apoptosis; Brain Ischemia; Cells, Cultured; Disease Models, Animal; Dizocilpine Maleate; Embryo, Mammalian; Glucose; Hippocampus; Hypoxia; Imidazoles; Indoles; L-Lactate Dehydrogenase; Neurons; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Receptor-Interacting Protein Serine-Threonine Kinases; Tumor Necrosis Factor-alpha; Up-Regulation

In vitro experiments have demonstrated that preconditioning primary neuronal cultures by temporary application of NMDA receptor antagonists induces long-term tolerance against lethal insults. In the present study we tested whether similar effects also occur in brain submitted to ischemia in vivo and whether the potential benefit outweighs the danger of enhancing the constitutive apoptosis in the developing brain.. Memantine in pharmacologically relevant doses of 5 mg/kg or (+)MK-801 (3 mg/kg) was administered i.p. 24, 48, 72 and 96 h before 3-min global forebrain ischemia in adult Mongolian gerbils or prior to hypoxia/ischemia in 7-day-old rats. Neuronal loss in the hippocampal CA1 in gerbils or weight deficit of the ischemic hemispheres in the rat pups was evaluated after 14 days. Also, the number of apoptotic neurons in the immature rat brain was evaluated.. In gerbils only the application of (+)MK-801 24 h before ischemia resulted in significant prevention of the loss of pyramidal neurons. In rat pups administration of (+)MK-801 at all studied times before hypoxia-ischemia, or pretreatment with memantine or with hypoxia taken as a positive control 48 to 92 h before the insult, significantly reduced brain damage. Both NMDA receptor antagonists equally reduced the number of apoptotic neurons after hypoxia-ischemia, while (+)MK-801-evoked potentiation of constitutive apoptosis greatly exceeded the effect of memantine.. We ascribe neuroprotection induced in the immature rats by the pretreatment with both NMDA receptor antagonists 48 to 92 h before hypoxia-ischemia to tolerance evoked by preconditioning, while the neuroprotective effect of (+)MK-801 applied 24 h before the insults may be attributed to direct consequences of the inhibition of NMDA receptors. This is the first report demonstrating the phenomenon of inducing tolerance against hypoxia-ischemia in vivo in developing rat brain by preconditioning with NMDA receptor antagonists.

Topics: Animals; Animals, Newborn; Asphyxia Neonatorum; Brain; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gerbillinae; Immunohistochemistry; Ischemic Preconditioning; Male; Memantine; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate

It is established that the new compound, 9-[2-(4-isopropylphenoxy)ethyl]adenine (9-IPE-adenine) in a dose of 10 mg/kg per day produces neuroprotective effect in rats with brain ischemia model. 9-IPE-adenine decreased the neurologic deficiency 1.2 times more effectively (p < 0.05) than the reference drug mexidol in analogous dose, and had equal effect with this drug at 25 mg/kg per day on the neurologic deficiency and survival of animals. Electrophysiological studies in hippocampal slices in rats showed that 9-IPE-adenine depressed orthodromic population spikes in CA1 area by 42 ± 4%. Non-competitive antagonist of NMDA receptor complex MK-801, in contrast to D-AP5 (competitive NMDA receptor antagonist) and CNQX (competitive AMPA receptor antagonist), enhanced the depressive effect of the new drug more than two times. These ese results are indicative of the ability of 9-IPE-adenine to modulate the ion channel of NMDA receptor complex.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Action Potentials; Adenine; Animals; Brain Ischemia; CA1 Region, Hippocampal; Dizocilpine Maleate; Drug Administration Schedule; Excitatory Amino Acid Antagonists; Male; Neuroprotective Agents; Picolines; Rats; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Tissue Culture Techniques

Evidence suggests that pre-ischeamic conditioning (PIC) offers protection against a subsequent ischeamic event. Although some brain areas such as the hippocampus have received much attention, the receptor mechanisms of PIC in other brain regions are unknown. We have previously shown that 10 min oxygen and glucose deprivation (OGD) evokes tolerance to a second OGD event in the caudate. Here we further examine the effect of length of conditioning event on the second OGD event. Caudate mouse brain slices were superfused with artificial cerebro-spinal fluid (aCSF) bubbled with 95%O(2)/5%CO(2). OGD was achieved by reducing the aCSF glucose concentration and by bubbling with 95%N(2)/5%CO(2). After approximately 5 min OGD a large dopamine efflux was observed, presumably caused by anoxic depolarisation. On applying a second OGD event, 60 min later, dopamine efflux was delayed and reduced. We first examined the effect of varying the length of the conditioning event from 5 to 40 min and found tolerance to PIC increased with increasing duration of conditioning. We then examined the receptor mechanism(s) underlying PIC. We found that pre-incubation with either MK-801 or 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) reduced tolerance to the second OGD event. These data suggest that either N-methyl-D-aspartate (NMDA) or adenosine A(1) receptor activation evokes PIC in the mouse caudate.

Topics: Adenosine A1 Receptor Antagonists; Animals; Brain Ischemia; Caudate Nucleus; Dizocilpine Maleate; Dopamine; Glucose; Heart Arrest; Ischemic Preconditioning; Male; Mice; Mice, Inbred C57BL; Oxygen; Receptor, Adenosine A1; Receptors, N-Methyl-D-Aspartate; Signal Transduction; Tetrazolium Salts; Xanthines

Previous studies in our laboratory have shown that mixed lineage kinase 3 (MLK3) can be activated following global ischemia. In addition, other laboratories have reported that the activation of MLK3 may be linked to the accumulation of free radicals. However, the mechanism of MLK3 activation remains incompletely understood. We report here that MLK3, overexpressed in HEK293 cells, is S-nitrosylated (forming SNO-MLK3) via a reaction with S-nitrosoglutathione, an exogenous nitric oxide (NO) donor, at one critical cysteine residue (Cys-688). We further show that the S-nitrosylation of MLK3 contributes to its dimerization and activation. We also investigated whether the activation of MLK3 is associated with S-nitrosylation following rat brain ischemia/reperfusion. Our results show that the administration of 7-nitroindazole, an inhibitor of neuronal NO synthase (nNOS), or nNOS antisense oligodeoxynucleotides diminished the S-nitrosylation of MLK3 and inhibited its activation induced by cerebral ischemia/reperfusion. In contrast, 2-amino-5,6-dihydro-6-methyl-4H-1,3-thiazine (an inhibitor of inducible NO synthase) or nNOS missense oligodeoxynucleotides did not affect the S-nitrosylation of MLK3. In addition, treatment with sodium nitroprusside (an exogenous NO donor) and S-nitrosoglutathione or MK801, an antagonist of the N-methyl-D-aspartate receptor, also diminished the S-nitrosylation and activation of MLK3 induced by cerebral ischemia/reperfusion. The activation of MLK3 facilitated its downstream protein kinase kinase 4/7 (MKK4/7)-JNK signaling module and both nuclear and non-nuclear apoptosis pathways. These data suggest that the activation of MLK3 during the early stages of ischemia/reperfusion is modulated by S-nitrosylation and provides a potential new approach for stroke therapy whereby the post-translational modification machinery is targeted.

Topics: Animals; Brain Ischemia; Dizocilpine Maleate; Enzyme Activation; Enzyme Inhibitors; HEK293 Cells; Humans; Male; MAP Kinase Kinase Kinases; Mitogen-Activated Protein Kinase Kinase Kinase 11; Neuroprotective Agents; Nitric Oxide Synthase Type I; Oligodeoxyribonucleotides, Antisense; Protein Multimerization; Protein Processing, Post-Translational; Rats; Rats, Sprague-Dawley; Reperfusion Injury; S-Nitrosoglutathione; Thiazines

Previous studies suggested that activated c-Src promote the tyrosine phosphorylation of NMDA receptor subunit NR2A, and thus aggravate the injury induced by transient cerebral ischemia/reperfusion (I/R) in rat hippocampus CA1 region. In this study, we examined the effect of nitric oxide (NO) on the activation of c-Src and the tyrosine phosphorylation of NMDA receptor NR2A subunit. The results show that S-nitrosylation and the phosphorylation of c-Src were induced after cerebral I/R in rats, and administration of nNOS inhibitor 7-NI, nNOS antisense oligonucleotides and exogenous NO donor sodium nitroprusside diminished the increased S-nitrosylation and phosphorylation of c-Src during cerebral I/R. The cysteine residues of c-Src modified by S-nitrosylation are Cys489, Cys498, and Cys500. On the other hand, NMDAR antagonist MK-801 could attenuate the S-nitrosylation and activation of c-Src. Taken together, the S-nitrosylation of c-Src is provoked by NO derived from endogenous nNOS, which is activated by Ca(2+) influx from NMDA receptors, and promotes the auto-phosphorylation at tyrosines and further phosphorylates NR2A. The molecular mechanism we outlined here is a novel postsynaptic NMDAR-nNOS/c-Src-mediated signaling amplification, the 'NMDAR-nNOS → NO → SNO-c-Src → p-c-Src → NMDAR-nNOS' cycle, which presents the possibility as a potential therapeutic target for stroke treatment.

Topics: Amino Acid Motifs; Animals; Apoptosis; Brain Ischemia; Cysteine; Dizocilpine Maleate; Enzyme Activation; HEK293 Cells; Hippocampus; Humans; Indazoles; Male; Neuroprotective Agents; Nitric Oxide Donors; Nitric Oxide Synthase Type I; Nitroprusside; Phosphorylation; Protein Processing, Post-Translational; Protein Structure, Tertiary; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Reperfusion Injury; S-Nitrosoglutathione; src-Family Kinases

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a glycolytic enzyme, plays an important role in glycolysis. It was reported that GAPDH undergoes S-nitrosylation, which facilitated its binding to Siah1 and resulted in nuclear translocation and cell apoptosis. The results of this study show that GAPDH S-nitrosylation, Siah1 binding, translocation to nucleus, and concomitant neuron death occur during the early stages of reperfusion in the rat four-vessel occlusion ischemic model. N-Methyl-D-aspartate receptor antagonist MK801, neuronal nitric oxide synthase inhibitor 7-nitroindazole, or monoamine oxidase-B inhibitor (R)-(-)-deprenyl hydrochloride could inhibit GAPDH S-nitrosylation and translocation and exert neuroprotective effects.

Topics: Active Transport, Cell Nucleus; Analysis of Variance; Animals; Apoptosis; Brain Ischemia; CA1 Region, Hippocampal; Cell Nucleus; Dizocilpine Maleate; Enzyme Inhibitors; Glyceraldehyde-3-Phosphate Dehydrogenases; Humans; Indazoles; Male; Monoamine Oxidase; Neuroprotective Agents; Nitric Oxide; Nitric Oxide Synthase Type I; Nitroso Compounds; Nuclear Proteins; Pyramidal Cells; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Reperfusion Injury; Selegiline; Ubiquitin-Protein Ligases; Vitamin B 12

Deficiency of zinc, which modulates glutamate release, might increase ischemic vulnerability of the brain. We examined effects of dietary zinc deficiency for 2 weeks on ischemic vulnerability in several brain regions using dynamic positron autoradiography technique and [18F]2-fluoro-2-deoxy-d-glucose with rat brain slices. In the normal diet group, the cerebral glucose metabolic rate (CMRglc) was not significantly different from that of the ischemia-unloaded control even after the loading of ischemia for 45 min. However, in the zinc-deficient diet group, CMRglc was significantly lower than that of the ischemia-unloaded control after loading of ischemia for 45 min. With treatment of MK-801 (NMDA receptor antagonist) from the start of ischemia loading, CMRglc was not significantly different from that of the ischemia-unloaded control. These findings, obtained for all analyzed brain regions, suggest that dietary zinc deficiency increased ischemic vulnerability in the brain, and that glutamate might contribute to this effect through activation of the NMDA receptor.

Topics: Animals; Autoradiography; Brain Ischemia; Cerebral Cortex; Diet; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Fluorodeoxyglucose F18; Glucose; Radionuclide Imaging; Rats; Rats, Wistar; Zinc

Both oxidative stress and excessive activation of glutamate receptors are implicated as major causes of ischemic brain injury. However, the existing N-methyl-D-aspartate (NMDA) receptor antagonists have not exerted good clinical outcome, most likely because they do not protect neurons against oxidative stress. Thus, more effective glutamate antagonists and antioxidants are needed for the treatment of ischemic stroke. In previous study, SP-8203, derived from earth worms, showed the blocking effect of NMDA receptor. We provided evidence that SP-8203 could also suppress the oxidative stress in this study. In vitro, 250 μM H2O2 was treated to SH-SY5Y cells after the pre-treatment of SP-8203 (2, 20 and 200 μM). SP-8203 significantly suppressed H2O2-induced cell death and reactive oxygen species production. In addition, we investigated the effects of SP-8203 in middle cerebral artery (MCA) occluded rat model. SP-8203 (5 and 10 mg/kg) was administered intraperitoneally to rats before and after the MCA occlusion and was injected daily for 10 days. After 10 days, SP-8203 remarkably reduced brain infarct volume and lipid peroxidation products in the MCA-occluded rats but MK-801 didn't. Moreover, SP-8203 significantly improved neurological deficits such as shortening of latency time in Rota rod performance. However, MK-801 didn't improve behavioral deficits. Therefore, SP-8203 may be more effective for multiple-target mechanisms of ischemic stroke.

Topics: Acetamides; Animals; Antioxidants; Behavior, Animal; Brain Ischemia; Cell Death; Cell Line; Disease Models, Animal; Dizocilpine Maleate; Glutamic Acid; Humans; Hydrogen Peroxide; Lipid Peroxidation; Male; Motor Activity; Neurons; Neuroprotective Agents; Oxidative Stress; Quinazolinones; Rats; Rats, Wistar; Superoxide Dismutase

CaN induces the apoptosis in neurons, but the influence of CIPC and the intervention of pretreament with Ca(2+)-regulated factors, such as nimodipine, MK801 and cyclosporine A, on CaN expression is not clear. We also do not know whether cbl-b takes part in the induction of ischemia or induces an expression change of cbl-b in CIPC. So we will discuss the effect of CIPC, pretreatment with nimodipine, MK801 and cyclosporine A on the expression of the CaN, cbl-b and p-AKT in the hippocampus neurons. In our study, we established rat models including sham, ischemia, CIPC, nimodipine, MK801 and cyclosporine A. The neurological deficit scores were processed. The right hippocampus was removed and stained with TTC, and the volume of cerebral infarction was calculated. The apoptotic neurons were detected by TUNEL staining. The expressions of CaN, cbl-b and p-AKT at the protein level were examined by Western blotting, and the transcription of cbl-b by RT-PCR, respectively. The results showed that the neurological deficit scores, the volume of the cerebral infarction, the numbers of the apoptotic neurons, the protein expression of CaN, cbl-b and the transcription of cbl-b were the highest in the ischemia and MK801 groups, there were no difference between the two groups(P>0.05); these factors in CIPC group were all lower than those in the ischemia group(P<0.05); and much lower in the nimodipine and cyclosporine A group than those in the CIPC group (among them, the volume of the cerebral infarction in the nimodipine and cyclosporine A groups P<0.01, the expression of CaN in nimodipine group P<0.01, others were P<0.05), but no significant difference existed between the nimodipine and cyclosporine A groups(P>0.05). The expression of p-AKT was the lowest in the ischemia and MK801 groups, and there was no difference between the two groups (P>0.05), This factor was higher in CIPC group than that in the ischemia group (P<0.05); it was the highest in the nimodipine and cyclosporine A groups among these groups (the nimodipine group P<0.01, the cyclosporine A group P<0.05), no significant difference existed between the nimodipine and cyclosporine A groups (P>0.05. Continuous ischemia increases the expression of CaN, and the transcription and the protein expression of cbl-b. Cbl-b reduces the phosphorylated expression of AKT, ultimately activating apoptosis. CIPC inhibits above process and reduces the expression of CaN and cbl-b, and increases the expression of p-AKT, thereby inh

Topics: Animals; Blotting, Western; Brain Ischemia; Calcineurin; Calcium; Calcium Signaling; Cyclosporine; Dizocilpine Maleate; Hippocampus; In Situ Nick-End Labeling; Ischemic Preconditioning; Male; Neurons; Neuroprotective Agents; Nimodipine; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-cbl; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction

The purpose of this study was to investigate neuroprotective efficiency of N-methyl D-aspartate (NMDA) receptor (NMDAR) blockade on the neuronal damage in the less studied and allegedly less affected CA3 hippocampus and striatum in the Mongolian gerbil model of global cerebral ischemia. The common carotid arteries of gerbils were occluded for 5, 10 or 15 minutes. Gerbils were given a low dose of non-competitive NMDA antagonist (MK-801, 3 mg/kg i.p.) or saline immediately after the occlusion in normothermic conditions. Neuronal damage was examined on 4th, 14th and 28th day after reperfusion. The effect of NMDAR blockade was followed in vivo by monitoring the neurological status of whole animals or at the cellular level by standard light- and confocal microscopy on brain slices. Increased duration of cerebral ischemia resulted in a progressive loss of striatal and CA3 hippocampal neurons. The most beneficial NMDAR blockade effect was observed when the neuronal damage was most severe - on the 28th day after 15-min ischemia. As judged by morphological and neurological data, the effect of ischemia is also apparent in the presumed less vulnerable regions (CA3 and striatum) which are functionally important in stroke plasticity. So, NMDAR blockade in normothermic conditions showed neuroprotective efficiency.

Topics: Animals; Brain; Brain Ischemia; CA3 Region, Hippocampal; Corpus Striatum; Dizocilpine Maleate; Gerbillinae; Male; Neuroprotective Agents; Receptors, N-Methyl-D-Aspartate; Reperfusion

N-Methyl-D-aspartate receptors (NMDARs) are essential mediators of synaptic plasticity under normal physiological conditions. During brain ischemia, these receptors are excessively activated due to glutamate overflow and mediate excitotoxic cell death. Although organotypical hippocampal slice cultures are widely used to study brain ischemia in vitro by induction of oxygen and glucose deprivation (OGD), there is scant data regarding expression and functionality of NMDARs in such slice cultures. Here, we have evaluated the contribution of NMDARs in mediating excitotoxic cell death after exposure to NMDA or OGD in organotypical hippocampal slice cultures after 14 days in vitro (DIV14). We found that all NMDAR subunits were expressed at DIV14. The NMDARs were functional and contributed to cell death, as evidenced by use of the NMDAR antagonist MK-801 (dizocilpine). Excitotoxic cell death induced by NMDA could be fully antagonized by 10 μM MK-801, a dose that offered only partial protection against OGD-induced cell death. Very high concentrations of MK-801 (50-100 μM) were required to counteract cell death at long delays (48-72 h) after OGD. The relative high dose of MK-801 needed for long-term protection after OGD could not be attributed to down-regulation of NMDARs at the gene expression level. Our data indicate that NMDAR signaling is just one of several mechanisms underlying ischemic cell death and that prospective cytoprotective therapies must be directed to multiple targets.

Topics: Animals; Blotting, Western; Brain Ischemia; Cattle; Cell Death; Chickens; Dizocilpine Maleate; Gene Expression Regulation; Glucose; Hydrogen-Ion Concentration; Models, Biological; N-Methylaspartate; Neurotoxins; Organ Culture Techniques; Oxygen; Protein Subunits; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Reproducibility of Results; RNA, Messenger; Signal Transduction

A new series of ionone derived allylic alcohols have been evaluated for anti-ischemic activity. Out of them, 12f and 13b decreased infarct volume to 23.98+/-4.7 mm3 and 93.98+/-24.8 mm3 as compared to ischemic group.

Topics: Animals; Apoptosis; Brain Edema; Brain Ischemia; Cell Survival; Drug Design; Male; Molecular Structure; Neurons; Neuroprotective Agents; Norisoprenoids; Rats; Rats, Sprague-Dawley; Stereoisomerism; Structure-Activity Relationship

Although caffeinol (a combination of a low dose of caffeine and ethanol) was shown to robustly reduce stroke damage in experimental models and is now in clinical evaluation for treatment of ischemic stroke, little is known about the potential mechanism of its action.. We used an in vivo excitotoxicity model based on intracortical infusion of N-methyl-D-aspartate (NMDA) and a model of reversible focal ischemia to demonstrate NMDA receptor inhibition as a potential mechanism of caffeinol anti-ischemic activity.. Caffeinol reduced the size of excitotoxic lesion, and substitution of ethanol in caffeinol with the NMDA antagonists CNS-1102 and MK-801 but not with MgSO(4) produced treatment with strong synergistic effect that was at least as robust in reducing ischemic damage as caffeinol. This NMDA receptor antagonist and caffeine combination demonstrated a long window of opportunity, activity in spontaneously hypertensive rats, and, unlike caffeinol, was fully effective in animals chronically pretreated with ethanol.. Our study suggests that antiexcitotoxic properties may underlie some of the anti-ischemic effect of caffeinol. This study provides strong evidence that the anti-ischemic effect of NMDA receptor blockers in general can be dramatically augmented by caffeine, thus opening a possibility for new use of NMDA-based pharmacology in the treatment of stroke.

Topics: Animals; Brain Ischemia; Caffeine; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Combinations; Drug Synergism; Ethanol; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Guanidines; Male; N-Methylaspartate; Nerve Degeneration; Neuroprotective Agents; Neurotoxins; Phosphodiesterase Inhibitors; Rats; Rats, Inbred SHR; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Time Factors; Treatment Outcome

The use of neuroprotective agents has been under investigation for the treatment of ischaemic brain stroke. In this study, we examined the effects of rosiglitazone and MK-801, two potential neuroprotectants, on thromboembloic focal stroke in hyperthermic rats. The animals were assigned into groups of rosiglitazone, MK-801 and control, all under both normothermic and hyperthermic conditions. A focal ischaemia was induced by injection of preformed clot into the origin of the middle cerebral artery. The animals were assessed by measuring infarct size and brain oedema and also evaluating neurological deficit and seizure activity. Rosiglitazone improved infarct volume and neurological deficit in both normo- (36%) and hyperthermic (63%) animals; but MK-801 only improved normothermic animals. Our results do not support the use of MK-801 in hyperthermic conditions of brain stroke but suggest that rosiglitazone may preserve its efficiency even in hyperthermia.

Topics: Animals; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Fever; Male; Neuroprotective Agents; Rats; Rats, Wistar; Rosiglitazone; Thiazolidinediones

We investigated MK-801 effect on ischemia-induced oxidative stress-the most important factor that exacerbates brain damage by reperfusion. The common carotid arteries of gerbils were occluded for 5, 10, or 15 min. Immediately after the occlusion, MK-801 (3 mg/kg i.p.) or saline were given in normothermic conditions. The MK-801 effects were followed in vivo by monitoring the neurological status of animals and at the intracellular level by standard biochemical assays. We investigated nitric oxide levels, superoxide production, superoxide dismutase activity, index of lipid peroxidation (ILP), and reduced glutathione content in hippocampus, striatum, forebrain cortex, and cerebellum. The measurements took place at different times (1, 2, 4, 7, 14, and 28 days) after reperfusion. Increased duration of cerebral ischemia resulted in a progressive induction of oxidative stress. Our results revealed pattern of dynamic changes in each oxidative stress parameter level which corresponded with ischemia duration in all tested brain structures. Most sensitive oxidative stress parameters were ILP and superoxide production. Our study confirmed spatial distribution of ischemia-induced oxidative stress. Tested brain structures showed different sensitivity to each oxidative stress parameter. As judged by biochemical and neurological data, applied MK-801 showed neuroprotective efficiency by reduction of ischemia-induced oxidative stress in brain.

Topics: Animals; Brain; Brain Ischemia; Carotid Artery, Common; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gerbillinae; Glutathione; Lipid Peroxidation; Male; Nitric Oxide; Nitrites; Oxidative Stress; Reperfusion; Superoxide Dismutase; Superoxides

Brain function is highly dependent on cerebral blood flow (CBF). The precise mechanisms by which blood flow is controlled by NIR laser irradiation on the central nervous system (CNS) have not been elucidated. In this study, we examined the effect of 808 nm laser diode irradiation on CBF in mice.. We examined the effect of NIR irradiation on CBF at three different power densities (0.8, 1.6 and 3.2 W/cm(2)) and directly measured nitric oxide (NO) in brain tissue during NIR laser irradiation using an amperometric NO-selective electrode. We also examined the contribution of NO and a neurotransmitter, glutamate, to the regulation of CBF by using a nitric oxide synthase (NOS) inhibitor, N(g)-nitro-L-arginine methyl ester hydrochloride (L-NAME), and an N-methyl-D-aspartate (NMDA) receptor blocker, MK-801, respectively. We examined the change in brain tissue temperature during NIR laser irradiation. We also investigated the protection effect of NIR laser irradiation on transient cerebral ischemia using transient bilateral common carotid artery occlusion (BCCAO) in mice.. We showed that NIR laser irradiation (1.6 W/cm(2) for 15-45 minutes) increased local CBF by 30% compared to that in control mice. NIR laser irradiation also induced a significant increase in cerebral NO concentration. In mice that received L-NAME, NIR laser irradiation did not induce any increase in CBF. Mice administered MK-801 showed an immediate increase but did not show a delayed additional increase in local CBF. The increase in brain tissue temperature induced by laser irradiation was estimated to be as low as 0.8 degrees C at 1.6 W/cm(2), indicating that the heating effect is not a main mechanism of the CBF increase in this condition. Pretreatment with NIR laser irradiation improved residual CBF and reduced the numbers of apoptotic cells in the hippocampus.. Our data suggest that NIR laser irradiation is a promising experimental and therapeutic tool in the field of cerebral circulation research.

Topics: Animals; Apoptosis; Body Temperature; Brain; Brain Ischemia; Carotid Stenosis; Cerebrovascular Circulation; Dizocilpine Maleate; Enzyme Inhibitors; Lasers; Male; Mice; Mice, Inbred C57BL; Neuroprotective Agents; NG-Nitroarginine Methyl Ester; Nitric Oxide; Radiation Dosage

N-methyl-D-aspartate receptors (NMDARs) play an important role in cell survival versus cell death decisions during neuronal development, ischemia, trauma, and epilepsy. Coupling of neurons by electrical synapses (gap junctions) is high or increases in neuronal networks during all these conditions. In the developing CNS, neuronal gap junctions are critical for two different types of NMDAR-dependent cell death. However, whether neuronal gap junctions play a role in NMDAR-dependent neuronal death in the mature CNS was not known. Using Fluoro-Jade B staining, we show that a single intraperitoneal administration of NMDA (100 mg/kg) to adult wild-type mice induces neurodegeneration in three forebrain regions, including rostral dentate gyrus. However, the NMDAR-mediated neuronal death is prevented by pharmacological blockade of neuronal gap junctions (with mefloquine, 30 mg/kg) and does not occur in mice lacking neuronal gap junction protein, connexin 36. Using Western blots, electrophysiology, calcium imaging, and gas chromatography-mass spectrometry in wild-type and connexin 36 knockout mice, we show that the reduced level of neuronal death in knockout animals is not caused by the reduced expression of NMDARs, activity of NMDARs, or permeability of the blood-brain barrier to NMDA. In wild-type animals, this neuronal death is not caused by upregulation of connexin 36 by NMDA. Finally, pharmacological and genetic inactivation of neuronal gap junctions in mice also dramatically reduces neuronal death caused by photothrombotic focal cerebral ischemia. The results indicate that neuronal gap junctions are required for NMDAR-dependent excitotoxicity and play a critical role in ischemic neuronal death.

Topics: Animals; Apoptosis; Blood-Brain Barrier; Brain Ischemia; Cells, Cultured; Connexins; Dentate Gyrus; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gap Junction delta-2 Protein; Gap Junctions; Male; Mefloquine; Mice; Mice, Inbred C57BL; Mice, Knockout; N-Methylaspartate; Nerve Degeneration; Neurons; Neurotoxins; Patch-Clamp Techniques; Photochemistry; Receptors, N-Methyl-D-Aspartate; Rose Bengal

Glutamate is a major excitatory neurotransmitter in the central nervous system and plays a significant role in the pathophysiology of ischemic stroke. During acute ischemic cerebrovascular disease, glutamate efflux in the CNS produces excitotoxicity in neurons and may mediate forms of stress in other tissues expressing glutamate ionotropic (N-methyl-D-aspartate (NMDA)) receptors, e.g., cerebral endothelial cells. While endothelial cell stress in response to glutamate has been reported (oxidant stress, loss of barrier function), changes in protein expression produced by glutamate (an agonist of metabotropic and NMDA receptors) have not been documented. Here, we have examined how exposure of human cerebral endothelial cells to glutamate, in the presence and absence of the NMDA receptor antagonist MK-801, can alter the proteomic profile of cerebral endothelial cells. We found several important changes in the proteins expressed by cerebral endothelial cells in response to glutamate. Interestingly, MK-801 itself had some direct effects on cerebral endothelial cells. Taken together, our findings demonstrate that cerebral endothelial cells respond to glutamate by altering their protein expression profile. We assume that protein alterations found in the cerebral endothelial proteome, in response to glutamate and which were blocked by MK-801, may be important vascular targets in better understanding the pathogenesis of ischemic stroke.

Topics: Animals; Brain Ischemia; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Electrophoresis, Gel, Two-Dimensional; Endothelial Cells; Excitatory Amino Acid Antagonists; Glutamic Acid; Humans; Neuroprotective Agents; Protein Array Analysis; Proteome

A photochemical ring model of ischaemia was introduced in the middle of the nineteen eighties. Irradiation by a laser or arc lamp followed by intravenous injection of rose bengal resulted in thrombosis of pial and superficial cortical vessels. This ring model imitated focal ischaemic damage in humans. In our experiment twenty-seven Wistar rats of both sexes weighing 250-300 grams were examined. A photochemical ring model based on irradiation of the area of parietal bone 4 mm posteriorly to the bregma and 4 mm laterally from the sagittal suture was applied. A ring-shaped light beam with a wavelength of 510-540 nm with 5 mm diameter was generated by a high pressure discharge lamp at a power of 400 W. Two groups of rats treated and untreated with MK-801 and two rings of the thickness of 0.35 mm and 0.5 mm were used in the experiment. Morphological examination was performed in animals sacrificed 1 and 4 days after the irradiation. On formalin-fixed and paraffin-embedded slices HE staining method and immunoreaction with antibodies to ubiquitin were applied. Our material confirmed well known information about the dynamics of infarct breakdown, ischaemic-induced angiogenesis, glial reaction and other typical changes described previously in handbooks and numerous papers. In the experiment, morphological changes were more intensive after the irradiation by 0.5 mm than 0.35 mm irradiating rings and 4 days than one day after the irradiation. A surprising finding observed in some of the examined animals was more intensive neuronal damage after treatment with MK-801. Another unpredicted discovery was intensive morphological alterations found in CA4 and CA3 hippocampal sectors. Moreover, these alterations were not limited to the damaged hemisphere, but were also observed contralaterally. In some of the rats, ischaemic and necrotic cells were additionally found within both parasagittal areas. We connect this atypical localization of the ischaemic changes with dispersion of light emitted by the used lamp. Dispersed light also leads to thrombotic occlusion of the meningeal arteries in the parasagittal area. Among these arteries, thrombosis in pericallosal and penetrating arteries was present. Our experiment demonstrated that if a non-laser lamp is used, brain areas distant from the necrotic ring must be carefully investigated.

Topics: Animals; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Hippocampus; Lasers; Male; Rats; Rats, Wistar

Previous work has demonstrated that ischemic preconditioning neuroprotection is associated with inhibition of JNK pathway activation. The present study was designed to examine the hypothesis that the suppression of JNK3 activation by preconditioning is mediated by NMDA receptors and crosstalk between ERK1/2 and JNK3. Preconditioning (3 min ischemia) 2 days before global cerebral ischemia (8-min) markedly decreased neuronal degeneration in hippocampus CA1, an effect abolished by pretreatment with the NMDA receptor antagonist, MK-801. Furthermore, preconditioning abolished cerebral ischemia-induced JNK3 activation and enhanced ERK1/2 activation, an effect reversed by MK-801. Due to the inverse relationship between ERK1/2 and JNK3 activation following preconditioning, we hypothesized that ERK1/2 may regulate JNK3 activation following preconditioning. In support of this contention, pretreatment with the MEK inhibitor, PD98059 significantly attenuated preconditioning-induced ERK1/2 phosphorylation, and strongly reversed preconditioning down-regulation of JNK3 phosphorylation. This finding suggests that ERK1/2 signaling is responsible for preconditioning-induced down-regulation of JNK3 activation. Western blot analysis and immunohistochemistry further demonstrated that preconditioning, in an NMDA-dependent manner, enhanced activation of the pro-survival factors, p-CREB and Bcl-2, while attenuating activation of putative pro-death factors, p-c-Jun and Fas-L in the hippocampus CA1. As a whole, the study demonstrates that preconditioning attenuation of pro-death JNK3 in the hippocampus CA1 following global cerebral ischemia is mediated by NMDA receptor-induced crosstalk between ERK1/2 and JNK3. The ERK1/2-mediated reduction of JNK3 activation leads to enhanced pro-survival signaling (P-CREB and Bcl-2 induction) and attenuation of pro-death signaling (p-c-Jun and Fas-L), with subsequent induction of ischemic tolerance.

Topics: Analysis of Variance; Animals; Brain Ischemia; CREB-Binding Protein; Disease Models, Animal; Dizocilpine Maleate; Down-Regulation; Enzyme Activation; Flavonoids; Gene Expression Regulation, Enzymologic; Hippocampus; In Situ Nick-End Labeling; Ischemic Preconditioning; Male; Mitogen-Activated Protein Kinase 10; Mitogen-Activated Protein Kinase 3; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Signal Transduction; Time Factors

We report in this article for the first time the neuroprotective effects of unconjugated TAT carrier peptide against a mild excitotoxic stimulus both in vitro and in vivo. In view of the widespread use of TAT peptides to deliver neuroprotectants into cells, it is important to know the effects of the carrier itself. Unconjugated TAT carrier protects dissociated cortical neurons against NMDA but not against kainate, suggesting that TAT peptides may interfere with NMDA signaling. Furthermore, a retro-inverso form of the carrier peptide caused a reduction in lesion volume (by about 50%) in a rat neonatal cerebral ischemia model. Thus, even though TAT is designed merely as a carrier, its own pharmacological activity will need to be considered in the analysis of TAT-linked neuroprotectant peptides.

Topics: Age Factors; Animals; Animals, Newborn; Brain Ischemia; Cell Death; Cells, Cultured; Cerebral Cortex; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Agonists; Kainic Acid; L-Lactate Dehydrogenase; N-Methylaspartate; Neurons; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; tat Gene Products, Human Immunodeficiency Virus; Time Factors

It is well documented that exitotoxicity induced by N-methyl-D-aspartate (NMDA) receptor activation plays a pivotal role in delayed neuronal death in the hippocampal CA1 region after transient global ischemia. However, the effect of gamma-aminobutyric acid (GABA) receptor activation is uncertain in ischemia brain injury. The aim of this study was to investigate whether the enhancement of GABA receptor activity could inhibit NMDA receptor-mediated nitric oxide (NO) production by neuronal NO synthase (nNOS) in brain ischemic injury. The results showed that both the GABA(A) receptor agonist muscimol and the GABA(B) receptor agonist baclofen had neuroprotective effect, and the combination of two agonists could significantly protect neurons against death induced by ischemia/reperfusion. Coapplication of muscimol with baclofen not only enhanced nNOS (Ser847) phosphorylation but also increased the interaction of nNOS with PSD95 at 6 hr and 1 day of reperfusion. Interestingly, the inhibitors of calcineurin and PP1/PP2A could enhance nNOS phosphorylation at Ser847 site at 1 day of reperfusion after ischemia but not at 6 hr of reperfusion. From these data, we conclude that GABA receptor activation could exert its neuroprotective effect through increasing nNOS (Ser847) phosphorylation by different mechanisms at 6 hr and 1 day of reperfusion. The increased interaction of nNOS and postsynaptic density-95 induced by GABA agonists is responsible for nNOS (Ser847) phosphorylation at both time points, but at 1 day of reperfusion the inhibition of protein phosphatase activity by GABA agonists also contributes to the neuroprotection. Our results suggest that GABA receptor agonists may serve as a potential and important neuroprotectant in therapy for ischemic stroke.

Topics: Animals; Baclofen; Blotting, Western; Brain Ischemia; Disks Large Homolog 4 Protein; Dizocilpine Maleate; GABA Agonists; Immunohistochemistry; Immunoprecipitation; In Situ Nick-End Labeling; Intracellular Signaling Peptides and Proteins; Male; Membrane Proteins; Muscimol; Neurons; Neuroprotective Agents; Nitric Oxide Synthase Type I; Phosphoprotein Phosphatases; Phosphorylation; Rats; Rats, Sprague-Dawley; Receptors, GABA

Dark neurons have plagued the interpretation of brain tissue sections, experimentally and clinically. Seen only when perturbed but living tissue is fixed in aldehydes, their mechanism of production is unknown. Since dark neurons are seen in cortical biopsies, experimental ischemia, hypoglycemia, and epilepsy, we surmised that glutamate release and neuronal transmembrane ion fluxes could be the perturbation leading to dark neuron formation while the fixation process is underway. Accordingly, we excised biopsies of rat cortex to simulate neurosurgical production of dark neurons. To ascertain the role of glutamate, blockade of N-methyl-D-aspartate (NMDA) and non-NMDA receptors was done prior to formaldehyde fixation. To assess the role of transmembrane sodium ion (and implicitly, water) fluxes, tetraethylammonium (TEA) was used. Blockade of NMDA receptors with MK-801 and non-NMDA receptors with the quinoxalinediones (CNQX and NBQX) abolished dark neuron formation. More delayed exposure of the tissue to the antagonist, CNQX, by admixing it with the fixative directly, allowed for some production of dark neurons. Aminophosphonoheptanoate (APH), perhaps due to its polarity, and TEA, did not prevent dark neurons, which were abundant in control formaldehyde fixed material unexposed to either receptor or ion channel antagonists. The results demonstrate a role for the pharmacologic subtypes of glutamate receptors in the pathogenetic mechanism of dark neuron formation. Our results are consistent with the appearance of dark neurons in biopsy where the cerebral cortex has been undercut, and rendered locally ischemic and hypoglycemic, as well as in epilepsy, hypoglycemia, and ischemia, all of which lead to glutamate release. Rather than a pressure-derived mechanical origin, we suggest that depolarization, glutamate release or receptor activation are more likely mechanisms of dark neuron production.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Artifacts; Biopsy; Brain Ischemia; Cerebral Cortex; Dizocilpine Maleate; Epilepsy; Excitatory Amino Acid Antagonists; Glutamic Acid; Hypoglycemia; Male; Neurons; Potassium Channel Blockers; Quinoxalines; Rats; Rats, Wistar; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Tetraethylammonium

The purpose of this study was to investigate the temporal pattern of NMDA receptors antagonist-MK-801 on motor behaviour parameters in gerbils submitted to different duration of global cerebral ischemia. The common carotid arteries of gerbils were occluded for 5, 10 or 15min. Gerbils were given MK-801 (3mg/kg i.p.) or saline immediately after the occlusion in normothermic conditions prior to testing. Motor activity was registered 1, 2, 4, 7, 14, 21 and 28 days after reperfusion during 60min by open field test. At the same time, the effect of NMDA receptor blockade was followed in vivo by monitoring the neurological status of whole animals or at the cellular level by standard light and confocal microscopy on brain slices. Post-ischemic gerbils quickly developed hypermotor response with the most intensity in animals submitted to 15min ischemia. MK-801 administrated immediately after ischemia significantly decreased this hyperactivity. In all ischemic-treated animals, behavioural suppression by MK-801 was observed already 1 day after occlusion and was lasting as far as observed ischemia-dependent hypermotor responses. Beneficial effect of MK-801 was also confirmed by morphological and neurological status data. These findings suggest that sustained ischemia-induced hyperactivity is related to abnormalities in NMDA glutamatergic function, as well as its manifestation could be completely abolished by NMDA receptor blockade immediately after ischemic insult.

Topics: Analysis of Variance; Animals; Behavior, Animal; Brain Ischemia; Corpus Striatum; Disease Models, Animal; Dizocilpine Maleate; Gerbillinae; Male; Motor Activity; Movement Disorders; Neurologic Examination; Neuroprotective Agents; Reperfusion; Stereotyped Behavior; Time Factors

Topics: Animals; Brain Infarction; Brain Ischemia; Depressive Disorder; Dizocilpine Maleate; Eliminative Behavior, Animal; Neuroprotective Agents; Rats; Rats, Wistar

Damage to oligodendrocytes caused by glutamate release contributes to mental or physical handicap in periventricular leukomalacia, spinal cord injury, multiple sclerosis, and stroke, and has been attributed to activation of AMPA/kainate receptors. However, glutamate also activates unusual NMDA receptors in oligodendrocytes, which can generate an ion influx even at the resting potential in a physiological [Mg2+]. Here, we show that the clinically licensed NMDA receptor antagonist memantine blocks oligodendrocyte NMDA receptors at concentrations achieved therapeutically. Simulated ischaemia released glutamate which activated NMDA receptors, as well as AMPA/kainate receptors, on mature and precursor oligodendrocytes. Although blocking AMPA/kainate receptors alone during ischaemia had no effect, combining memantine with an AMPA/kainate receptor blocker, or applying the NMDA blocker MK-801 alone, improved recovery of the action potential in myelinated axons after the ischaemia. These data suggest NMDA receptor blockers as a potentially useful treatment for some white matter diseases and define conditions under which these blockers may be useful therapeutically. Our results highlight the importance of developing new antagonists selective for oligodendrocyte NMDA receptors based on their difference in subunit structure from most neuronal NMDA receptors.

Topics: Action Potentials; Animals; Animals, Newborn; Brain Ischemia; Central Nervous System; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Interactions; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; In Vitro Techniques; Memantine; Myelin Basic Protein; N-Methylaspartate; Oligodendroglia; Patch-Clamp Techniques; Protein Binding; Rats; Receptors, N-Methyl-D-Aspartate

Cerebral ischemia due to secondary injuries plays an important role in the high mortality rate of acute subdural hematoma (SDH). Although promising results were obtained from experimental works with excitatory amino acid (EAA) antagonists which inhibit the excitotoxic mechanism in the development of cerebral ischemia, these agents could not be used clinically due to their psychomimetic side effects. Memantine, also an EAA antagonist, has been used for a long time in the treatment of different neurodegenerative diseases; however, it was not used in treatment of acute subdural hematoma before. This study has been designed to investigate the development of cerebral ischemia and ischemic edema under experimental acute subdural hematoma and the effect of memantine (Sigma M-9292) and MK-801 (Sigma M-107) in the treatment of ischemia.. Forty-two adult female Sprague-Dawley rats were divided into two groups: Group A for investigation of ischemia related to SDH and its treatment, and Group B for investigation of cerebral edema. Both groups were further divided into five subgroups, i.e. for sham operations, formation of SDH and treatment with saline, MK-801 and memantine. Treatment of cerebral edema could not be investigated because formation of cerebral edema could not be proven statistically. For evaluation of ischemia, the ratio of ischemic area/the total brain area was calculated as percentages in coronal slices of the rats' brains.. In all of the evaluated slices, statistical analysis showed that treatment with MK-801 as well as memantine reduced ischemia caused by SDH.. Our study showed that memantine, which is already considered as a safe treatment alternative for other central nervous system (CNS) diseases, can be useful in the treatment of acute SDH as well.

Topics: Animals; Brain Edema; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Functional Laterality; Hematoma, Subdural, Acute; Memantine; Rats; Rats, Sprague-Dawley; Statistics, Nonparametric

Hematopoietic progenitor kinase 1 (HPK1) is a hematopoietic cell-restricted member of the Ste20 serine/threonine kinase super family. We recently reported that HPK1 is involved in c-Jun NH2-terminal kinase (JNK) signaling pathway by sequential activation of MLK3-MKK7-JNK3 after cerebral ischemia. Here, we used 4-amino-5-(4-chlorophenyl)-7-(t-butyl) pyrazolo [3,4-d] pyrimidine (PP2) and MK801 to investigate the events upstream of HPK1 in ischemic brain injury. Immunoprecipitation and immunoblot results showed that PP2 and MK801 significantly decreased the activation of Src, HPK1, MLK3, JNK3 and c-Jun, respectively, during ischemia/reperfusion. Histology and TUNEL staining showed PP2 or MK801 protects against neuron death after brain ischemia. We speculate that this unique signaling pathway through the tyrosine phosphorylation of HPK1 promotes ischemic brain injury by activated Src via N-methyl-d-aspartate receptor and, ultimately, the activation of the MLK3-MKK7-JNK3 pathway after cerebral ischemia.

Topics: Animals; Brain Ischemia; Cytoprotection; Dizocilpine Maleate; Hippocampus; Male; MAP Kinase Kinase Kinases; Mitogen-Activated Protein Kinase 10; Mitogen-Activated Protein Kinase Kinase Kinase 11; Neuroprotective Agents; Phosphorylation; Protein Kinase Inhibitors; Protein Serine-Threonine Kinases; Pyrimidines; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction; src-Family Kinases; Tyrosine

Recombinant tissue plasminogen activator (rt-PA) treatment improves functional outcome after acute ischemic stroke, inducing reperfusion by its thrombolytic activity. Conversely, there is evidence that rt-PA can mediate neuronal damage after ischemic brain injury in vivo. In addition to other mechanisms, enhancement of N-methyl-D-aspartate (NMDA) receptor signalling has been proposed to underlie rt-PA-mediated neurotoxicity. However, the role of poly(ADP-ribose) polymerase-1 (PARP-1) activation, which mediates postischemic excitotoxic cell death, in rt-PA-mediated aggravation of ischemic brain injury has not been established and was therefore addressed in this study. After permanent focal cerebral ischemia, intravenous rt-PA application significantly increased early postischemic PARP-1 activation within ischemic hemispheres and infarct volumes compared with control mice without affecting cerebral blood flow. Rt-PA induced increase in PARP-1 activation, and infarct volumes could be blocked by the PARP inhibitor 3-aminobenzamide. Moreover, the rt-PA-induced increase in PARP-1 activation was also prevented by the NMDA antagonist MK-801. In summary, we demonstrate that rt-PA treatment enhances postischemic PARP-1 activation, which contributes to rt-PA induced aggravation of ischemic brain injury in vivo. Furthermore, we provide evidence that NMDA receptor activation is required for rt-PA-mediated effects on postischemic PARP-1 activation.

Topics: Animals; Benzamides; Blood Gas Analysis; Blood Pressure; Blood-Brain Barrier; Brain; Brain Infarction; Brain Ischemia; Dizocilpine Maleate; Enzyme Activation; Ischemic Attack, Transient; Male; Mice; Mice, Inbred C57BL; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerase Inhibitors; Poly(ADP-ribose) Polymerases; Receptors, N-Methyl-D-Aspartate; Recombinant Proteins; Tissue Plasminogen Activator

The long-term effects of the 5-HT(2A) receptor antagonist ketanserin on deficits in sensorimotor integration (limb placing tests) following transient focal cerebral ischemia in rats were compared to the effects of the NMDA antagonist MK-801. Middle cerebral artery occlusion was induced in conscious rats by microinjection of endothelin-1 in the vicinity of the artery (EMCAO model). The EMCAO/vehicle rats exhibited impaired tactile and proprioceptive limb placing. In contrast to ketanserin, MK-801 exerted severe early behavioral disturbances, but both drugs significantly improved the neurological scores much earlier than the spontaneous recovery of function occurred. The present results suggest that pharmacotherapy by means of ketanserin lacking the severe side effects of the NMDA antagonists can be used to enhance functional recovery after stroke.

Topics: Analysis of Variance; Animals; Behavior, Animal; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Endothelin-1; Extremities; Ketanserin; Male; Neurologic Examination; Neuroprotective Agents; Platelet Aggregation Inhibitors; Rats; Rats, Wistar; Severity of Illness Index

To explore the action mechanism of Tongxinluo Capsule (TXL) in protecting brain from ischemic damage.. SD rats were divided into five groups randomly, the sham operation group, the model group, the MK-801 group, the large and low dosage TXL groups (TXLL and TXLS). After the middle cerebral arterial obstructive (MCAO) model was established, peritoneal injection of MK-801 0.5 mg/kg per day was given to the MK-801 group, and 1.0 g/(kg x d) and 0.5 g/(kg x d) of TXL powder was administered in twice via gastrogavage to the two TXL groups respectively. The nerve cell apoptosis rate, protein and mRNA expressions of Caspase-3, p53 and heat shock protein (HSP70) were observed using flow cytometry, Western blot and RT-PCR technique.. Both TXL and MK-801 could obviously lower the apoptosis rate in model rat (P < 0.05, P < 0.01), TXLL showed the optimal effect. Caspase-3, p53 protein and mRNA expression in the model group were obviously higher than those in the sham operated group. As compared with the model group, the expressions of Caspase-3 and p53 were lower and those of HSP70 and mRNA were higher in the two TXL and MK-801 groups (P < 0.05 or P < 0.01).. TXL displays it brain protective effect through reducing nerve cell apoptosis rate in MCAO model rats, the mechanism may be related to its actions in inhibiting apoptosis related factors Caspase-3 and p53, and promoting stress protecting factor HSP70.

Topics: Animals; Apoptosis; Arterial Occlusive Diseases; Blotting, Western; Brain Ischemia; Capsules; Caspase 3; Cerebral Arteries; Disease Models, Animal; Dizocilpine Maleate; Drugs, Chinese Herbal; Male; Neurons; Neuroprotective Agents; Random Allocation; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tumor Suppressor Protein p53

Neuregulin-1 (NRG-1) is a growth factor with potent neuroprotective capacity in ischemic stroke. We recently showed that NRG-1 reduced neuronal death following transient middle cerebral artery occlusion (tMCAO) by up to 90% with an extended therapeutic window. Here, we examined the neuroprotective potential of NRG-1 using a permanent MCAO ischemia (pMCAO) rat model. NRG-1 reduced infarction in pMCAO by 50% when administered prior to ischemia. We previously demonstrated using gene expression profiling that pMCAO was associated with an exaggerated excitotoxicity response compared to tMCAO. Therefore, we examined whether co-treatment with an inhibitor of excitotoxicity would augment the effect of NRG-1 following pMCAO. Both NRG-1 and the N-methyl-D-aspartate (NMDA) antagonist MK-801 similarly reduced infarct size following pMCAO. However, combination treatment with both NRG-1 and MK-801 resulted in greater neuroprotection than either compound alone, including a 75% reduction in cortical infarction compared to control. Consistent with these findings, NRG-1 reduced neuronal death using an in vitro ischemia model and this effect was augmented by MK-801. These results demonstrate the efficacy of NRG-1 in pMCAO rat focal ischemia model. Our findings further indicate the potential clinically relevance of NRG-1 alone or as a combination strategy for treating ischemic stroke.

Topics: Animals; Brain Infarction; Brain Ischemia; Cell Death; Cell Line, Tumor; Disease Models, Animal; Dizocilpine Maleate; Glucose; Hypoxia; Male; Neuregulin-1; Neuroblastoma; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Tetrazolium Salts

Cerebral ischemia can induce both the increase of excitation and the decrease of inhibition, which leads to neuronal excitotoxicity. Since glutamatergic and GABAergic transmissions work by each counterbalancing the function of the other, enhancing GABAergic activity should balance excessive glutamatergic excitation. But the potential mechanisms underlying these effects are obscure. Here, we used two GABA agonists, muscimol and baclofen, and performed immunoblotting, immunoprecipitation and histology analysis to evaluate the neuroprotective effects by stimulating GABA receptors in rat four-vessel occlusion (4-VO) ischemic model, and to investigate the potential mechanism. Our results indicate that whether in global cerebral ischemia in vivo, or in oxygen glucose deprivation (OGD) in vitro, coapplication of muscimol with baclofen can protect neurons from neuronal death through down-regulating the function of N-methyl-d-aspartic acid (NMDA) receptors via attenuating the tyrosine phosphorylation of NR2A subunit. We further elucidate that the phosphorylation level of Src kinase and the interaction among Src, post-synaptic density protein 95 and NR2A were also suppressed by coapplication of muscimol with baclofen. Both MK-801, a specific antagonist of NMDA receptors, and chelerythrine, an inhibitor of protein kinase C (PKC), could down-regulate the phosphorylation of NR2A via inhibiting the activation of Src and PKC respectively. These results suggest that the modified pattern of dynamic balance between excitation and inhibition by coactivation of the GABA receptors in cerebral ischemia can attenuate the excitatory NMDAR via inhibiting a novel postsynaptic NMDAR/Src-mediated signal amplification, the 'NMDAR-Ca(2+) --> PKC --> Src --> NMDAR-Ca(2+)' cycle.

Topics: Analysis of Variance; Animals; Apoptosis; Baclofen; Brain Ischemia; Cerebral Infarction; Disease Models, Animal; Dizocilpine Maleate; Drug Interactions; Excitatory Amino Acid Antagonists; GABA Agents; In Situ Nick-End Labeling; Male; Muscimol; Neurons; Rats; Rats, Sprague-Dawley; Receptors, GABA; Receptors, N-Methyl-D-Aspartate; Reperfusion; src-Family Kinases; Tyrosine

In vitro studies suggest that glutamate receptor activation is important in the genesis of post-hypoxic preterm brain injury, but there are limited data on post-hypoxic N-methyl-D-aspartate (NMDA) receptor activation. We therefore examined an infusion of the specific, non-competitive NMDA receptor antagonist dizocilpine (2 mg kg(-1) bolus plus 0.07 mg kg(-1) h(-1) i.v.) from 15 min to 4 h after severe hypoxia-ischemia induced by umbilical cord occlusion for 25 min in fetal sheep at 70% of gestation. Dizocilpine suppressed evolving epileptiform transient activity in the first 6 h after reperfusion (2.3 +/- 0.9 versus 9.3 +/- 2.3 maximal counts min(-1), P < 0.05) and mean EEG intensity up to 11 h after occlusion (P < 0.05). Fetal extradural temperature transiently increased during the dizocilpine infusion (40.1 +/- 0.2 versus 39.3 +/- 0.1 degrees C, P < 0.05). After 3 days recovery, treatment was associated with a significant reduction in neuronal loss in the striatum (31 +/- 7 versus 58 +/- 2%, P < 0.05), expression of cleaved caspase-3 (111+/-7 versus 159 +/- 10 counts area(-1), P < 0.05) and numbers of activated microglia (57 +/- 9 versus 92 +/- 16 counts area(-1), P < 0.05); there was no significant effect in other regions or on loss of immature O4-positive oligodendrocytes. In conclusion, abnormal NMDA receptor activation in the first few hours of recovery from hypoxia-ischemia seems to contribute to post-hypoxic striatal damage in the very immature brain.

Topics: Animals; Brain Ischemia; Corpus Striatum; Disease Models, Animal; Dizocilpine Maleate; Electroencephalography; Female; Gestational Age; Hypoxia; Microglia; Neuroprotective Agents; Pregnancy; Pyramidal Cells; Sheep

We observed recently that elevated plasma cysteine levels are associated with poor clinical outcome in acute stroke patients. In a rat stroke model, cysteine administration increased the infarct volume apparently via its conversion to hydrogen sulfide (H2S). We therefore investigated the effects of H2S and the inhibition of its formation on stroke.. Cerebral ischemia was studied in a rat stroke model created by permanent occlusion of the middle cerebral artery (MCAO). The resultant infarct volume was measured 24 hours after occlusion.. Administration of sodium hydrosulfide (NaHS, an H2S donor) significantly increased the infarct volume after MCAO. The NaHS-induced increase in infarct volume was abolished by the administration of dizolcilpine maleate (an N-methyl-d-aspartate receptor channel blocker). MCAO caused an increase in H2S level in the lesioned cortex as well as an increase in the H2S synthesizing activity. Administration of 4 different inhibitors of H2S synthesis reduced MCAO-induced infarct volume dose dependently. The potency of these inhibitors in effecting neuroprotection in vivo appeared to parallel their potency as inhibitors of H2S synthesis in vitro. It also appeared that most of the H2S synthesizing activity in the cortex results from the action of cystathionine beta-synthase.. The present results strongly suggest that H2S plays a part in cerebral ischemic damage after stroke. Inhibition of H2S synthesis should be investigated for its potential as a novel neuroprotective stroke therapy.

Topics: Air Pollutants; Animals; Brain Injuries; Brain Ischemia; Cerebral Cortex; Cysteine; Dizocilpine Maleate; Dose-Response Relationship, Drug; Humans; Hydrogen Sulfide; Infarction, Middle Cerebral Artery; Male; Models, Statistical; Neuroprotective Agents; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; Stroke; Time Factors; Treatment Outcome

Whether cannabinoids act as neuroprotectants or, on the contrary, even worsen neuronal damage after cerebral ischemia is currently under discussion. We have previously shown that treatment with the cannabinoid (CB1) receptor antagonist SR141716A reduces infarct volume by approximately 40% after experimental stroke. Since it is suggested that SR141716A may exert neuroprotection besides its cannabinoid receptor-blocking effect, we addressed the question whether SR141716A may act via modulation of postischemic ligand binding to excitatory NMDA and/or alpha-amino-3-hydroxy-5-methyl-4-isoxazole-proprionic acid (AMPA) receptors. For this purpose, rats (n = 12) were treated with either intravenous saline (control) or CB1 receptor antagonist SR141716A (1 mg/kg) 30 min after permanent middle cerebral artery occlusion. Five hours after ischemia, quantitative receptor autoradiography was performed using [(3)H]CP 55,940, [(3)H]MK-801, and [(3)H]AMPA for labeling of CB1, NMDA, and AMPA receptors, respectively. Ligand binding was analyzed within the infarct core, cortical penumbra, and corresponding areas of the contralateral hemisphere and compared to that of sham-operated rats (n = 5). Both in ischemic controls and SR141716A-treated rats [(3)H]CP 55,940 ligand binding was not specifically regulated in the cortical penumbra or contralateral cortex. Importantly, reduced infarct volumes in SR141716A-treated rats were associated with maintained [(3)H]MK-801 binding to excitotoxic NMDA receptors in the penumbra, compared to a decrease in the control group. In summary, our data suggest that SR141716A may possess additional intrinsic neuroprotective properties independent of receptor-coupled pathways or due to action as a partial agonist.

Topics: Animals; Autoradiography; Binding, Competitive; Brain Ischemia; Cyclohexanols; Dizocilpine Maleate; Down-Regulation; Excitatory Amino Acid Antagonists; Male; Neuroprotective Agents; Piperidines; Pyrazoles; Rats; Rats, Wistar; Receptor, Cannabinoid, CB1; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Rimonabant

Ginkgo biloba exerts many pharmacological actions. It possesses antioxidant properties, the ability of neurotransmitter/receptor modulation and antiplatelet activation factor. This research is designed to investigate the neuroprotective effects of long-term treatment with EGb761 (a standard form of the extract of Ginkgo biloba leaf) in combination with MgSO(4), FK506, or MK-801 on the infarct volume of male gerbils' brain induced by unilateral middle cerebral artery occlusion (MCAO). Thirty-five gerbils fed a standard diet were intragastrically given water or EGb761 (100 mg/kg/day) for one week. Five randomized groups were established: control (n = 7), EGb761 (n = 8), EGb761 + MgSO(4) (n = 7), EGb761 + FK506 (n = 7), and EGb761 + MK-801 (n = 6). The three drug-combination groups were injected with MgSO(4) (90 mg/kg), FK506 (0.5 mg/kg), or MK-801 (1 mg/kg), respectively 30 min before MCAO. Gerbils were anesthetized and craniectomized to expose the right middle cerebral artery (MCA). The right MCA was constricted with an 8-0 suture to produce a permanent ligation for 24 hours. Postmortem infarct volumes were determined by quantitative image analysis of 2,3,5-triphenyltetrazolium chloride (TTC)-stained brain sections. Results showed that the total infarct volumes of the four treated groups either EGb761 alone or in combination with drugs were lower than the control group by 36.1% (EGb761 alone), 40.3% (EGb761 + MgSO(4)), 35.3% (EGb761 + FK506), and 56.4% (EGb761 + MK-801), respectively (p < 0.01). The main affected areas of the brain in the four treated groups were significantly focused between 4 and 6 mm from the frontal pole, when compared to the control group (p < 0.01). All animals in the five groups had infarctions in both cortex and subcortex. These results indicate that long-term pre-treatment of EGb761 administered either alone or in combination with drugs significantly effective neuroprotection on infarct volume in gerbil ischemic brains.

Topics: Animals; Brain Infarction; Brain Ischemia; Dizocilpine Maleate; Drug Therapy, Combination; Gerbillinae; Ginkgo biloba; Infarction, Middle Cerebral Artery; Magnesium Sulfate; Male; Neuroprotective Agents; Plant Extracts; Random Allocation; Tacrolimus; Treatment Outcome

To observe the effects of Ginkgo biloba extract (GBE) on N-methyl-D-aspartate (NMDA) excitotoxicity and focal cerebral ischemia, and further explore the neuroprotective mechanisms of GBE.. Neonatal SD rat hippocampus was taken out to make into cell suspension. immunohistochemistry with neuron nucleoprotein monoclonal antibody (NeuN) was used to calculate the percentage of NeuN positive cells. Twelve days after incubation the suspension of neurons were randomly divided into 4 groups: normal control group (exposed to normal saline for 15 min and then to DMEM without NMDA and glycine for 24 h), NMDA group (exposed to culture fluid with NMDA of the terminal concentration of 100 micromol/L and glycine of the terminal concentration of 10 micromol/L for 15 min and then to DMEM without NMDA and glycine for 24 h), MK-801 group (exposed to MK-801, an NMDAR antagonist, for 2 min, to culture fluid with NMDA for 15 min, and then to DMEM without NMDA and glycine for 24 h), and GBE pretreatment group (exposed to GBE of the terminal concentration of 150 microg/ml for 3 d, culture fluid with NMDS for 15 min, and then to DMEM without NMDA and glycine foe 24 h). Trypan blue staining was used to calculate the survival rate of the neurons. The lactic dehydrogenase (LDH) level in the supernatant of cultured cell suspension was detected. Whole-cell patch clamp recording was carried out to evaluate the modulatory effects of GBE on NMDA-activated currents in the rat hippocampal neurons. 108 SD rats were randomly divided into 5 groups: sham operation group (n = 12), standard middle cerebral artery occlusion (MCAO) group (n = 24, undergoing MCAO and then reperfusion), MK-801 acute administration group (n = 24, undergoing MCAO and immediate peritoneal administration of MK-801 1 mg/kg), GBE acute administration group (n = 24, undergoing peritoneal injection of GBE 100 mg/kg immediately after the MCAO), and GBE pretreatment group (n = 24, undergoing peritoneal administration of GBE every day for 7 days before the MCAO). The 4 groups were re-divided into 4 subgroups with 3 approximately 4 rats: 0.5 h ischemia, and 3 h, 1 d, and 7 d ischemia-reperfusion (IR) subgroups. The neurological symptoms were evaluated by Longa's scoring after the rats became conscious. The rats were killed at different time-points, their brains were taken out to undergo 2, 3, 5-triphenyl-tetrazolium chloride staining, the areas of cerebral infarction were calculated, and immunohistochemistry was used to evaluate the contents of NeuN and microtubule-associated protein (MAP-2).. The cell viability of the GBE group was 85% +/- 5%, significantly higher than that of the NMDA group (39.8% +/- 2.1%, P < 0.01), and significantly lower than that of the MK-801 group (93.8% +/- 2.7%, P < 0.05). The LDH efflux of the GBE group was 87 U/L +/- 8 U/L, significantly lower than that of the NMDA group (138 U/L +/- 12 U/L, P < 0.01) and significantly higher than that of the MK-801 group (47 U/L +/- 7 U/L, P < 0.05). The inward current (I(NMDA)) of the NMDA group was significantly activated, The inhibitory rate of the NMDA-activated I(NMDA) of the GBE group was 40% +/- 17%, significantly lower than that of the MK-801 group (78% +/- 18%, P < 0.05); After washing out with standard extracellular solution, the I(NMDA) could recover to 91% +/- 8% in the GBE group, but not in the MK-801 group (P < 0.05), which indicated that GBE had lower affinity to NMDA receptor than MK-801. The Longa's scores of the 3 h and 24 h IR subgroups of the GBS pretreatment group were all significantly lower than those of the corresponding subgroups of the standard MCAO and GBE acute administration groups. The symptoms of the MK-801 were the most severe. Cerebral infarction began to appear in the 1-day subgroups. The cerebral infarction areas of the 1 d subgroups of the GBF pretreatment and MK-801 groups were 11.5% +/- 1.3% and 6.5% +/- 0.9% respectively, both significantly smaller than those of the MCAO and GBE acute administration groups (24.5% +/- % and 22.9% +/- 1.3% respectively, both P < 0.01), however, there was no significant difference in the cerebral infarction area between the GBE acute administration and MCAO group. It was true too for the cerebral infarction areas of the 7 d subgroups. Except in the control group, loss of NeuN positive neuron was seen in all groups, especially the MCAO and GBE acute administration groups. Except in the control group, the MAP-2 positive neurons were decreased in all groups, especially the MCAO and GBE acute administration groups, and 1 day and 7 days after the IR MAP-2 positive neurons were almost unseen in the MCAO and GBE acute administration groups, however, could be seen in small amounts in the GBE and MK-801 groups (all P < 0.01).. GBE pretreatment protects the neurons from excitotoxicity induced by over-activated NMDA receptor and focal cerebral ischemia, which can be explained by the mild blocking effect of GBE on NMDA receptor with low affinity, comparing with MK-801, and GBE is expected to interfere in excitotoxicity in clinic without neurotoxic behaviors.

Topics: Animals; Animals, Newborn; Brain Ischemia; Cell Survival; Cells, Cultured; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Ginkgo biloba; Glycine; Hippocampus; Immunohistochemistry; Male; Membrane Potentials; Microtubule-Associated Proteins; N-Methylaspartate; Nerve Tissue Proteins; Neurons; Nuclear Proteins; Plant Extracts; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Reperfusion Injury

We investigated potential contribution of K+ channel activity to regulation of ischemia-induced neuronal injury, using cerebrocortical slice cultures. Exposure of cultures to a glucose-free conditioning solution containing sodium azide and 2-deoxyglucose caused neuronal cell death as assessed by cellular uptake of propidium iodide, which was prevented by MK-801, an N-methyl-D-aspartate (NMDA) receptor antagonist. Application of tetraethylammonium markedly exacerbated ischemic neuronal injury. Charybdotoxin, a blocker of large-conductance Ca(2+)-activated K+ (BK(Ca)) channels, also augmented ischemic injury, whereas AM 92016, a blocker of delayed rectifier K+ channels, and dequalinium, a blocker of small-conductance Ca(2+)-activated K+ channels, had no significant effect. In addition, tetraethylammonium and charybdotoxin were effective in augmenting NMDA-induced neuronal injury. These results present unprecedented evidence for the ability of tetraethylammonium to enhance ischemic neuronal death, and suggest that BK(Ca) channels constitute an endogenous system to protect cortical neurons from ischemic injury, via prevention of NMDA receptor over-activation.

Topics: Animals; Animals, Newborn; Brain Ischemia; Cell Death; Cerebral Cortex; Charybdotoxin; Dequalinium; Dizocilpine Maleate; Dose-Response Relationship, Drug; N-Methylaspartate; Neurons; Neuroprotective Agents; Organ Culture Techniques; Rats; Rats, Wistar; Sotalol; Tetraethylammonium; Time Factors

Status epilepticus and cerebral ischemia stimulate persistent neurogenesis in the adult brain, but both conditions cause neuronal damage. We determined whether spreading depression, a common epiphenomenon of these conditions, stimulates persistent neurogenesis.. We analyzed the effect of KCl-induced spreading depression on persistent neurogenesis and the spatio-temporal distribution of cells exhibiting immunohistochemical markers for divided and early committed neurons (new neurons) in the adult rat brain.. After induction of spreading depression for 48 hours, the density of mitotic cells, divided cells, and new neurons in the subventricular zone increased at days 1 to 3, days 3 to 6, and day 6, respectively (P<0.05). The divided cell density in the rostral migratory stream and the stream size increased at day 12 (P<0.001). Vehicle (saline) infusion or induction of spreading depression for 4 hours only did not increase the divided cell density, but the latter increased new neuron density in the subventricular zone (P<0.001). Double-labeled new neuron-like cells also appeared in the caudate putamen or cortex in ectopic fashion at day 3, with dramatic increases at days 6 and 12. Administration of the NMDA receptor antagonist, MK-801, which inhibits the propagation of spreading depression, abolished the increase in new neurons in the subventricular zone and the appearance of ectopic new neuron-like cells after 48-hour KCl infusion. There was no neuronal damage, as evidenced by mature neuron density, neurite density, and apoptotic cell appearance after spreading depression for 48 hours.. Spreading depression has the potential to stimulate persistent neurogenesis or to produce ectopic new neuron-like cells.

Topics: Animals; Antimetabolites; Apoptosis; Biomarkers; Brain; Brain Ischemia; Bromodeoxyuridine; Cell Differentiation; Cell Division; Central Nervous System; Cerebral Cortex; Cortical Spreading Depression; Dizocilpine Maleate; Glial Fibrillary Acidic Protein; Immunohistochemistry; In Situ Nick-End Labeling; Membrane Potentials; Microscopy, Confocal; Neurons; Potassium Chloride; Putamen; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Status Epilepticus; Stem Cells; Time Factors

Statins exert beneficial effects in brain diseases including stroke. Here, we investigated whether oral prophylactic atorvastatin provides long-term neuroprotection and functional recovery in permanent middle cerebral artery occlusion (pMCAO), and whether cerebral hemodynamics are affected. Male Long-Evans rats were treated with 10 mg/kg oral atorvastatin for 14 days and subjected to pMCAO. Cerebral hemodynamics were measured by bolus tracking MRI and laser Doppler flowmetry (LDF). Infarct volume was quantified at 1 week by T2-MRI and at 3 weeks by histology. Rats were also subjected to neuroscoring and cylinder test. The number of animals per group was 10. The infarct volumes were 100.8 +/- 8.2 and 47.3 +/- 5.5 mm(3) in vehicle, and 68.7 +/- 11.0 and 28.6 +/- 3.82 mm(3) in atorvastatin group at 7 and 21 days post-ischemia, respectively (mean +/- SEM). Atorvastatin significantly reduced infarct volume both at 7 and 21 days (P = 0.04 and 0.03, respectively, 1-way ANOVA). Interestingly, no improvement in cerebral hemodynamic parameters was observed in atorvastatin treated animals. The vehicle group recovered normal neuroscore at day 13, whereas atorvastatin group recovered already at day 10 after pMCAO. All treatment groups preferred to use the unaffected forelimb for rearing in Cylinder test, whereas the defected forelimb use was minimal in all groups. These results suggest that oral atorvastatin protects cerebral tissue against the subsequent pMCAO without influencing cerebral hemodynamic parameters, and it may well be that persons with ongoing atorvastatin treatment benefit in the incidence of stroke.

Topics: Animals; Atorvastatin; Behavior, Animal; Blood Circulation; Brain Infarction; Brain Ischemia; Dizocilpine Maleate; Heptanoic Acids; Infarction, Middle Cerebral Artery; Magnetic Resonance Imaging; Male; Motor Activity; Neuroprotective Agents; Psychomotor Performance; Pyrroles; Rats; Rats, Long-Evans; Recovery of Function; Staining and Labeling; Time; Time Factors

We have isolated from rat cerebral cortex an endogenous Na(+), K(+)-ATPase inhibitor, termed endobain E, which modulates glutamatergic N-methyl-d-aspartate (NMDA) receptor. This endogenous factor allosterically decreases [(3)H]dizocilpine binding to NMDA receptor, most likely acting as a weak channel blocker. In the present study we investigated whether endobain E is present in the cerebral cortex of rats subjected to ischemia and modulates NMDA receptor exposed to the same conditions. Ischemia-reperfusion was carried out by bilateral occlusion of common carotid arteries followed by a 15-min reperfusion period. Elution profile of brain soluble fraction showed that endobain E is present in cerebral cortex of ischemia-reperfusion rats. On assaying its effect on synaptosomal membrane Na(+), K(+)-ATPase activity and [(3)H]dizocilpine binding to cerebral cortex membranes prepared from animals without treatment, it was found that the endogenous modulator isolated from ischemia-reperfusion rats was able to inhibit both enzyme activity and ligand binding. On the other hand, endobain E prepared from rats without treatment also decreased binding to cerebral cortex or hippocampal membranes obtained from animals exposed to ischemia-reperfusion. Since ischemia decreases tissue pH and NMDA receptor activity varies according to proton concentration, pH influence on endobain E effect was tested. Endobain E ( approximately 80 mg original tissue) decreased [(3)H]dizocilpine binding 25% at pH 7.4 or 8.0 but 90% at pH 6.5. These results demonstrate that endobain E is present and also able to modulate NMDA receptor in the short-term period that follows cerebral ischemia and that its effect depends on proton concentration, suggesting greater NMDA receptor modulation by endobain E at low pH, typical of ischemic tissues.

Topics: Animals; Brain Ischemia; Cerebral Cortex; Disease Models, Animal; Dizocilpine Maleate; Hydrogen-Ion Concentration; Intracellular Membranes; Male; Ouabain; Protein Binding; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Reperfusion Injury; Sodium-Potassium-Exchanging ATPase; Subcellular Fractions

Topics: Acetylcholine; Adenosine A3 Receptor Antagonists; Animals; Brain; Brain Ischemia; Calcium Channels; Central Nervous System; Dihydropyridines; Dizocilpine Maleate; Electrophysiology; Inosine; Ischemia; Neuroprotective Agents; Rats; Receptors, N-Methyl-D-Aspartate

Glutamate is a major excitatory neurotransmitter in the mammalian central nervous system and initiates the events leading to ischemic brain damage. Glutamate receptor antagonists are being used to reduce neuronal damage observed after hypoxia and ischemia. The glutamate receptor antagonist, (+)-5-methyl-10,11-dihydro-5H-dibenzo-(a,d)-cyclohepten-5,10-imine maleate (MK-801) crosses the blood-brain barrier readily and produces a non-competitive use-dependent blockade of the N-methyl-D-aspartate subtype of glutamate receptor. The aim of this study was to investigate effects of MK-801 administered before and just after the onset of ischemia in rats on nitrite and cyclic guanosine monophosphate (cGMP) levels. Focal cerebral ischemia in rats was produced by permanent occlusion of right middle cerebral artery (MCAO). Nitrite and cGMP levels were measured in both cortex and cerebellum at 0, 10, and 60 min following MCAO. The same parameters were measured in rats treated with MK-801 (0.5 mg/kg, i.p.) 30 min before or just after MCAO. Ipsilateral cortical nitrite levels were increased relative to contralateral cortex after MCAO. No significant changes were observed in cerebellum. The cGMP concentrations in both sides of the cortex and cerebellum were increased at 10 and 60 min compared with 0 min values. cGMP level in the ipsilateral cortex was higher than contralateral cortex, whereas the opposite was found for the cerebellum. MK-801 treatment before or just after MCAO decreased significantly nitrite and cGMP production. Our data indicate that MK-801 treatment before or just after focal ischemia prevents the increase in NO and cGMP production.

Topics: Animals; Brain; Brain Ischemia; Cyclic GMP; Disease Models, Animal; Dizocilpine Maleate; Male; Nitrites; Rats; Time Factors

Effects of high and moderate affinity uncompetitive NMDA receptor antagonists (+)MK-801 and memantine on ischemic tolerance were compared in relation to telemetrically controlled brain temperature. The tolerance to an injurious 3 min test of global forebrain ischemia in Mongolian gerbils was induced 48 h earlier by 2 min preconditioning ischemia. Normothermic preconditioning was virtually harmless, and greatly reduced neurodegeneration evoked by test ischemia. In hyperthermic animals it was injurious and failed to induce tolerance. Memantine (5 mg/kg) and (+)MK-801 (3 mg/kg) injected i.p. 1 h before preconditioning did not inhibit ischemic tolerance in the normothermic gerbils, while in hyperthermic animals treated with (+)MK-801 ischemic tolerance was partially restored. Subchronic 3 day infusion of memantine (30 mg/kg/day) significantly decreased neurodegeneration, and preconditioning in the normothermic gerbils further reduced neuronal damage. Hyperthermia exacerbated preconditioning ischemia and in this way reduced expression of tolerance, while (+)MK-801 partially reversed this effect. Our results do not confirm previous reports on the role of NMDA receptors in the induction of ischemic tolerance in gerbils.

Topics: Animals; Body Temperature; Brain; Brain Ischemia; Carotid Arteries; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gerbillinae; Hippocampus; Ischemic Preconditioning; Male; Memantine; Neuroprotective Agents; Receptors, N-Methyl-D-Aspartate; Telemetry

Death-associated protein kinase (DAPK) is a calcium calmodulin-regulated serine/threonine protein kinase involved in ischemic neuronal death. In situ hybridization experiments show that DAPK mRNA expression is up-regulated in brain following a global ischemic insult and down-regulated in ischemic tissues after focal ischemia. DAPK is inactive in normal brain tissues, where it is found in its phosphorylated state and becomes rapidly and persistently dephosphorylated and activated in response to ischemia in vivo. A similar dephosphorylation pattern is detected in primary cortical neurons subjected to oxygen glucose deprivation or N-methyl-D-aspartate (NMDA)-induced toxicity. Both a calcineurin inhibitor, FK506, and a selective NMDA receptor antagonist, MK-801, inhibit the dephosphorylation of DAPK after in vitro ischemia. This indicates that DAPK could be activated by NMDA receptor-mediated calcium flux, activation of calcineurin, and subsequent DAPK dephosphorylation. Moreover, concomitantly to dephosphorylation, DAPK is proteolytically processed by cathepsin after ischemia. Furthermore, a selective DAPK inhibitor is neuroprotective in both in vitro and in vivo ischemic models. These results indicate that DAPK plays a key role in mediating ischemic neuronal injury.

Topics: Animals; Apoptosis Regulatory Proteins; Brain Ischemia; Calcium-Calmodulin-Dependent Protein Kinases; Cells, Cultured; Death-Associated Protein Kinases; Dizocilpine Maleate; Enzyme Activation; Enzyme Inhibitors; Glucose; Male; Oxygen; Phosphorylation; Rats; Rats, Wistar; RNA, Messenger; Tacrolimus

We have investigated in more detail our previous observations on a form of ischaemic pre-conditioning "metabolic adaptation", i.e.--that sequential metabolic insults (hypoxia followed 40 min later by combined hypoxia + hypoglycaemia, or vice versa) are less injurious (monitored by increased [Ca2+]i and decreased PCr) than the immediate combined insult. We have now observed that the "adaptation" occurs between 10 and 20 min. Pre-treatment of the tissues with 10 microM-MK801 showed that it had no effect on the increase in [Ca2+]i caused by the sequential insult and only partially blocked the increase observed by exposure to the immediate combined insult. Exposure to both the delayed and immediate combined insults with low extracellular Ca2+ resulted in a two-fold increase in [Ca2+]i, similar to the increase observed with normal extracellular Ca2+ in the presence of MK801. The results are discussed in terms of the possible origins of the increases in [Ca2+]i.

Topics: Animals; Brain; Brain Ischemia; Calcium; Calcium Channel Blockers; Dizocilpine Maleate; Egtazic Acid; Guinea Pigs; In Vitro Techniques; Ischemic Preconditioning; Magnetic Resonance Imaging; Nifedipine; Receptors, N-Methyl-D-Aspartate

In vitro studies suggested that tissue plasminogen activator (t-PA) may aggravate ischemic injury by enhancing N-methyl-D-aspartate (NMDA) receptor signalling. It remained unclear whether NMDA signalling is also relevant for t-PA toxicity in vivo. We herein examined effects of intravenous t-PA (10 mg/kg), administered alone or in combination with the NMDA antagonist MK-801 (0.2 mg/kg), following 90 min of middle cerebral artery occlusion in mice. In our study, MK-801 alone, administered intraperitoneally, neither affected infarct volume nor brain swelling at 24 h after reperfusion. t-PA significantly increased infarct size, in accordance with previous findings. t-PA-induced ischemic injury was completely abolished and brain swelling markedly reduced when t-PA-treated animals received additional MK-801 injections. To elucidate how t-PA influences brain damage, we examined actions of t-PA on the expression of NO synthases by immunohistochemistry, showing that t-PA does not influence neuronal NO synthase, but increases inducible NO synthase in ischemic areas. The effect of t-PA on inducible NO synthase levels was completely reversed after cotreatment with MK-801. Our study provides in vivo evidence in a model of focal cerebral ischemia that t-PA-induced brain injury involves an NMDA receptor-dependent mechanism.

Topics: Analysis of Variance; Animals; Blood Gas Analysis; Brain Edema; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Drug Interactions; Excitatory Amino Acid Agonists; Immunohistochemistry; Laser-Doppler Flowmetry; Male; Mice; Mice, Inbred C57BL; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type II; Tissue Plasminogen Activator

The spatial memory impairment and expression of apoptotic cells in hippocampal CA1 cells were investigated in rats using single and repeated ischemia models. The neuroprotective and memory-improving effect of YM-90K, an alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionate (AMPA) receptor antagonist, was compared to MK-801, an N-methyl-D-aspartate (NMDA) receptor antagonist. Twice-repeated ischemia, but not single ischemia, impaired the spatial memory and increased expression of apoptotic cells. YM-90K, given before and 6 h after the second reperfusion, significantly improved the memory and reduced the apoptotic cells 7 days after the second reperfusion in repeated ischemia. MK-801 neither improved the spatial memory nor reduced apoptotic cells. The present study showed that delayed expression of apoptotic cells is mediated by mechanisms involving AMPA receptors, but not by NMDA receptor, during the late phase after reperfusion. YM-90K could provide neuroprotective activity and improve the spatial memory impaired by repeated ischemia.

Topics: Animals; Apoptosis; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hippocampus; Male; Maze Learning; Memory Disorders; Nerve Degeneration; Neurons; Neuroprotective Agents; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Recovery of Function; Reperfusion Injury

We investigated levels and compositions of N-acylethanolamines (NAEs) and their precursors, N-acyl phosphatidylethanolamines (N-acyl PEs), in a rat stroke model applying striatal microdialysis for glutamate assay. Rats (n = 18) were treated with either intravenous saline (control), NMDA receptor antagonist MK801 (1 mg/kg), or CB1 receptor antagonist SR141716A (1 mg/kg) 30 min after permanent middle cerebral artery occlusion (MCAO). MK801 significantly attenuated the release of glutamate in the infarcted striatum (79 +/- 22 micromol/L) as compared with controls (322 +/- 104 micromol/L). The administration of CB1 antagonist SR141716A had no statistically significant effect on glutamate release (340 +/- 89 micromol/L), but reduced infarct volume at 5 h after MCAO significantly by approximately 40%, whereas MK801 treatment resulted in a non-significant (18%) reduction of infarct volume. In controls, striatal and cortical NAE concentrations were about 30-fold higher in the infarcted than in the non-infarcted hemisphere, whereas ipsilateral N-acyl phosphatidylethanolamine (N-acyl PE) levels exceeded contralateral levels by only a factor of two to three. Treatment with MK801 or SR141716A, or glutamate release in the infarcted tissue, had no significant effect on these levels. NAE accumulation during acute stroke may be due to increased synthesis as well as decreased degradation, possibly by inhibition of fatty acid amide hydrolase (FAAH).

Topics: Acute Disease; Animals; Arachidonic Acids; Brain Ischemia; Cerebral Cortex; Corpus Striatum; Disease Models, Animal; Dizocilpine Maleate; Endocannabinoids; Ethanolamines; Excitatory Amino Acid Antagonists; Extracellular Fluid; Male; Microdialysis; Phospholipids; Piperidines; Polyunsaturated Alkamides; Pyrazoles; Rats; Rats, Wistar; Receptor, Cannabinoid, CB1; Receptors, N-Methyl-D-Aspartate; Rimonabant; Signal Transduction; Stroke

Focal brain infarcts are surrounded by extended perilesional zones that comprise the partially ischemic penumbra but also completely non-ischemic cortex of the remote ipsilateral hemisphere. To delineate the impact of lesion-associated vs. remote processes on transcriptional programming after focal ischemia, we used cDNA array analysis, quantitative real-time polymerase chain reaction and immunohistochemistry in the photothrombosis model of circumscribed cortical ischemia in rats. At an early stage of 4 h after ischemia, gene induction occurred to a similar extent in the ischemic infarct and remote non-ischemic cortex of the ipsilateral hemisphere. Among the genes induced in non-ischemic cortex we found the NGF-inducible genes PC3, VGF and Arc, the transcriptional regulators I kappa B-alpha and Stat3, and the beta-chemokine MIP-1 alpha (CCL3). At 3 days, the spatial pattern of gene expression had changed dramatically with brain fatty acid-binding protein as the only gene significantly induced in non-ischemic ipsilateral cortex. In contrast, numerous genes were exclusively regulated at the lesion site, comprising genes involved in cell cycle regulation, proteolysis, apoptosis, lipid homeostasis and anti-inflammatory counter-regulation. Cortical spreading depression was identified as the main mechanism underlying gene induction in remote non-ischemic cortex. Our data demonstrate a dynamic spatiotemporal pattern of gene induction, which may contribute to delayed progression of damage or, alternatively, mediate neuroprotection, tissue remodeling and functional compensation.

Topics: AIDS-Related Complex; Animals; Brain Ischemia; Carrier Proteins; Cathepsin K; Cathepsins; Cdc20 Proteins; Cell Cycle Proteins; Cerebral Cortex; Cerebral Infarction; Cortical Spreading Depression; Dizocilpine Maleate; DNA-Binding Proteins; Drug Interactions; Excitatory Amino Acid Antagonists; Fatty Acid-Binding Protein 7; Fatty Acid-Binding Proteins; Functional Laterality; Gene Expression Profiling; Gene Expression Regulation; HSP70 Heat-Shock Proteins; Immunohistochemistry; Male; Nerve Tissue Proteins; Nuclear Receptor Subfamily 4, Group A, Member 1; Oligonucleotide Array Sequence Analysis; Phosphoric Monoester Hydrolases; Potassium Chloride; Proprotein Convertase 1; Rats; Rats, Wistar; Receptors, Cytoplasmic and Nuclear; Receptors, Steroid; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; STAT3 Transcription Factor; Time Factors; Trans-Activators; Transcription Factors; Transcription, Genetic; Transcriptional Activation

Tolerance against ischemia can be induced in the CA1 region of the hippocampus of the brain. In gerbils tolerance evolvement is blocked by the NMDA-antagonist MK-801. To examine this mechanism in rats, MK-801 was administered i.p. 1 h prior to tolerance inducing ischemia. Body temperature and activity were monitored before and after ischemia, and show that MK-801 results in hyperthermia immediately after the injection, the post-ischemic body temperature remain elevated until 5 h post-ischemia in spite of the animals being less active than control animals. Histology shows that pre-treatment with MK-801 does not affect the CA1 neuronal density, and we thus conclude that for the used rat model, MK-801 does not affect development of ischemic tolerance.

Topics: Animals; Brain; Brain Ischemia; Dizocilpine Maleate; Fever; Male; Rats; Rats, Wistar

Characterising the mechanisms of cell death following focal cerebral ischaemia has been hampered by a lack of an in vitro assay emulating both the apoptotic and necrotic features observed in vivo. The present study systematically characterised oxygen-glucose-deprivation (OGD) in primary rat cortical neurones to establish a reproducible model with components of both cell-death endpoints. OGD induced a time-dependent reduction in cell viability, with 80% cell death occurring 24 h after 3 h exposure to 0% O2 and 0.5 mM glucose. Indicative of a necrotic component to OGD-induced cell death, N-methyl-D-aspartate (NMDA) receptor inhibition with MK-801 attenuated neuronal loss by 60%. The lack of protection by the caspase inhibitors DEVD-CHO and z-VAD-fmk suggested that under these conditions neurones did not die by an apoptotic mechanism. Moderating the severity of the insult by decreasing OGD exposure to 60 min did not reduce the amount of necrosis, but did induce a small degree of apoptosis (a slight reduction in cell death was observed in the presence of 10 uM DEVD-CHO). In separate experiments purported to enhance the apoptotic component, cells were gradually deprived of O2, exposed to 4% O2 (as opposed to 0%) during the OGD period, or maintained in serum-containing media throughout. While NMDA receptor antagonism significantly reduced cortical cell death under all conditions, a caspase-inhibitor sensitive component of cell death was not uncovered. These studies suggest that OGD of cultured cortical cells models the excitotoxic, but not the apoptotic component of cell death observed in vivo.

Topics: Amino Acid Chloromethyl Ketones; Animals; Apoptosis; Brain Ischemia; Caspases; Cell Hypoxia; Cell Survival; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glucose; Models, Biological; N-Methylaspartate; Neurons; Oligopeptides; Oxygen; Rats; Rats, Sprague-Dawley; Staurosporine; Time Factors

The excitatory actions of corticotropin-releasing hormone (CRH) in the brain and the neuroprotective effects of CRH antagonists in models of ischemia suggest a role for this peptide in the cascade of events leading to cellular damage. The present study aimed to characterize endogenous activation of CRH in discrete brain regions following global ischemia. Time-dependent changes in CRH concentrations were assessed in 10 brain regions including hippocampal, parahippocampal, and hypothalamic regions as well as the amygdala and the frontal cortex at three post-ischemic intervals: 4, 24, and 72 h (Experiment 1). The impact of pretreatment with a neuroprotective dose of the NMDA antagonist (5R,10S)-(+)-5-Methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate (MK-801; hydrogen maleate) on 24-h ischemia-induced CRH concentrations in the 10 brain regions was also determined (Experiment 2). In vivo microdialysis was used to assess dynamic fluctuations in CRH release at the dorsal hippocampus (CA1 pyramidal layer) and central nucleus of the amygdala (CeA; Experiment 3). Our findings revealed a rapid elevation of CRH concentrations at the piriform cortex (Pir) and hypothalamic nuclei following global ischemia. This was followed by decreased CRH concentrations at the amygdala, the frontal cortex (FC), the CA3, and the hypothalamus 24-h post-ischemia. MK-801 reversed the decreases in the hypothalamic nuclei but not in the other brain regions. Seventy-two hours post-ischemia, CRH levels returned to control values in all regions except the dentate gyrus (DG) where elevated CRH levels were observed. In vivo, a significant increase in CRH release in response to global ischemia was found at the CeA with no alterations at the CA1. These findings support brain region-specific ischemia-induced CRH alterations and suggest that CRH actions to mediate neuronal damage at the hippocampal CA1 layer may be indirect.

Topics: Animals; Brain Chemistry; Brain Ischemia; Corticotropin-Releasing Hormone; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Ischemia; Male; Radioimmunoassay; Rats; Rats, Wistar; Time Factors

Neuroprotective action of MK-801 administrated pre- and postischemically, in vivo or in vitro, respectively, was studied on hippocampal slices using decapitation ischemia model. Recovery of orthodromic population spikes in CA1 region was measured during postischemic incubation of the slices with oxygenated artificial cerebrospinal fluid (ACSF). The ability of postischemically applied MK-801 to restore the electrical activity dramatically depended on the timing of its application during the reoxygenation period. When applied in vitro, together with the start of reoxygenation, MK-801 was as effective as in the case of in vivo administration before the ischemia. The delay in the in vitro administration for only a few minutes led to a dramatic decrease in the drug effectiveness. When applied in 30 min after the start of reoxygenation, MK-801 was totally ineffective. The dose/response relationship between MK-801 concentration and the amplitude of recovered orthodromic population spikes of hippocampal pyramidal neurons is logarithmic. The ED(50) value for the action of "postischemic" MK-801 is ca. 10(-5) M. Preischemic in vivo application of the drug [intraperitoneal (i.p.) injection 15 min prior to decapitation] results in ED(50) ca. 0,2 mg/kg. The slope of both dose/concentration-response curves is similar. The time course of population spike recovery after 90-min ischemia is identical for pre- and postischemic action of MK-801 (estimated for ED(50) in both cases). These data allow to suggest that "preischemic" MK-801 is predominantly active as a neuroprotector only after ischemia, within a short therapeutic window at the start of the reoxygenation period.

Topics: Animals; Brain Ischemia; Dizocilpine Maleate; Dose-Response Relationship, Drug; Functional Laterality; Hippocampus; In Vitro Techniques; Membrane Potentials; Neurons; Neuroprotective Agents; Rats; Rats, Wistar; Time Factors

The polyamine sites on the NMDA receptor complex offer a therapeutic target for focal ischaemia, potentially devoid of most side effects associated with NMDA antagonists. In this study, we investigated the effect of a novel polyamine antagonist, N(1)-dansyl-spermine (0.5-10 mg kg(-1)) in a permanent focal cerebral ischaemia model in mice, and compared its effect to that of MK-801 (0.3-3 mg kg(-1)) following administration 30 min prior to ischaemia. A battery of histological and behavioural tests was employed following permanent middle cerebral artery occlusion to assess any neuroprotective effect. Following middle cerebral artery occlusion, N(1)-dansyl-spermine (1-5 mg kg(-1)) and MK-801 (1 or 3 mg kg(-1)) caused a comparable and significant reduction in the percentage hemisphere lesion volume. Similarly, both drugs significantly reduced oedema and neurological deficit score to a similar extent. Locomotor activity in MCAO mice was not significantly improved by MK-801 or N(1)-dansyl-spermine, although N(1)-dansyl-spermine induced a trend towards significant improvement. Significant improvement in rotarod performance was observed at neuroprotective doses with both drugs. Upon comparison of the profile of effects, N(1)-dansyl-spermine at least matched the effectiveness of MK-801 as a neuroprotective agent in this model. In addition, in sham-operated control mice, N(1)-dansyl-spermine was well tolerated, in contrast to the pronounced adverse effects of MK-801 on locomotor activity and rotarod performance. In conclusion, this study has shown that N(1)-dansyl-spermine is as effective a neuroprotective drug as MK-801 in this model. Moreover, in contrast to MK-801, N(1)-dansyl-spermine could be a promising therapeutic candidate for stroke as it is well tolerated at neuroprotective doses in sham-operated animals.

Topics: Analysis of Variance; Animals; Behavior, Animal; Brain Damage, Chronic; Brain Ischemia; Dansyl Compounds; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Male; Mice; Motor Activity; Neuroprotective Agents; Polyamines; Receptors, N-Methyl-D-Aspartate; Rotarod Performance Test; Spermine

Oxygen and glucose deprivation (OGD) in cell cultures is generally studied in a medium, such as artificial cerebrospinal fluid (CSF), with an ion composition similar to that of the extracellular fluid of the normal brain (2 to 4 mmol/L K+, 2 to 3 mmol/L Ca2+; pH 7.4). Because the distribution of ions across cell membranes dramatically shifts during ischemia, the authors exposed mouse organotypic hippocampal tissue cultures to OGD in a medium, an ischemic cerebrospinal fluid, with an ion composition similar to the extracellular fluid of the brain during ischemia (70 mmol/L K+, 0.3 mmol/L Ca2+; pH 6.8). In ischemic CSF, OGD induced a selective and delayed cell death in the CA1 region, as assessed by propidium iodide uptake. Cell death was glutamate receptor dependent since blockade of the N-methyl-D-aspartate and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors mitigated cell damage. Hyperglycemia aggravates ischemic brain damage whereas glucose in artificial CSF prevents oxygen deprivation-induced damage. The authors demonstrate that glucose in ischemic CSF significantly exacerbates cell damage after oxygen deprivation. This new model of "ischemia" can be useful in future studies of the mechanisms and treatment of ischemic cell death, including studies using genetically modified mice.

Topics: Animals; Brain Ischemia; Calcium; Cerebrospinal Fluid; Culture Techniques; Dizocilpine Maleate; Electrophysiology; Excitatory Amino Acid Antagonists; Glucose; Hippocampus; Hydrogen; Hypoxia; Ions; Mice; Mice, Inbred BALB C; Potassium; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate

Although stroke is a major cause of death and disability in the elderly, the inhibitory effects of neuroprotectants in acute stroke have been investigated using experimental cerebral ischemic models of young animals. Recent clinical trials have found that few neuroprotectants are effective. These observations indicate that effects in the clinical setting do not always reflect data from young animals. Thus, we compared the effects of the NMDA receptor antagonist MK-801 and of the AMPA receptor antagonist NBQX [2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinixaline] on ischemic cerebral damage in the photothrombosis model of aged and young rats. MK-801 administered immediately after MCA occlusion significantly (P<0.05) reduced the extent of cerebral damage in young, but not in aged, rats and the effects of NBQX were similar. In separate experiments, we evaluated brain damage after microinjecting NMDA or kainic acid into the cortex using a stereotaxic apparatus. We found no significant differences in focal cerebral damage caused by NMDA between young and aged rats. On the other hand, kainic acid caused all of the aged rats tested to die, but none of the young rats. Our observations indicate that NMDA and AMPA receptor antagonists are less effective in aged, than in young, rats and that cerebral damage by receptor agonists depends on the type of receptor, such as NMDA and AMPA.

Topics: Aging; Animals; Brain Ischemia; Cerebral Infarction; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Infarction, Middle Cerebral Artery; Kainic Acid; Male; Nerve Degeneration; Neurons; Neuroprotective Agents; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Telencephalon

We have previously shown that exogenous application of brain-derived neurotrophic factor (BDNF) reduces infarct volume in the cortical ischemic penumbra after experimental focal ischemia [Stroke 31 (2000) 2212-2217]. Since BDNF is known to modulate the expression and function of various neurotransmitter receptors, we addressed the question whether BDNF may act via modification of postischemic ligand binding to excitatory NMDA and AMPA and/or inhibitory GABA(A) receptors, respectively. Transient focal cerebral ischemia was induced in male Wistar rats for 2 h using the suture occlusion technique. A period of 30 min after occlusion of the middle cerebral artery, BDNF (300 microg/kg per hour in vehicle; n=5) or vehicle alone (n=5) was continuously infused intravenously for 3 h. Using quantitative receptor autoradiography, postischemic ligand binding of [(3)H]MK-801, [(3)H]AMPA and [(3)H]muscimol was analyzed in the ischemic core, the ischemic cortical penumbra and corresponding regions of the contralateral hemisphere. Transient focal ischemia caused a significant reduction of [(3)H]muscimol binding to GABA(A) receptors within the ischemic cortical penumbra of placebo-treated rats. This was largely prevented by exogenous application of BDNF. [(3)H]MK-801 and [(3)H]AMPA binding values were also reduced in the cortical penumbra and the corresponding area of the contralateral hemisphere. Our data suggest that the neuroprotective effect of BDNF against ischemic damage in the cortical penumbra may be mediated in part by maintained activity of the inhibitory GABAergic system which likely counteracts glutamate induced excitotoxicity.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Binding, Competitive; Brain Ischemia; Brain-Derived Neurotrophic Factor; Cerebral Cortex; Cerebral Infarction; Dizocilpine Maleate; Down-Regulation; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; GABA Agonists; Male; Muscimol; Neurons; Neuroprotective Agents; Rats; Rats, Wistar; Receptors, GABA-A; Receptors, N-Methyl-D-Aspartate; Tritium

A strong linkage between adrenergic and glutamatergic systems exists in the CNS but it is still unclear whether the excessive release of noradrenaline under ischemic conditions is modulated by excitatory amino acids. We studied the effect of selective glutamate receptor antagonists on the release of [3H]noradrenaline evoked by glucose and oxygen deprivation in hippocampal CA1, CA3 and dentate gyrus subregions. The release of glutamate, aspartate and GABA was measured by HPLC. Omission of oxygen and glucose increased the release of [3H]noradrenaline as well as the release of amino acids. Maximum effect on noradrenaline release was observed in CA1 region. The relative increase of the release after 30 min energy deprivation (R(2)) versus the basal release under normal conditions (R(1)), i.e. the R(2)/R(1) ratio was 7.1+/-1.0, 3.87+/-0.4 and 3.26+/-0.27 for CA1, CA3 and dentate gyrus, respectively. The [3H]noradrenaline outflow in response to glucose and oxygen deprivation was abolished at low temperature, but not by Ca(2+) removal, suggesting a cytoplasmic release process. In CA1 and CA3 [3H]noradrenaline release was significantly attenuated by MK-801, an NMDA receptor antagonist. The AMPA receptor antagonist GYKI-53784 had no effect in CA3, but partly reduced noradrenaline release in CA1. Our results suggest that ionotropic glutamate receptors seem to be implicated in the massive cytoplasmic release of noradrenaline in CA1 what may contribute to its selective vulnerability.

Topics: Animals; Benzodiazepines; Brain Ischemia; Chromatography, High Pressure Liquid; Dentate Gyrus; Dizocilpine Maleate; Hippocampus; Kinetics; Male; Norepinephrine; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate

During cerebral ischemia, excess of glutamate release and dysfunction of its high affinity transport induce an accumulation of extracellular glutamate, which plays an important role in neuronal death. The authors studied the relationship among propofol neuroprotection, glutamate extracellular concentrations, and glutamate transporter activity in a model of ischemic cortical cell cultures.. Thirteen-day-old primary cortical neuronal-glial cultures were exposed to a 90-min combined oxygen-glucose deprivation (OGD) in an anaerobic chamber, followed by reoxygenation. Propofol was added only during the OGD period, and its effect was compared to that of the N-methyl-d-aspartate receptor antagonist dizocilpine (MK-801). Twenty-four hours after the injury, cell death was quantified by lactate dehydrogenase release and cell viability by reduction of 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT). Extracellular concentrations of glutamate in culture supernatants and glutamate uptake were performed at the end of OGD period by high-performance liquid chromatography and incorporation of l-[3H]glutamate into cells, respectively.. At clinically relevant concentrations (0.05-10 microm), propofol offered protection equivalent to that of MK-801. It significantly reduced lactate dehydrogenase release and increased the reduction of MTT. At the end of the ischemic injury, propofol was able to reverse the OGD-induced increase in glutamate extracellular concentrations and decrease of glutamate uptake. The inhibition of the glial GLT1 transporter by 3-methyl-glutamate did not further modify the effect of propofol on glutamate uptake, suggesting that GLT1 was not the major target of propofol.. Propofol showed a neuroprotective effect in this in vitro model of OGD, which was apparently mediated by a GLT1-independent restoration of the glutamate uptake impaired during the injury.

Topics: Amino Acid Transport System X-AG; Amino Acids; Anesthetics, Intravenous; Animals; Brain Ischemia; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Extracellular Space; Female; Gangliosides; Glucose; Glutamic Acid; Immunohistochemistry; L-Lactate Dehydrogenase; Microscopy, Phase-Contrast; Neurons; Neuroprotective Agents; Oxygen; Pregnancy; Propofol; Rats; Rats, Wistar; Tetrazolium Salts; Thiazoles

Biochemical changes in rat brain acetylcholinesterase (AChE) activity and molecular isoform distribution were studied using the middle cerebral artery occlusion model. After 24 h of permanent focal ischemia, AChE specific activity from infarcted cortex was found to decrease compared with contralateral cortex. Also a decrease in the minor monomeric (G1) form of AChE is response to ischemia was observed. Ischemia-induced AChE decrease was partially diminished by pre-treatment with dizocilpine (MK-801), a neuroprotective NMDA receptor blocker.

Topics: Acetylcholinesterase; Animals; Brain; Brain Chemistry; Brain Ischemia; Dizocilpine Maleate; Functional Laterality; Male; Neuroprotective Agents; Protein Isoforms; Rats; Rats, Inbred F344

The effect of MK-801 on ischemic neuronal damage was studied in a rat model of permanent focal cerebral ischemia in terms of ipsilateral and contralateral cortical and cerebellar tissue lipid peroxides. Forty-five male Swiss Albino rats were assigned into one of four groups: sham operated as controls, subjected to right middle cerebral artery occlusion (MCAO) or injection of MK-801 (0.5 mg/kg i.p.) either 30 min before or just after right MCAO. Changes in lipid peroxides were expressed as nmol malondialdehyde (MDA) and conjugated diene (CD)/mg protein. MDA values after 60 min of ischemia relative to contralateral cortex and CD levels in 0, 10 and 60 min after ischemia were found to be higher in ipsilateral cortex than those in contralateral cortex. On the other hand, contralateral cerebellar MDA levels in 0 and 60 min of ischemia and CD levels in 0, 10 and 60 min after ischemia were found to be higher than those in ipsilateral cerebellum. Pharmacological inhibition of glutamate receptor by MK-801 before or just after permanent MCAO decreased significantly the MDA and CD levels in both cortex and cerebellum. Although no significant differences found in MDA values between rats pre- and posttreated with MK-801, CD levels in posttreated group seemed significantly lower than those in pretreated group. On the whole, these results suggest that MDA and CD represent early biochemical marker of lipid peroxidation in ischemic tissues, reflecting the radical-mediated tissue damage.

Topics: Alkenes; Animals; Brain Chemistry; Brain Ischemia; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Lipid Peroxidation; Male; Malondialdehyde; Rats; Receptors, N-Methyl-D-Aspartate

Cellular homeostatic adaptation to cerebral ischemia is complex and contains changes in receptor mediated gene expression and signaling pathways. The proteins of the immediate early genes c-Fos and c-Jun are thought to be involved in coupling neuronal excitation to target gene expression, due to formation of heterodimers and binding to the AP1 promotor region. We used an in vitro model to compare ischemia induced c-Fos and c-Jun expression in rat neuronal cell cultures and nerve growth factor (NGF) differentiated PC 12 cells. Since activation of glutamate receptors is known to mediate ischemic injury we determined the effect of the noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist MK 801 on c-Fos and c-Jun expression in both cell culture systems during ischemia. Neuron rich cultures and NGF differentiated PC 12 cells were exposed to sublethal in vitro ischemia using an hypoxic chamber flushed with argon/CO2 (95 %/5%). C-Fos and c-Jun mRNA expression was analyzed by competitive reverse transcription-polymerase chain reaction using glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as internal standard. One hour of in vitro ischemia significantly increased c-Fos and c-Jun mRNA levels in both cell culture systems. In neuron rich cultures a 10-fold (c-Fos) and 7-fold (c-Jun) mRNA increase was observed. The mRNA rise was less pronounced in PC 12 cells (5.5-fold and 2-fold) for c-Fos and c-Jun, respectively. The addition of MK 801 significantly reduced the expression of c-Fos and c-Jun mRNA in neuronal cultures, whereas no effect was detectable in PC 12 cells. Since MK 801 failed to reduce the c-Fos and c-Jun expression in NGF differentiated PC 12 cells different signaling pathways may initiate c-Fos and c-Jun expression in both cell culture systems.

Topics: Animals; Brain Ischemia; Cell Death; Dizocilpine Maleate; Down-Regulation; Excitatory Amino Acid Antagonists; Fetus; Gene Expression Regulation; In Vitro Techniques; Neurons; PC12 Cells; Proto-Oncogene Proteins c-fos; Proto-Oncogene Proteins c-jun; Rats; Rats, Inbred Lew; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Signal Transduction; Up-Regulation

The influence of 20 min global cerebral ischemia on the free arachidonic acid (FAA) level and Na+,K+-ATPase activity in the rat hippocampus at different time points after ischemia was examined. In addition, the effect of MK-801 on mentioned parameters was studied. Animals were exposed to 20 min global cerebral ischemia and were sacrificed immediately, 0.5, 1, 2, 6, 24, 48, 72, and 168 h after ischemic procedure. The level of the FAA and the Na+,K+-ATPase activity was measured during all reperfusion periods examined. Various doses of MK-801 (0.3, 1.0, 3.0, and 5.0 mg/kg) had been injected 30 min before ischemic procedure started. It was found that 20 min global cerebral ischemia induces a statistically significant increase of the FAA level immediately after ischemia and during the first 0.5 h of reperfusion. After a transient decrease, the level of FAA level increased again after 24 and 168 h of recirculation. Treatment with 3.0 mg/kg of MK-801 significantly prevented the FAA accumulation immediately and 0.5 h after ischemic insult while application of 5.0 mg/kg of MK-801 exerted a protective effect during the first 24 h. Global cerebral ischemia induces the significant decline in the Na+,K+-ATPase activity in the hippocampus starting from 1 to 168 h of reperfusion. Maximal inhibition was obtained 24 h after the ischemic damage. Application of 3.0 mg/kg of MK-801 exerted statistically significant protection during the first 24 h while the treatment with 5.0 mg/kg of MK-801 prevented fall in enzymatic activity during all reperfusion periods examined. Our results suggest that, in spite of different and complex pathophysiological mechanisms involved in the increase of FAA level and the decrease of the Na+,K+-ATPase activity, blockade of NMDA receptor subtype provides a very important strategy for the treatment of the postischemic excitotoxicity.

Topics: Animals; Arachidonic Acid; Brain Ischemia; Dizocilpine Maleate; Hippocampus; Neuroprotective Agents; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Sodium-Potassium-Exchanging ATPase

Cerebral ischemia activates ERK and Akt pathways. We studied whether these activations were affected by treatment with the protective growth factor transforming growth factor-alpha (TGF-alpha), and whether they were mediated through N-methyl D-aspartate (NMDA) receptors. The middle cerebral artery was occluded in rats and signaling was studied 1 h later. Noncompetitive NMDA receptor antagonist MK-801 was injected i.p. before the occlusion, whereas in other rats TGF-alpha was given intraventricularly before and after occlusion. Ischemia caused ERK phosphorylation in the nucleus, localized in the endothelium and neurons. Phosphorylation of ERK was prevented by TGF-alpha, but it was enhanced in the nucleus and cytoplasm by MK-801. Also, MK-801 but not TGF-alpha increased p-Akt. Results suggest that preventing ERK activation is related to the protective effect of TGF-alpha, whereas the protective effect of MK-801 is associated with activation of pro-survival Akt. While results support that NMDA receptor signaling precludes Akt activation, we did not find evidence to support that it underlies ischemia-induced ERK phosphorylation. This study illustrates that neuroprotection results from a fine balance between death and survival signaling pathways.

Topics: Animals; Blotting, Western; Brain Ischemia; Densitometry; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Immunohistochemistry; Male; Mitogen-Activated Protein Kinases; Neuroprotective Agents; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Signal Transduction; Transforming Growth Factor alpha

To investigate whether the protective effect of therapy with different combined neuroprotectant agents was better than that of single agent on focal cerebral ischemia.. The right middle cerebral artery in the rats was occluded with suture occlusion technique. The rats were divided into five groups treated with FDP (50 mg/kg, n = 10), MK-801 (1 mg/kg, n = 10) and NAC (150 mg/kg, n = 10) singly, or in combination, respectively, by intraperitoneal infusion 30 minutes after vessel occlusion. The rats were weighed and assessed neurologically, based on a 5-point scale, six and 24 hours after focal cerebral ischemia. The expression of anti-apoptotic protein bcl-2 was observed with SDS-PAGE protein electrophoresis and Western blot technique.. The optical density of bcl-2 increased more distinctly in the rats treated with combined neuroprotective agents than that with any single agent six and 24 hours after cerebral ischemia, with a statistically significant difference (P < 0.05).. Treatment with combined neuroprotectant agents could un-regulate the anti-apoptotic protein bcl-2 more distinctly than that with any single agents. Combined use of neuroprotectants might be more effective than that of single agent in protecting rats' brain from ischemia.

Topics: Acetylcysteine; Actins; Animals; Brain Ischemia; Dizocilpine Maleate; Drug Therapy, Combination; Fructosediphosphates; Male; Molecular Weight; Neuroprotective Agents; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Wistar

Computer-assisted procedures are used to measure infarct areas in animal stroke models, but this approach usually follows the less objective manual tracing of the boundaries of the infarct. Building on previously reported methodology using scanned images of triphenyltetrazolium chloride (TTC)-stained rat brains in the intraluminal thread model, we developed an objective method to assess ischemic damage in both the mouse and rat brains. The unique addition to our approach is the use of sham-treated animals, which thereby permits the removal of normal brain white matter from the ipsilateral injured brain. All brain sections per animal were scanned simultaneously using a Microtek Scanmaker 4 flatbed scanner. Color segmentation on full color images of 2 mm coronal brain sections was performed. Using Image Pro Plus (4.0) and color segmentation, ischemic and normal white matter areas were measured in the green channel and the entire brain area in the red channel. The percent of unstained tissue was calculated for sham-treated animals and for those with cerebral ischemia. By subtracting the average unstained area of the sham-treated group from the average unstained area from the ischemic group, the ischemic area was calculated. This methodology was validated using mouse and rat permanent and transient, focal ischemia models and MK-801 in the permanent ischemia models. MK-801, dosed at 3 mg/kg i.p. prior to the injury, reduced the injury by 75% in the mouse and 44% in the rat permanent occlusion models. The benefits of this methodology include: objectivity of the analysis of the ischemic injury, use of readily available software so that costs can be contained and removal of normal subcortical white matter from the calculation. This method should allow more consistent evaluation of changes in the infarct size, therefore, resulting in reduced variability and higher productivity.

Topics: Animals; Brain Infarction; Brain Ischemia; Carotid Arteries; Coloring Agents; Diagnostic Imaging; Disease Models, Animal; Dizocilpine Maleate; Image Processing, Computer-Assisted; Ischemic Attack, Transient; Laser-Doppler Flowmetry; Male; Mice; Mice, Inbred C57BL; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Stroke

General anesthetics reduce neuronal death caused by focal cerebral ischemia in rodents and by in vitro ischemia in cultured neurons and brain slices. However, in intact animals, the protective effect may enhance neuronal survival for only several days after an ischemic injury, possibly because anesthetics prevent acute but not delayed cell death. To further understand the mechanisms and limitations of volatile anesthetic neuroprotection, the authors developed a rat hippocampal slice culture model of cerebral ischemia that permits assessment of death and survival of neurons for at least 2 weeks after simulated ischemia.. Survival of CA1, CA3, and dentate gyrus neurons in cultured hippocampal slices (organotypic slice culture) was examined 2-14 days after 45 min of combined oxygen-glucose deprivation at 37 degrees C (OGD). Delayed cell death was serially measured in each slice by quantifying the binding of propidium iodide to DNA with fluorescence microscopy.. Neuronal death was greatest in the CA1 region, with maximal death occurring 3-5 days after OGD. In CA1, cell death was 80 +/- 18% (mean +/- SD) 3 days after OGD and was 80-100% after 1 week. Death of 70 +/- 16% of CA3 neurons and 48 +/- 28% of dentate gyrus neurons occurred by the third day after OGD. Both isoflurane (1%) and the N-methyl-D-aspartate antagonist MK-801 (10 microm) reduced cell death to levels similar to controls (no OGD) for 14 days after the injury. Isoflurane also reduced cell death in CA1 and CA3 caused by application of 100 but not 500 microm glutamate. Cellular viability (calcein fluorescence) and morphology were preserved in isoflurane-protected neurons.. In an in vitro model of simulated ischemia, 1% isoflurane is of similar potency to 10 microm MK-801 in preventing delayed cell death. Modulation of glutamate excitotoxicity may contribute to the protective mechanism.

Topics: Anesthetics, Inhalation; Animals; Brain Ischemia; Cell Death; Cell Survival; Dizocilpine Maleate; Glutamic Acid; In Vitro Techniques; Isoflurane; Neuroprotective Agents; Rats; Rats, Sprague-Dawley

Behavioral and morphological changes were examined for up to 9 days after moderate cerebral ischemia caused by slow compression of a specific brain area in the sensorimotor cortex of Sprague-Dawley rats. Functional deficits after the cerebral ischemia were assessed by daily beam-walking tests, whereas morphological changes were verified using Nissl staining on day 1, 2, 3, 5, and 9, respectively. Rats exposed to cerebral ischemia displayed impaired beam walking performance. Mild hypothermia prevented both the compression-produced functional deficits and the brain damage. Younger (5 weeks) animals showed less neurological deficits than older (9 weeks) animals. Histological examination revealed a pronounced increase in the number of injured pyramidal neurons from day 1 to day 3 in the primarily damaged brain region. Between day 3 and day 5, the number of injured cells remained constant, whereafter there was a slow decline of thionin-positive neurons as examined on day 9. The noncompetitive NMDA receptor antagonist, dizocilpine (MK-801; 3 mg/kg, i.p.), did not alter the neurological impairment on day 1, but improved thereafter the rate of functional recovery and reduced the number of damaged cells. The AMPA receptor antagonist, LY326325 (15 or 30 mg/kg; i.p.), dose-dependently diminished the neurological deficits on day 1, enhanced the rate of recovery, and reduced the number of injured neurons over time. Our data suggest that short-lasting extradural compression of a well-defined brain area in the sensorimotor cortex is a highly reproducible model with a high success rate for the study of functional and morphological consequences after cerebral ischemia as well as for the evaluation of the therapeutic potential of novel, neuroprotective pharmacological agents.

Topics: Animals; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Dura Mater; Excitatory Amino Acid Antagonists; Hypothermia, Induced; Isoquinolines; Male; Motor Activity; Motor Cortex; Neuroprotective Agents; Postural Balance; Pressure; Rats; Rats, Sprague-Dawley; Receptors, Glutamate; Recovery of Function; Somatosensory Cortex; Tetrazoles

These studies have addressed the role of caspase-3 activation in neuronal death after cerebral ischemia in different animal models. The authors were unable to show activation of procaspase-3 measured as an induction of DEVDase (Asp-Glu-Val-Asp) activity after focal or transient forebrain ischemia in rats. DEVDase activity could not be induced in the cytosolic fraction of the brain tissue obtained from these animals by exogenous cytochrome c/dATP and Ca2+. However, the addition of granzyme B to these cytosolic fractions resulted in a significant activation of DEVDase, confirming that the conditions were permissive to analyze proteolytic cleavage of the DEVD-AMC (7-amino-4-methyl-coumarin) substrate. Consistent with these findings, zVal-Ala-Asp-fluoromethylketone administered after focal ischemia did not have a neuroprotective effect. In contrast to these findings, a large increase in DEVDase activity was detected in a model of hypoxic-ischemia in postnatal-day-7 rats. Furthermore, in postnatal-day-7 animals treated with MK-801, in which it has been suggested that excessive apoptosis is induced, the authors were unable to detect activation of DEVDase activity but were able to induce it in vitro by the addition of cytochrome c/dATP and Ca2+ to the cytosolic fraction. Analysis of cytochrome c distribution did not provide definitive evidence for selective cytochrome c release in the permanent focal ischemia model, whereas in the transient model a small but consistent amount of cytochrome c was found in the cytosolic fraction. However, in both models the majority of cytochrome c remained associated with the mitochondrial fraction. In conclusion, the authors were unable to substantiate a role of mitochondrially derived cytochrome c and procaspase-3 activation in ischemia-induced cell death in adult brain, but did see a clear induction of caspase-3 in neonatal hypoxia.

Topics: Amino Acid Chloromethyl Ketones; Animals; Animals, Newborn; Brain; Brain Ischemia; Caspase 3; Caspases; Cell Death; Cytochrome c Group; Dizocilpine Maleate; Enzyme Activation; Granzymes; Humans; Infarction, Middle Cerebral Artery; Male; Mice; Mice, Inbred C57BL; Neurons; Neuroprotective Agents; Peptide Hydrolases; Protein Precursors; Rats; Rats, Sprague-Dawley; Rats, Wistar; Serine Endopeptidases

The single-cell gel electrophoresis (comet assay) was used to evaluate the possibility of detecting single-strand breaks of brain DNA in the early phase of ischemia. Four hours after occlusion of the middle cerebral artery (MCAO) in rats, the percentage of DNA migrating into the comet tail (indicating the presence of breaks) increased from 11.4 +/- 4.70 to 34.7 +/- 9.2 (means +/- SD) in the caudate and from 9.9 +/- 4.3 to 42.8 +/- 14.1 in the cortex. Interestingly, a subpopulation of cells exhibiting higher resistance to the ischemic insult was present in the caudate putamen, but not in the cortex. Administration of MK801, an N-methyl-d-aspartate (NMDA) glutamate receptor antagonist, (1 mg/kg subcutaneously, 10 minutes before MCAO), reduced the ischemia-induced DNA breaks and the infarct volume, suggesting that excessive stimulation of NMDA receptors contributes to the formation of both DNA damage and infarct volume. In contrast, DPQ, an inhibitor of poly(ADP-ribose) polymerase (PARP) (10 mg/kg intraperitoneally, 2 hours before and 1 hour after MCAO), reduced the infarct volume but not DNA damage, suggesting that the neuroprotective actions of PARP inhibitors occur at a later step of the processes leading to postischemic neuronal death.

Topics: Animals; Brain Ischemia; Comet Assay; Dizocilpine Maleate; DNA; DNA Damage; DNA, Single-Stranded; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Infarction, Middle Cerebral Artery; Isoquinolines; Male; Neuroprotective Agents; Piperidines; Poly(ADP-ribose) Polymerase Inhibitors; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

Excitotoxic activation of glutamate receptors is currently thought to play a pivotal role in delayed neuronal death (DND) of highly vulnerable CA1 neurons in the gerbil hippocampus after transient global ischemia. Postischemic degeneration of these neurons can be prevented by "preconditioning" with a short sublethal ischemic stimulus. The present study was designed to test whether ischemic preconditioning is associated with specific alterations of ligand binding to excitatory glutamate and/or inhibitory gamma-aminobutyric acid (GABA)A receptors compared with ischemia severe enough to induce DND.. With the use of quantitative receptor autoradiography, postischemic ligand binding of [3H]MK-801 and [3H]alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) to excitatory N-methyl-D-aspartate (NMDA) and AMPA receptors as well as [3H]muscimol to inhibitory GABA(A) receptors in hippocampal subfields CA1, CA3, and the dentate gyrus were analyzed in 2 experimental paradigms. Gerbils were subjected to (1) a 5-minute ischemic period resulting in DND of CA1 neurons and (2) a 2.5-minute period of ischemia mediating tolerance induction.. [3H]MK-801 and [3H]AMPA binding values to excitatory NMDA and AMPA receptors showed a delayed decrease in relatively ischemia-resistant CA3 and dentate gyrus despite maintained neuronal cell density. [3H]Muscimol binding to GABA(A) receptors in CA1 neurons was transiently but significantly increased after preconditioning but not after global ischemia with consecutive neuronal death.. Downregulation of ligand binding to glutamate receptors in relatively ischemia-resistant CA3 and dentate gyrus neurons destined to survive suggests marked synaptic reorganization processes despite maintained structural integrity. More importantly, upregulation of binding to inhibitory GABA(A) receptors in the hippocampus indicates a relative shift between inhibitory and excitatory neurotransmission that we suggest may participate in endogenous postischemic neuroprotection.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Autoradiography; Binding, Competitive; Brain Ischemia; Cell Count; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Gerbillinae; Hippocampus; Immunohistochemistry; Ischemic Preconditioning; Ligands; Male; Muscimol; Neurons; Receptors, GABA-A; Tritium; Up-Regulation

Harmful effects of peri-infarct depolarizations (PIDs) may depend on recurrent Ca(2+) influx. Thus far, few studies have documented the relevance of PIDs in gyrencephalic animals, and the progressive nature of this process has not been investigated over extended periods. We therefore studied in prolonged focal ischemia in cats spatial and temporal profiles of extracellular calcium ([Ca(2+)](o)) shifts in relation to direct current (DC) potential, nitric oxide (NO) concentration and regional cerebral blood flow alterations, and final pathological outcome.. In halothane-anesthetized cats receiving either vehicle (n=12) or MK-801 treatment (5 mg/kg IV; n=10), the left middle cerebral artery was permanently occluded. Laser-Doppler probes, ion-selective microelectrodes, and NO electrodes measured simultaneously regional cerebral blood flow, DC potential, electrocorticogram, [Ca(2+)](o), and NO concentrations in ectosylvian and suprasylvian gyri of the left cerebral cortex.. Persistent depolarization immediately after middle cerebral artery occlusion occurred in 10 ectosylvian and 4 suprasylvian gyri of vehicle-treated animals and in 9 ectosylvian and 3 suprasylvian gyri of MK-801-treated animals. PIDs associated with transient decreases of [Ca(2+)](o) were detected in suprasylvian gyri of only 4 vehicle-treated animals, of which 3 developed recurrent PIDs. Electrocorticogram was suppressed during PIDs, and electrocorticogram recovery worsened in a stepwise manner with consecutive depolarizations. PID duration increased slightly with ongoing ischemia and evolved to persistent depolarization at a final stage. NO transients were not detected during PID, and regional cerebral blood flow transients were not pronounced. Infarction was larger with initial persistent depolarization than with PID and was smallest in MK-801-treated animals.. PID is not a common finding in peri-infarct zones in cats, and it is suppressed by the N:-methyl-D-aspartate antagonist MK-801. However, if repeated PIDs are generated, they result in a stepwise, progressive breakdown of neuronal function and ion homeostasis, probably contributing to the growth of infarction in focal cerebral ischemia. Recurrent Ca(2+) influx is a mechanism that presumably contributes to this process.

Topics: Animals; Blood Flow Velocity; Brain Ischemia; Calcium; Cats; Cerebral Cortex; Cerebrovascular Circulation; Cortical Spreading Depression; Disease Models, Animal; Disease Progression; Dizocilpine Maleate; Electroencephalography; Female; Homeostasis; Infarction, Middle Cerebral Artery; Ion Transport; Laser-Doppler Flowmetry; Male; Membrane Potentials; Microelectrodes; Neuroprotective Agents; Nitric Oxide

The aim of this study is to determine experimentally whether N-methyl-D-aspartate (NMDA) receptor is involved in nitrotyrosine formation in rat brain subjected to focal ischemia-reperfusion, by using the NMDA receptor antagonist MK-801. Halothane-anesthetized and artificially ventilated rats were given MK-801 (5 mg/kg, i.p.) or vehicle prior to 2 h of focal cerebral ischemia followed by 0.5 h of reperfusion. The brain was then removed and divided into four sections, cortical ischemic core, peri-ischemic cortex, lateral caudate-putamen and non-ischemic cortex. Tissue nitrotyrosine was measured by means of hydrolysis/HPLC. MK-801 significantly attenuated nitrotyrosine formation in the lateral caudate-putamen. We conclude that nitrotyrosine formation required activation of NMDA receptors, at least in part.

Topics: Animals; Brain Ischemia; Calcium Channels; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Male; Neostriatum; Neuroprotective Agents; Nitric Oxide; Nitric Oxide Synthase; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Reperfusion Injury; Tyrosine

Levetiracetam (LEV) is a new antiepileptic drug with a promising preclinical profile involving both anticonvulsant and antiepileptogenic effects in kindling models. The latter stimulated the present study to compare its neuroprotective properties with the potent and selective, non-competitive NMDA antagonist, MK-801, in the rat middle cerebral artery occlusion model. Twenty-four hours after a transient occlusion of 90 minutes the animals were sacrificed and infarct volume and lesion distribution were determined from stained coronal sections. LEV was administered by intraperitoneal (i.p.) bolus injections of 5.5, 11, 22 and 44 mg x kg(-1), 30 minutes before occlusion followed by a continuous 24 hour i.p. infusion of 1.25, 2.6, 5.1 and 10.2 mg x kg(-1) per hour, respectively. LEV administration did not alter body temperature but reduced the infarct volume by 33% (P< 0.05) at the highest dose tested. An i.p. bolus injection of 0.04, 0.12 and 0.4 mg x kg(-1) of MK-801 followed by continuous i.p. infusion of 0.036, 0.108 and 0.36 mg x kg(-1) per hour, reduced the infarct volume by 49, 51 and 74% (P< 0.05), respectively. However, only the highest dose of MK-801 induced a significant reduction in the infarct volume (P< 0.05) and this was associated with hypothermia. These results suggest that LEV possesses neuroprotective properties which may be relevant for its antiepileptogenic action.

Topics: Animals; Anticonvulsants; Arterial Occlusive Diseases; Body Temperature; Brain; Brain Ischemia; Cerebrovascular Circulation; Disease Models, Animal; Dizocilpine Maleate; Levetiracetam; Male; Middle Cerebral Artery; N-Methylaspartate; Neuroprotective Agents; Piracetam; Rats; Rats, Wistar

1. The aim of this work was to evaluate the role of leukotrienes in brain damage in vivo in a model of focal cerebral ischaemia in the rat, obtained by permanent occlusion of middle cerebral artery. 2. A significant (P < 0.01) elevation of LTC(4), LTD(4) and LTE(4) (cysteinyl-leukotrienes) levels occurred 4 h after ischaemia induction in the ipsilateral cortices of ischaemic compared to sham-operated animals (3998 +/- 475 and 897 +/- 170 fmol g(-1) tissue, respectively, P < 0.01). 3. The NMDA receptor antagonist MK-801 and the adenosine A(2A) receptor antagonist SCH 58261 were administered in vivo at doses known to reduce infarct size and compared with the leukotriene biosynthesis inhibitor MK-886. 4. MK-886 (0.3 and 2 mg kg(-1) i.v.) and MK-801 (3 mg kg(-1) i.p.) decreased cysteinyl-leukotriene levels (-78%, P < 0.05; -100%, P < 0.01; -92%, P < 0.01, respectively) 4 h after permanent occlusion of the middle cerebral artery, whereas SCH 58261 (0.01 mg kg(-1) i.v.) had no significant effects. 5. MK-886 (2 mg kg(-1) i.v.) was also able to significantly reduce the cortical infarct size by 30% (P < 0.05). 6. We conclude that cysteinyl-leukotriene formation is associated with NMDA receptor activation, and that it represents a neurotoxic event, the inhibition of which is able to reduce brain infarct area in a focal ischaemic event.

Topics: Animals; Brain Ischemia; Cerebral Cortex; Cysteine; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Indoles; Leukotrienes; Lipoxygenase Inhibitors; Male; Neuroprotective Agents; Purinergic P1 Receptor Antagonists; Pyrimidines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Receptors, Purinergic P1; Triazoles

Effects of intrauterine hypoxia-ischemia (HI) on expression of the NMDA receptor subunits as well as on [3H]MK-801 binding of the NMDA receptor were studied in 1-day to 30-day old rat brain. Intrauterine HI conditions were achieved on gestation day 17 by clamping the uterine vasculature for 30 min followed by removal of the clamps to permit reperfusion. As determined by reverse-transcriptase polymerase chain reaction, prenatal HI significantly reduced mRNA expression of the NRI subunit of the NMDA receptor in the hippocampus of 4, 8, and 30-day old rat brains. NR2A and NR2B subunit mRNAs were expressed in the hippocampus and the cortex of both the control and the prenatal HI rat brains. Intrauterine HI did not significantly affect expression of either the NR2A or NR2B subunit mRNA. Consistent with the RT-PCR data, protein expression of the NRI subunit in the hippocampus, but not the cortex, of 21-day old prenatal HI rat brains was significantly decreased as compared to the control rat brain. Intrauterine HI also significantly reduced binding affinity, but not the number of binding sites, of the NMDA receptor to [3H]MK-801, a noncompetitive antagonist of the NMDA receptor, in the hippocampus of 21-day old rat brain. The overall results suggest that prenatal HI-induced reduction of NRI expression and the altered binding ability of the NMDA receptor in the young rat brain may contribute to other long-lasting effects of intrauterine HI that we reported previously.

Topics: Aging; Animals; Animals, Newborn; Brain; Brain Ischemia; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Fetal Diseases; Hypoxia; Pregnancy; Prenatal Exposure Delayed Effects; Rats; Receptors, N-Methyl-D-Aspartate; RNA, Messenger

The objective of this study was to investigate whether hypothermia would suppress secondary damage in the chronic postischemic stage, in terms of glutamate excitotoxicity.. Gerbils underwent 5 minutes of ischemia via bilateral common carotid artery occlusion. Seven groups were studied, as follows: 1) ischemia without treatment group; 2) intraischemic hypothermia group; 3) postischemic hypothermia group (32 degrees C for 4 h); 4) MK-801 treatment group (2 mg/kg, every other day for 1 mo); 5) postischemic hypothermia with MK-801 treatment for 1 week group (2 mg/kg, every other day); 6) postischemic hypothermia with MK-801 treatment for 1 month group (2 mg/kg, every other day); and 7) sham-treated control group. One month after ischemia, histological changes in hippocampal CA1 neurons (assessed using hematoxylin and eosin staining) and memory function (assessed using an eight-arm radial maze) were studied. Extracellular glutamate concentrations were monitored by microdialysis during ischemia and hypothermia. Staining of microglia was performed 1 week and 1 month after ischemia.. MK-801 alone, postischemic hypothermia alone, and postischemic hypothermia with MK-801 treatment for 1 week failed to prevent ischemic neuronal damage and memory function decreases 1 month after the insult (P < 0.05 versus control). However, the postischemic hypothermia with MK-801 treatment for 1 month group exhibited significant protective effects (not significant [P > 0.05] compared with the control group). Extracellular glutamate levels for the intraischemic hypothermia group were significantly low, compared with the postischemic hypothermia group. There was no microglial activation in the postischemic hypothermia at 1 week and 1 month after ischemia groups.. Postischemic hypothermia and long-term intermittent administration of MK-801 demonstrated significant neuronal protection, indicating that long-term glutamatergic activation, with changes in N-methyl-D-aspartate receptors, plays a role in neuronal damage in the chronic postischemic stage.

Topics: Animals; Brain; Brain Ischemia; Carotid Stenosis; Dizocilpine Maleate; Extracellular Space; Gerbillinae; Glutamates; Hippocampus; Hypothermia; Male; Maze Learning; Memory Disorders; Neuroprotective Agents; Receptors, N-Methyl-D-Aspartate; Space Perception

The effects of citicoline and/or low dose of MK-801 (sufficient to prevent the development of seizures) on survival, neurological and behavioral recovery following transient hyperglycemic-oligemic-hypoxic insult have been evaluated in mice. Neurological recovery was assessed semi-quantitatively on the third and the 10th day after the insult, and behavioral tests evaluating spontaneous locomotor activity, motor coordination and spontaneous alternation performance were performed on day 10. Neither drug given alone did influence survival rate, but the combination of MK-801 and higher citicoline dose decreased mortality on day 10. Behavioral performance was markedly compromised by the insult. Citicoline, but not MK-801, slightly but significantly improved behavioral outcome in all three tests.. when brain ischemic insult is complicated with acute hyperglycemia, post-treatment with citicoline combined with MK-801 in low anti-convulsive dose improves survival and neurological recovery, and citicoline but not MK-801 enhances behavioral recovery.

Topics: Animals; Behavior, Animal; Brain Ischemia; Cytidine Diphosphate Choline; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hyperglycemia; Hypoxia; Male; Mice; Motor Activity; Nervous System

This study characterized the neuroprotective and behavioral effects of (3aS,6aS)-6a-naphtalen-2-ylmethyl-5-methyliden-hexahydro-cyclopenta[c]furan-1-on (BAY 36-7620), a novel, selective and systemically active metabotropic glutamate (mGlu)(1) receptor antagonist. In the rat, neuroprotective effects were obtained in the acute subdural hematoma model (efficacy of 40-50% at 0.01 and 0.03 mg/kg/h, i.v. infusion during the 4 h following surgery); whereas in the middle cerebral artery occlusion model, a trend for a neuroprotective effect was obtained after triple i.v. bolus application of 0.03-3 mg/kg, given immediately, 2 and 4 h after occlusion. Hypothermic effects were mild and only obtained at doses which were considerably higher than those at which maximal neuroprotective efficacy was obtained, indicating that the neuroprotective effects are not a consequence of hypothermia. BAY 36-7620 protected against pentylenetetrazole-induced convulsions in the mouse (MED: 10 mg/kg, i.v.). As assessed in rats, BAY 36-7620 was devoid of the typical side-effects of the ionotropic glutamate (iGlu) receptor antagonists phencyclidine and (+)-5-methyl-10,11-dihydroxy-5H-dibenzo(a,d)cyclohepten-5,10-imine (MK-801). Thus, BAY 36-7620 did not disrupt sensorimotor gating, induce phencyclidine-like discriminative effects or stereotypical behavior, or facilitate intracranial self-stimulation behavior. Although behavioral stereotypies and disruption of sensorimotor gating induced by amphetamine or apomorphine were not affected by BAY 36-7620, the compound attenuated some behavioral effects of iGlu receptor antagonists, such as excessive grooming or licking, and their facilitation of intracranial self-stimulation behavior. It is concluded that mGlu(1) receptor antagonism results in neuroprotective and anticonvulsive effects in the absence of the typical side-effects resulting from antagonism of iGlu receptors.

Topics: Animals; Anticonvulsants; Behavior, Animal; Body Temperature; Brain Ischemia; Cerebral Cortex; Discrimination, Psychological; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Exploratory Behavior; Infarction, Middle Cerebral Artery; Male; Mice; Naphthalenes; Neural Inhibition; Neuroprotective Agents; Phencyclidine; Rats; Rats, Sprague-Dawley; Rats, Wistar; Receptors, Metabotropic Glutamate; Reflex, Startle; Seizures; Self Stimulation; Stereotyped Behavior

Although accumulating evidence indicates that cAMP response element-binding protein (CREB) phosphorylation mediates not only synaptic plasticity but also survival of certain neurons, it remains uncertain whether CREB phosphorylation induced after metabolic insult leads to CRE-mediated gene transcription and is involved in cell survival or not. In the present study, we clarified that (1) CREB phosphorylation and ischemic tolerance induced after preconditioning ischemia in the hippocampal neurons was abolished by MK801 administration in gerbil global ischemia model, (2) CREB phosphorylation induced after exposure to glutamate in cultured neurons was inhibited by removal of extracellular calcium, by MK801 and by an inhibitor of calcium-calmodulin-dependent protein kinase (CaMK) II and IV, (3) inhibitor of CaMK II-IV or CRE-decoy oligonucleotide suppressed upregulation of BCL-2 expression and accelerated neuronal damage after exposure to glutamate, and (4) CREB phosphorylation induced in the hippocampal neurons after ischemia and in cultured neurons after exposure to glutamate was followed by CRE-mediated gene transcription in transgenic mice with a CRE-LacZ reporter. Our results suggest that CREB phosphorylation in neurons after ischemia and exposure to glutamate is induced by NMDA receptor-gated calcium influx and subsequent activation of CaMK II-IV and that CREB phosphorylation after metabolic stress might show a neuroprotective response through CRE-mediated gene induction.

Topics: Animals; Brain Ischemia; Calcium; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Calcium-Calmodulin-Dependent Protein Kinase Type 4; Calcium-Calmodulin-Dependent Protein Kinases; Cell Survival; Cells, Cultured; Cyclic AMP Response Element-Binding Protein; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Gene Expression Regulation; Genes, Reporter; Gerbillinae; Glutamic Acid; Hippocampus; Ischemic Preconditioning; Mice; Mice, Inbred C57BL; Mice, Transgenic; Neurons; Oligonucleotides; Phosphorylation; Proto-Oncogene Proteins c-bcl-2; Signal Transduction; Transcriptional Activation

Ischemic injury of immature oligodendrocytes is a major component of the brain injury associated with cerebral palsy, the most common human birth disorder. We now report that cultured immature oligodendrocytes [O4(+)/galactoceramide (GC)(-)] are exquisitely sensitive to ischemic injury (80% of cells were dead after 25.5 min of oxygen and glucose withdrawal). This rapid ischemic cell death was mediated by Ca(2+) influx via non-NMDA glutamate receptors. The receptors were gated by the release of glutamate from the immature oligodendrocytes themselves via reverse glutamate transport and included a significant element of autologous feedback of glutamate from cells onto their own receptors. High (> or = 100 microM) extracellular glutamate was protective against ischemic injury as a result of non-NMDA glutamate receptor desensitization. Other potential pathways of Ca(2+) influx, such as voltage-gated Ca(2+) channels, NMDA receptors, or the Na(+)-Ca(2+) exchanger, did not significantly contribute to ischemic Ca(2+) influx or cell injury. Release of Ca(2+) from intracellular stores was also not an important factor. In agreement with previous studies, more mature oligodendrocytes (O4(-)/GC(+)) were found to be less sensitive to ischemic injury than were the immature cells studied here.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Animals, Newborn; Brain Ischemia; Calcium; Cell Death; Cell Size; Cells, Cultured; Cellular Senescence; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Extracellular Space; Feedback; Glutamic Acid; Necrosis; Oligodendroglia; Rats; Rats, Long-Evans; Receptors, Glutamate

The endogenous cannabinoids (endocannabinoids) anandamide and 2-arachidonylglycerol increased cell viability in cerebral cortical neuron cultures subjected to 8 h of hypoxia and glucose deprivation. This effect was observed at nanomolar concentrations, was reproduced by a non-hydrolyzable analog of anandamide, and was unaltered by CB1 or CB2 cannabinoid receptor antagonists. Like synthetic cannabinoids, endocannabinoids can protect neurons from hypoxic injury, and may represent endogenous neuroprotective molecules in cerebral ischemia.

Topics: Animals; Arachidonic Acids; Brain Ischemia; Cannabinoid Receptor Modulators; Cannabinoids; Cell Hypoxia; Cell Survival; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Endocannabinoids; Excitatory Amino Acid Antagonists; Glycerides; Neurons; Neuroprotective Agents; Polyunsaturated Alkamides; Rats; Rats, Sprague-Dawley

The mechanisms of ischemic neuronal death have been focused on glutamate receptor activation and subsequent elevation of intracellular Ca2+ concentration. The purpose of this study was to evaluate the effects of dizocilpine, an NMDA receptor antagonist, pretreatment on Fos expression and parvalbumin (PV, calcium binding protein) immunoreactivity in the hippocampus of the mongolian gerbil after global ischemic insults. The number of PV-immunoreactive (PV-ir) neurons in CA1 were significantly decreased from 1 day after cerebral ischemia, while dizocilpine pretreatment completely suppressed the loss of PV-ir neurons in CA1. Dizocilpine pretreatment also protected the structural loss of microtubule-associated protein 2 immunoreactivity in CA1 after ischemic insults. In addition, dizocilpine pretreatment increased Fos expression in both hippocampal CA3 and CA4 after 3 hr ischemic reperfusion as compared to that of the saline pretreated group. Subsequently, the Fos-defined cellular activity of PV-ir neurons was slightly increased by dizocilpine pretreatment in the hippocampal area. This study demonstrated that NMDA receptor mediated calcium influx was associated with the loss of PV-ir neurons in CA1 hippocampal region, and that dizocilpine pretreatment increased Fos expression and the neuronal activity of PV-ir neurons in the non-vulnerable region of hippocampus after cerebral ischemia. Based on this data, we conclude that the protective effect of dizocilpine may be induced by the regulation of calcium overload, or by the upregulation of a neuroregenerative initiator such as Fos protein.

Topics: Animals; Brain Ischemia; Buffers; Calcium; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gerbillinae; Hippocampus; Microtubule-Associated Proteins; Parvalbumins; Proto-Oncogene Proteins c-fos

In primary neuronal-astrocyte cultures from mouse brain, ischemic conditions were simulated by combined oxygen-glucose deprival (OGD) for 2 hrs. This treatment resulted in near complete neuronal damage 24 hrs. later and was accompanied by DNA degradation and apoptotic nuclear morphology. Since caspases are key enzymes in the propagation and execution of programmed cell death, we evaluated the effect of the caspase inhibitor z-VAD-fmk. Damage following 2 hrs. OGD could be reduced by up to 56% with z-VAD-fmk (p<0.05). DNA-fragmentation and caspase activation has been also reported in an in vivo model of cerebral ischemia imitating human stroke. In this model the middle cerebral artery (MCA) is permanently occluded resulting in focal cerebral ischemia and subsequent infarction. Since z-VAD.fmk does not penetrate the blood-brain barrier it was applied intraventricularly as a bolus injection given 30 min. before MCA occlusion which was followed by 24 hrs. of infusion. Infarct volume was determined 48 hrs. after MCA occlusion by means of in vivo magnetic resonance imaging. Z-VAD.fmk dose dependently reduced infarct volume reaching a significant decrease of the cortical infarct by 45% when given as a 120 ng bolus followed by 40 ng/hr. infusion (p<0.05). In summary, our study supports the concept that caspase inhibitors are beneficial in brain ischemia.

Topics: Amino Acid Chloromethyl Ketones; Animals; Apoptosis; Astrocytes; Brain; Brain Ischemia; Caspase Inhibitors; Dizocilpine Maleate; Enzyme Inhibitors; Glucose; Immunohistochemistry; In Situ Nick-End Labeling; L-Lactate Dehydrogenase; Magnetic Resonance Imaging; Mice; Neuroprotective Agents; Rats; Rats, Inbred F344; Stroke

Acute subdural hematoma (SDH) is the most common mass lesion in severe head injury, and brain ischemia is the leading pathophysiological mechanism in the development of secondary brain damage following SDH. Hypothermia has been employed as an effective neuroprotective procedure in clinical and laboratory studies on cerebral ischemic and contusional injuries. In the present study, we used a rat acute SDH model to assess the effect of hypothermia on the intracranial pressure (ICP) and also on the brain edema formation at 4 h after hematoma induction. Mild (34 degrees C) and moderate (32 degrees C) hypothermia did not significantly affect the ICP or cerebral perfusion pressure, but they were associated with a significant lower cortical brain edema formation beneath the hematoma (81.09 +/- 0.49%, p<0.05; and 80.88 +/- 0.17%, p<0.01) when compared with the normothermic control group (81.65 +/- 0.52%). This reduction in brain edema formation was comparable to the results of MK-801 treatment (80.95 +/- 0.35%, p<0.01). The present findings indicate that hypothermia represents a potent neuroprotective strategy. The possible protective mechanisms of hypothermic protection afforded in this rat acute SDH model are discussed.

Topics: Animals; Brain; Brain Edema; Brain Ischemia; Cerebrovascular Circulation; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hematoma, Subdural; Hyperthermia, Induced; Intracranial Pressure; Male; Rats; Rats, Sprague-Dawley

A simple and reproducible animal model of global ischemia, induced by decapitation in 30-day-old Wistar rats, has been developed. It allows to perform electrophysiological analysis of the postischemic reperfusion period in the brain slices. Periods of ischemia up to 40 min increase population spikes measured in the CA1 area of the hippocampus during 2-5 h of reperfusion. Thus after 30-min decapitation-induced ischemia (at t(ischem)=25 degrees C), the mean amplitude of the recorded maximum orthodromic population spikes was 159% of the control obtained in the non-ischemic animals. Longer ischemic episodes result in the depression of the population spikes. After 2 h of ischemia, the amplitude of population spikes was about 89% of control. After 3 h of decapitation ischemia, the neurons could not be reactivated. The duration of ischemic episode needed for the irreversible depression of the electrical activity of the brain neurons drastically depends on the temperature at which the ischemic brain is maintained. Thus, only 2 h were needed at 30 degrees C as compared to nearly 3 h at 25 degrees C. We have found that intraperitoneal injection of neuroprotectors which precedes decapitation enables reactivation of the post-ischemic neurons even after very long periods of global ischemia. Thus, MK-801, a non-competitive NMDA receptors antagonist, or NBQX, a blocker of AMPA receptors, administrated 15 min before the long-term (90 min) decapitation ischemia (30 degrees C), induced dose-dependent recovery of population spike with ED(50) values 0.2 mg/kg and 3 mg/kg respectively. Our results demonstrate that, in spite of the high vulnerability of hippocampal neurons to hypoxia and ischemia, their electrical activity can be restored after prolonged (more then 1 h) decapitation ischemia. Administration of NMDA or AMPA antagonists enhances recovery.

Topics: Action Potentials; Animals; Brain Ischemia; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Hippocampus; Neurons; Neuroprotective Agents; Quinoxalines; Rats; Rats, Wistar; Time Factors

Fresh rat brain slices were incubated with [18F]2-fluoro-2-deoxy-D-glucose ([18F]FDG) in oxygenated Krebs-Ringer solution at 36 degrees C, and serial two-dimensional time-resolved images of [18F]FDG uptake in the slices were obtained on imaging plates. The fractional rate constant of [18F]FDG (proportional to the cerebral glucose metabolic rate) from pre-loading of ischemia (O(2) and glucose deprivation)/hypoxia (O(2) deprivation) to the reperfused/reoxygenated post-loading phase was quantitatively evaluated by applying the Gjedde-Patlak graphical method to the image data. Against ischemia an N-methyl-D-aspartate antagonist and hypothermia, but not a free radical scavenger, showed a protective effect when administered during ischemia, whereas no such effect was achieved with any of the above agents when administered after reperfusion. Against hypoxia, there was no protective effect with any of the above agents when administered during hypoxia, although an effect was noted with each when administered after reoxygenation. Excitatory amino acids during ischemia loading were found to be the main factor in the neuronal damage associated with ischemia, while in hypoxia, excitatory amino acids working in tandem with free radicals immediately after reoxygenation were implicated.

Topics: Animals; Brain; Brain Ischemia; Cyclic N-Oxides; Dizocilpine Maleate; Fluorodeoxyglucose F18; Glucose; Glutamic Acid; Hypothermia; Hypoxia, Brain; Male; Neuroprotective Agents; Nitrogen Oxides; Radiopharmaceuticals; Rats; Rats, Sprague-Dawley

The ability of the NMDA receptor antagonist, MK-801, to protect myelinated axons after focal cerebral ischemia has been examined. Amyloid precursor protein (APP) immunocytochemistry was used to assess the anatomic extent of axonal injury, and conventional histopathology was used to assess the volume of ischemic damage to neuronal perikarya. The middle cerebral artery was permanently occluded in 16 cats. The cats were treated with either vehicle or MK-801 as a 0.5-mg/kg bolus at 15 minutes before middle cerebral artery occlusion, followed by an infusion of 0.14 mg/kg per hour. After 6 hours, the animals were killed and the brains processed for histology and immunocytochemistry. The volume of neuronal necrosis was determined from 16 preselected coronal levels of the brain. The circumscribed zones of APP accumulation in axons were mapped onto images at the same 16 coronal levels, and quantitative analysis was performed using a transparent counting grid, randomly placed over each image. The histologic appearance and anatomic location of axons with increased APP immunoreactivity was similar in animals treated with vehicle and MK-801. MK-801 failed to reduce the hemispheric APP score significantly. In vehicle-treated animals, there was a significant association between the volume of neuronal necrosis and the amount of APP immunoreactivity. MK-801 significantly reduced the slope of the association between the volume of neuronal necrosis and the amount of APP immunoreactivity compared with that observed in vehicle-treated animals. As a result, the ratio of hemispheric APP score and volume of neuronal necrosis was significantly increased with MK-801 treatment. The inability of NMDA receptor antagonists to protect axons may limit their functional efficacy in improving functional outcome after stroke.

Topics: Amyloid beta-Protein Precursor; Animals; Axons; Brain; Brain Ischemia; Cats; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Immunohistochemistry; Neuroprotective Agents; Receptors, N-Methyl-D-Aspartate; Treatment Failure

We sought to prolong the window for stroke treatment using synergistic combinatorial therapy. We used the intraluminal filament occlusion model in rats to cause focal cerebral ischemia and a quantal bioassay to measure efficacy. The GABA agonist muscimol and the glutamate antagonist MK-801 were used alone and in combination at various times after ischemia onset. At progressively longer treatment delay intervals (30, 60, 75, 120, 240, and 360 min), higher doses of the single drugs were required to achieve neuroprotection. In contrast, the combination 1.0 mg/kg muscimol plus 0.5 mg/kg MK-801 was effective at all delay intervals studied except the longest (P < 0.05 at each time). After 240 min from ischemia onset, the combination was more effective than either single agent (P < 0.05 for each drug dose), suggesting synergism. The neuroprotective effect could not be demonstrated using morphometry. The treatment effects were probably not due to hypothermia because brain temperatures recorded in awake, unregulated subjects remained normo- or slightly hyperthermic following all treatments. Awake subjects kept on a heating pad exhibited mild brain hyperthermia. The combination caused a drop and MK-801 caused a significant increase in mean arterial blood pressure (main effects F(5,172) = 29, P < 0.0001). The combination of a GABA agonist and glutamate antagonist appears to possess synergistic neuroprotective effects when treatment is delayed up to 240 min following the onset of cerebral ischemia. Temperature regulation causes hyperthermia in awake subjects. The quantal bioassay is one method suitable for studies of synergistic stroke therapy.

Topics: Animals; Blood Pressure; Body Temperature; Brain Ischemia; Dizocilpine Maleate; Drug Therapy, Combination; Excitatory Amino Acid Antagonists; GABA Agonists; Male; Muscimol; Rats; Rats, Sprague-Dawley; Time Factors

We have evaluated the neuroprotective effects of the decahydroisoquinoline LY377770, a novel iGlu5 kainate receptor antagonist, in two models of cerebral ischaemia. Global ischaemia, induced in gerbils by bilateral carotid artery occlusion (BCAO) for 5 min, produced a large increase in locomotor activity at 96 hr post-occlusion and a severe loss of CA1 cells in the hippocampus histologically at 120 hr post-occlusion. LY377770 (80 mg/kg i.p. 30 min before or 30 min after BCAO followed by 40 mg/kg i.p. administered at 3 and 6 hr after the initial dose) attenuated the ischaemia-induced hyperactivity and provided (92%) and (29%) protection in the CA1 cells respectively. This protection was greater than that seen with maximally tolerated doses of other glutamate receptor antagonists (CGS19755, CPP, MK-801, ifenprodil, eliprodil, HA-966, ACEA1021, L701,324, NBQX, LY293558, GYKI52466 and LY300164). Focal ischaemia was induced by infusing 200 pmol of endothelin-1 (Et-1) adjacent to the middle cerebral artery and LY377770 was administered at 80 mg/kg i.p. immediately, 1 or 2 hr post-occlusion followed by 40 mg/kg i.p. 3 and 6 hr after the first dose. The infarct volume, measured 72 hr later, was reduced by LY377770 when given immediately (P<0.01), at 1 hr (P<0.05) but not significantly at 2 hr post-occlusion. Reference compounds, LY293558 (20 mg/kg i.p. and then 10 mg/kg as above) and MK-801 (2.5 mg/kg i.p. ), both administered immediately post-occlusion produced significant (P<0.05) but somewhat less neuroprotection. In parallel microdialysis studies, LY377770 (75 mg/kg i.p.) attenuated ischaemia-induced increases in extracellular levels of glutamate, but not of dopamine. In conclusion, these results indicated that iGlu5 kainate receptors play a central role in ischaemic brain damage following global and focal cerebral ischaemia. LY377770 is a novel, soluble, systemically active iGlu5 antagonist with efficacy in global and focal ischaemia, even when administered post-occlusion. LY377770 may therefore be useful as a neuroprotectant in man.

Topics: Animals; Brain Ischemia; Carotid Stenosis; Cell Death; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gerbillinae; Hippocampus; In Situ Nick-End Labeling; Isoquinolines; Male; Microdialysis; Motor Activity; Neuroprotective Agents; Quinoxalines; Rats; Rats, Sprague-Dawley; Rats, Wistar; Receptors, Kainic Acid; Tetrazoles

A prominent feature of cerebral ischemia is the excessive intracellular accumulation of both Na(+) and Ca(2+), which results in subsequent cell death. A large number of studies have focused on pathways involved in the increase of the intracellular Ca(2+) concentration [Ca(2+)](i), whereas the elevation of intracellular Na(+) has received less attention. In the present study we investigated the effects of inhibitors of different Na(+) channels and of the Na(+)/Ca(2+) exchanger, which couples the Na(+) to the Ca(2+) gradient, on ischemic damage in organotypic hippocampal slice cultures. The synaptically evoked population spike in the CA1 region was taken as a functional measure of neuronal integrity. Neuronal cell death was assessed by propidium iodide staining. The Na(+) channel blocker tetrodotoxin, and the NMDA receptor blocker MK 801, but not the AMPA/kainate receptor blocker NBQX prevented ischemic cell death. The novel Na(+)/Ca(2+) exchange inhibitor 2-[2-[4-(4-nitrobenzyloxy)phenyl]ethyl]isothiourea methanesulfonate (KB-R7943), which preferentially acts on the reverse mode of the exchanger, leading to Ca(2+) accumulation, also reduced neuronal damage. At higher concentrations, KB-R7943 also inhibits Ca(2+) extrusion by the forward mode of the exchanger and exaggerates neuronal cell death. Neuroprotection by KB-R7943 may be due to reducing the [Ca(2+)](i) increase caused by the exchanger.

Topics: Animals; Brain Ischemia; Cell Death; Culture Techniques; Dizocilpine Maleate; Electrophysiology; Hippocampus; Homeostasis; Hypoglycemia; Hypoxia; Neurons; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Sodium; Sodium Channel Blockers; Sodium Channels; Sodium-Calcium Exchanger; Tetrodotoxin; Thiourea

The effect of diethylmaleate administration on ascorbic acid release following cerebral ischemia was investigated in anesthetized rat brain cortex. Cerebral ischemia, induced by ligating bilateral common carotid arteries and unilateral middle cerebral artery, significantly increased the extracellular ascorbic acid levels. Diethylmaleate (4 mmoles/kg, i.p.), which has been shown in earlier studies to decrease the ischemia-induced glutamate release, significantly reduced the ischemia-induced ascorbic acid release. The ischemia-induced ascorbic acid release was unaffected by perfusing NMDA receptor antagonist MK 801 (75 microM). Additionally, elevated extracellular glutamate levels, achieved by either externally applied glutamate solutions or by perfusing L-trans-pyrrolidine-2,4-dicarboxylate (PDC) (31.4 mM and 15.7 mM) to inhibit the glutamate uptake transporter, also significantly increased the extracellular ascorbic acid levels. These results suggested that ascorbic acid release in cerebral ischemia might be related to the elevated extracellular glutamate levels, which occurs following cerebral ischemia.

Topics: Amino Acid Transport System X-AG; Anesthesia; Animals; Ascorbic Acid; Brain Chemistry; Brain Ischemia; Carrier Proteins; Cerebral Cortex; Dicarboxylic Acids; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Extracellular Space; Glutamate Plasma Membrane Transport Proteins; Glutamic Acid; Glutathione; Infarction, Middle Cerebral Artery; Male; Maleates; Microdialysis; Neurotransmitter Uptake Inhibitors; Pyrrolidines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Symporters

Inflammatory mediators are involved in the pathogenesis of focal ischemic brain damage. In this study we used quantitative reverse transcriptase-polymerase chain reaction to analyze the spatiotemporal pattern of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and inducible nitric oxide synthase (iNOS) expression in focal ischemia of the rat brain. Focal ischemia of the rat parietal cortex was induced noninvasively by photothrombosis of cortical microvessels. In a proportion of the animals NMDA receptor signaling was blocked by the noncompetitive receptor antagonist MK-801. Within 4 h after ischemia we found induction of TNF-alpha and IL-1beta mRNA not only in the infarcts but also in all representative tissue samples removed from noninfarcted frontal, lateral, and occipital cortex of the ipsilateral, but not contralateral hemisphere. Contrastingly, the expression of iNOS mRNA remained restricted to the evolving infarcts. Pretreatment with MK-801 strongly inhibited remote cytokine expression (mean reduction by 80% relative to vehicle treated animals at 4 h; P<0.001) whereas in the lesions only partial reductions in the expression of IL-1beta and iNOS mRNA were found. Our data for the first time demonstrate remote cytokine induction following focal brain ischemia and suggest that NMDA receptor-mediated signaling can activate inflammatory gene expression independently from the occurrence of neuronal cell death.

Topics: Animals; Brain Ischemia; Cerebral Cortex; Cortical Spreading Depression; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gene Expression Regulation, Enzymologic; Infarction, Middle Cerebral Artery; Inflammation; Interleukin-1; Intracranial Thrombosis; Male; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Photochemistry; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Tumor Necrosis Factor-alpha

N-(1-Naphthyl)-N'-(3-[(125)I]-iodophenyl)-N'-methylguanidine ([(125)I]-CNS 1261) was synthesized as a potential radioligand to image N-methyl-D-aspartate (NMDA) receptor activation. [(125)I]-CNS 1261 was prepared by radioiodination of N-(1-naphthyl)-N'-(3-tributylstannylphenyl)-N'-methylguanidine using Na(125)I and peracetic acid. [(125)I]-CNS 1261 uptake in vivo reflected NMDA receptor distribution in normal rat brain, whereas in ischemic rat brain, uptake was markedly increased in areas of NMDA receptor activation. Radiolabeled CNS 1261 appears to be a good candidate for further development as a single photon emission computed tomography tracer in the investigation of NMDA receptor activation in cerebral ischemia.

Topics: Animals; Autoradiography; Binding, Competitive; Brain; Brain Chemistry; Brain Ischemia; Chromatography, High Pressure Liquid; Dizocilpine Maleate; Guanidines; Iodine Radioisotopes; Ligands; Male; Molecular Structure; Radioligand Assay; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Tissue Distribution; Tomography, Emission-Computed, Single-Photon

The effect of N-methyl-D-aspartate (NMDA) and 2-(aminomethyl)phenylacetic acid/kainate (AMPA/kainate) glutamate receptors on dentate cell proliferation and hippocampal synapsin-I induction was examined after global ischemia. Cell proliferation was assessed using BrdU labeling, and synaptic responses were assessed using synapsin-I expression. Systemic glutamate receptor antagonists (MK-801 and NBQX) increased BrdU-labeled cells in the dentate subgranular zone (SGZ) of control adult gerbils (30% to 90%, P < 0.05). After global ischemia (at 15 days after 10 minutes of ischemia), most CA1 pyramidal neurons died, whereas the numbers of BrdU-labeled cells in the SGZ increased dramatically (>1000%, P < 0.0001). Systemic injections of MK801 or NBQX, as well as intrahippocampal injections of either drug, when given at the time of ischemia completely blocked the birth of cells in the SGZ and the death of CA1 pyramidal neurons at 15 days after ischemia. Glutamate receptor antagonists had little effect on cell birth and death when administered 7 days after ischemia. The induction of synapsin-I protein in stratum moleculare of CA3 at 7 and 15 days after global ischemia was blocked by pretreatment with systemic or intrahippocampal MK-801 or NBQX. It is proposed that decreased dentate glutamate receptor activation--produced by glutamate receptor antagonists in normal animals and by chronic ischemic hippocampal injury--may trigger dentate neurogenesis and synaptogenesis. The synapsin-I induction in mossy fiber terminals most likely represents re-modeling of dentate granule cell neuron presynaptic elements in CA3 in response to the ischemia. The dentate neurogenesis and synaptogenesis that occur after ischemia may contribute to memory recovery after hippocampal injury caused by global ischemia.

Topics: Animals; Brain Ischemia; Cell Division; Dentate Gyrus; Dizocilpine Maleate; Down-Regulation; Excitatory Amino Acid Antagonists; Gerbillinae; Glutamic Acid; Male; Memory; Microinjections; Neurons; Neuroprotective Agents; Quinoxalines; Receptors, AMPA; Receptors, Glutamate; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Synapsins

N-methyl-D-aspartate (NMDA) receptor antagonists have been found to be protective after cerebral ischemia. However most of these drugs have limited value as neuroprotectives in clinical therapy because of their side effects. Memantine is a noncompetitive NMDA receptor antagonist and it has been used for the treatment of various cerebral disorders with relatively few side effects. We investigated the beneficial effects of Memantine and compared its effect with MK-801 in a temporary focal cerebral ischemia model. As cerebral ischemia model three hours middle cerebral artery occlusion (MCAO) with intraluminal thread and three hours reperfusion was used. 78 male Spraque-Dawley rats were divided into three groups as follows: Control (Saline), treatment 1 (MK-801), and treatment 2 (Memantine) groups. In the treated groups, 15 minutes after MCAO, MK-801 and Memantine were administered in amounts of 1 mg/kg and 10 mg/kg intraperitoneally respectively. After a 3 hour period of reperfusion, the animals were examined for neurological deficits and then killed. The following values were measured; cerebral water content, blood brain barrier (BBB) permeability at the core and periphery of the ischemic hemisphere and contralateral hemisphere and infarct volumes. The severity of neurological deficit (p < 0.001) and infarct volume (p < 0.001) was reduced in both Memantine and MK-801 treated groups compared with saline treated groups. Memantine attenuated brain edema formation and BBB permeability at the periphery (p < 0.01), MK-801 both at the core (p < 0.05) and the periphery (p < 0.01) of the ischemia. These results demonstrated that the NMDA receptor antagonists Memantine and MK-801 were neuroprotective when given 15 min after MCAO in temporary focal cerebral ischemia.

Topics: Animals; Brain Edema; Brain Ischemia; Dizocilpine Maleate; Dopamine Agents; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Infusions, Parenteral; Male; Memantine; Rats; Rats, Sprague-Dawley

An in vitro model of ischemia was developed and characterized using the acute rat hippocampal slice preparation. Neuroprotective concentrations of several competitive and noncompetitive glutamate subtype-selective antagonists (CGS-19755, MK-801, YM90K and GYKI-52466) were initially determined in anoxia-enhanced agonist-induced excitotoxicity experiments. Concentrations which proved to be effective in these studies were subsequently tested for their effectiveness against an ischemic episode. Ischemia was defined as a 30-min exposure to aglycemic media ending in 5 min of concurrent anoxia, a protocol which was arrived at by empirically determining the effect of various hypoglycemic and anoxic insults on the ability of hippocampal slices to retain their electrophysiological viability. Exposure to such an ischemic episode resulted in a loss of viability by most slices, an effect which was strongly dependent on extracellular calcium. AMPA antagonists applied alone produced no neuroprotective effect in the present model of in vitro ischemia, while NMDA antagonists applied alone had a modest neuroprotective effect. In contrast, the coapplication of 10 microM MK-801 and 300 microM GYKI-52466, noncompetitive NMDA and AMPA receptor antagonists, respectively, resulted in almost complete neuroprotection. This protection was comparable to that obtained by withholding extracellular calcium, indicating that the toxic effects of glutamate receptor overstimulation can be accounted for solely by calcium influx. The effect of this combination treatment on the survival rate of hippocampal slices was synergistic, that is greater than the sum of the effects of the individual compounds. The results indicate that neuroprotection against acute ischemic insults may require a combination therapy approach.

Topics: Animals; Anti-Anxiety Agents; Benzodiazepines; Brain Ischemia; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hypoxia; Male; Neuroprotective Agents; Pipecolic Acids; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate

Murine cortical cultures containing both neurons and glia (days in vitro 13-15) were exposed to periods of oxygen-glucose deprivation (5-30 min) too brief to induce neuronal death. Cultures "preconditioned" by sublethal oxygen-glucose deprivation exhibited 30-50% less neuronal death than controls when exposed to a 45-55 min period of oxygen-glucose deprivation 24 hr later. This preconditioning-induced neuroprotection was specific in that neuronal death induced by exposure to excitotoxins or to staurosporine was not attenuated. Neuroprotection was lost if the time between the preconditioning and severe insult were decreased to 7 hr or increased to 72 hr and was blocked if the NMDA antagonist 100 microM 3-((D)-2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid was applied during the preconditioning insult. This was true even if the duration of preconditioning was increased as far as possible (while still remaining sublethal). A similar preconditioning effect was also produced by sublethal exposure to high K+, glutamate, or NMDA but not to kainate or trans-1-aminocyclopentane-1, 3-dicarboxylic acid.

Topics: Animals; Brain Ischemia; Cell Death; Cell Hypoxia; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Fetus; Glucose; Mice; N-Methylaspartate; Neurons; Neuroprotective Agents; Piperazines; Receptors, N-Methyl-D-Aspartate; Time Factors

Neuroprotection against cerebral ischemia can be realized if the brain is preconditioned by previous exposure to a brief period of sublethal ischemia. The present study was undertaken to test the hypothesis that nitric oxide (NO) produced from the neuronal isoform of NO synthase (NOS) serves as a necessary signal for establishing an ischemia-tolerant state in brain. A newborn rat model of hypoxic preconditioning was used, wherein exposure to sublethal hypoxia (8% oxygen) for 3 hours renders postnatal day (PND) 6 animals completely resistant to a cerebral hypoxic-ischemic insult imposed 24 hours later. Postnatal day 6 animals were treated 0.5 hour before preconditioning hypoxia with the nonselective NOS inhibitor L-nitroarginine (2 mg/kg intraperitoneally). This treatment, which resulted in a 67 to 81% inhibition of calcium-dependent constitutive NOS activity 0.5 to 3.5 hours after its administration, completely blocked preconditioning-induced protection. However, administration of the neuronal NOS inhibitor 7-nitroindazole (40 mg/kg intraperitoneally) before preconditioning hypoxia, which decreased constitutive brain NOS activity by 58 to 81%, was without effect on preconditioning-induced cerebroprotection, as was pretreatment with the inducible NOS inhibitor aminoguanidine (400 mg/kg intraperitoneally). The protective effects of preconditioning were also not blocked by treating animals with competitive [3-(2-carboxypiperazin-4-yl)propyl-1-phosphonate; 5 mg/kg intraperitoneally] or noncompetitive (MK-801; 1 mg/kg intraperitoneally) N-methyl-D-aspartate receptor antagonists prior to preconditioning hypoxia. These findings indicate that NO production and activity are critical to the induction of ischemic tolerance in this model. However, the results argue against the involvement of the neuronal NOS isoform, activated secondary to a hypoxia-induced stimulation of N-methyl-D-aspartate receptors, and against the involvement of the inducible NOS isoform, but rather suggest that NO produced by the endothelial NOS isoform is required to mediate this profound protective effect.

Topics: Animals; Animals, Newborn; Brain Ischemia; Calcium; Dizocilpine Maleate; Enzyme Inhibitors; Guanidines; Hypoxia; Indazoles; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Nitroarginine; Oxygen; Piperazines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

The reduction of the apparent diffusion coefficient (ADC) of brain tissue water in acute cerebral ischemia, as measured by diffusion-weighted magnetic resonance imaging, is generally associated with the development of cytotoxic edema. However, the underlying mechanism is still unknown. Our aim was to elucidate diffusion changes in the intracellular environment in cytotoxic edematous tissue. The ADC of intracellular metabolites was measured by use of diffusion-weighted 1H-magnetic resonance spectroscopy after (1) unilateral N-methyl-D-aspartate (NMDA) injection and (2) cardiac arrest-induced global ischemia in neonatal rat brain. The distinct water ADC drop early after global ischemia was accompanied by a significant reduction of the ADC of all measured metabolites (P < 0.01, n = 8). In the first hours after excitotoxic injury, the ADC of water and the metabolites taurine and N-acetylaspartate dropped significantly (P < 0.05, n = 8). At 24 and 72 hours after NMDA injection brain metabolite levels were diminished and metabolite ADC approached contralateral values. Administration of the NMDA-antagonist MK-801 1.5 hours after NMDA injection completely normalized the water ADC but not the metabolite ADC after 1 to 2 hours (n = 8). No damage was detected 72 hours later and, water and metabolite ADC had normal values (n = 8). The contribution of brain temperature changes (calculated from the chemical shift between the water and N-acetylaspartate signals) and tissue deoxygenation to ischemia-induced intracellular ADC changes was minor. These data lend support to previous suggestions that the ischemia-induced brain water ADC drop may partly be caused by reduced diffusional displacement of intracellular water, possibly involving early alterations in intracellular tortuosity, cytoplasmic streaming, or intracellular molecular interactions.

Topics: Animals; Animals, Newborn; Body Water; Brain Diseases; Brain Ischemia; Diffusion; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Intracellular Fluid; N-Methylaspartate; Rats; Rats, Wistar

There has been growing evidence that the breakdown of cytoskeletal proteins is an important biochemical change leading to ischemic neuronal death. In the present study, we investigated species differences in the susceptibility of fodrin to calpain activation induced by cerebral ischemia in gerbils, rats, and mice. In vivo fodrin proteolysis and degradation of microtubule-associated protein 2 after complete ischemia occurred more rapidly in the hippocampus and cerebral cortex of the gerbil brain than in the corresponding area of the rat and mouse brain. The N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 injected intraperitoneally before ischemia did not diminish fodrin degradation in the gerbil hippocampus. In vivo fodrin proteolysis was inhibited at 33 degrees C and enhanced at 41 degrees C compared with proteolysis at 37 degrees C during ischemia. However, in vitro fodrin proteolysis after addition of Ca2+ into the crude membrane fraction did not show any differences among three species. Although it is highly unlikely that the difference in the sensitivity of NMDA receptor or the sensitivity of calpain activation to calcium was the crucial determinant of susceptibility of fodrin degradation in the gerbil brain, the present study clearly demonstrated that fodrin in the gerbil brain was more susceptible to calpain activation induced by ischemia than that in the rat and mouse brains. Enhanced proteolysis may be one of the reasons neurons in the gerbil brain are highly vulnerable to ischemia.

Topics: Animals; Blotting, Western; Brain; Brain Ischemia; Calcium Chloride; Calpain; Carrier Proteins; Cell Membrane; Dizocilpine Maleate; Enzyme Activation; Gerbillinae; Immunohistochemistry; Male; Mice; Mice, Inbred C57BL; Microfilament Proteins; Microtubule-Associated Proteins; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Species Specificity; Temperature; Time Factors

Incubation of rat striatal slices in the absence of oxygen (anoxia), glucose (aglycemia), or oxygen plus glucose (ischemia) caused significant increases in dopamine (DA) release. Whereas anoxia decreased extracellular 3,4-dihydroxyphenylacetic acid levels by 50%, aglycemia doubled it, and ischemia returned this aglycemia-induced enhancement to its control level. Although nomifensine, a DA uptake blocker, completely protected the slices against anoxia-induced DA depletion, aglycemia- and ischemia-induced increases were not altered. Moreover, hypothermia differentially affected DA release stimulated by anoxia, aglycemia, and ischemia. Involvement of glutamate in DA release induced by each experimental condition was tested by using MK-801 and also by comparing the glutamate-induced DA release with that during anoxia, aglycemia, or ischemia. MK-801 decreased the anoxia-induced DA depletion in a dose-dependent manner. This treatment, however, showed a partial protection in aglycemic conditions but failed to improve ischemia-induced DA depletion. Like anoxia, DA release induced by exogenous glutamate was also sensitive to nomifensine and hypothermia. These results indicate that anoxia enhances DA release by a mechanism involving both the reversed DA transporter and endogenous glutamate. Partial or complete lack of effect of nomifensine, hypothermia, or MK-801 in the absence of glucose or oxygen plus glucose also suggests that experimental conditions, such as the degree of anoxia/ischemia, may alter the mechanism(s) involved in DA depletion.

Topics: 3,4-Dihydroxyphenylacetic Acid; Animals; Biological Transport; Brain Ischemia; Corpus Striatum; Dizocilpine Maleate; Dopamine; Dopamine Uptake Inhibitors; Dose-Response Relationship, Drug; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Female; Glucose; Glutamic Acid; Hypothermia; Hypoxia; Male; Neurons; Nomifensine; Organ Culture Techniques; Ouabain; Oxygen; Rats; Rats, Sprague-Dawley; Sodium-Potassium-Exchanging ATPase; Tritium

MRI was used for the in vivo evaluation of unilateral hypoxic-ischemic brain injury and the evaluation of MK-801 in the neonatal rat.. T2-weighted scans were obtained during the acute phase of HI injury and 3 months later. Histology was performed to correlate MRI signal changes with pathology. Finally, the effectiveness of MK-801 to limit brain injury was regionally assessed in vivo using T2-weighted MRI.. Injury visualized by MRI at 72 hours after hypoxia correlated strongly with histopathologic analysis. Transient injury was identified. MK-801 significantly reduced the lesion extent at the level of the hippocampus. Patterns of unilateral versus bilateral neonatal brain injury were found to differ.. The study demonstrates unique patterns of brain injury not seen in adult animal hypoxia-ischemia studies, and the sensitivity of the corpus callosum to hypoxia-ischemia. MK-801, although neuroprotective, did not offer any selective neuroprotective benefit.

Topics: Animals; Animals, Newborn; Brain; Brain Ischemia; Dizocilpine Maleate; Hypoxia, Brain; Magnetic Resonance Imaging; Neuroprotective Agents; Rats; Rats, Wistar

Numerous studies over the past three decades have used rodent models of cerebral ischemia. To measure the postischemic outcome, the majority of these studies used histopathology as the method of choice both quantitatively and qualitatively. No functional measure of postischemic outcome has been proved to correlate well with the histopathological one. The rat chest compression model of cardiac-arrest-induced global cerebral ischemia was used in the present study. Two separate measures of neuronal damage at 7 days postischemia were performed: (a) histologically, by counting normal pyramidal cell bodies in the mid-CA1 hippocampal region of the rat brain, in hematoxylin-eosin-stained, paraffin-embedded 6-microm sections, and (b) electrophysiologically, by counting the number of 400 microm hippocampal slices in which it was possible to evoke a normal (>/=10 mV) CA1 population spike by orthodromic stimulation of the Schaffer collaterals. The correlation between these two measures was tested in the following groups of rats: (a) control, untreated group, (b) MK-801-treated groups (0.03 to 1.0 mg/kg given i.p. shortly after ischemia), (c) diltiazem-treated (DILT) groups 1.0 to 30 mg/kg, given i.p. shortly after ischemia, and (d) a group treated with a combination of the two drugs together (0.1 mg/kg MK-801+3.0 mg/kg DILT given i.p. shortly after ischemia). The two measures of postischemic outcome were highly correlated in all groups studied. Both MK-801 and DILT exhibited a dose-dependent neuroprotective effect. When administered together, a synergy between the neuroprotective effect of MK-801 and DILT was observed. At the doses used, minimal or no side effects of either MK-801 or DILT were observed.

Topics: Animals; Brain Ischemia; Diltiazem; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Synergism; Electrophysiology; Excitatory Amino Acid Antagonists; Heart Arrest; Hippocampus; Male; Neuroprotective Agents; Organ Culture Techniques; Pyramidal Cells; Rats; Rats, Long-Evans; Vasodilator Agents

We have recently demonstrated that inducible nitric oxide synthase (iNOS) is expressed in rat forebrain slices exposed to oxygen and glucose deprivation (OGD). Now, we have found that the expression of iNOS after OGD is time-dependent since 20 min of OGD produces the appearance of iNOS at earlier times than 10 min of OGD. OGD also causes neurotoxicity in this model, as revealed by the increase in excitatory amino acid, neuron specific enolase and lactate dehydrogenase (LDH) efflux to the incubation solution. Finally, the administration of the NMDA receptor antagonist MK-801 (100 nM) inhibits both the expression of iNOS and the release of LDH. Our findings demonstrate that this method may be considered an useful in vitro model of ischemia-reperfusion to determine the therapeutic role of neuroprotective tools.

Topics: Animals; Brain Ischemia; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Excitatory Amino Acids; Glucose; Hypoxia; In Vitro Techniques; L-Lactate Dehydrogenase; Male; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Phosphopyruvate Hydratase; Prosencephalon; Rats; Rats, Sprague-Dawley; Reperfusion Injury

Spreading depression is a wave of sustained depolarization challenging the energy metabolism of the cells without causing irreversible damage. In the ischaemic brain, sreading depression-like depolarization contributes to the evolution of ischaemia to infarction. The depolarization is propagated by activation of N-methyl-D-aspartate receptors, but changes in signal transduction downstream of the receptors are not known. Because protein phosphorylation is a general mechanism whereby most cellular processes are regulated, and inhibition of N-methyl-D-aspartate receptors or protein kinase C is neuroprotective, the expression of protein kinase C subspecies in spreading depression was examined. Cortical treatment with KCl induced an upregulation of protein kinase Cdelta and zeta messenger RNA at 4 and 8 h, whereas protein kinase Calpha, beta, gamma and epsilon did not show significant changes. The gene induction was the strongest in layers 2 and 3, and was followed by an increased number of protein kinase Cdelta-immunoreactive neurons. Protein kinase Cdelta and zeta inductions were inhibited by pretreatment with an N-methyl-D-aspartate receptor antagonist, dizocilpine maleate, which also blocked spreading depression propagation, and with dexamethasone, which acted without blocking the propagation. Quinacrine, a phospholipase A2 inhibitor, reduced only protein kinase C5 induction. In addition, N(G)(-nitro-L-arginine methyl ester, a nitric oxide synthase inhibitor, did not influence protein kinase Cdelta or zeta induction, whereas 6-nitro-7-sulphamoylbenzo[f]quinoxaline-2,3-dione, an alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate/kainate receptor antagonist, and the cyclo-oxygenase inhibitors indomethacin and diclophenac tended to increase gene expression. The data show that cortical spreading depression induces Ca2(+)-independent protein kinase C subspecies delta and zeta, but not Ca(2+)-dependent subspecies, through activation of N-methyl-D-aspartate receptors and phospholipase A2. Even though the signal pathway is similar to the induction described previously in ischaemia for genes implicated in delayed neuronal death, the gene inductions observed here are not necessarily pathogenetic, but may represent a general reaction to metabolic stress.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Brain Ischemia; Cortical Spreading Depression; Cyclooxygenase Inhibitors; Dexamethasone; Diclofenac; Disease Susceptibility; Dizocilpine Maleate; Enzyme Induction; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Gene Expression Regulation, Enzymologic; Indomethacin; Isoenzymes; Male; Nerve Tissue Proteins; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Parietal Lobe; Phospholipases A; Phospholipases A2; Promoter Regions, Genetic; Protein Kinase C; Protein Kinase C-delta; Quinacrine; Quinoxalines; Rats; Rats, Wistar; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Signal Transduction; Transcriptional Activation

Following focal brain lesions, complex adaptive processes take place in remote intact areas. The present study examines changes in NMDA (N-methyl-D-aspartate), AMPA ((+/-)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid) and kainate receptors following focal photothrombotic ischemic lesions using quantitative receptor autoradiography. Increases in binding density of NMDA receptors were seen in both hemispheres for up to 30 days. In the contralateral hemisphere, this increase of NMDA receptors occurred as early as 4 h after lesion whereas it appeared with a delay for 14 days on the lesioned side. Binding density of [3H]AMPA and [3H]kainate was unchanged. We suggest that the translational process is differentially regulated by spreading depressions. The delayed up-regulation of NMDA receptor binding on the lesioned side may be due to a translation block similar to that previously described for GABA(A) receptor subunits.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Autoradiography; Brain Ischemia; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Intracranial Thrombosis; Kainic Acid; Light; Male; Radiation Injuries, Experimental; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Time Factors; Up-Regulation

Recent studies have shown that a brief 'pre-conditioning' ischaemic insult reduces the hippocampal cell death caused by a subsequent more severe test insult. In the present studies, we have examined the effects of the non-competitive NMDA receptor antagonist ((5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5, 10-imine, MK-801) a competitive NMDA receptor antagonist, LY202157, AMPA receptor antagonist ((3S,4aR,6R,8aR)-6-[2-(1(2)H-tetrazole-5-yl)]decahydroiso quinoline-3-carboxylic acid, LY293558), a non-competitive AMPA receptor antagonist ((-)-1-(4-amino-phenyl)-4-methyl-7,8-methylenedioxy-4,5-dihydro-3-acetyl -2,3-benzodiazepine, LY300164), and a mixed NMDA/AMPA receptor antagonist, LY246492, in a gerbil model of ischaemic tolerance. Ischaemic tolerance was induced by subjecting gerbils to a 2-min 'pre-conditioning' ischaemia (bilateral carotid occlusion) 2 days prior to a 3-min test ischaemia. The effects of MK-801 (2 mg/kg i.p.), LY293558 (20 mg/kg i.p., followed by 4 x 10 mg/kg at 3 h intervals), LY300164 (4 x 10 mg/kg i.p. at 1 h intervals), LY246492 (40 mg/kg i.p., followed by 4 x 20 mg/kg i.p. at 3 h intervals) and LY202157 (30 mg/kg i.p., followed by 4 x 15 mg/kg i.p. at 2 h intervals) were then examined in this model. Initial dosing commenced 30 min prior to the 2-min 'pre-conditioning' ischaemia. Results indicated that a 2-min 'pre-conditioning' ischaemia produced ischaemic tolerance in all cases. The non-competitive NMDA receptor antagonist, MK-801, produced a significant (P < 0.01) reduction in the induced tolerance, while the competitive NMDA receptor antagonist, LY202157, also attenuated (P < 0.05) the induction of tolerance. In contrast, two AMPA receptor antagonists (LY293558 and LY300164) and a mixed NMDA/AMPA receptor antagonist (LY246492) had no effect on the induction of tolerance. These results suggest that NMDA receptor activation, but not AMPA receptor activation is involved in the phenomenon of ischaemic tolerance.

Topics: Animals; Brain Ischemia; Cell Count; Cell Survival; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gerbillinae; Hippocampus; Ischemic Preconditioning; Isoquinolines; Male; Neurons; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Tetrazoles

We have investigated the neuroprotective effects of combining an NMDA or AMPA receptor antagonist with a nitric oxide synthase (NOS) inhibitor in the gerbil model of global cerebral ischaemia. Ischaemia was induced by occlusion of the common carotid arteries for 5 min. (5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,1 0-imine (MK-801, 2.5 mg/kg i.p.) or (3S,4aR,6R,8aR)-6-[2-(1(2)H-tetrazole-5-yl)]decahydroisoq uinoline-3-carboxylic acid (LY293558, 20 mg/kg i.p.) and 7-nitroindazole (25 mg/kg i.p.) or N-[4-(2-[[(3-chlorophenyl)methyl]amino]ethyl) phenyl]-2-thiophenecarboximidamide dihydrochloride (ARL17477, 25 mg/kg i.p.) were administered alone or in combination (i.e., MK-801 with 7-nitroindazole or ARL17477 or LY293558 with 7-nitroindazole or ARL17477). In the present studies, both MK-801 and LY293558 provided significant degree of neuroprotection, while 7-nitroindazole and ARL17477 also provided some neuroprotection, which failed to reach significance in every case. However, the combination of MK-801 with 7-nitroindazole or ARL17477 provided 21% or 44% greater protection than the total protection or either alone. Likewise, the combination of LY293558 with 7-nitroindazole or ARL17477 provided 14.5% and 35% greater protection than total protection of either compound alone. These results indicate that several pathways contribute to ischaemic cell death and combining excitatory amino antagonists and NOS inhibitors provides greater protection than either alone. Therefore, combination therapy should be considered as an approach for treating ischaemic conditions.

Topics: Amidines; Animals; Brain Ischemia; Carotid Arteries; Dizocilpine Maleate; Drug Synergism; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Gerbillinae; Hippocampus; Indazoles; Isoquinolines; Male; Neuroprotective Agents; Nitric Oxide Synthase; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Tetrazoles

The mechanism of glutamate release from cultured cerebellar granule neurones in response to a chemical model of ischaemia (10 mM 2-deoxyglucose plus 1 mM sodium cyanide) was investigated. In the first 2 min of ischaemia, release of preloaded D-[3H]aspartate could be extensively attenuated by tetanus toxin and bafilomycin A1 and was dependent on the activation of Ca2+ channels sensitive to the "Q" type Ca2+ channel antagonist, omega-conotoxin-MVIIC. During this period, ATP/ADP ratios fell rapidly. The extent of release in the first 2 min was comparable to that evoked by 2-min depolarization by 50 mM KCl. Free Ca2+ concentrations, determined in neurites and somata, did not increase until after 2 min. The neurite increase in cellular Ca2+ precedes that of the cell somata. Release of D-[3H]aspartate was partially inhibited by the NMDA receptor antagonist MK-801, which also delayed the increase in free Ca2+ concentration. Prolonging the period of ischaemia to 6 and 10 min produced no further increase in the apparently exocytotic component of release, but initiated an extensive nonexocytotic release of the amino acid. Studies with the synaptic vesicle membrane probe FM1-43 in which released amino acid was removed by superfusion indicated that Ca2+-dependent exocytosis was delayed in this system. It is concluded that chemical ischaemia initiates an initial exocytotic followed by nonexocytotic release and that the former is facilitated by NMDA receptor activation. These events occur in cells that are still able to exclude propidium iodide, indicating that cell death has not yet occurred.

Topics: Adenosine Diphosphate; Adenosine Triphosphate; Animals; Anti-Bacterial Agents; Aspartic Acid; Brain Ischemia; Calcium; Calcium Channel Blockers; Calcium Channels; Cells, Cultured; Cerebellum; Cytoplasm; Deoxyglucose; Dizocilpine Maleate; Exocytosis; Fluorescent Dyes; Glutamic Acid; Macrolides; Neurons; omega-Conotoxins; Peptides; Pyridinium Compounds; Quaternary Ammonium Compounds; Rats; Receptors, N-Methyl-D-Aspartate; Sodium Cyanide; Tetanus Toxin; Time Factors

Recent clinical trials with non-competitive and competitive N-methyl-D-aspartate (NMDA) receptor antagonists in patients with stroke have shown that these patients develop more adverse effects, particularly psychomimetic effects such as hallucinations and agitation, than normal volunteers at equivalent doses. We therefore examined whether such increased adverse effect potential of NMDA antagonists also occurs in a rat model of permanent focal ischemia. For this purpose, the right middle cerebral artery was occluded under halothane anesthesia, and behavioral alterations in response to the non-competitive NMDA antagonist, MK-801 (dizocilpine), were recorded after recovery from anesthesia. Behavioral alterations in ischemic rats were compared with those in sham-lesioned rats in a blinded fashion. MK-801 (0.4 mg/kg) induced psychomimetic-like stereotyped behaviors which were about twice as intense in ischemic than in non-ischemic rats. A similar trend for enhanced adverse effects was seen with the competitive NMDA antagonist CGS 19755 (Selfotel). Although more NMDA antagonists have to be tested to draw definite conclusions, the present data may indicate that enhanced sensitivity of stroke patients to adverse effects of NMDA antagonists can be predicted by use of a focal ischemia model in rats, thus allowing use of this model for developing novel cytoprotective strategies targeted to minimize glutamatergic excitotoxicity with reduced adverse effect potential.

Topics: Animals; Behavior, Animal; Binding, Competitive; Brain; Brain Ischemia; Cerebrovascular Disorders; Dizocilpine Maleate; Injections, Intraperitoneal; Male; Pipecolic Acids; Piperazines; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate

Three fenamates (flufenamate, meclofenamate and mefenamate) were examined for their protective effect on neurons under ischemic (glucose/oxygen deprivation) or excitotoxic conditions, using the isolated retina of chick embryo as a model. Retinal damage was evaluated by histology and lactate dehydrogenase assay. Whole-cell recording was used to examine the direct effect of the fenamates on glutamate receptor-mediated currents. The fenamates protected the retina against the ischemic or excitotoxic insult. Part of the neuroprotection by the fenamates derived from inhibition of N-methyl-D-aspartate receptor-mediated currents. However, kainate receptor-mediated currents were not blocked by the fenamates, which nonetheless reduced kainate receptor-mediated retinal damage. Our results raise the possibility that fenamates may serve as lead structures in the development of novel therapeutic agents against brain ischemia.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Brain Ischemia; Chick Embryo; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Flufenamic Acid; Glucose; Kainic Acid; Meclofenamic Acid; Mefenamic Acid; N-Methylaspartate; Neuroprotective Agents; Neurotoxins; Organ Culture Techniques; ortho-Aminobenzoates; Oxygen; Patch-Clamp Techniques; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Retinal Ganglion Cells

The effect of neuroprotective drugs on the early and late electrophysiological manifestations of photothrombotic occlusion of distal branches of middle cerebral artery was studied in rats treated with MK-801 and Cerebrolysin (CL). DC potentials were recorded from the irradiated cortex (ischemic core), from the adjacent penumbra zone and from remote intact cortex. Irradiation elicited after a few minutes of spontaneous spreading depression (SD) waves followed during 10-15 min by focal ischemic depolarization (FID) developing in the irradiated cortex and spreading into the perifocal areas. While the core FID amplitude reached about 30 mV and decayed during subsequent 2 h to 10-13 mV, FID in the penumbra zone was broken by periods of partial repolarization and returned during 30-90 min almost to baseline. At the same time, generation of spontaneous SD waves almost stopped. MK-801 (0.5 mg/kg, i.p., 45 min after ischemia) blocked SD waves, but did not shorten penumbra FID, the decay of which was slowed down to the rate found in the ischemic core. CL treatment (2.5 ml/kg, i.p. , 1 h after ischemia) did not influence FID in the acute phase of the experiment, but its 10-day administration facilitated post-ischemic recovery indicated by higher amplitude of evoked SD waves penetrating into the former penumbra zone. Morphological examination showed that the volume of total and partial necrosis was increased in the MK-801 group and marginally reduced in the CL group. It is suggested that the absence of the SD-induced hyperperfusion episodes in MK-801-treated rats may accelerate perifocal thrombotization in this model of focal ischemia.

Topics: Amino Acids; Animals; Brain; Brain Ischemia; Dizocilpine Maleate; Electrophysiology; Intracranial Embolism and Thrombosis; Male; Neuroprotective Agents; Radiation Injuries, Experimental; Rats; Rats, Inbred Strains

Focal cerebral ischemia in rats produces elevated levels of tumor necrosis factor (TNF)alpha in the ischemic brain region. To better understand the modulation of TNF during brain ischemia processes we carried out studies in a model of permanent middle cerebral artery occlusion (MCAo) in the rat. In non-treated ischemic animals, the maximum expression of TNF was observed at 12 h (246.1+/-33 U/g) in the ischemic cortex and declined reaching near zero levels 24 h after MCAo. Given 10 min after MCAo, MK 801 (3 mg/kg, i.p.), a non-competitive NMDA receptor antagonist, exerted significant neuroprotection as measured by 47% reduction of total volume of infarction (P < 0.01 vs. ischemic-control). At the high dose of 3 mg/kg i.p., dexamethasone (DEX), which is known to reduce brain edema, decreased infarct size by 50% (P < 0.01 vs. ischemic-control). Both MK 801 and DEX reduced TNF production in the ipsilateral cortex of ischemic animals by 61 and 73%, respectively (P < 0.01 vs. ischemic-control). The data indicate that TNF levels increase after brain infarction, whereas they are reduced by neuroprotective agents, such as MK 801 and DEX, which act on different cellular levels.

Topics: Animals; Brain Ischemia; Cerebral Cortex; Cerebral Infarction; Dexamethasone; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Male; Rats; Rats, Sprague-Dawley; Time Factors; Tumor Necrosis Factor-alpha

This study attempted to ascertain whether heatstroke-induced ischemia is associated with augmented striatal glutamate release and can be attenuated by NMDA receptor antagonists. Mean arterial pressure (MAP), striatal cerebral blood flow (CBF), striatal glutamate release and striatal neuronal damage score were assessed in saline-treated rats and in rats treated with NMDA receptor antagonists. Heatstroke was induced by exposing the animals to a high ambient temperature; the moment at which MAP and CBF began to decrease from their peak levels was taken as the onset of heatstroke. During onset of heatstroke, rats displayed higher values of colonic temperature, striatal glutamate release and striatal neuronal damage score, and lower values of MAP and striatal blood flow compared with normothermic control rats. The decreased MAP, the diminished striatal blood flow, the augmented striatal glutamate release and the increased striatal neuronal damage score during onset of heatstroke were significantly attenuated by pretreatment with an NMDA receptor antagonist such as MK-801 or ketamine. In addition, the survival time (interval between onset of heatstroke and death) of the rats was extended by pretreatment with one of these two NMDA receptor antagonists. These results suggest that marked accumulation of glutamate in the striatum is important for the development of ischemic damage to striatal neurons during the onset of heatstroke.

Topics: Animals; Body Temperature; Brain Ischemia; Cerebrovascular Circulation; Corpus Striatum; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Heat Stroke; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate

The present study was designed to evaluate the effects of pretreatment with a combination of desmethyl tirilazad (21-aminosteroid) plus dizocilpine maleate (N-methyl-D-aspartate receptor antagonist) and nimodipine (calcium channel antagonist) on constitutive nitric oxide synthase (cNOS) activity and cyclic guanosine monophosphate (cGMP) levels in brain homogenates of rats subjected to global cerebral transient ischemia induced by bilateral clamping of the carotids for 30 min and reduction of arterial pressure (to 50-60 mm Hg) by intravenous infusion of trimethaphan (30 mg/kg). Our results show that cerebral ischemia produced an increase in cNOS activity and cGMP levels in brain homogenates. Pretreatment with desmethyl tirilazad or dizocilpine maleate or nimodipine individually significantly suppressed (p < 0.01) the increase in cNOS activity and cGMP levels induced by cerebral ischemia, which may be related to their neuroprotective effect. Similar results were obtained with pretreatment by a combination of desmethyl tirilazad plus dizocilpine maleate plus nimodipine.

Topics: Animals; Brain; Brain Ischemia; Cyclic GMP; Dizocilpine Maleate; Enzyme Inhibitors; Male; Neuroprotective Agents; Nimodipine; Nitric Oxide Synthase; Pregnatrienes; Rats; Rats, Wistar

It is hypothesized that after various types of brain injury, blood-brain barrier (BBB) opening and vasogenic edema result from excessive neuronal release of glutamate and stimulation of capillary N-methyl-d-aspartate (NMDA) receptors linked to polyamine (putrescine) synthesis in endothelial cells. We produced cerebral ischemia in rats and measured BBB opening 6 h later as the increase in regional transfer constants (Ki) for blood to brain diffusion of [3H]sucrose. Such BBB opening was not mitigated by drugs which block NMDA receptors (MK801 or AR-R 15896AR) or polyamine synthesis (difluoromethylornithine). These results question generality of the capillary NMDA receptor/polyamine hypothesis.

Topics: Animals; Biogenic Polyamines; Blood-Brain Barrier; Brain Ischemia; Dizocilpine Maleate; Eflornithine; Excitatory Amino Acid Antagonists; Male; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

In this study, the N-Methyl-D-Aspartate (NMDA) receptor-dependent nitric oxide and cyclic GMP (cGMP) synthesis in the course of reperfusion after 5 min of ischemia in gerbil brain hemispheres and cerebellum were investigated. Moreover, the role of the neuronal isoform of nitric oxide (NO) synthase (nNOS) in liberation of NO in postischemic brain and the involvement of NO in membrane lipoperoxidations activated during reperfusion were evaluated. Enhancement of Ca2+/calmodulin-regulated NOS activity and cGMP level in brain hemispheres and in cerebellum during reperfusion was found to be coupled to the activation of the NMDA receptor. cGMP concentration 40% above the control level was observed to persist up to 7 days after ischemia. The amount of conjugated double bounds in membrane lipids and the level of thiobarbituric acid reactive substances were increased exclusively in brain hemispheres, indicating activation of lipid peroxidation. The NMDA receptor antagonist, MK-801, eliminated, and a rather selective nNOS inhibitor, 7-Nitroindazole (7-NI) attenuated, NMDA receptor-evoked enhancement of NOS activity and cGMP level in brain hemispheres and in cerebellum during reperfusion. Moreover, 7-NI decreased significantly membrane lipid peroxidation during the early time of reperfusion. Histological examination demonstrated that 7-NI protects against death a selected population of neuronal cells in CA1 layer of hippocampus. It is suggested that NMDA receptor dependence of NO release during reperfusion is responsible for the degeneration of some populations of neurons and that the effect is mediated by activation of free radical formation and lipid peroxidation. Moreover, in cerebellum, ischemia-evoked activation of glutamatergic system stimulates NO-dependent signal transmission. Our results indicated that 7-NI has a significant ameliorating effect on biochemical alterations evoked by ischemia, suggesting nNOS inhibitors as a potential therapeutic agents in reperfusion injury.

Topics: Animals; Brain Ischemia; Cerebellum; Cerebral Cortex; Cyclic GMP; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Gerbillinae; Indazoles; Lipid Peroxidation; Male; Membrane Lipids; Neuroprotective Agents; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Prosencephalon; Receptors, N-Methyl-D-Aspartate; Reperfusion Injury; Signal Transduction; Thiobarbituric Acid Reactive Substances

The purpose of the present study was to compare the characteristics of the photochemical-induced thrombotic occlusion model and the thermocoagulated occlusion model of the middle cerebral artery in rats. We evaluated the neuroprotective effects of a NMDA receptor antagonist, (+)-MK-801 (dizocilpine, (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cycloheptan-5,10-imine), an alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor antagonist, YM90K (6-(1H-imidazol-1-yl)-7-nitro-2,3(1H,4H)-quinoxalinedione monohydrochloride), a Ca2+ channel antagonist, S-312-d (S-(+)-methyl-4,7-dihydro-3-isobutyl-6-methyl-4-(3-nitrophenyl)-thieno[2 ,3-b]pyridine-5-carboxylate), the radical scavengers, MCI-186 (3-methyl-1-phenyl-2-pyrazolin-5-one) and EPC-K1 (L-ascorbic acid 2-[3,4-dihydro-2,5,7,8-tetramethyl-2-(4,8,12-trimethyl-tridecyl)-2H-1-be nzopyran-6yl-hydrogen phosphate] potassium salt), and a calcineurin inhibitor, FK506 (tacrolimus, Prograf). Although all tested agents in the present study attenuated the brain damage in the photochemical-induced thrombotic occlusion model, the radical scavengers did not attenuate the brain damage in the thermocoagulated occlusion model. The time course of brain damage and brain edema formation in the two models was examined. The time course of brain damage was not different in the two models, but the time course of brain edema was quite different. Brain edema formation in the photochemical-induced thrombotic occlusion model was significantly greater (P < 0.01) than that in the thermocoagulated occlusion model at all time point studied until 24 h after occlusion of the middle cerebral artery. The present study suggests that the photochemical-induced thrombotic occlusion model has characteristics of both permanent ischemia and ischemia-reperfusion.

Topics: Animals; Brain Edema; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Electrocoagulation; Immunosuppressive Agents; Injections, Intraperitoneal; Injections, Intravenous; Intracranial Embolism and Thrombosis; Male; Neuroprotective Agents; Quinoxalines; Rats; Rats, Sprague-Dawley; Tacrolimus

An anti-ischemic effect of the delta-sleep-inducing peptide (DSIP) was found in rats. The DSIP effect was more obvious than that of the MK-801. The data obtained is discussed considering a possible use of the DSIP for brain stroke prophylaxis.

Topics: Animals; Brain Ischemia; Delta Sleep-Inducing Peptide; Disease Models, Animal; Dizocilpine Maleate; Injections, Intraperitoneal; Male; Neuroprotective Agents; Rats; Rats, Wistar

Many glutamate antagonists have been reported to have a neuroprotective effect against ischemic brain damage; however, some of them have been also reported to induce hypothermia that confers remarkable neuroprotection against the damage. In order to avoid the confounding effects of hypothermia, we assembled a telemeter-based brain temperature control system that allows continuous monitoring and regulating of brain temperature during an ischemic insult and in the post-ischemic period in conscious and freely moving animals. Experiments were performed in gerbils that were subjected to administration of MK-801 (3, 5, and 10 mg/kg) and/or to 5-min ischemia. The system monitored continuous changes in brain temperature and regulated brain temperature at normothermic levels, revealing that a neuroprotective effect of 3 mg/kg MK-801 against ischemia-induced delayed hippocampal CA1 neuronal death was mainly due to hypothermia, whereas a high dose of MK-801 (5 and 10 mg/kg) produced a neuroprotective effect even when the brain temperature was maintained at normothermic levels. These results indicate that this system is very useful to test potential antiischemic agents, especially when the agents have hypothermic side effects.

Topics: Animals; Body Temperature Regulation; Brain; Brain Ischemia; Cell Count; Cell Death; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gerbillinae; Hippocampus; Hypothermia, Induced; Male; Neurons

This report describes the initial clinical assessment of (+)-3-[123I]Iodo-MK-801 and its potential to provide single photon emission tomographic (SPET) images in vivo of NMDA receptor activation during cerebral ischaemia. Multiple SPET images were obtained in the 120 min after the administration of 150 MBq of (+)-3-[123I]Iodo-MK-801 to five patients with cerebral ischaemia (due to cerebral haemorrhages) and to five normal volunteers. In normal subjects, (+)-3-[123I]Iodo-MK-801 has a rapid uptake into the brain. The tracer has a high non-specific retention in the central nervous system due to its lipophilicity, which was made evident by the retention of tracer in the cerebellum and white matter (brain areas with few NMDA receptors). In all patients with cerebral haemorrhages, the initial uptake of (+)-3-[123I]Iodo-MK-801 into the ipsilateral hemisphere was markedly reduced, consistent with a reduced level of cerebral blood flow. In two of five patients, relatively increased tracer retention at later time points (60-120 min after tracer administration) could be seen in cortical areas adjacent to the site of the haemorrhage, consistent with activated NMDA receptors. In three of the patients, no relatively enhanced tracer retention could be identified. Using (+)-3-[123I]Iodo-MK-801, it may be possible to image excessive glutamate (NMDA) receptor activation during an ischaemic episode in living human patients. The utility of (+)-3-[123I]Iodo-MK-801 as a SPET ligand for assessing modest alterations in NMDA receptor activity may ultimately be limited by its lipophilicity and consequent high non-specific binding.

Topics: Aged; Brain; Brain Ischemia; Cerebral Hemorrhage; Dizocilpine Maleate; Gamma Cameras; Humans; Iodine Radioisotopes; Receptors, N-Methyl-D-Aspartate; Subarachnoid Hemorrhage; Tissue Distribution; Tomography, Emission-Computed, Single-Photon

Cerebral arteriolar dilation to N-methyl-D-aspartate (NMDA) is drastically reduced by anoxic stress. The effects of anoxic stress on cerebrovascular dilation to activation of other types of glutamate receptors are unknown. The purpose of this study was to examine the effects of ischemia on cerebral arteriolar responses to kainate in anesthetized piglets.. Arteriolar responses to 5 x 10(-5) mol/L and 10(-4) mol/L kainate were evaluated before and 10 minutes after total, global ischemia. Ischemia was induced by increasing intracranial pressure. We measured pial arteriolar diameters (approximately 100 microns) using a cranial window and intravital microscopy.. Before ischemia, kainate dilated arterioles by 16 +/- 2% at 5 x 10% mol/L and 30 +/- 2% at 10(-4) mol/L (mean +/- SEM; n = 6). After ischemia, the diameter of arterioles increased by 17 +/- 3% and 26 +/- 3% to 5 x 10% and 10(-4) mol/L kainate, respectively (P > .05). We also investigated the mechanisms involved in mediating arteriolar dilation to kainate. Intravenous administration of N omega-nitro-L-arginine methyl ester (L-NAME) (15 mg/kg) (n = 7) or indomethacin (10 mg/kg) (n = 6) individually reduced arteriolar dilation to kainate by approximately one half. Coadministration of L-NAME and indomethacin almost completely eliminated arteriolar dilation to kainate (n = 10). Administration of theophylline (20 mg/kg IV) did not affect dilator responses to kainate (n = 7). Blockade of NMDA receptors by MK801 had minimal effects on arteriolar dilation to kainate (n = 6).. There are three main findings from this study: (1) kainate is a potent dilator agent in the neonatal cerebral circulation; (2) nitric oxide and prostaglandins both participate in the vasodilator response to kainate; and (3) in contrast to NMDA, cerebral arteriolar dilator responses to kainate are resistant to ischemic stress.

Topics: Animals; Animals, Newborn; Arterioles; Brain Ischemia; Cerebral Arteries; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Indomethacin; Kainic Acid; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Prostaglandins; Reperfusion Injury; Swine; Theophylline; Vasodilator Agents

Hypothermia after hypoxia-ischaemia (HI) confounds the interpretation of the effects of neuroprotective drug intervention. The effect of 0.5 mg/kg of dizocilpine (MK-801) administered after HI on rectal temperature at 2-36 h and on brain damage 2 weeks after the insult was evaluated in the immature rat. In pups kept at an ambient temperature of 21 degrees C, MK-801 lowered the temperature by 1.1 degrees C and reduced the brain damage by 45%. In pups held at an ambient temperature of 33 degrees C, MK-801 treatment afforded a 34% reduction of brain damage without lowering the rectal temperature. In conclusion, the neuroprotection offered by MK-801 does not depend on systemic hypothermia in this model.

Topics: Animals; Animals, Newborn; Body Temperature; Brain Ischemia; Dizocilpine Maleate; Female; Hypothermia; Hypoxia, Brain; Male; Neuroprotective Agents; Rats; Rats, Wistar; Rectum

The present study covers both the effects of MK-801, a noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist, and pentobarbital on cholinergic terminal damage and delayed neuronal death (DND) in ischemic gerbil. To study the above effects, in vivo microdialysis, immunohistochemical, and morphological techniques were used. MK-801 (3 mg/kg) or pentobarbital (50 mg/kg) were injected intraperitoneally 1 h or 30 min before 5 min ischemia, respectively. Each estimation was then carried out 4, 7, or 14 days after ischemia. Ischemia induced a significant decrease in acetylcholine (ACh) release and a disappearance of choline acetyltransferase (ChAT)-immunoreactivity in the hippocampus in addition to inducing DND. On day 4, MK-801 protected ischemia-induced DND in the hippocampal CA1 subfield. However, MK-801 had no effect against the decrease in ACh release in spite of protection of the decrease in ChAT-immunoreactivity. On day 7 and 14, no protective effect of MK-801 was observed in all estimations. It became clear that the mechanism of cholinergic terminal dysfunction is different from that involved in pyramidal cell death, i.e., excitative neurotoxicity induced by overabundant extracellular glutamate. Pentobarbital also provided protection against DND. However, protective effects of pentobarbital on the decrease in ACh release and the low ChAT-immunoreactivity were incomplete. Our present study indicated a limitation on the efficacy of NMDA receptor antagonist and barbiturate against cerebral ischemia.

Topics: Acetylcholine; Animals; Brain Ischemia; Cell Death; Choline O-Acetyltransferase; Dizocilpine Maleate; Drug Evaluation, Preclinical; Excitatory Amino Acid Antagonists; Female; Gerbillinae; Hippocampus; Male; Nerve Degeneration; Neurons; Neuroprotective Agents; Pentobarbital; Presynaptic Terminals

We previously demonstrated that 2-iminothiazolidine-4-carboxylic acid (2-ICA), formed by cyanide reacting with cysteine, caused glutamate antagonist-sensitive seizures when injected i.c.v. (intracerebroventricular) in mice and produced hippocampal CA1 damage following i.c.v. infusion in rats. In this study, the ability of either 2-ICA, glutamate, proline or NMDA (N-methyl-D-aspartate) injected i.c.v. to produce hippocampal lesions sensitive to glutamate antagonists was compared in mice. Hippocampal CA1 damage was observed 5-days following either a seizure (3.2 mumol) or subseizure (1.0 mumol) dose of 2-ICA. Glutamate (3.2 mumol) or proline (10 mumol) also produced hippocampal damage; glutamate damage was primarily to the CA1 subfield, whereas proline damaged neurons throughout the entire hippocampal formation. NMDA (3.2 nmol) caused seizure activity in all animals with a 50% lethality. No hippocampal damage was observed in surviving mice. Neither MK-801 (dizocilpine maleate) nor CNQX (6-cyano-7-nitroquinoxaline-2,3-dione) pretreatment prevented hippocampal lesions produced by 2-ICA. In contrast, MK-801 significantly reduced the frequency of mice displaying glutamate hippocampal lesions, but failed to block seizures produced by glutamate. MK-801 also protected neurons in the CA2-3 zone and the dentate gyrus, but not in the CA1 region of proline-injected mice. Finally, pretreatment with the mixed metabotropic glutamate receptor (mGluR)1/mGluR2 antagonist-agonist (S)-4-carboxy-3-hydroxyphenylglycine (CHPG) prevented hippocampal damage produced by the mGluR1 agonist (RS)-3,5-dihydroxyphenylglycine (DHPG), but did not protect against 2-ICA hippocampal lesions. These results show that 2-ICA hippocampal CA1 damage is not mediated through ionotropic or metabotropic glutamate receptors. 2-ICA hippocampal damage may represent a neurotoxicity that is distinct from excitotoxic-mediated cell death.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Brain Ischemia; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Glutamic Acid; Hippocampus; Injections, Intraventricular; Male; Neuroprotective Agents; Neurotoxins; Proline; Rats; Rats, Sprague-Dawley; Seizures

The neuroprotective activity of the novel, selective glycine antagonist GV150526 was assessed in the middle artery occlusion (MCAo) model of focal ischemia. Postischemia administration of GV150526 (3 mg/kg i.v.) up to 6 h post-MCAo resulted in a significant reduction of the infarct volume measured histologically 24 h later. The neuronal protection by GV150526 was accompanied by functionally significant protection determined by somatosensory evoked potential (SEP) responses recorded from the primary somatosensory cortex of rats under urethane anesthesia. Experimental occlusion of the MCA 7 days prior to electrophysiological testing induced a clear reduction in the SEP amplitude. GV150526 (3mg/kg, i.v.) was able to protect SEP responses recorded from the hindpaw cortical field in two groups of animals treated either 1 (n = 9) or 6 h (n = 10) post-MCAo. SEP responses recorded from the forepaw cortical field, an area closer to the core of the ischemic damage, were significantly protected only in the group treated 1 h post-MCAo. Histological evaluation of the rat brain regions showed a correlated decrease in the ischemic area of GV150526-treated groups. The volumes of the ischemic brains of both GV150526 groups were statistically different from the MCAo group (P < 0.05). These findings demonstrate that GV150526 is able to prevent the ischemic damage assessed histologically and affect the functional correlates of the ischemia evaluated by the electrophysiological SEP measurements.

Topics: Animals; Arterial Occlusive Diseases; Brain Ischemia; Cerebral Infarction; Disease Models, Animal; Dizocilpine Maleate; Electric Stimulation; Evoked Potentials, Somatosensory; Excitatory Amino Acid Antagonists; Forelimb; Glycine Agents; Hindlimb; Indoles; Male; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Somatosensory Cortex

In vitro data suggest that low tissue pH reduces, whereas extracellular alkalosis potentiates, cerebral anoxic injury via excitotoxic mechanisms. We tested the hypothesis that in vivo metabolic alkalemia potentiates defects in energy metabolism after global incomplete cerebral ischemia (12 min) and reperfusion (180 min) by an N-methyl-D-aspartate (NMDA) receptor-mediated mechanism. Brain ATP, phosphocreatine, and intracellular pH (pHi) were measured by 31P magnetic resonance spectroscopy in anesthetized dogs treated with 1) preischemic intravenous carbicarb buffer (NaHCO3+Na2CO3, Carb, n = 7); 2) carbicarb infusion plus NMDA receptor antagonist MK-801 (MK-801 + Carb, n = 7); 3) an osmotically equivalent volume of 5% NaCl (NaCl, n = 8); or 4) equivalent volume of 0.9% NaCl (Sal, n = 3). Sagittal sinus pH was raised to 7.82 +/- 0.04 before and 7.65 +/- 0.03 during ischemia in Carb vs. 7.72 +/- 0.01 and 7.60 +/- 0.01 in MK-801+Carb, 7.25 +/- 0.02 and 7.15 +/- 0.03 in NaCl, and 7.31 +/- 0.00 and 7.26 +/- 0.01 in Sal, respectively. Ischemic cerebral blood flow (CBF, radiolabeled microspheres), pHi, and ATP reduction were similar among groups. By 180 min of reperfusion, recovery of ATP was greater in MK-801+Carb (104 +/- 6% of baseline), NaCl (93 +/- 6%), and Sal (94 +/- 6%) than in Carb (47 +/- 6%). Intraischemic pHi was similar among groups, and pHi recovery did not vary among groups despite differences in sagittal sinus pH. In Carb, CBF was restored but with delayed hypoperfusion. We conclude that extracellular alkalosis is deleterious to postischemic CBF and energy metabolism, acting by NMDA receptor-mediated mechanisms.

Topics: Adenosine Triphosphate; Alkalies; Animals; Blood Pressure; Brain; Brain Ischemia; Carbonates; Cerebrovascular Circulation; Cranial Sinuses; Dizocilpine Maleate; Dogs; Drug Combinations; Evoked Potentials, Somatosensory; Hydrogen-Ion Concentration; Male; Osmolar Concentration; Receptors, N-Methyl-D-Aspartate; Reperfusion; Sodium Bicarbonate; Sodium Chloride

Glial inclusions containing the microtubule-associated protein tau are present in a variety of chronic neurodegenerative conditions. We now report a rapid and time-dependent increase of tau immunoreactivity within oligodendrocytes after focal cerebral ischemia in the rat. The number of tau positive oligodendrocytes in the ipsilateral subcortical white matter increased six- to eightfold by 40 minutes after permanent middle cerebral artery occlusion (MCAO). Tau was detected using antibodies that label both the N- and C-terminal of the protein, suggesting accumulation of full-length protein within these cells. Pretreatment with the spin trap agent alpha-phenyl-tert-butyl-nitrone (PBN)(100mg/kg) reduced the number of tau-positive oligodendrocytes by 55% in the subcortical white matter of the ischemic hemisphere compared with untreated animals at 40 minutes after MCAO. In contrast, pretreatment with glutamate receptor antagonists MK-801 (0.5 mg/kg) or 2,3-dihydroxy-6-nitro-7-sulpfamoyl-benzo(f)quinoxaline (NBQX) (2 x 30 mg/kg), failed to reduce the number of tau-positive oligodendrocytes after 40 minutes of ischemia. The results indicate that oligodendrocytes respond rapidly to an ischemic challenge and that free radical-mediated mechanisms are involved in the cascade leading to increased tau immunoreactivity.

Topics: Animals; Brain Ischemia; Caudate Nucleus; Cerebral Cortex; Cyclic N-Oxides; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Free Radicals; Immunohistochemistry; Male; Microtubule-Associated Proteins; Nitrogen Oxides; Oligodendroglia; Quinoxalines; Rats; Rats, Inbred F344; Spin Labels; tau Proteins

The cellular mechanisms underlying the neuroprotective action of the immunosuppressant FK506 in experimental stroke remain uncertain, although in vitro studies have implicated an antiexcitotoxic action involving nitric oxide and calcineurin. The present in vivo study demonstrates that intraperitoneal pretreatment with 1 and 10 mg/kg FK506, doses that reduced the volume of ischemic cortical damage by 56-58%, did not decrease excitotoxic damage induced by quinolinate, NMDA, and AMPA. Similarly, intravenous FK506 did not reduce the volume of striatal quinolinate lesions at a dose (1 mg/kg) that decreased ischemic cortical damage by 63%. The temporal window for FK506 neuroprotection was defined in studies demonstrating efficacy using intravenous administration at 120 min, but not 180 min, after middle cerebral artery occlusion. The noncompetitive NMDA receptor antagonist MK801 reduced both ischemic and excitotoxic damage. Histopathological data concerning striatal quinolinate lesions were replicated in neurochemical experiments. MK801, but not FK506, attenuated the loss of glutamate decarboxylase and choline acetyltransferase activity induced by intrastriatal injection of quinolinate. The contrasting efficacy of FK506 in ischemic and excitotoxic lesion models cannot be explained by drug pharmacokinetics, because brain FK506 content rose rapidly using both treatment protocols and was sustained at a neuroprotective level for 3 d. Although these data indicate that an antiexcitotoxic mechanism is unlikely to mediate the neuroprotective action of FK506 in focal cerebral ischemia, the finding that intravenous cyclosporin A (20 mg/kg) reduced ischemic cortical damage is consistent with the proposed role of calcineurin.

Topics: Animals; Brain Ischemia; Cerebrovascular Circulation; Choline O-Acetyltransferase; Dizocilpine Maleate; Glutamate Decarboxylase; Injections, Intraperitoneal; Male; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Tacrolimus

In the present study we have examined the effects of the small organic molecules: NNC 09-0026 ((-)-trans-1-butyl-4-(4-dimethylaminophenyl)-3-[(4-trifluoromethyl-ph eno xy) methyl] piperidine dihydrochloride); SB 201823-A (4-[2-(3,4-dichlorophenoxy)ethyl]-1-pentyl piperidine hydrochloride); NS 649 (2-amino-1-(2,5-dimethoxyphenyl)-5-trifluoromethyl benzimidazole); CNS 1237 (N-acenaphthyl-N'-4-methoxynaphth-1-yl guanidine) and riluzole on human omega-conotoxin sensitive N-type voltage-dependent Ca2+ channel currents (ICa) expressed in HEK293 cells, on Na+ channel currents (INa) in acutely isolated cerebellar Purkinje neurones in vitro and in the gerbil model of global cerebral ischaemia in vivo. Estimated IC50 values for steady-state inhibition of ICa were as follows; NNC 09-0026, 1.1 microM; CNS 1237, 4.2 microM; SB 201823-A, 11.2 microM; NS 649, 45.7 microM and riluzole, 233 microM. Estimated IC50 values for steady-state inhibition of Na+ channel currents were as follows: NNC 09-0026, 9.8 microM; CNS 1237, 2.5 microM; SB 201823-A, 4.6 microM; NS 649, 36.7 microM and riluzole, 9.4 microM. In the gerbil model of global cerebral ischaemia the number of viable cells (mean +/- S.E.M.) per 1 mm of the CA1 was 215 +/- 7 (sham operated), 10 +/- 2 (ischaemic control), 44 +/- 15 (NNC 09-0026 30 mg/kg i.p.), 49 +/- 19 (CNS 1237 30 mg/kg i.p.), 11 +/- 2 (SB 201823-A 10 mg/kg i.p.), 17 +/- 4 (NS 649 50 mg/kg i.p.) and 48 +/- 18 (riluzole 10 mg/kg i.p.). Thus NNC 09-0026, CNS 1237 and riluzole provided significant neuroprotection when administered prior to occlusion while SB 201823-A and NS 649 failed to protect. These results indicate that the Ca2+ channel antagonists studied not only inhibited human N-type voltage-dependent Ca2+ channels but were also effective blockers of rat Na+ channels. Both NNC 09-0026 and CNS 1237 showed good activity at both Ca2+ and Na+ channels and this may contribute to the observed neuroprotection.

Topics: Animals; Brain Ischemia; Calcium Channel Blockers; Calcium Channels; Cell Line; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gerbillinae; Guanidines; Humans; Male; Mollusk Venoms; Neuroprotective Agents; omega-Conotoxins; Peptides; Piperidines; Rats; Receptors, N-Methyl-D-Aspartate; Riluzole; Sodium Channel Blockers; Sodium Channels; Tetrodotoxin

In order to investigate the time course change of local cerebral blood flow (1CBF) and local cerebral glucose metabolism (ICGM) and the effect of MK-801 (dizocilpine), an NMDA receptor antagonist on glucose metabolism in a middle cerebral artery occlusion-reperfusion model, 14C-Iodo-antipyrine and 14C-Deoxyglucose autoradiographic method have been used. The 1CBF was reduced to 0-10% of the control level in the ischaemic core and to 12-40% in the ischaemic penumbra between 60 and 120 min after the onset of the ischaemia. In the ischaemic core, the marked hyperfusion appeared at 15 min and maintained about 30 to 45 min following reperfusion. In the ischaemic penumbra, the hyperfusion during reperfusion was not found. Hypermetabolism occurred at 30 min and reached to the peak at 60 min after the middle cerebral artery (MCA) occlusion both in the ischaemic core and in the penumbra. The shift from hyper- to hypometabolism was observed during the ischaemia. The reperfusion following 2 hours of MCA occlusion facilitated the decrease of cerebral glucose metabolism in the ischaemic region. The pretreatment of MK-801 (0.4 mg/kg) inhibited both increased glucose metabolism during the ischaemia and decreased glucose metabolism during the reperfusion. The effect of limiting decreased glucose metabolism during the reperfusion by MK-801 was remarkable in the ischaemic penumbra. These findings support the hypothesis that excitation-induced hypermetabolism play a major role in the ischaemic insult following focal cerebral vascular occlusion.

Topics: Animals; Autoradiography; Blood Glucose; Brain; Brain Ischemia; Dizocilpine Maleate; Energy Metabolism; Male; Neuroprotective Agents; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Regional Blood Flow; Reperfusion Injury

The authors studied the effects of pre- and postischemic administration of dizocilpine maleate (MK-801) on collateral and regional cerebral blood flow (CBF). The ischemic penumbra appears to benefit most from the neuroprotective effects of MK-801. The precise mechanism by which MK-801 provides this neuroprotection remains controversial. Alterations in CBF have been demonstrated with MK-801 administration, but whether the response is an increase or decrease in flow has remained unclear. A left-sided craniectomy was performed in 20 dogs. A branch of the middle cerebral artery (MCA) was cannulated and collateral blood supply-dependent tissue (CDT) was identified using the "shadow flow" technique. Regional CBF was measured using radiolabeled microspheres. Six dogs received MK-801 (1 mg/kg administered intravenously) before they underwent MCA branch occlusion; the remaining 14 dogs received MK-801 after they underwent MCA occlusion. Cerebral blood flow and vascular pressures were measured 30 and 60 minutes after MK-801 administration. In animals that received MK-801 before MCA occlusion, CBF did not change significantly from baseline values before or after occlusion. In contrast, in animals that received MK-801 after MCA occlusion, CBF was significantly reduced in all regions of the brain, including the CDT. Collateral blood supply-dependent tissue showed a 51.7% reduction in flow, whereas normal CBF was reduced by 29.7%. The MK-801 induced cerebral vasoconstriction in both groups. The neuroprotective effects of MK-801 do not appear to be caused by the augmentation of collateral or global cerebral circulation and, in fact, may block the glutamate-mediated vasodilation that occurs during ischemia.

Topics: Animals; Blood Pressure; Brain Ischemia; Cerebral Arteries; Cerebrovascular Circulation; Collateral Circulation; Dizocilpine Maleate; Dogs; Excitatory Amino Acid Antagonists; Injections, Intravenous; Microspheres; Neuroprotective Agents; Radionuclide Imaging; Radiopharmaceuticals; Receptors, N-Methyl-D-Aspartate; Vascular Resistance; Vasoconstrictor Agents; Vasodilation

Injuries to certain parts of the brain may induce neuronal death in distant areas innervated by the sites of the primary lesion. Such characteristic pathological changes, known as anterograde transneuronal degeneration, may occur at the next and more distant synaptic levels and play a part in the slow progression of some types of system degeneration. Delayed transneuronal degeneration of the substantia nigra pars reticulata (SNr) is one example of this form of cell death, and it occurs as a consequence of a neostriatal lesion caused by focal ischemia, Huntington's disease, or experimental axon-sparing injections of neurotoxin. Ever since the demonstration by Saji and Reis that the administration of GABA receptor agonist effectively prevented delayed transneuronal degeneration of the SNr, the degeneration of nigral reticulata cells has been attributed to the loss of striatal inhibition (Fig. 1A). The latter process severely upset the balance of membrane potential of nigral reticulata cells, producing an effect resembling excitotoxicity. In this report, we describe a continuous intraventricular MK-801 infusion technique that is useful in clarifying the role of glutamatergic action via N-methyl-D-aspartate (NMDA) receptor subclasses involved in exo-focal postischemic death of the SNr.

Topics: Animals; Atrophy; Body Temperature; Brain Ischemia; Cell Survival; Corpus Striatum; Dizocilpine Maleate; Infusion Pumps; Injections, Intraventricular; Male; Nerve Degeneration; Neurons; Neuroprotective Agents; Rats; Rats, Wistar; Rectum; Substantia Nigra

AMPA antagonists have been shown to be remarkably neuroprotective in models of global ischemia, but the data in focal ischemia remain controversial. We, therefore, studied the dose-response characteristics and the time window of efficacy of the AMPA antagonist NBQX in a rat model of permanent focal ischemia. NBQX 40, 60 or 100 mg/kg i.v., substantially reduced infarct size. Neuroprotection was maintained when the initiation of drug administration was withheld for 15, 45 or 90 min after permanent middle cerebral artery occlusion. Furthermore, NBQX did not induce heat shock protein in cingulate cortex, as do some N-methyl-D-aspartate antagonists. Thus, the compound is a potent neuroprotectant in focal ischemia and has an unusually long time window of effectiveness.

Topics: Animals; Brain; Brain Ischemia; Cerebral Arteries; Cerebral Infarction; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Heat-Shock Proteins; Male; N-Methylaspartate; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA

A series of 1,2,3,4-tetrahydroisoquinoline derivatives were synthesized and evaluated for anticonvulsant activity against intracerebro-ventriculas (i.c.v.) N-methyl-D-aspartate (NMDA)-induced seizures in mice. Among these compounds, (+)-1-methyl-1-phenyl-1,2,3,4-tetrahydroisoquinoline hydrochloride ((+)-1a, FR115427) was the most effective anticonvulsant, and also protected CA1 hippocampal neurons from ischemia-induced neuronal degeneration in rats at 32 mg/kg i.p. In addition, (+)-1a showed anti-hypoxic activity in mice at 3.2-32 mg/kg i.p. The absolute configuration at the C-1 position of the isoquinoline ring was determined to be S by a single-crystal X-ray analysis of (+)-1a (+)-di-p-toluoyl-D-tartrate. Structure-activity relationships with regard to the anticonvulsant activity of this series of compounds are discussed, and the three-dimensional structures of (S)-(+)-1a and MK801 are compared.

Topics: Animals; Anticonvulsants; Brain Ischemia; Crystallography, X-Ray; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hippocampus; Hypoxia; Isoquinolines; Magnetic Resonance Spectroscopy; Male; Mice; Mice, Inbred ICR; Molecular Structure; Neuroprotective Agents; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Seizures; Stereoisomerism; Structure-Activity Relationship; Tetrahydroisoquinolines

We assessed the efficacy of recombinant human interleukin-1 receptor antagonist (rhIL-1ra) on brain injury and edema formation after permanent middle cerebral artery occlusion (MCAo) in the rat. Previous studies showed that low amounts of rhIL-1ra injected directly into the brain significantly decreased infarct size after MCAo or excitotoxic injury in rats. Peripheral administration of rhIL-1ra (100 mg/kg sc at 0, 4, 8, 12, and 18 h after MCAo) significantly inhibited infarct size, by 46% (P < 0.05), measured at 24h. This was greater than the effect of MK801 administered immediately after MCAo (4 mg/kg ip, 0 h) which did not significantly reduce infarct size. rhIL-1ra (100 mg/kg also significantly inhibited cerebral edema formation by 49% (p< 0.05 measured 24 h after MCAo, but did not reduce edema formation measured 2 h after MCAo, but did not reduce edema formation measured 2 h after MCAo. Inhibition of infarction by rhIL-1ra was dependent on dose and time of administration. Together the results demonstrate that peripherally administered rhIL-1ra at high doses is able to mimic the efficacy of low dose of rhIL-1ra administered directly into the brain in a rodent model of stroke and that protection observed with rhIL-1ra was better than that offered by MK801 in this model.

Topics: Animals; Blood Glucose; Brain Damage, Chronic; Brain Edema; Brain Ischemia; Cerebral Infarction; Dizocilpine Maleate; Humans; Injections, Intravenous; Injections, Subcutaneous; Male; Rats; Rats, Sprague-Dawley; Receptors, Interleukin-1; Recombinant Proteins

To assess the effects of the novel sigma ligand JO 1994 on behavioural, histological and autoradiographical changes following global ischaemia, the Mongolian gerbil was used. Three experiments were carried out and in each case ischaemia was induced by bilateral carotid occlusion (BCO) for 5 min. In the first experiment we examined the effects of JO 1994 administered at doses of 0.25, 0.5 and 1 mg/kg i.p. 1 h before 5 min BCO on histological parameters 96 h after surgery. In the second experiment the effects of JO 1994 administered at doses of 2.5, 5, 10 and 20 mg/kg i.p. 1 h before 5 min BCO on locomotor activity 24, 48 and 72 h after surgery and on histological parameters 96 h after surgery was examined. In the third experiment the effects of JO 1994 (2.5 and 5 mg/kg i.p.), BMY 14802 (1 and 10 mg/kg i.p.) and MK-801 (2.5 mg/kg i.p.) administered 30 min, 6, 24, 48, 72, 96 and 120 h post-surgery on the densities of M1 and M2 muscarinic receptors in 35 brain regions, 7 days after surgery was examined. Results indicated that 5 min bilateral carotid occluded animals were hyperactive 24, 48 and 72 h after surgery. JO 1994 attenuated this hyperactivity. Extensive neuronal death was observed in the CA1 layer of the hippocampus in 5 min BCO animals 96 h after surgery. The low doses of JO 1994 (0.25, 0.5 and 1 mg/kg) had no effect on the ischaemia-induced cell death. However JO 1994 (2.5, 5, 10 and 20 mg/kg i.p.) protected against the neuronal death of cells in the CA1 layer (P < 0.01-0.03). There was a large loss of M1 and M2 receptors in the CA1 regions of the hippocampus. MK-801, BMY 14802 and JO 1994 provided significant (P < 0.01) protection against this ischaemia-induced receptor loss.

Topics: Animals; Anti-Anxiety Agents; Autoradiography; Behavior, Animal; Brain; Brain Chemistry; Brain Ischemia; Cell Death; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gerbillinae; Male; Motor Activity; Muscarinic Antagonists; Neuroprotective Agents; Organic Chemicals; Pyrimidines; Quinuclidinyl Benzilate; Receptors, sigma

Glutamate receptors are numerous on the ischemia vulnerable CA-1 pyramidal cells. Postischemic use of the AMPA antagonist NBQX has shown up to 80% protection against cell death. Three aspects of this were studied: In the first study, male Wistar rats were given NBQX (30 mg/kg x 3) either 20 hours or immediately (0 h) before 12 min of 4-vessel occlusion with hypotension. After six days of reperfusion comparison with an untreated group showed almost full protection in the 0 h group (4% cell loss, p < 0.001) but only slight protection in the 20 h group (62% cell loss, p < 0.05). After 12 min of ischemia in the present model, eosinophilic CA-1 cells are seen from day 2 on. Since there could be a late, deleterious calcium influx via NMDA receptors, one group of ischemic rats was given MK-801 (5 mg/kg i.p.) 24 hours after ischemia. However, quantitation 6 days later of remaining CA-1 cells showed no protection. In the third study referred here, two groups of ischemic rats were given NBQX (30 mg/kg x 3) immediately after ischemia. The groups survive for six and 21 days, respectively. Counting of CA-1 pyramidal cells showed an equal, significant protection in both groups (approx 20% cell loss).

Topics: Animals; Brain Damage, Chronic; Brain Ischemia; Brain Mapping; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Administration Schedule; Excitatory Amino Acid Antagonists; Glutamic Acid; Hippocampus; Male; Neurons; Premedication; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate

Post-treatment with the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 reduces hypoxic-ischemic brain injury in immature animals. To elucidate possible mechanisms, cerebral glucose utilization (CMRglc) and cerebral blood flow (CBF) were measured 1-5 h after hypoxia-ischemia and administration of MK-801 in 7-day-old rats. After 100 min of unilateral hypoxia-ischemia, half of the pups were injected with MK-801. CMRglc was assessed by the [14C]deoxyglucose (2-DG) method. The brains were analyzed either by autoradiography or for energy metabolites and chromatographic separation of 2-DG-6-phosphate and 2-DG. CBF was measured by the autoradiographic [14C]iodoantipyrine method. Mean CMRglc in the cerebral cortex was increased ipsilaterally after hypoxia-ischemia to 15 +/- 3.3 mumol 100 g-1 min-1 (p < 0.01) and areas with CMRglc > 20 mumol 100 g-1 min-1 amounted to 8.0 +/- 7.7 mm2 in the ipsilateral hemisphere compared with 1.2 +/- 1.6 mm2 contralaterally (p < 0.001). Treatment with MK-801 decreased CMRglc bilaterally (p < 0.05) and reduced ipsilateral areas with increased CMRglc by 64% (p < 0.01). CBF was unaltered after hypoxia-ischemia and by MK-801 treatment. In conclusion, regional glucose hyperutilization in the parietal cortex after hypoxia-ischemia was attenuated by MK-801; this may have relevance to the neuroprotective effect of NMDA-receptor antagonists in this model.

Topics: Animals; Autoradiography; Brain; Brain Ischemia; Deoxyglucose; Dizocilpine Maleate; Female; Glucose; Glucose-6-Phosphate; Glucosephosphates; Hypoxia, Brain; Kinetics; Male; Parietal Lobe; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate

In vitro combinations of MK-801, an NMDA receptor antagonist, and alpha-phenyl-tert-butyl-nitrone (PBN), a free radical scavenger, have been tested for possible additive neuroprotective effects against anoxia/hypoglycemia (Ax/Hg)-induced neuronal damage. Rat organotypic hippocampal slice cultures were exposed to Ax/Hg for different lengths of time to vary the severity of the insult. Cell death (CD) was assessed using propidium iodide fluorescence and expressed as a percentage of the total neuronal cells present. Pretreatment with PBN alone (500 microM) provided significant protection against moderate ischemic injury and reduced CD from 65% in controls to 2% in the treated group (P < 0.003). A longer ischemic exposure time caused more neuronal damage, which was only slightly reduced by PBN, but significantly reduced by MK-801 (30 microM) (4% CD with MK-801 vs 75% CD in controls; P < 0.0003). With a further increase in the time of ischemic exposure, MK-801 was still protective (33% CD with MK-801 vs 90% CD in controls; P < 0.002), although the combination MK-801 + PBN was more efficient (7% CD with combination, P < 0.01 compared to MK-801 alone). With yet a further increase in the ischemic exposure, PBN or MK-801 alone was not protective; however, a combination of the two still provided significant protection (64% CD with combination vs 100% CD with MK-801 alone; P < 0.01). PBN was protective when administered up to 2 h after Ax/Hg (66% CD in controls vs 36% CD with PBN 500 microM; P < 0.007). The combination MK-801 + PBN was able to increase the therapeutic window up to 3 h (61% CD in controls vs 41% with PBN alone vs 7% with MK-801 + PBN; P < 0.002 compared to PBN alone). In conclusion, the combination of MK-801 and PBN increases both the efficacy and the time window of protection against ischemia.

Topics: Animals; Brain Ischemia; Cyclic N-Oxides; Dizocilpine Maleate; Dose-Response Relationship, Drug; Free Radical Scavengers; Hippocampus; Nitrogen Oxides; Organ Culture Techniques; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

The neuroprotective activity of basic fibroblast growth factor (bFGF) in combination with the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 was evaluated in organotypic hippocampal slice cultures. Oxygen/glucose deprivation produced neuronal damage which was assessed using propidium iodide fluorescence. Treatment with increasing doses of bFGF demonstrated significant neuroprotection that was optimal at 10 ng ml-t. This effect was diminished at higher concentrations. MK-801, at the optimal concentration of 30 microM, demonstrated greater neuroprotective efficacy than bFGF. However, bFGF significantly enhanced the protection conferred by MK-801 alone. These results suggest that neurotrophic factors such as bFGF may augment the neuroprotective effects of NMDA antagonists against ischemic neuronal injury.

Topics: Animals; Brain Ischemia; Cell Survival; Dizocilpine Maleate; Drug Synergism; Excitatory Amino Acid Antagonists; Fibroblast Growth Factor 2; Hippocampus; Neurons; Neuroprotective Agents; Organ Culture Techniques; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

In rodents, postischemic hypothermia can provide robust and long-term functional and histological neuroprotection, even when intervention is delayed for several hours following ischemia. This generates a need to follow temperature precisely for many hours, perhaps several days if a hypothermic effect is to be studied or excluded. Such protracted temperature control (> 24 h) is difficult and often lethal when performed under general anesthesia. In awake animals, manual temperature control is safer, but exceedingly time consuming and tedious, and is impractical for large experiments. The present method allows for continuous brain temperature measurement and control in free-moving rats and gerbils. Brain temperature was measured by wireless AM probes while feedback regulation was achieved by servo-control of a lamp, fan and water misting system. Hypothermia was easily induced and maintained for 24 h at 32 degrees C in both gerbils and rats. Gerbils also tolerated 24 h at 32 degrees C followed by 24 h at 34 degrees C. This 'exposure technique' is capable of safely producing lengthy periods of mild hypothermia in rats and gerbils. Furthermore, this method can clamp temperature when temperature-altering drugs are given. For example, temperature was maintained in MK-801 drugged gerbils. The system is, therefore, eminently suitable for drug neuroprotection studies in brain ischemia.

Topics: Animals; Automation; Body Temperature; Brain; Brain Ischemia; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gerbillinae; Hypothermia, Induced; Ischemic Preconditioning; Neurophysiology; Rats; Rats, Sprague-Dawley; Telemetry; Temperature

The present study was designed to evaluate whether the neuroprotective efficacy of MK-801 in focal cerebral ischemia was dependent on strain and/or vendor differences. MK-801 (0.12 mg/kg i.v. bolus followed by 0.108 mg/kg/h infusion or 0.60 mg/kg i.v. bolus followed by 0.540 mg/kg/h infusion) or saline was administered just after intraluminal middle cerebral artery occlusion. Administration of 0.540 mg/kg/h MK-801 provided strain/line-dependent neuroprotection in the following rank order: Simonsen Laboratories Sprague-Dawley rats > Simonsen Laboratories Wistar rats > Taconic Laboratories Sprague-Dawley rats. After 0.108 mg/kg/h MK-801 treatment, Simonsen Laboratories Wistar rats were the only strain/line that were significantly neuroprotected. These results indicate that the neuroprotective effect of an experimental drug may be influenced by rat strain and vendor differences.

Topics: Animals; Animals, Laboratory; Body Temperature; Brain Ischemia; Cerebral Arteries; Cerebral Cortex; Dizocilpine Maleate; Dose-Response Relationship, Drug; Ligation; Male; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Rats, Wistar

To evaluate the role played by nitric oxide in global cerebral ischaemia we examined the effects of 7-nitroindazole and a sodium salt of 7-nitroindazole (inhibitors of neuronal nitric oxide (NO) synthase) and NG-nitro-L-arginine methyl ester (a more general inhibitor of NO synthase) in the gerbil model of cerebral ischaemia. Four experiments were carried out. In the first experiment, animals were either sham-operated, subjected to 5 min bilateral carotid occlusion (BCAO) or administered 7-nitroindazole or NG-nitro-L-arginine methyl ester immediately after occlusion followed by three further doses at 3, 6 and 24 h post-occlusion. In the second experiment, we examined the effects of a sodium salt of 7-nitroindazole, which is more soluble than 7-nitroindazole, using the same protocol. In the third experiment, the effects of the sodium salt of 7-nitroindazole administered at 10 mg/kg at 0, 3, 6, 24, 27, 30, 33, 52, 55, 72, 75 and 78 h post-occlusion or at 0.05 mg/h for 72 h via mini-pumps were evaluated. In separate experiments, we examined the effects of three reference compounds dizocilpine (MK-801), 2, 3-dihydroxy-6-nitro-7-sulphamoyl-benz(F)-quinoxaline (NBQX) and eliprodil using the same model. Extensive neuronal death was observed in the CA1 layer of the hippocampus in 5 min bilateral carotid occluded animals 5 days after surgery. Both 7-nitroindazole and NG-nitro-L-arginine methyl ester provided significant neuroprotection (P < 0.01) against this neuronal death. The sodium salt of 7-nitroindazole showed no protection when administered up to 12 times post-occlusion, but did provide significant (P < 0.01) neuroprotection when administered via mini-pump. The neuroprotection was similar to that provided by MK-801 and eliprodil, but not as good as that observed with NBQX. These results indicate that nitric oxide plays a role in ischaemic cell death and that selective neuronal nitric oxide synthase inhibitors can protect against ischaemic brain damage.

Topics: Animals; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Enzyme Inhibitors; Gerbillinae; Indazoles; Male; Neuroprotective Agents; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Piperidines; Quinoxalines

We investigated electrophysiological responses induced by ischemia-like insult (anoxia and aglycemia, AA) in the rat hippocampal CA1 pyramidal cells in an in vitro slice preparation devoid of glial metabolism. In the slice treated with fluorocitrate (100 microM), a glia-specific metabolic inhibitor, 10 min AA induced hyperexcitation as evidenced by an appearance of multiple population spikes evoked by stimulation of the Schaffer collateral/commissural pathway in the CA1 region prior to elimination of the response. Readministration of oxygen and glucose failed to restore the population spike amplitude. Intracellular recordings revealed that 10 min AA induced slow EPSPs with relative long duration. The induction of the slow EPSPs was followed by a rapid membrane depolarization with a large amplitude. When the fluorocitrate-treated slice was exposed to MK-801 (10 microM), a non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist, 10 min AA failed to induce either the hyperexcitation of synaptic responses or the rapid depolarization. Furthermore, synaptic responses were fully restored after readministration of oxygen and glucose. In contrast, neither the synaptic hyperexcitation nor the rapid depolarization was observed during 10 min AA in the hippocampal CA1 pyramidal cells of the control slice. In addition, an irreversible synaptic failure associated with AA was not induced in the control slice. These results strongly suggest that fluorocitrate increases NMDA receptor-dependent AA-induced damage in the hippocampal slice by interfering glial spatial buffering of K+.

Topics: Animals; Brain Ischemia; Citrates; Dizocilpine Maleate; Evoked Potentials; Excitatory Amino Acid Antagonists; Hippocampus; Hypoxia, Brain; In Vitro Techniques; Male; Membrane Potentials; Neuroglia; Neuroprotective Agents; Pyramidal Cells; Rats; Rats, Wistar; Synaptic Transmission

We have characterized the induction of the mitogen-inducible form of cyclo-oxygenase, COX-2, during focal cerebral ischemia following permanent middle cerebral artery occlusion (MCAO) in the rat. Marked unilateral induction of COX-2 mRNA was detected in ischemic regions ipsilateral to the occlusion. A significant increase in COX-2 mRNA was detected in "core" and "penumbra" regions of the cerebral cortex between 4 and 24 h after occlusion; this was most marked at 4 h in the penumbra region, in which a 19-fold increase above untreated control levels was detected. A smaller but significant induction was also detected at 4 h in the caudate. A correlation was demonstrated between the extent of COX-2 mRNA induction in cortical regions at 4 h and the severity of tissue damage subsequently detected at 24 h post MCAO. MK-801 significantly attenuated the induction of COX-2 mRNA in the penumbra region at 4 h. The demonstration of COX-2 induction following experimental ischemia highlights the importance of this reaction and its products and by-products, for example, free radicals, in the tissue response to this insult.

Topics: Animals; Brain Ischemia; Cyclooxygenase 2; Dizocilpine Maleate; Isoenzymes; Male; Neuroprotective Agents; Prostaglandin-Endoperoxide Synthases; Rats; Rats, Sprague-Dawley; RNA, Messenger

We investigated the temporo-spatial expression of astrocyte glial fibrillary acidic protein (gfap) and sulfated glycoprotein 2 (sgp-2) mRNAs in comparison to 70-kDa heat shock protein (hsp70) mRNA by in situ hybridisation in rats subjected to permanent occlusion of the middle cerebral artery (MCA). Gfap mRNA started to increase in the cingulate cortex of the lesioned hemisphere 6 h after MCA occlusion and gradually spread over the lateral part of the ipsilateral cortex and the striatum from 12 h to 3 days, peaking at 3 days after MCA occlusion. Gfap mRNA also increased in the contralateral cingulate cortex and corpus callosum at 12 and 24 h. Hsp70 mRNA increased markedly in the ipsilateral cortex adjacent to the ischemic lesion, and slightly within the lesion area from 3 to 24 h and disappeared after 3 days. By 7 days, gfap and sgp-2 mRNAs were increased markedly in the peri-infarct area, and in the ipsilateral thalamus parallel with the delayed neuronal damage, whereas the widespread increase of gfap mRNA in the ipsilateral hemisphere declined. Post-occlusion treatment with the glutamate receptor antagonists MK-801 and NBQX slightly attenuate the induction of gfap but did not qualitatively affect the topical expression pattern. Within the cingulate cortex MK-801 treatment resulted in a significant decrease of the signal intensity at all survival times, reflecting most likely an attenuation of lesion-induced spreading depression like depolarization waves by MK-801. The area of hsp70 expression was reduced by both MK-801 and NBQX, most likely reflecting the decrease of the lesion area by both treatment regimens. Our study thus revealed an early and widespread increase of gfap mRNA in the non-ischemic area including the contralateral hemisphere starting between 3 and 6 h, and a delayed circumscribed expression in the peri-infarct border zone after 1 week. Comparison with the expression of hsp70 mRNA suggests that the absence of an early gfap mRNA induction in the peri-lesion zone reflects an impairment of astrocytic function which may be of importance for infarct growth during the early evolution of the pathological process.

Topics: Animals; Astrocytes; Brain Ischemia; Clusterin; Dizocilpine Maleate; Electroencephalography; Excitatory Amino Acid Antagonists; Glial Fibrillary Acidic Protein; Glycoproteins; HSP70 Heat-Shock Proteins; In Situ Hybridization; Male; Molecular Chaperones; Quinoxalines; Rats; Rats, Inbred F344; RNA, Messenger; Time Factors

Induction of chemical anoxia, using sodium azide in cerebellar granule cells maintained in primary culture, was evaluated as an in vitro assay for screening of potential neuroprotective compounds. The purpose of this study was to evaluate sodium azide as an alternative to cyanide salts, compounds which, despite their unfavorable characteristics, are often used in assays for chemical anoxia. The viability of neuronal cultures after treatment with azide, with or without preincubation with calcium channel blockers, tetrodotoxin (TTX), or glutamate receptor antagonists, was monitored by subsequent incubation with the tetrazolium dye MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide), followed by isopropanol extraction and spectrophotometric quantification of cellularly reduced MTT. The azide-induced degeneration of neurons was shown to be dependent on the concentration as well as on the duration of incubation with submaximal concentrations of azide. Incubation of the neurons with nifedipine, a blocker of L-type voltage-sensitive calcium channels (L-VSCC), or with the noncompetitive N-methyl-D-aspartate (NMDA) subtype glutamate receptor antagonist MK-801, prior to addition of submaximal concentrations of azide, significantly attenuated azide-induced neuronal death. Blockers of N-type and Q-type VSCC (omega-conotoxin MVIIA and MVIIC, respectively) and the P-type VSCC blocker omega-agatoxin IVA had no effect in this assay. The sodium channel blocker TTX was without effect when added to neurons under depolarizing conditions, but potently and effectively protected cells when experiments were performed in a nondepolarizing buffer. The results show that chemical anoxia induced by incubation of cultured neurons with azide leads to detrimental effects, which may be quantitatively monitored by the capability of the cells to reduce MTT. This procedure is a suitable method for screening of compounds for possible protective effects against neuronal death induced by energy depletion. In addition, the results suggest involvement of L-type VSCC as well as of glutamate receptors in the pathways leading to neuronal degradation induced by energy depletion in cerebellar granule neurons. This would further support the notion that these pathways might be important in neurodegeneration induced by cerebral ischemia or anoxia.

Topics: Animals; Azides; Brain Ischemia; Cells, Cultured; Cerebellum; Dizocilpine Maleate; Dose-Response Relationship, Drug; Female; Hypoxia; Mice; Mice, Inbred Strains; Mutagens; Nifedipine; Pregnancy; Sodium Azide

Acidosis is a universal response of tissue to ischemia. In the brain, severe acidosis has been linked to worsening of cerebral infarction. However, milder acidosis can have protective effects. As part of our investigations of the therapeutic window in our neuronal tissue culture model of ischemia, we investigated the effects of acidosis during recovery from brief simulated ischemia. Ischemic conditions were simulated in dissociated cortical cultures by metabolic inhibition with potassium cyanide to block oxidative metabolism and 2-deoxyglucose to block glycolysis. Lowering the extracellular pH (pH0) to 6.2 during metabolic inhibition had no effect on injury, as measured by lactate dehydrogenase release from cultures after 24 h of recovery. Lowering the pH0 during the first hour of recovery, in contrast, had profound protective effects. When the duration of metabolic inhibition was lengthened to 30 min, most of the protective effects of the NMDA receptor antagonist MK-801 were lost. However, the protective effects of acidosis were unchanged. This suggested that the protective effects of extracellular acidosis could be due to more than blockade of NMDA receptors. Intracellular acidosis might be responsible. To test this, recovery of intracellular pH (pH1) was slowed by incubation with blockers of Na+/H+ exchangers at normal pH0. The two compounds tested, dimethylamiloride and harmaline, had protective effects when present during recovery from metabolic inhibition. Measurements of pH1 confirmed that the blockers slowed recovery from intracellular acidosis; more rapid pH1 recovery was correlated with injury. The protective effects of acidosis could be reversed by brief incubation with the protonophore monensin, which rapidly normalized pH1. These results are the first demonstration of the protective effects of blocking Na+/H+ exchange in a model of cerebral ischemia. The protective effects of acidosis appear to arise either from suppressing pH-sensitive mechanisms of injury or from blocking sodium entry due to Na+/H+ exchange.

Topics: Acidosis; Amiloride; Animals; Brain Ischemia; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Extracellular Space; Female; Hydrogen; Hydrogen-Ion Concentration; Ionophores; Neuroprotective Agents; Pregnancy; Protons; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Sodium; Sodium-Hydrogen Exchangers

Diffusion-weighted single voxel experiments conducted at b-values up to 1 x 10(4) smm-2 yielded biexponential signal attenuation curves for both normal and ischemic brain. The relative fractions of the rapidly and slowly decaying components (f1, f2) are f1 = 0.80 +/- 0.02, f2 = 0.17 +/- 0.02 in healthy adult rat brain and f1 = 0.90 +/- 0.02, f2 = 0.11 +/- 0.01 in normal neonatal rat brain, whereas the corresponding values for the postmortem situation are f1 = 0.69 +/- 0.02, f2 = 0.33 +/- 0.02. It is demonstrated that the changes in f1 and f2 occur simultaneously to those in the extracellular and intracellular space fractions (fex, f(in)) during: (i) cell swelling after total circulatory arrest, and (ii) the recovery from N-methyl-D-aspartate induced excitotoxic brain edema evoked by MK-801, as measured by changes in the electrical impedance. Possible reasons for the discrepancy between the estimated magnitude components and the physiological values are presented and evaluated. Implications of the biexponential signal attenuation curves for diffusion-weighted imaging experiments are discussed.

Topics: Age Factors; Animals; Animals, Newborn; Brain Edema; Brain Ischemia; Diffusion; Dizocilpine Maleate; Electric Impedance; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Extracellular Space; Intracellular Fluid; Magnetic Resonance Spectroscopy; N-Methylaspartate; Phantoms, Imaging; Rats; Rats, Wistar; Water

Following cerebral ischemia, the extracellular concentration of excitatory amino acids increases, and the excitatory cell death may play an important role contributing to ischemic neuronal damage. Although sequential metabolic changes in permanent local cerebral ischemia have been reported, the effect of reperfusion in local cerebral ischemia on glucose metabolism is less clear. In order to investigate the time course change of glucose metabolism in a middle cerebral artery occlusion-reperfusion model and the effect of dizocilpin (MK-801) on glucose metabolism, the 14C-Deoxyglucose method was used. Hypermetabolism occurred at 30 min after the middle cerebral artery (MCA) occlusion, and reached a peak at 60 min after ischemia in both ischemic core and penumbra. The shift from hyper- to hypometabolism was observed after the ischemia. The reperfusion facilitated the decrease of cerebral glucose metabolism in the ischemic region following 2 h of MCA occlusion. The pretreatment of MK-801 (0.4 mg kg-1) inhibited both increased glucose metabolism during ischemia and decreased glucose metabolism during reperfusion. These findings support the hypothesis that excitation-induced hyper-metabolism plays a major role in the ischemic insult following focal cerebral vascular occlusion.

Topics: Animals; Arterial Occlusive Diseases; Blood Gas Analysis; Blood Glucose; Body Temperature; Brain Chemistry; Brain Ischemia; Cerebral Arteries; Disease Models, Animal; Dizocilpine Maleate; Glucose; Male; Neuroprotective Agents; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Reperfusion Injury; Time Factors

Because of ontogenic influences on the pathophysiologic mechanisms of brain injury in the perinatal brain, and in particular, the incomplete development of adenosine receptor systems, we investigated the potential for adenosine to provide cerebro-protection in a well established newborn rat model of hypoxia-ischemia. Fifteen litters of postnatal d 7 animals were subjected to unilateral carotid ligation and exposure to hypoxia (8% oxygen) for 3 h. Immediately after hypoxia-ischemia, animals received either the adenosine deaminase inhibitor deoxycoformycin (DCF; 2.5 mg/kg intraperitoneally) or the adenosine uptake inhibitor propentofylline (PPF; 10 mg/kg intraperitoneally); paired littermates received an equivalent volume of normal saline. On postnatal d 14, injury or protection was assessed by differences in hemispheric weights, morphometric determinations of infarct area, and histopathologic analyses. DCF resulted in a 34% (p = 0.02) and 31% (p = 0.03) reduction in hemispheric weight disparities and infarct area, respectively; for PPF, these reductions were 46% (p = 0.03) and 32% (p = 0.04), respectively. Light microscopic examinations of striatum, thalamus, hippocampus, and cortex revealed that both drugs significantly improved histologic scores as well. Measurements in six separate litters indicated that neither drug significantly reduced core body temperature for at least 6 h postadministration. These findings indicate that potentiation of endogenous adenosine levels in the perinatal brain can significantly ameliorate brain injury. Each of these treatment strategies was effective even when administered after the hypoxic-ischemic insult. Thus, further investigations of adenosinergic therapies are warranted in this and other perinatal models of cerebral ischemia to elucidate in detail their potential for clinical application.

Topics: Adenosine; Animals; Animals, Newborn; Body Temperature; Brain; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Enzyme Inhibitors; Neuroprotective Agents; Organ Size; Pentostatin; Rats; Rats, Sprague-Dawley; Xanthines

The therapeutic time window for N-methyl-D-aspartate (NMDA) antagonists, non-NMDA antagonists, and glutamate release inhibitors in focal models of ischemia appears to be about 1-2 h. In contrast, a free radical spin trap was found to have an improved therapeutic window. We compared the therapeutic time windows of the NMDA antagonist dizolcilpine maleate (MK-801), the glutamate release inhibitor lamotrigine, and the free radical spin trap n-tert-butyl-alpha-(2-sulfophenyl)-nitrone (S-PBN) against striatal lesions produced by the mitochondrial toxin malonate, which produces histotoxic hypoxia. Lamotrigine exerted neuroprotective effects when administered at 1 h before malonate injections. MK-801 protected at 1 h before and 1 h after malonate injections, whereas S-PBN showed efficacy when administered up to 6 h after malonate injections. Striatal injections of malonate produced a rapid increase in lactate production and early changes in diffusion-weighted imaging as assessed by magnetic resonance imaging. Therefore, the time course to evolve a lesion in our model of histotoxic hypoxia is comparable with that of other models of focal ischemia. These findings provide direct evidence that a free radical spin trap has an improved therapeutic window compared to an NMDA antagonist and a glutamate release inhibitor. This could be a therapeutic advantage in the treatment of clinical stroke patients.

Topics: Animals; Benzenesulfonates; Brain Ischemia; Corpus Striatum; Dizocilpine Maleate; Dose-Response Relationship, Drug; Free Radicals; Lamotrigine; Magnetic Resonance Imaging; Male; Malonates; Oxygen; Rats; Rats, Sprague-Dawley; Spin Trapping; Time Factors; Triazines

In contrast to intraischemic hypothermia, immediate postischemic hypothermia (30 degrees C) has been shown to delay but not chronically protect the CA1 hippocampus from transient global forebrain ischemia. The inability of a relatively short postischemic hypothermic period to protect chronically might involve a delayed or secondary injury mechanism. We determined whether delayed treatment with the noncompetitive N-methyl-D-aspartate receptor antagonist MK-801 (dizocilpine), alone or in combination with immediate postischemic hypothermia, would chronically protect histopathologically. Wistar rats underwent 10 min of normothermic forebrain ischemia induced by bilateral common carotid artery occlusion plus hypotension (50 mg Hg). Four ischemia groups were studied after normothermic (37 degrees C) ischemia: no treatment; 3 h of immediate postischemic hypothermia (30 degrees C); delayed MK-801 treatment (4 mg/kg) on postischemic days 3, 5, and 7; and postischemic hypothermia combined with multiple MK-801 treatments. Two months after the ischemic insult, rats were perfusion-fixed for quantitative histopathological assessment. Postischemic hypothermia alone or MK-801 treatment alone failed to protect the CA1 hippocampus chronically. However, immediate postischemic hypothermia combined with delayed MK-801 treatment led to significant increases in normal CA1 neuron counts per microscopic field compared with normothermic ischemia. For example, neuronal counts within the hippocampal CA1 areas were 58 +/- 39 (mean +/- SD) in normothermic ischemic rats compared with 395 +/- 198 in rats treated with postischemic hypothermia and MK-801. Chronic survival also led to pronounced striatal damage. Within the dorsolateral striatum, significant protection was documented with either postischemic hypothermia alone or delayed MK-801 treatment alone.

Topics: Animals; Brain Ischemia; Cell Count; Cell Survival; Dizocilpine Maleate; Hypothermia, Induced; Male; Neurons; Neuroprotective Agents; Prosencephalon; Rats; Rats, Wistar

Experimental studies of stroke in animal models have traditionally relied on histological endpoints for the measurement of neuroprotection. In this study, we used in vivo and dynamic MRI to quantify the neuroprotective effects of the non-competitive NMDA antagonist MK801. Four hours of occlusion followed by 6 h of reperfusion was performed in a rabbit model of focal cerebral ischemia. Spin-echo T2-weighted (T2W) MRI was used to quantify ischemic lesion volumes. Hemispheric measurements of perfusion deficits were assessed by using dynamic susceptibility-contrast MRI to map the first-pass transit of injected GdDTPA. Histological correlates of infarction were quantified using tetrazolium staining. Animals treated with 2 mg/kg MK801 infused immediately post-occlusion (n = 6) were compared with untreated controls (n = 8). T2W MRI scans obtained after 6 h of reperfusion showed high-intensity lesions in the ischemic basal ganglia and cortex. MK801-treated animals showed significantly decreased lesion volumes compared to untreated controls (7.3 +/- 3.2% treated vs 20.7 +/- 4.8% control, p < 0.05). Lesion volumes measured with MRI were significantly correlated with tetrazolium-defined infarct volumes (r = 0.766, p = 0.004). Dynamic MRI demonstrated the phenomenon of delayed hypoperfusion in the ischemic hemisphere during the late reperfusion phase; relative cerebral blood volume (rCBV) was 45.2 +/- 10.3% in untreated animals. MK801 slightly improved these deficits although the differences did not reach statistical significance (rCBV = 77.0 +/- 9.7%, p = 0.128).(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Brain Ischemia; Cerebrovascular Disorders; Coloring Agents; Disease Models, Animal; Dizocilpine Maleate; Magnetic Resonance Imaging; N-Methylaspartate; Neurons; Neuroprotective Agents; Rabbits; Tetrazolium Salts

Brain microdialysis was used to study changes in the glutamate and aspartate extracellular concentrations in the striatum of conscious rats submitted to 30 minutes cerebral ischaemia, using the four-vessel occlusion model. Perfusion of the N-methyl-D-aspartate (NMDA) receptor channel blockers, dizocilpine (MK-801; 75 microM) and Mg2+ (2.5 mM), inhibited the ischaemia-induced accumulation of glutamate and aspartate. The AMPA/kainate receptor antagonist, 2,3-dihydroxy-6-nitro-7-sulfamylbenzo (F) quinoxaline (NBQX; 15 microM and 450 microM) had no effect on glutamate and aspartate levels during ischaemia. On the other hand, omission of Ca2+ from the perfusing solution did not alter the increases in glutamate and aspartate induced by ischaemia. These results suggest that the glutamate and aspartate accumulation in four-vessel occlusion ischaemia is mediated by activation of NMDA receptors in a Ca2+ independent manner.

Topics: Animals; Aspartic Acid; Brain Ischemia; Calcium; Corpus Striatum; Dizocilpine Maleate; Glutamic Acid; Magnesium; Male; Microdialysis; Prosencephalon; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate

1. Ischaemia was induced by 5 min of deprivation of glucose and an additional 5 min of deprivation of glucose and oxygen from Mg(2+)-free artificial cerebrospinal fluid in vitro. 2. During the ischaemic period, 11 +/- 1.5% of the total [3H]-dopamine ([3H]-DA) was released into the incubation medium. 3. Ischaemia-evoked release of [3H]-DA from striatal slices was attenuated by tetrodotoxin (TTX), MgSO4, dizocilpine, ketamine, 6,7-dinitroquinoxaline-2,3-dione (DNQX) or carbetapentane. 4. Release of [3H]-DA was attenuated by verapamil, omega-conotoxin GVIA and dantrolene. 5. Nomifensin inhibited the ischaemia-induced release of [3H]-DA. 6. Omission of Ca(2+) from incubation media potentiated ischaemia-evoked [3H]-DA release. The inhibitory effect of nomifensin was potentiated in Ca(2+)-free incubation media. 7. These results suggest that ischaemia induces release of [3H]-DA by dual mechanisms; one is Ca(2+)-dependent exocytosis and the other is reversal of transporter.

Topics: Animals; Brain Ischemia; Calcium; Calcium Channel Blockers; Corpus Striatum; Culture Techniques; Cyclopentanes; Dantrolene; Dizocilpine Maleate; Dopamine; Ketamine; Magnesium Sulfate; Nomifensine; omega-Conotoxin GVIA; Peptides; Quinoxalines; Rats; Tetrodotoxin; Verapamil

The CA1 region of hippocampus is selectively vulnerable to a variety of insults, including hypoxia-ischemia and Alzheimer's disease, but the basis of this regional susceptibility is poorly understood. We examined the regional hippocampal sensitivity to mitochondrial metabolic disruption induced by malonate, an inhibitor of succinate dehydrogenase. The CA1 region was exquisitely sensitive to malonate and the dentate gyrus was extremely resistant; the CA3 region had intermediate sensitivity. This pattern of vulnerability is reminiscent of hypoxic-ischemic damage. Malonate damage was blocked by the N-methyl-D-aspartic acid (NMDA) antagonist, MK-801, but regional susceptibility to malonate did not correlate with the density of NMDA receptors. Instead, malonate toxicity was inversely correlated with activity of succinate dehydrogenase. Our results suggest that regional metabolic capacity may help to determine sensitivity to metabolic/excitotoxic insults such as hypoxia-ischemia.

Topics: Animals; Autoradiography; Brain Ischemia; Dizocilpine Maleate; Hippocampus; Hypoxia; Hypoxia, Brain; Male; Malonates; N-Methylaspartate; Rats; Rats, Sprague-Dawley

The present study was inspired by two previous findings from the laboratory. The first was that dizocilpine maleate (MK-801) fails to reduce infarct size when the middle cerebral artery (MCA) is permanently occluded by an intraluminal filament technique in rats. In seeking the reasons for this we measured temperature and found that the body temperature of occluded animals increases to 39.0-39.5 degrees C during the first 2-3 h. In order to explore whether the rise in temperature was responsible for the lack of effect of MK-801, two groups of animals were studied, both containing animals which were subjected to 2 h of transient MCA occlusion and given MK-801 15 min before, as well as 6 and 24 h after ischemia. In one group, temperature was allowed to rise spontaneously during ischemia (39.0-39.5 degrees C). In the other, body temperature was maintained close to normal during ischemia, and for the first 6 h postischemically, by cooling of the ambient air. Infarct volume was assessed by triphenyltetrazolium chloride staining after 48 h of recovery. The results showed that MK-801 failed to reduce infarct size in animals whose body temperature rose during ischemia. In contrast, the drug markedly reduced infarct volume in temperature-controlled animals; in fact, 5/8 animals had no infarcts but selective neuronal damage only. The results suggest that amelioration of focal ischemic damage cannot be expected if body and brain temperature is allowed to rise above normal.

Topics: Animals; Body Temperature; Brain Ischemia; Cerebral Infarction; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Fever; Male; Rats; Rats, Wistar

Cerebral ischemia in the gerbil results in early hippocampal changes, which include transient activation and/or translocation of protein kinase C (PKC), increased enzymatic activity of ornithine decarboxylase (ODC), and elevated DNA binding ability of activator protein-1 (AP1). The time-course of all three of these postischemic responses was found to be almost parallel, peaking at 3 hr after the ischemic insult. The effectiveness of known modulators of postischemic morphological outcome (MK-801, L-NAME, and gingkolides BN 52020 and BN 52021) in counteracting the induction of PKC, ODC, and AP1 formation was tested. These drugs were administrated as followed: MK-801 (a noncompetitive inhibitor of NMDA channel), 0.8 mg/kg i.p., 30 min before ischemia, and 5 min after the insult; L-NAME (competitive inhibitor of NO synthase), 10 mg/kg i.p., 30 min before ischemia, and 5 mg/kg, 5 min after ischemia; BN52020 and BN52021 (inhibitors of platelet-activating factor: PAF receptors) were administered as a suspension in 5% ethanol in water by oral route, 10 mg/kg for 3 days before ischemia. Three of these drugs, MK-801, L-NAME, and BN52021, significantly reduced ischemia-elevated activity of PKC and ODC, whereas AP1 formation was only partially attenuated. Our observations implicate the existence of different mechanism(s) for postischemic PKC and ODC activation, which in turn is engaged in AP1 induction.

Topics: Animals; Blood Platelets; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Gerbillinae; Hippocampus; N-Methylaspartate; Nitric Oxide; Protein Kinase C; Transcription Factor AP-1

A large body of evidence exists to demonstrate that excitatory amino acids (EAA) and their receptors are involved in the pathophysiological mechanisms linking several acute brain insults, such as cerebral ischemia, to neuronal degeneration and death. Accordingly, the use of EAA receptor antagonists can be beneficial in attenuating or preventing the neuronal irreversible damage subsequent to various neuropathological syndromes. We have investigated the effect of 15 min of simulated ischemic conditions, i.e., oxygen/glucose deprivation, on hippocampal slices preparation measuring, as neurotoxicity indexes, both the amino acids efflux in the incubation medium, detected by HPLC, and the inhibition of protein synthesis, evaluated as 3H-Leucine incorporation into proteins. Accumulation of neurotransmitter amino acids was measured in the medium during the "ischemic" period. Glutamate increased 30-fold over the basal level while aspartate was sevenfold and GABA 12-fold higher than in normal conditions. After a reoxygenation period of 30 min, the rate of protein synthesis of hippocampal slices subjected to "ischemia" was reduced to 35-50% of controls. The non-competitive NMDA antagonist MK-801 (100 microM) and the competitive NMDA antagonist CGP 39551 (100-250 microM) as well as the non-NMDA receptor antagonists NBQX (100 microM) and AP3 (300 microM) were unable to counteract the metabolic impairment when they were present alone in the incubation fluid during simulated "ischemia." An incomplete, but highly significant (p < 0.001), protection from protein synthesis impairment was achieved in the presence of an equimolar concentration (100 microM) of MK-801 and NBQX.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 2-Amino-5-phosphonovalerate; Amino Acids; Animals; Brain Ischemia; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glucose; Hippocampus; In Vitro Techniques; Ionophores; Leucine; Male; Nerve Tissue Proteins; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate

Substantiating evidence has raised the possibility that sigma ligands may have therapeutic potential as neuroprotective agents in brain ischemia. It has been suggested that the neuroprotective capacity of sigma ligands is related primarily to their affinity for the NMDA receptor complex and not to any selective action at the sigma binding site. However, sigma specific ligands, devoid of significant affinity for the NMDA receptor, are also neuroprotective via an inhibition of the ischemic-induced presynaptic release of excitotoxic amino acids. In the present study, we have investigated the potential neuroprotective effect of a comprehensive series of sigma ligands, with either significant (sigma/PCP) or negligible (sigma) affinity for the PCP site of the NMDA receptor, in order to delineate a selective sigma site-dependent neuroprotective effect. For this aim, we have employed two different neuronal culture toxicity paradigms implicating either postsynaptic-mediated neurotoxicity, (brief exposure of cultures to a low concentration of NMDA or Kainate) or pre- and postsynaptic mechanisms (exposure to hypoxic/hypoglycemic conditions). Only sigma ligands with affinity for the NMDA receptor [(+) and (-) cyclazocine, (+) pentazocine, (+) SKF-10047, ifenprodil and haloperidol] were capable of attenuating NMDA-induced toxicity whereas the sigma [(+)BMY-14802, DTG, JO1784, JO1783, and (+)3-PPP] and kappa-opioid [CI-977, U-50488H] ligands, with very low affinity for the NMDA receptor, were inactive. The rank order of potency, based on the 50% protective concentration (PC50) value, of sigma/PCP ligands against NMDA-mediated neurotoxicity correlates with their affinity for the PCP site of the NMDA receptor, and not with their affinity for the sigma site. In addition sigma/PCP, sigma or kappa-opioid ligands failed to attenuate kainate-mediated neurotoxicity. On the other hand, sigma/PCP, sigma and kappa-opioid ligands were potent inhibitors of hypoxia/hypoglycemia-induced neurotoxicity, although their neuroprotective potency did not correlate with their affinity for either the sigma or PCP binding sites. In conclusion, the ability of sigma and kappa-opioid ligands to attenuate hypoxia/hypoglycemia, but not NMDA or kainate-induced toxicity, suggests that these drugs exert their neuroprotective role by a predominantly presynaptic mechanism possibly by inhibiting ischemic-mediated glutamate release.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Anti-Arrhythmia Agents; Antihypertensive Agents; Benzofurans; Brain Ischemia; Cell Death; Cells, Cultured; Dizocilpine Maleate; Hypoxia; Kainic Acid; Ligands; N-Methylaspartate; Neurons; Pyrrolidines; Rats; Rats, Sprague-Dawley; Receptors, Opioid, kappa; Receptors, Phencyclidine; Receptors, sigma

Ischemia-induced neuronal injury can be reduced by glutamate antagonists acting at the N-methyl-D-aspartate (NMDA) receptor. 7-Chlorokynurenic acid and the recently synthesized compound Acea 1021 block NMDA receptors by acting at the strychnine-insensitive glycine site. The anti-ischemic properties of these compounds were tested by evaluating their ability to reduce CA1 neuronal damage in hippocampal slice cultures deprived of oxygen and glucose. Acea 1021 and 7-chlorokynurenic acid significantly reduced CA1 injury produced by oxygen and glucose deprivation in a dose-dependent manner. The neuroprotective effect of these compounds was reversed by the addition of glycine. The phencyclidine site NMDA antagonist MK-801 also provided significant protection to CA1 neurons against the same insult, and this protection was not affected by the addition of glycine. These results indicate that Acea 1021 and 7-chlorokynurenic acid can provide protection to CA1 neurons against ischemia-induced injury by a glycine-sensitive mechanism.

Topics: Animals; Brain Ischemia; Cell Death; Dizocilpine Maleate; Glycine; Glycine Agents; Hippocampus; Kynurenic Acid; Neurons; Organ Culture Techniques; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Glycine; Receptors, N-Methyl-D-Aspartate

The present study was carried out to compare the neuroprotective effect of the novel noncompetitive NMDA antagonist, FR115427, with that of(+)MK-801 in rat focal cerebral ischemia. Focal cerebral ischemia was produced by permanent occlusion of the left middle cerebral artery (MCA). Drugs were administered intraperitoneally immediately after ischemia and once a day for 6 successive days. FR115427 (10 mg/kg, i.p.) significantly improved neurologic deficit at 1 day after ischemia and reduced total infarct volume (54%) at 7 days after ischemia. Although FR115427 (10 mg/kg, s.c.) produced neuronal vacuolization similar to (+)MK-801, FR115427 did not produce adverse effects such as a loss of body weight, mortality, and hypothermia, in contrast to (+)MK-801. These results suggest that FR115427 may be useful in the treatment of stroke.

Topics: Animals; Brain Diseases; Brain Ischemia; Dizocilpine Maleate; Isoquinolines; Male; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Tetrahydroisoquinolines

We determined the ability of a new benzomorphan derivative [2R-[2 alpha, 3(R*),6 alpha]]-1,2,3,4,5,6-hexahydro-3-(2-methoxypropyl)- 6,11,11-trimethyl-2,6-methano-3-benzazocin-9-ol hydrochloride (BIII 277 CL) to inhibit the N-methyl-D-aspartic acid (NMDA) receptor-channel complex in vitro and in vivo. BIII 277 CL potently displaced [3H]MK-801 binding from the NMDA receptor-channel complex in synaptosomal membrane preparations from rat brain cortex (Ki = 4.49 nmol/l). It was much less effective at displacing [3H]dihydromorphine, [3H]naloxone and [3H]ditolyguanidine binding in similar membrane preparations: the Ki values were 3323, 8031 and 1017 nmol/l, respectively. BIII 277 CL did not exhibit any marked affinities for a variety of other central neurotransmitter receptors. BIII 277 CL antagonized NMDA-induced [3H]noradrenaline release (EC50 = 1.7 mumol/l) and NMDA-induced inhibition of protein synthesis in rat hippocampal slices (EC50 = 3.0 mumol/l). In mice, BIII 277 CL prevented NMDA-induced lethality (ID50 = 0.54 mg/kg s.c.) and, as expected, also caused disturbances in motor coordination in the same dose range (ED50 = 0.47 mg/kg s.c.). The duration of BIII 277 CL was much shorter than than of (+)MK-801 in both tests. Finally, BIII 277 CL (0.3 mg/kg s.c. 5 times over 24 h) reduced the cortical infarct area in mice that had been subjected previously to focal cerebral ischemia by unilateral occlusion of the middle cerebral artery. In summary, these results indicate that BIII 277 CL is a potent and specific ion-channel blocker of the NMDA receptor-channel complex which could be used for the treatment of acute thromboembolic stroke in humans.

Topics: Animals; Benzomorphans; Brain Ischemia; Dizocilpine Maleate; Guinea Pigs; In Vitro Techniques; Ion Channels; Male; Mice; Motor Activity; Protein Binding; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Synaptic Membranes

Hypothermia reduces ischemic brain damage, confounding interpretation of the neuroprotective effects of drugs. Specifically, the neuroprotectant MK-801 has been shown to cause hypothermia. Some have claimed that when body temperature is maintained, MK-801 is not a neuroprotectant, whereas others claim it retains its neuroprotective activity. MK-801 was evaluated for neuroprotective properties in free-regulating as well as temperature-maintained gerbils receiving 5 or 10 min of bilateral carotid occlusion. After 10 min of ischemia, free-regulating animals exhibited significant hypothermia (as low as 32 degrees C) and showed significant neuroprotection after 3 mg/kg IP MK-801. When a hyperthermic body temperature (38.5 degrees C) was maintained, no reduction in brain damage was evident after up to 10 mg/kg IP MK-801, even when occlusion time was reduced to 5 min. However, when a normothermic body temperature (36.5 degrees C) was maintained, 10 mg/kg IP MK-801 significantly reduced brain damage after 5 min of ischemia. Thus, although a higher dose of the drug is required, MK-801 can reduce ischemic brain damage in the absence of hypothermia. The need for this high dose suggests that mechanisms other than NMDA receptor complex antagonism may be involved in the neuroprotective actions of MK-801.

Topics: Animals; Body Temperature; Brain; Brain Ischemia; Dizocilpine Maleate; Dose-Response Relationship, Drug; Female; Gerbillinae; Hypothermia, Induced; Neuroprotective Agents; Temperature

A novel experimental model of stroke has been developed using the powerful vasoconstrictor peptide, endothelin-1, to occlude the middle cerebral artery (MCA) of anaesthetised rats. Intracerebral microinjections of endothelin-1 were administered under stereotaxic guidance adjacent to the MCA, and after 3 days rats were perfusion fixed for histopathological determination of ischaemic brain damage. The pattern of brain damage noted using this model was similar to that reported following permanent surgical occlusion of the MCA. Brain damage was apparent in the dorsal and lateral neocortex (98 +/- 12 mm3) and striatum (32 +/- 3 mm3) ipsilateral to the insult. Rats anaesthetised with halothane and barbiturate exhibited a similar volume of brain damage. However, infarct volume increased when the duration of halothane anaesthesia was extended from 5 to 180 min post-occlusion. Neuroprotection studies demonstrated that dizocilpine (5 mg/kg, i.p.), administered 30 min prior to MCA occlusion, reduced the volume of cortical brain damage by 51% (P < 0.05) but did not alter the volume of striatal brain damage. The present results demonstrate that microinjections of endothelin-1 adjacent to the rat MCA result in a reproducible pattern of focal cerebral infarction which is sensitive to the duration of anaesthesia and can be reduced by dizocilpine.

Topics: Anesthesia; Animals; Brain Ischemia; Cerebral Arteries; Cerebral Cortex; Cerebral Infarction; Cerebrovascular Disorders; Disease Models, Animal; Dizocilpine Maleate; Endothelins; Excitatory Amino Acid Antagonists; Injections, Intra-Arterial; Male; Microinjections; Rats; Rats, Sprague-Dawley; Stereotaxic Techniques

The effects of acute hyperglycaemia and streptozotocin-induced diabetes on infarct size were measured 48 h after middle cerebral artery occlusion (MCAO) in Fischer 344 rats. Both hyperglycaemia (+46%) and diabetes (+68%) increased infarct volume when compared to normoglycaemic rats. Insulin-treated diabetic rats exhibited an infarct size similar to that observed in normoglycaemic rats. Neuroprotection has been difficult to demonstrate in pathological conditions that increase infarct volume such as chronic arterial hypertension. However, administration of the non-competitive NMDA antagonist, dizocilpine (MK-801), after MCAO, reduced the volume of ischaemic damage (by 33-48%) in all groups. The present findings indicate (a) that the detrimental effects of experimental diabetes on infarct volume are largely attributed to hyperglycaemia; and (b) dizocilpine was as neuroprotective in hyperglycaemia and diabetic conditions as in normoglycaemic rats.

Topics: Animals; Brain Ischemia; Cerebral Arteries; Cerebral Infarction; Diabetes Mellitus, Experimental; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hyperglycemia; Hypoglycemic Agents; Insulin; Male; Rats; Rats, Inbred F344; Stereotaxic Techniques

The present study was designed to determine whether postischemic hypothermia, delayed MK-801 (dizocilpine) administration, or a combination of these treatments can provide lasting neurobehavioral protection following transient global ischemia in rats. Rats were subjected to 10 min of normothermic (37 degrees C) ischemia induced by 2-vessel occlusion and hypotension (50 mmHg) or sham procedures. Ischemia was followed by either: (a) 3 h at normothermic brain temperatures, (b) 3 h of postischemic brain hypothermia at 30 degrees C, (c) hypothermia coupled with MK-801 (4 mg/kg, i.p.) on postischemic days 3, 5 and 7, or (d) postischemic MK-801 treatment alone. Neurobehavioral evaluation 6-8 weeks following surgery showed that normothermic ischemia (NI) was associated with water maze navigational deficits, including performance on a simple place task involving finding a hidden platform maintained in one position for 6 days, and a learning set task in which the platform was moved to a different location each day (both P's < 0.02 vs. sham). NI was also associated with increased locomotion in an open field (P < 0.01 vs. sham). A combination of postischemic hypothermia and delayed MK-801 injections provided partial protection from ischemic-associated hyperactivity in the open field (P < 0.02 vs. NI), and robust protection from simple place task deficits (P < 0.02 vs. NI). Evidence for significant protective effects of MK-801 or hypothermia alone was observed in the learning set, during the final trial blocks each day. These results provide further evidence for neuroprotective effects of these treatments at chronic survival intervals, and indicate that the therapeutic window for attenuating ischemic damage is considerably longer than has heretofore been appreciated.

Topics: Animals; Behavior, Animal; Brain Ischemia; Cell Count; Cerebral Cortex; Disease Models, Animal; Dizocilpine Maleate; Hippocampus; Hypothermia; Locomotion; Male; Rats; Rats, Wistar

1. We have recently developed a new model of transient focal ischaemia in the rat utilising topical application of endothelin-1 to the left middle cerebral artery (MCA). In order to validate this approach the present study assessed the neuroprotective efficacy of the NMDA receptor antagonist dizocilpine (MK-801) in the endothelin-1 model. The anti-ischaemic efficacy of the nitric oxide (NO) synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME) was subsequently evaluated, and contrasted with its efficacy against permanent focal ischaemia, to determine the utility of the endothelin-1 model for identification of novel pharmacoprotective agents. 2. MK-801 (0.12 mg kg-1 bolus, 108 micrograms kg-1 h-1 infusion i.v., either 1 or 2.5 h pre-transient MCA occlusion (MCAO)) induced hypotension that persisted for approximately 1.5 h so that mean arterial blood pressure (MABP) at the time of MCAO was significantly lower in the 1 h group compared with control (MABP: 86 +/- 11, 68 +/- 6 and 84 +/- 4 mmHg (mean +/- s.d.) for saline, 1 h MK-801 and 2.5 h MK-801 groups respectively). The 2.5 h pretreatment schedule resulted in significant reduction (71%) in the volume of hemispheric damage (assessed 4 h post onset of ischaemia) while the 1 h pretreatment schedule did not (volumes of hemispheric damage: 59 +/- 38, 51 +/- 51 and 17 +/- 28 mm3 for saline, 1 h and 2.5 h MK-801 groups). 3. Thus the considerable neuroprotective effect of MK-801 in the endothelin-1 model of transient focal cerebral ischaemia was highly sensitive to drug-induced hypotension. This result is in contrast to previous studies of permanent MCAO where MK-801-induced hypotension did not compromise its neuroprotective action.4. L-NAME (3 mg kg-1, i.v. 30 min pre-MCAO) moderately, but significantly, reduced (16%) the volume of ischaemic damage 4 h post-permanent MCA occlusion, whereas the 29% reduction in volume of damage achieved in the model of transient focal ischaemia did not attain significance due to the greater variability associated with this model. L-NAME did not significantly alter MABP in either model.5. The modest neuroprotection achieved with NO synthase inhibition suggests NO is of relatively minor importance as a mediator of neurotoxicity following permanent focal cerebral ischaemia. In addition the comparable efficacy of L-NAME against transient focal ischaemia suggests the presence of reperfusion does not enhance the contribution of NO to neuronal injury in the acute (4 h) phase foll

Topics: Amino Acid Oxidoreductases; Animals; Arginine; Blood Pressure; Brain; Brain Ischemia; Cerebral Arteries; Dizocilpine Maleate; Endothelins; Ischemic Attack, Transient; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

In the present study we utilise the potent vasoconstrictor properties of endothelin-1 (Et-1) in a new model of middle cerebral artery occlusion in the anaesthetized rat. We evaluate the reproducibility of the model and examine the neuroprotective efficacy of the potent anti-ischaemic agent, MK801. Adult male SD rats received MK801 (5 mg/kg, n = 7) or saline vehicle (n = 7) 30 mins prior to the microinjection of Et-1 (60 pmol in 3 microliters) via a 31-g cannula stereotaxically positioned 0.5 mm above the middle cerebral artery. Three days after the injection of Et-1, rats were perfusion fixed, the brain removed, cryostat sectioned and processed for histological staining. Sections at eight predetermined levels were examined by light microscopy and the volume of infarction calculated. Following administration of Et-1, saline-pretreated rats exhibited a pattern of ischaemic damage similar to that previously reported following permanent occlusion of the rat middle cerebral artery. This pattern was characterised by a large volume of infarction covering the dorsal and lateral neocortex (98 +/- 12 mm3) and striatum (32 +/- 3 mm3) ipsilateral to the insult. Power analysis predicted a group size of 7 would be required for a 50% reduction in ischaemic damage to be recorded as statistically significant at the 5% level. Pretreatment with MK801 reduced cortical tissue damage by 51% (P = 0.026) but did not significantly alter either the pattern or volume of infarction (33 +/- 4 mm3; P = 0.95) in the striatum.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Anesthetics; Animals; Brain; Brain Ischemia; Cerebral Arteries; Dizocilpine Maleate; Endothelins; Injections; Male; Rats; Rats, Sprague-Dawley; Receptors, Endothelin; Stereotaxic Techniques; Vasoconstriction

Middle cerebral artery (MCA) occlusion in rats induced c-fos and junB mRNA 4h later in all ipsilateral cortex outside the MCA distribution and in many subcortical structures: medial striatum; most of thalamus including medial and lateral geniculate nuclei: substantia nigra; and hippocampus. The N-methyl-D-aspartate (NMDA) antagonist, MK-801 (4 mg/kg, i.p.) inhibited c-fos and junB mRNA induction in the cortex, striatum, thalamus, and hippocampus but not in the substantia nigra. These data show that c-fos and junB mRNA induction in cortex, striatum, thalamus, hippocampus involves the activation of NMDA receptors whereas different receptors must be implicated in the induction in substantia nigra.

Topics: Animals; Brain Chemistry; Brain Ischemia; Cerebral Arteries; Cerebral Cortex; Dizocilpine Maleate; Gene Expression; Genes, fos; Genes, Immediate-Early; Genes, jun; Hippocampus; Male; Rats; Rats, Sprague-Dawley; RNA, Messenger; Substantia Nigra; Thalamus

In a model of perinatal hypoxic-ischemic brain damage, we examined the neuroprotective efficacy of posttreatment with the NMDA receptor antagonist MK-801 and the AMPA receptor antagonist NBQX. Unilateral brain damage developed in 95% of rat pups subjected to hypoxia-ischemia with a 27.8 +/- 1.2% weight deficit of the damaged hemisphere. MK-801 in doses of 0.3 and 0.5 mg/kg i.p. reduced the brain damage by 61% (p < 0.001) and 43% (p < 0.001), respectively. A higher dose of MK-801 (0.75 mg/kg) did not offer neuroprotection. Treatment with NBQX (40 mg/kg) reduced the hemispheric lesion by 28% (p < 0.05). In conclusion, posttreatment with both NBQX and low doses of MK-801 reduced perinatal brain damage. The NMDA receptor antagonist offered stronger neuroprotection which is in agreement with a proposed NMDA receptor hyperactivity around postnatal day 7 in rats.

Topics: Animals; Animals, Newborn; Brain Damage, Chronic; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Female; Hypoxia, Brain; Injections, Intraperitoneal; Male; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Severity of Illness Index; Treatment Outcome

Focal cerebral ischemia was induced in rats by permanent occlusion of the left middle cerebral artery (MCA). The cerebroprotective properties of the competitive NMDA antagonist CGP 40116 were evaluated in that model and compared to the neuroprotective effects of MK 801, D-CPPene and CGS 19755 under the same experimental conditions. Infarct volume was assessed using in vivo magnetic resonance imaging. The rank order of potency for the NMDA antagonists tested was MK 801 > CGP 40116 approximately D-CPPene > CGS 19755. CGP 40116 dose-dependently reduced the volume of cortical infarction, with an ED50 of 11 mg/kg i.v., and its cerebroprotective efficacy was comparable to that of MK 801. Neuroprotection by CGP 40116 was still apparent when treatment was started 30 minutes after MCA occlusion. It is concluded that CGP 40116 is an effective cerebroprotectant with potential clinical utility for amelioration of focal cerebral ischemic damage.

Topics: 2-Amino-5-phosphonovalerate; Animals; Blood Pressure; Brain Ischemia; Dizocilpine Maleate; Dose-Response Relationship, Drug; Magnetic Resonance Imaging; Male; Neuroprotective Agents; Pipecolic Acids; Piperazines; Rats; Rats, Inbred F344; Receptors, N-Methyl-D-Aspartate

In this study Mongolian gerbils were submitted to a normothermic bilateral carotid ligation lasting 5 min. A noncompetitive antagonist of NMDA receptors, MK-801, 0.8 mg/kg, was injected i.p. 30 min before ischemia, or the ganglioside GM1, 30 mg/kg, was given i.p. for 3 days, twice a day. The morphology of the hippocampal CA1 neurones and the brain content of cyclooxygenase metabolites of arachidonic acid: prostaglandin 6-keto PGF1 alpha and thromboxane Tx B2 were studied. Untreated ischemia induced the accumulation in brain of the 6-keto PGF1 alpha and Tx B2 immunoreactive materials, and resulted in a lesion of 70% of CA1 neurones. In the MK-801- and GM1-pretreated groups the postischemic levels of Tx B2 were significantly decreased. However MK-801 and GM1 did not prevent damage to the CA1 neurones in gerbils normothermic after ischemia, whereas a partial neuroprotection was observed in hypothermic, MK-801 treated gerbils. The results of this study indicate that NMDA receptors may participate in the mechanism of postischemic release of eicosanoids in brain. They also confirm a potential modulatory role of gangliosides. These results are discussed in terms of the involvement of cyclooxygenase metabolites of arachidonic acid in the mechanism of a selective delayed neuronal damage to the hippocampus CA1 after ischemia.

Topics: 6-Ketoprostaglandin F1 alpha; Animals; Brain; Brain Ischemia; Dizocilpine Maleate; Female; G(M1) Ganglioside; Gerbillinae; Hippocampus; Male; Prostaglandins; Thromboxane B2

Neuronal damage induced by ischemia involves various changes in neurotransmission. Nitric oxide (NO), a putative neurotransmitter and/or neuromodulator has some role in this neuronal damage. In the present study, the effect of NO on the terminal site of dopamine (DA) neurons in the rat striatum was examined using the microdialysis technique. First perfusion with sodium nitroprusside (SNP) as an NO donor increased extracellular DA (10 mM, 460%; 1 mM, 140%) in the striatum and decreased its metabolites. Pretreatment with tetrodotoxin (TTX, 5 microM), (+)-5-methyl-10,11-dihydro-5H-dibenzo [a,d] cyclohepten-5,10-imine hydrogen maleato (MK801, 1 microM) or muscimol (1 microM) inhibited SNP-induced increases in extracellular DA and decreases in DOPAC (TTX, complete block; MK801, 75% inhibition; muscimol, 80% inhibition). Second, extracellular NO, DA and DOPAC were measured in the gerbil striatum following 10 minutes of forebrain ischemia produced by occluding both carotid arteries. Occlusion of the carotid arteries also caused increases in extracellular NO and DA in the gerbil striatum (NO, 3000%; DA, 2800%). These findings suggest that NO-facilitated DA release occurs via interaction between glutamatergic and dopaminergic neurons. These changes are probably partially involved in the neurodegenerative phenomena following ischemia. It is also shown that simultaneous measurements of NO and DA using this technique may be useful in assessing ischemic changes in vivo.

Topics: 3,4-Dihydroxyphenylacetic Acid; Animals; Brain Ischemia; Chromatography, High Pressure Liquid; Corpus Striatum; Dizocilpine Maleate; Dopamine; Gerbillinae; Male; Microdialysis; Nitric Oxide; Nitroprusside; Rats; Rats, Wistar; Tetrodotoxin

The excitotoxic effects of glutamate can be blocked almost completely with gamma-aminobutyric acid (GABA), an inhibitory neurotransmitter, in cell culture, tissue slices, and in some animal models. After stroke in rats, we showed previously that an agonist of GABA, muscimol, was as neuroprotective as MK-801, an antagonist of glutamate. To obtain further neuroprotection and to avoid the side effects associated with high doses of MK-801, we wanted to assess the efficacy of the two agents in combination.. Treatment was administered 5 minutes after embolic cerebral ischemia in Sprague-Dawley rats. The subjects were rated using a neurological evaluation 48 hours later. Visual-spatial learning was measured 8 to 10 weeks after stroke, after which we measured the volume of each cerebral hemisphere and several large cerebral compartments. Treatment groups included saline (n = 27), MK-801 1.0 mg/kg (n = 23), muscimol 1.0 mg/kg (n = 17), and both agents together using a dose of 0.5 mg/kg each (n = 25).. A probit analysis of the neurological ratings revealed a protective effect of muscimol used alone (MK-801 potency ratio, 2.0; P = NS; muscimol potency ratio, 4.0; P < .05) and a protective effect of the combination (potency ratio, 5.0; P < .05). Focal ischemia caused a moderate to severe delay in the acquisition of visual-spatial information, which was completely eliminated by the combination treatment but only partially ameliorated with MK-801 or muscimol alone. Ischemia reduced the cerebral hemisphere volume from 0.42 mm3 to 0.34 mm3 (P < .0001), the volume density of cortex from 22% to 17% of total cerebral volume (P < .01), and that of hippocampus from 4.3% to 3.0% (P < .05). Only the combination was neuroprotective, as measured by the ratio of the lesioned to the contralateral hemisphere volume (P = .013). The combination treatment and MK-801 protected the hemisphere volume, the cortex, and the hippocampus and reduced the size of visible infarction.. Combination therapy, using a glutamate antagonist and a GABA-A agonist, appeared to protect the brain and ameliorate a defect in learning behavior after stroke. The combination may have been more effective than either agent used alone, although further study of higher doses is needed.

Topics: Animals; Brain; Brain Ischemia; Cerebral Infarction; Dizocilpine Maleate; Drug Combinations; Excitatory Amino Acid Antagonists; gamma-Aminobutyric Acid; Glutamic Acid; Learning; Male; Microspheres; Motor Activity; Muscimol; Organ Size; Rats; Rats, Sprague-Dawley; Reaction Time

In the present study the in vitro and ex vivo distributions of [3H]dizocilpine binding sites in mouse brain after middle cerebral artery occlusion (MCA-O) were compared using receptor autoradiography. The distribution patterns of [3H]dizocilpine binding sites obtained in vitro and ex vivo in normal mouse brain were the same with the highest densities occurring in the hippocampus and cerebral cortex. MCA-O had little or no effect on the in vitro binding density for at least 24 hr post-ischaemia. However after 2-3 days binding densities in the region of infarct were significantly reduced compared to the contralateral cerebral cortex. Further reductions occurred after 5-7 days. By contrast ex vivo [3H]dizocilpine binding was reduced in the infarcted area by 78.7 +/- 4% within 2 hr of the ischaemic insult and at all subsequent times binding was reduced by more than 75%. Ex vivo binding after ischaemia was always less than 30% of in vitro binding and this decrease was apparent within 2 hr of the ischaemic insult whereas in vitro binding was maintained at control levels for at least 24 hr. The neuroprotective activity of the NMDA antagonists dizocilpine and CGP 37849 in this model at different times after MCA-O was assessed. The time scale for receptor access following MCA-O is discussed and it is suggested that although the population of NMDA receptors is maintained in the infarct region for some days access to them in vivo may be sufficiently impaired within 2 or 4 hr of ischaemic insult to reduce the neuroprotective activity of NMDA antagonists after this time.

Topics: 2-Amino-5-phosphonovalerate; Animals; Autoradiography; Binding Sites; Brain; Brain Ischemia; Cerebral Arteries; Cerebral Infarction; Dizocilpine Maleate; In Vitro Techniques; Male; Mice; Receptors, N-Methyl-D-Aspartate

Several reports have indicated that the two glutamate receptor antagonists, dizocilpine (that binds to the phencyclidine recognition site of the NMDA (N-methyl-D-aspartate) receptor) and NBQX (2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline, that binds to the AMPA (alpha-amino-3-hydroxy-5-methyl-isoxazole) receptor), protect neurons against damage caused by hypoxia, ischemia or excitotoxicity. We, therefore, used a combination of these drugs to achieve enhanced neuroprotection. Primary cultures of rat hippocampal neurons were challenged by glutamate intoxication. Both dizocilpine and NBQX produced dose-dependent increases in the percentage of viable neurons. Combined treatment with both glutamate receptor antagonists had an over-additive neuroprotective effect. Simultaneous administration of dizocilpine and NBQX also had a pronounced neuroprotective effect in vivo in mice subjected to focal cerebral ischemia and rats with global forebrain ischemia. This suggest that such a combination may have therapeutic relevance.

Topics: Animals; Brain Ischemia; Cells, Cultured; Dizocilpine Maleate; Drug Synergism; Glutamates; Glutamic Acid; Hippocampus; Ischemic Attack, Transient; Male; Mice; Neurons; Quinoxalines; Rats; Rats, Inbred F344; Rats, Wistar; Receptors, AMPA

Hippocampal slices of rats at postnatal day 7 were submitted to superfusion with Ca(2+)- and Mg(2+)-free, bicarbonate buffered ion balanced medium, and perfusate concentrations of eicosanoids: thromboxane B2 and 6-keto prostaglandin F1 alpha were determined by the radioimmunoassay. It was noted that the permanent presence of Ca2+ increased the basal eicosanoid level, and in these conditions modulation of eicosanoid production was lost, whereas temporary, a 20 min application of 1.3 mM Ca2+ did not influence significantly eicosanoid release. A 20 min application of the anoxic/aglycemic medium containing calcium did not change the content of eicosanoids in superfusates. A significant stimulation of the thromboxane B2 and 6-keto prostaglandin F1 alpha release was noted provided the application of the experimental medium was accompanied by a 10 min arrest of superfusion. This effect was inhibited by MK-801 and quinacrine, suggesting an involvement of NMDA receptors and phospholipase A2. We propose that a model of anoxic/aglycemic superfusion with a stop flow period allows retention of endogenous glutamate in the extracellular fluid, resembling a similar effect during in vivo ischemia, whereas during a continuous superfusion glutamate is immediately washed out. Consequently, an application of the anoxic/aglycemic medium accompanied by a temporary arrest of superfusion represents more adequate in vitro model of ischemia than a constant superfusion with this medium. In these conditions NMDA receptors mediate eicosanoid release.

Topics: 6-Ketoprostaglandin F1 alpha; Animals; Arachidonic Acid; Brain Ischemia; Calcium; Culture Techniques; Dizocilpine Maleate; Eicosanoids; Hippocampus; Hypoxia; N-Methylaspartate; Quinacrine; Radioimmunoassay; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Thromboxane B2

This study addresses the issue of endpoint selection in the evaluation of neuroprotective drugs in experimental focal ischaemia. Previous work with the permanent middle cerebral artery (MCA) occlusion model in the rat has demonstrated that the ischaemic lesion does not acquire its final appearance until at least 28 days after the ictus. Therefore, the effect of the NMDA receptor blocker MK-801 (dizocilpine maleate) was evaluated both early (3 days) and late (28 days) after MCA occlusion to determine if the previously reported protective effect of a single post-ischaemic dose of MK-801 found in acute experiments remained after 28 days. Mk-801 (0.5 mg/kg, i.v.) or isotonic saline was randomly given to rats 30 min after MCA occlusion. Infarct volume and volume of ipsilateral and contralateral hemispheres were estimated from camera lucida drawings of 8 defined coronal histological sections of the brain. As expected, a 40% (p < 0.05) reduction of infarct size was found in MK-801 treated rats after 3 days. In animals evaluated 28 days after MCA occlusion, no significant difference in infarct size, total tissue loss (infarct volume+ipsilateral hemisphere atrophy) or remaining non-infarcted tissue (contralateral hemisphere--total tissue loss) was seen between the MK-801 and placebo treated rats. The results suggest that the single dose treatment with MK-801 postponed the evolution of the infarct, which at 3 days after MCA occlusion is still in progress, possibly by ameliorating oedema formation. It remains to be shown if a multiple dose treatment with NMDA receptor antagonists improves the final neuropathological outcome after experimental stroke.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Atrophy; Brain Edema; Brain Ischemia; Cerebral Cortex; Cerebral Infarction; Dizocilpine Maleate; Dominance, Cerebral; Image Processing, Computer-Assisted; Infusions, Intravenous; Male; Rats; Rats, Sprague-Dawley

The objective of this study was to explore how alterations in tissue pH during ischemia influence cell calcium uptake, as this is reflected in the extracellular calcium concentration (Ca2+e). Variations in pH were achieved by making animals hypo-, normo- or hyperglycemic prior to cardiac arrest ischemia or by increasing preischemic PCO2 in normoglycemic animals. For comparison, the N-methyl-D-aspartate (NMDA) receptor antagonist dizocilpine maleate (MK-801) was given prior to induction of ischemia. In some experiments the effect of acidosis on K+ efflux and Na+ influx were studied as well. In hypoglycemic subjects, the reduction of Ca2+e during ischemia was very rapid, 90% of the reduction occurring within 4.7 s. Normoglycemic animals showed a slower rate of reduction of Ca2+e. Hyperglycemic animals displayed an even slower rate of reduction and a biphasic response in which the initial, faster influx of Ca2+ was followed by a conspicuously slow one. This second phase led to a very gradual decrease in Ca2+e, a stable level being reached first after 6-7 min. This marked delay in calcium influx during ischemia was very similar in hypercapnic animals, who showed an extracellular pH during ischemia as low as hyperglycemic subjects. The effect of acidosis was duplicated by MK-801, suggesting that low pH reduces calcium influx by blocking NMDA-gated ion channels.

Topics: Animals; Blood Glucose; Brain; Brain Ischemia; Calcium; Carbon Dioxide; Cerebral Cortex; Dizocilpine Maleate; Heart Arrest; Hydrogen-Ion Concentration; Hyperglycemia; Hypoglycemia; Male; Parietal Lobe; Potassium; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Sodium

Recently, diffusion-weighted magnetic resonance imaging (DWI) has been shown to visualize acute ischemic lesions in the brain before changes are observable with conventional magnetic resonance imaging. However, the underlying mechanisms of these acute DWI changes are unclear and may include both reversible and irreversible damage. In this study, we demonstrate that acute DWI lesions may be reversed with MK801 therapy postischemia. Sprague-Dawley rats (n = 12) were subjected to middle cerebral artery occlusion and DWI scans were obtained beginning 60 min postocclusion. Distinct regions of hyperintensity were observed in the basal ganglia and cortex, corresponding with the expected distribution of ischemia in this model. After the first scan, animals were treated with MK801 (0.5 mg/kg i.v.) or normal saline and subsequently scanned again 30 and 60 min after treatment. In the control group, the area of hyperintense lesions continued to increase, by 55% in the cortex and 57% in the basal ganglia. MK801 therapy significantly (p < 0.01) reduced the area of damage by the third DWI scan at 60 min posttreatment (-50% cortex, -22% basal ganglia, -41% total hemisphere) compared to pretreatment scans. Tetrazolium (TTC) stains at 24 h confirmed that MK801 significantly reduced the volumes of infarction (p < 0.05). These results demonstrate that significant portions of the acute ischemic lesion on DWI are reversible with pharmacologic intervention.

Topics: Animals; Brain; Brain Ischemia; Diffusion; Dizocilpine Maleate; Image Processing, Computer-Assisted; Magnetic Resonance Imaging; Male; Rats; Rats, Sprague-Dawley; Time Factors

Excitatory amino acids and their receptors have been postulated to be involved in mediating ischemic neuronal damage. We occluded the bilateral carotid arteries for 5 min in gerbils to examine the effect of FR115427, a novel N-methyl-D-aspartate (NMDA) antagonist, on ischemic neuronal damage. FR115427 prevented hippocampal CA1 cell damage at a dose of 10 mg/kg and reduced spontaneous locomotor hyperactivity in gerbils after the development of ischemia at a dose of 32 mg/kg. The effective doses of MK801 were 3.2 mg/kg for preventing hippocampal CA1 cell damage and 1 mg/kg for reducing spontaneous locomotor hyperactivity. Moreover, we monitored the changes in body temperature of ischemic gerbils for 24 hr. The body temperature of ischemic gerbils significantly increased 1 hr after reperfusion. The pretreatment with FR115427 or MK801 prevented the hyperthermia provoked 1 hr after reperfusion in ischemic gerbils. In addition, the hypothermia was developed in gerbils treated with MK801 24 hr after reperfusion. However, FR115427 did not show hypothermia at any time. These results indicate that FR115427 has a protective effect against ischemic hippocampal CA1 cell damage after systemic administration, and this protective effect appears to be due to anti-NMDA activity.

Topics: Animals; Body Temperature; Brain Ischemia; Dizocilpine Maleate; Gerbillinae; Hippocampus; Isoquinolines; Male; Motor Activity; N-Methylaspartate; Pyramidal Cells; Receptors, N-Methyl-D-Aspartate; Tetrahydroisoquinolines

Previously we reported an improvement in histological outcome in cats treated with MK-801 shortly after the induction of temporary middle cerebral artery occlusion, and examined after 2 h of ischemia followed by 4 h of reperfusion. This study investigates the prolonged effects of the same drug treatment. Focal cerebral ischemia was produced in 34 cats by temporary occlusion of the left middle cerebral artery for 2 h. Stroke severity was determined using the ratio of the EEG amplitude from the ipsilateral to that of the contralateral hemisphere. Thirty minutes after the onset of ischemia, cats were treated i.v. with either 1 mg/kg MK-801 or saline. Electrocortical activity of the animals who survived were followed for 6 days postocclusion at which point they were sacrificed for histopathological analysis. Twelve of the animals died during recovery, of which 4 were MK-801 treated, and 8 were saline controls. The EEG ratios in the non-surviving animals were more depressed than in the animals that survived, whereas the depression in the EEG amplitude in both the treated and the control surviving animals was equal. Among the survivors no reduction in infarct size with MK-801 treatment was observed. Thus treatment with MK-801 in the middle cerebral artery occlusion model in the cat leads to a significant increase in the rate of survival (P < 0.05), but no prolonged improvement in late histopathology, in contrast with acute histological findings using this model. MK-801 treatment may be shifting the stroke model towards the survival of animals with larger infarcts. Histological recovery during prolonged reperfusion may eliminate the early neuroprotective effects seen with MK-801 treatment.

Topics: Analysis of Variance; Animals; Blood Pressure; Brain; Brain Ischemia; Cats; Dizocilpine Maleate; Electroencephalography; Male; Survival Analysis

Accumulation of glial fibrillary acidic protein xk(G-FAP) in reactive astrocytes is a characteristic neuropathologic feature of ischemic brain injury. We examined injury-induced changes in GFAP mRNA and protein in a well-characterized model of focal hypoxic-ischemic injury in perinatal rodent brain. Postnatal Day (PND) 7 rats underwent right carotid artery ligation followed by 2.5 h exposure to 8% oxygen, which results in injury to ipsilateral cortex, hippocampus, and striatum in the majority of animals. Using Northern analysis, we assayed GFAP mRNA in samples from the lesioned and contralateral hemispheres of animals killed 1 h to 14 days later, and from animals treated with the neuroprotective glutamate antagonist MK-801. GFAP immunoreactivity in tissue homogenates from the lesioned and contralateral hemispheres was also compared with an immunoblot assay. One and 4 h posthypoxia GFAP mRNA expression was barely detectable. In the lesioned cortex, increased GFAP mRNA was detected at 24 h postinjury; over the next 2 weeks GFAP mRNA was consistently higher (at least 2-fold) in lesioned than in contralateral cortex. In contrast, in lesioned hippocampus and striatum, consistent increases in GFAP mRNA were first detected on PND 12. Immunoassays of GFAP demonstrated early (PND 8) and sustained (to PND 21) up to 10-fold increases in lesioned cortex, hippocampus, and striatum. In this perinatal stroke model regionally specific increases in GFAP mRNA expression and GFAP immunoreactivity are detected in the first 2 weeks after hypoxic-ischemic injury; intrinsic properties of glia and/or neurons in different brain regions may influence the timing and magnitude of stimulation of this response.

Topics: Aging; Animals; Animals, Newborn; Blotting, Northern; Brain; Brain Ischemia; Cerebral Cortex; Corpus Striatum; Dizocilpine Maleate; Functional Laterality; Glial Fibrillary Acidic Protein; Hippocampus; Hypoxia, Brain; Immunohistochemistry; N-Methylaspartate; Organ Specificity; Rats; Rats, Sprague-Dawley; RNA, Messenger

Loss of cellular calcium homeostasis or the production of nitric oxide (NO) have been cited as possible mechanisms that may contribute to neuronal degeneration during ischemia. We therefore examined whether cellular calcium blockade, using the agent HA1077, was protective during anoxia in hippocampal neuronal cell cultures, and whether the in vitro effects of this drug were linked to the NO pathway. Administration of the agent during anoxia was neuroprotective in neuronal cell culture. In contrast, HA1077 did not protect hippocampal neurons during NO exposure. In addition, inhibition of NO synthesis in conjunction with HA1077 application during anoxia did not significantly increase survival beyond the maximum protection afforded by HA1077 alone. These results suggest that calcium may be an initial messenger in the ischemic cascade, but that subsequent neuronal degeneration is dependent upon the NO pathway.

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Animals; Brain Ischemia; Calcium; Cell Death; Cells, Cultured; Dizocilpine Maleate; Hippocampus; Homeostasis; Isoquinolines; Nitric Oxide; Nitroprusside; Rats; Rats, Sprague-Dawley

Transient ischemia-induced perturbations in calcium homeostasis have been proposed to lead to pathological activation of the cysteine protease calpain I and subsequent delayed neuronal death in the CA1 region of hippocampus. We report here on the design and characterization of antibodies selective for calpain-generated fragments of brain spectrin, and their use for immunoblot and immunohistochemical analyses of calpain activation following cerebral ischemia in the gerbil. Although spectrin was susceptible to degradation in vitro by many mammalian proteases, only calpain degraded spectrin to generate fragments immunoreactive with the antibodies. Following 5 min of global ischemia, immunoreactivity for calpain-degraded spectrin was rapidly (within 30 min) and markedly elevated in the perikarya and dendrites of several populations of forebrain neurons. The rapid calpain activation was completely prevented by the NMDA receptor antagonist MK-801. At later times postischemia, but prior to frank neuronal necrosis, calpain-degraded spectrin was restricted to hippocampal area CA1 pyramidal neurons. Silver impregnation histochemistry confirmed that neuronal damage was confined to area CA1. The results indicate that while nonpathological NMDA receptor stimulation can activate calpain, only those neurons showing sustained calpain activation are destined to die.

Topics: Amino Acid Sequence; Animals; Brain; Brain Ischemia; Calpain; Dizocilpine Maleate; Gerbillinae; Immunoblotting; Male; Molecular Sequence Data; Neurons; Spectrin; Time Factors; Tissue Distribution

The hippocampus demonstrates a regional pattern of vulnerability to ischemic injury that depends on its characteristic differentiation and intrinsic connections. We now describe a model of ischemic injury using organotypic hippocampal culture, which preserves the anatomic differentiation of the hippocampus in long-term tissue culture.. Ischemic conditions were modeled by metabolic inhibition. Cultures were briefly exposed to potassium cyanide to block oxidative phosphorylation and 2-deoxyglucose to block glycolysis. The fluorescent dye propidium iodide was used to observe membrane damage in living cultures during recovery.. 2-Deoxyglucose/potassium cyanide incubation resulted in dose-dependent, regionally selective neuronal injury in CA1 and the dentate hilus, which began slowly after 2 to 6 hours of recovery. Subsequent histological examination of cultures after 1 to 7 days of recovery demonstrated neuronal pyknosis that was correlated with the early, direct observation of membrane damage with propidium. Both propidium staining and histological degeneration were prevented by the noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 when administered 30 minutes after the end of the exposure to 2-deoxyglucose and potassium cyanide. Tetrodotoxin, which blocks voltage-dependent sodium channels, had protective effects that were greatest during the period of 2-deoxyglucose and potassium cyanide incubation but also produced protection against the mildest conditions of metabolic inhibition when administered after 30 minutes of recovery.. This in vitro model reproduced elements of the time course, regional vulnerability, and pharmacologic sensitivities of in vivo ischemic hippocampal injury. Inhibition of metabolism in organotypic culture provides a rapid, easily controlled injury and reproduces the in vitro pattern of hippocampal regional vulnerability to ischemia. It is the first in vitro model of ischemia to exhibit complete protection by delayed administration of an NMDA receptor antagonist during recovery from a brief insult. The protective effects of tetrodotoxin suggest that an early period of sodium entry into cells during and after ATP depletion may be responsible for the more prolonged period of toxic NMDA receptor activation.

Topics: Animals; Brain Ischemia; Deoxyglucose; Disease Models, Animal; Dizocilpine Maleate; Glycolysis; Hippocampus; Neurons; Organ Culture Techniques; Oxidative Phosphorylation; Potassium Cyanide; Pyramidal Tracts; Rats; Tetrodotoxin; Time Factors

Release of endogenous polyamines may contribute to neuronal loss in ischemia and related conditions. Primary cortical neurons were exposed to spermine and spermidine and subsequently assayed for [3H]ouabain binding to quantify neuronal loss. Neuronal survival was significantly decreased in the presence of spermine at 24 h (500 microM), 48 h (250 microM and 500 microM) and 72 h (10-500 microM) relative to controls. Co-application of 250 microM spermine and 10 microM dizocilpine for 48 h completely inhibited the effect of spermine alone. Spermidine exposure (10-500 microM) did not alter neuronal survival at any of the time points. These data indicate that the polyamine spermine is toxic to neurons in vitro and that toxicity is prevented by the NMDA-associated channel antagonist dizocilpine.

Topics: Animals; Brain Ischemia; Cell Survival; Cells, Cultured; Cerebral Cortex; Chick Embryo; Dizocilpine Maleate; Ion Channels; Nervous System; Neurons; Receptors, N-Methyl-D-Aspartate; Spermidine; Spermine

The ability of five agents (dizocilpine [MK-801], 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)-quinoxaline [NBQX], enadoline [CI-977], L-nitroarginine methyl ester [L-NAME] and BW 1003c87) with well defined, distinct pharmacological profiles and with established anti-ischemic efficacy, to modify neuronal damage has been examined in a simple in vivo model of glutamate excitotoxicity. Cortical lesions were produced in physiologically-monitored halothane-anesthetised rats by reverse dialysis of glutamate. The volume of the lesion was quantified histologically by image analysis of approximately 20 sections taken at 200 microm intervals throughout the lesion. The AMPA and NMDA receptor antagonists (NBQX and MK-801) and the inhibitor of nitric oxide synthase (L-NAME) significantly reduced the lesion volume by a similar extent (by approximately 30% from vehicle). Two agents (the kappa opioid agonist, CI-977 and the sodium channel blocker, BW 1003c87) which putatively inhibit the release of endogenous glutamate presynaptically, had dissimilar effects on lesion size. CI-977 failed to alter the amount of damage produced by exogenous glutamate, whereas BW 1003c87 reduced the lesion size by approximately 50%. Using this model, the neuroprotective effects of anti-ischemic drugs can be explored in vivo, uncomplicated in contrast to experimental ischemia by reduced oxygen delivery, drug effects on tissue blood flow and compromised energy generation. In consequence, additional mechanistic insight into anti-ischemic drug action in vivo can be obtained.

Topics: Amino Acid Oxidoreductases; Animals; Arginine; Benzofurans; Blood Pressure; Brain Ischemia; Cerebral Cortex; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamates; Glutamic Acid; Male; Microdialysis; Neurons; Neurotoxins; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Pyrimidines; Pyrrolidines; Quinoxalines; Rats; Rats, Sprague-Dawley; Time Factors

Immunoblot analysis using a phosphotyrosine-specific antibody was performed to investigate tyrosine phosphorylation of the mitogen activated protein (MAP) kinase in the rat brain. Epileptic seizures induced by systemic injection of bicuculline caused a rapid and transient stimulation of MAP kinase tyrosine phosphorylation in hippocampus and somatosensory cortex. This increase in tyrosine phosphorylation was markedly attenuated by the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801. In contrast, in the cerebellum, tyrosine phosphorylation of MAP kinase remained undetectable after bicuculline-induced seizures. These results demonstrate that generalized seizures stimulate tyrosine phosphorylation of MAP kinase in a regionally selective manner.

Topics: Animals; Bicuculline; Brain; Brain Ischemia; Dizocilpine Maleate; Enzyme Activation; Hippocampus; Immunoblotting; Mitogens; Phosphorylation; Phosphotyrosine; Protein-Tyrosine Kinases; Rats; Rats, Inbred Strains; Seizures; Somatosensory Cortex; Tyrosine

The acute anti-ischemic and anti-anoxic effects of dextrorphan (DX) were compared with those of dizocilpine (MK-801) in a variety of animal models, and in vivo and in vitro testings under anoxic conditions. DX reduced the incidence of death in ischemic mice and improved the rotarod performance of mice with brain ischemia. The ischemically-impaired memory of mice treated with DX markedly improved, as shown in the step-through type passive avoidance test, Morris water maze and in the habituation of exploratory behavior test. MK-801 likewise improved the water maze performance of the ischemically-impaired mice, but to a lesser extent. The step-through type passive avoidance performance of ischemic mice was not improved by MK-801. In the passive avoidance task with normal mice, DX, like MK-801, produced anterograde amnesia at doses higher than those needed to attenuate the behavioral effects of ischemia. DX, intravenously or centrally administered, markedly and dose-dependently reduced the incidence of death in mice receiving potassium cyanide (KCN). DX lessened the reduction in adenosine triphosphate (ATP) and increased lactate contents in mice dosed with KCN and also lessened the reduction in ATP in the TCA cycle and oxidative phosphorylation reactions caused by KCN (0.58 mmol/l), whereas MK-801 failed to show any effect on ATP formation pathways in vivo and in vitro, and failed to protect mice against KCN-induced lethal toxicity in vivo. In the in vitro studies, DX increased the adenylate kinase activity of the rat brain homogenate. DX was found to be a cerebroprotectant with anti-ischemic and anti-anoxic actions, the effects probably stemming from its N-methyl-d-aspartate receptor antagonistic property in cooperation with its ATP replenishing action.

Topics: Adenosine Triphosphate; Adenylyl Cyclases; Animals; Avoidance Learning; Behavior, Animal; Brain Ischemia; Dextrorphan; Dizocilpine Maleate; Exploratory Behavior; Hypoxia, Brain; Injections, Intraventricular; Male; Maze Learning; Mice; Mice, Inbred ICR; Mice, Inbred Strains; N-Methylaspartate; Nerve Degeneration; Oxidative Phosphorylation; Postural Balance; Potassium Cyanide; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate

The effect of the glutamate antagonist MK-801 on the ischemic threshold of energy metabolism and protein synthesis (CPS) was studied in rats submitted to 3 h occlusion of the left middle cerebral artery (MCA). Local blood flow and CPS were measured by double tracer autoradiography, and local ATP content by bioluminescence imaging. In untreated animals breakdown of energy metabolism occurred at flow values below 15 +/- 1 and CPS inhibition below 51 +/- 15 ml/100 g/min (means +/- S.D.). MK-801 treatment (3 mg/kg immediately after MCA occlusion) did not change the ischemic flow threshold of energy failure (16 +/- 3 ml/100 g/min) but lead to a highly significant decline of the perfusion threshold for the inhibition of CPS to 19 +/- 4 ml/100 g/min (P < 0.01). Our data demonstrate that MK-801 dramatically reduces the threshold for the suppression of protein synthesis which could explain previously reported therapeutical effects on the reduction of brain infarct size.

Topics: Adenosine Triphosphate; Animals; Autoradiography; Brain Ischemia; Cerebral Arteries; Cerebrovascular Circulation; Constriction; Differential Threshold; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Leucine; Male; Proteins; Rats; Rats, Inbred Strains

N-methyl-D-aspartate receptor antagonists are potent neuroprotectants in experimental focal cerebral ischemia, but behavioral and neuropathologic changes seen with these drugs in rodent models may limit the clinical utility of these compounds. Glycine's modulation of N-methyl-D-aspartate channel function offers another pharmacologic approach to excitotoxicity in ischemia. The potent glycine antagonist 7 Chlorothiokynurenic acid (7-Cl-Thio-Kyna) was studied in a permanent middle cerebral artery occlusion stroke model in the rat. The compound was effective, in a dose-dependent manner, in attenuating infarct size when administered before or after permanent middle cerebral artery occlusion. Its activity was mainly due to glycine antagonism inasmuch as 5 Chlorothiokynurenic acid, a compound having other pharmacological activities in common with 7-CI-Thio-Kyna (for instance the radical scavenger action), was inactive in this model. 7-Cl-Thio-Kyna did not produce cytological changes similar to MK 801.

Topics: Animals; Brain; Brain Chemistry; Brain Diseases; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Free Radical Scavengers; Heat-Shock Proteins; Immunohistochemistry; Kynurenic Acid; Male; Rats; Rats, Sprague-Dawley; Receptors, Glycine

Increases in mRNA levels for c-fos, c-jun, junB, hsp70 and NGFI-A were observed in the dentate gyrus of the hippocampus following 7 min ischaemia in the Mongolian gerbil. The response was rapid and transient (30 min to 4 hr) for NGFI-A, junB and c-fos mRNA. In contrast c-jun mRNA remained increased for several hours. Hsp70 increased in the dentate gyrus 1 hr after the insult, returned to control values at 4 hr and showed a secondary increase at 24 hr. At 24 hr increased hsp70 mRNA was observed in other regions of the CNS, i.e. CA1, CA2, CA3 and cortex. The non-competitive NMDA receptor antagonist, dizocilpine, attenuated the increases in IEG expression and was neuroprotective. In contrast the kappa opiate receptor agonist, enadoline, protected the CA1 neurones from degeneration but did not inhibit the increased levels of IEG mRNA. Increases in hsp70 mRNA were reduced to baseline by both enadoline and dizocilpine. These results suggest that inhibition of IEG expression is not a prerequisite for neuroprotection. However, hsp70 was predictive of neuronal protection and may be a useful assay in this and related models.

Topics: Animals; Autoradiography; Benzofurans; Brain Ischemia; Dizocilpine Maleate; DNA-Binding Proteins; Female; Gene Expression; Genes, Immediate-Early; Gerbillinae; Heat-Shock Proteins; Hippocampus; Pyrrolidines; Receptors, N-Methyl-D-Aspartate; Receptors, Opioid, kappa; RNA, Messenger; Transcription Factors

Neurons undergoing delayed neuronal death produced by hypoxia-ischaemia (HI) or status epilepticus (SE) showed a massive expression of c-Jun in their nuclei 24 h after the insult. With SE there was also a weaker induction of c-Fos and Jun B in dying neurons. SE induced in the presence of the NMDA antagonist MK-801 produced no delayed c-Jun expression in the hippocampus and nerve cell death did not occur in this region, although there was a delayed c-jun expression in the amygdala/piriform region, and cell death occurred in this area. Activation of central muscarinic receptors with pilocarpine, or block of D2 dopamine receptors with haloperidol, treatments which do not cause neuronal damage, strongly induced Fos and Jun B in hippocampal and striatal neurons, but only induced c-Jun very weakly. Thus, c-Jun may participate in the genetic cascade of events that produce programmed cell death in neurons.

Topics: Animals; Apoptosis; Brain Ischemia; Dizocilpine Maleate; Genes, fos; Genes, jun; Haloperidol; Hypoxia, Brain; Male; Neurons; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus; Transcription, Genetic

The effect of peri-infarct depolarizations on ischaemic injury was studied in rats submitted to 3 h occlusion of the left middle cerebral artery (MCA). The number of depolarizations varied from 1 to 8 and infarct volume from 37 to 159 mm3. The correlation between the two variables revealed a highly significant linear relationship (r = 0.800; p < 0.005), each depolarization accounting for an increase in infarct volume by about 13 mm3. The aggravating effect of repeated depolarizations was also demonstrated by the gradual increase in cortical DC shift duration, in EEG amplitude recovery time, and in EEG delta power with increasing number of depolarizations. Suppression of peri-infarct depolarizations is a rational approach for reducing the severity of ischaemic stroke.

Topics: Animals; Brain Ischemia; Cerebral Arteries; Cerebral Cortex; Cerebral Infarction; Cortical Spreading Depression; Dizocilpine Maleate; Electroencephalography; Male; Neurons; Rats; Rats, Inbred F344

Using [3H]MK-801, [3H]muscimol, [3H]cyclic AMP, and [3H]rolipram, we performed quantitative in vitro autoradiography to determine sequential alterations in the binding of N-methyl-D-aspartate and GABAA receptors, particulate cyclic AMP-dependent protein kinase, and cyclic AMP-selective phosphodiesterase, respectively, in the gerbil hippocampus following repeated brief ischemic insults. Changes from 1 h to 28 days after three 2-min ischemic insults at 1-h intervals were compared with those after 2 and 6 min of ischemia. We observed no alterations in the binding of all the four ligands throughout the observation period following 2 min of ischemia which produced no histological neuronal damage except for transient reductions in [3H]cyclic AMP binding during the early reperfusion period. [3H]Cyclic AMP binding in the CA1 subfield decreased one day after 6 min of ischemia and four days after three 2-min ischemic insults, and 62-65% of the binding was lost after 28 days. [3H]Rolipram binding in the CA1 subfield decreased one day after 6 min of ischemia and the binding was reduced by 45-50% after four and 28 days. Following three 2-min ischemic insults, [3H]rolipram binding decreased in the CA1 at one day, and decreased by 25-33% after 28 days. Both [3H]MK-801 and [3H]muscimol binding was preserved during the early reperfusion period after 6 min of ischemia and three 2-min ischemic insults. Reductions in [3H]MK-801 binding in CA1 were observed four days after ischemic insults when CA1 neuronal destruction was seen. After one month, approximately 50% of [3H]MK-801 binding was lost in CA1 in both groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 3',5'-Cyclic-AMP Phosphodiesterases; Animals; Autoradiography; Brain Ischemia; Cyclic AMP; Dizocilpine Maleate; Gerbillinae; Hippocampus; Histocytochemistry; Male; Muscimol; Phosphodiesterase Inhibitors; Protein Kinases; Pyrrolidinones; Rolipram

Excessive activation of the N-methyl-D-aspartate (NMDA) subtype of glutamate receptor has been implicated in the sequence of neurochemical events in cerebral ischaemia that results in irreversible neuronal damage. The effects of the NMDA antagonist MK-801 upon the amount of ischaemic brain damage has been assessed quantitatively in a cat and in a rat model of focal cerebral ischaemia. In chloralose-anaesthetised cats, focal cerebral ischaemia was produced by permanent occlusion of one middle cerebral artery (MCA) and the animal sacrificed 6 hours later. Pretreatment with the non-competitive NMDA antagonist, MK-801 (5 mg/kg, i.v.) reduced significantly the volume of ischaemic damage in the cerebral cortex by 57% compared to vehicle-treated cats. A similar degree of neuroprotection could be demonstrated in the cat MCA occlusion model if treatment with MK-801 was initiated 2 hours after the induction of ischaemia. In halothane-anaesthetised rats, focal cerebral ischaemia was produced by permanent MCA occlusion and the animals sacrificed 3 hours later. Pretreatment with MK-801 (0.5 mg/kg, i.v.) reduced the volume of ischaemic damage in the cerebral cortex by 38%; treatment with MK-801 initiated 30 minutes after MCA occlusion was equally effective in reducing cortical damage. In contrast to calcium entry blockers such as nimodipine in the rat MCA occlusion model, the improved histopathological outcome with MK-801 is not associated with improvement in cerebral tissue perfusion to the ischaemic tissue. The increasing evidence that NMDA receptor antagonists are beneficial in experimental focal cerebral ischaemia is reviewed.

Topics: Animals; Brain Damage, Chronic; Brain Ischemia; Cats; Caudate Nucleus; Cerebral Cortex; Cerebral Infarction; Dizocilpine Maleate; Dominance, Cerebral; Male; Nimodipine; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

We developed a sensitive and reliable gas chromatographic (GC) technique for the quantitative analysis of MK-801 in brain and serum and applied the technique to investigate tissue concentrations of neuroprotective and non-neuroprotective doses of MK-801 in a neonatal rat model of hypoxic-ischemia. Brain concentrations of MK-801 were a linear function of dose over 4 orders of magnitude. After administration of a neuroprotective dose of MK-801 (29.6 mumol/kg) to control rats, both serum and brain concentrations rose rapidly to approximately 300 nM and approximately 2000 nM, respectively, within 30 min. Approximately 60% of serum and 90% of brain MK-801 were bound leaving the free concentrations in both blood and brain at approximately 100-200 nM. After hypoxic-ischemia, serum MK-801 concentrations were not different from controls but brain concentrations were lowered by 32%. Free brain concentrations of MK-801 after hypoxic-ischemic were 124 nM after 29.6 mumol/kg while after 8.9 mumol/kg (the non-neuroprotective dose) they were 39 nM. In view of the slow kinetics of MK-801 channel blockade, which never reaches equilibrium during the current experiment, this difference in concentration would be expected to result in an 80-fold (or greater) difference in the current flux through NMDA receptor-operated ion-channels as the rate of NMDA receptor-operated ion-channel blockade is concentration dependent.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Animals, Newborn; Brain; Brain Ischemia; Chromatography, Gas; Dizocilpine Maleate; Female; Gas Chromatography-Mass Spectrometry; Hypoxia, Brain; Rats; Rats, Wistar; Tissue Distribution

The efficacy of GM1 ganglioside treatment in stroke was studied in a permanent middle cerebral artery occlusion model in the rat. A dose-dependent attenuation of infarct size at 24 hr was documented with the maximum effective dose halving the volume of the experimental stroke. Delayed administration at 5 min, but not 15 min, after vessel occlusion was as effective as preocclusion drug administration. Morphologic sparing was confined to the cortical penumbra; no protection in the ischemic core was found. Morphologically salvaged cortex was also metabolically preserved as demonstrated by quantitative measurement of glucose utilization. In vivo microdialysis demonstrated an attenuation of ischemic-induced glutamate release in the cortex with GM1 administration but no effect was found in the caudate. Hypotension did not occur even with doubling of the maximally effective dose of GM1. Accordingly, GM1 may be a safe and effective treatment for stroke.

Topics: Animals; Behavior, Animal; Brain; Brain Ischemia; Cerebral Cortex; Deoxyglucose; Dialysis; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; G(M1) Ganglioside; Male; Rats; Rats, Sprague-Dawley

New insights into the pathophysiology of the hypoxic-ischemic insult have opened the possibility of pharmacologic intervention in neonatal hypoxic-ischemic encephalopathy. It is now known that many neurons survive a hypoxic-ischemic insult but remain dysfunctional for hours, with profound alterations in cell function. A cascade of biochemical alterations occurs as a consequence of cellular ionic shifts, energy depletion, degradation of cell membrane phospholipids, and increased release of neurotransmitters. In addition, there are alterations in the metabolism of arachidonic acid and prostanoids and an excessive production of oxygen free radicals. The new therapeutic modalities are aimed at preventing or arresting the biochemical changes that occur in the period after hypoxia-ischemia. This review details the biochemical alterations associated with neonatal hypoxic-ischemic encephalopathy and discusses the possible use in newborns of pharmacologic agents currently undergoing extensive investigations in experimental animals and adult humans.

Topics: Asphyxia Neonatorum; Brain Ischemia; Calcium Channel Blockers; Dizocilpine Maleate; Free Radical Scavengers; Humans; Hypoxia, Brain; Infant, Newborn; Lipid Peroxides; Pregnatrienes; Reperfusion Injury

Cortical spreading depression (CSD) is a transient depression of neuronal activity that spreads across the cortical surface. In the present studies, we have investigated CSD activity in the penumbral zone following permanent middle cerebral artery (MCA) occlusion in the rat (n = 16/group), using double-barreled Ca(2+)-sensitive microelectrodes. Measurements of CSD activity were made for 3 h in each animal. During this time, a varying number of spontaneous CSDs were seen in the control group (total was 30, with a range of 0-7/rat). These CSDs were of varying duration: "small" (approximately 1 min) and "big" (5-45 min) CSDs. During a CSD, the extracellular [Ca2+] decreased to 0.11 +/- 0.07 mM (mean +/- SD). After 3 h, the extracellular [Ca2+] in the cortex (penumbral zone) was either normal (10/16 rats) or lowered to 0.5 mM (2/16 rats) or to 0.1 mM (4/16 rats). In the caudate nucleus (ischaemic core area), all rats had an extracellular [Ca2+] of approximately 0.1 mM when measured after the 3 h recording period. Neuropathological evaluation of the brains of the animals, which had been allowed to survive for 24 h after MCA occlusion, revealed ischaemic damage in the dorsolateral cortex and caudate nucleus. Administration of the noncompetitive NMDA antagonist, MK-801 (3 mg/kg i.p.), 30 min after MCA occlusion resulted in 24 and 29% reductions in the volume of hemispheric and cortical damage, respectively, which was highly significant (p less than 0.0001); no protection was seen against caudate damage.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Brain Ischemia; Calcium; Cerebral Arterial Diseases; Cerebral Cortex; Dizocilpine Maleate; Male; Rats; Rats, Inbred Strains; Synaptic Transmission; Tissue Survival

The present study reports on the acute effects of MK-801 on the histopathological outcome and blood flow changes during focal cerebral ischemia and reperfusion. In addition, acute changes in the EEG and blood pressure are also reported. In 16 halothane-anesthetized cats, the left middle cerebral artery (MCA) was occluded for 2 h followed by 4 h of reperfusion. Thirty minutes after the onset of ischemia, eight animals were treated with 1 mg/kg of MK-801, while eight animals received saline. Blood flow from the peripheral MCA territory was measured with H2 clearance. There was a comparable reduction in blood flow (down to 20% of control) in the ischemic gyri of the two groups followed by a partial recovery after recirculation. There was a similar decrease in the EEG amplitude over the ischemic central MCA territory in the treated and the untreated group. Treatment with MK-801 induced a burst suppression in the EEG and a transient drop (11.4 +/- 6.5 mm Hg) in the mean arterial pressure. The volume of early ischemic damage decreased by one-third in the MK-801-treated group compared to the untreated one, both in the total hemisphere (from 29 +/- 10 to 20 +/- 5%) and in the hemispheric cortex (range 36 +/- 8 to 24 +/- 13%). A major fraction of this improvement was localized to the middle and posterior parietal (mainly perifocal) regions of the MCA territory. These results show that in our model, MK-801 improves histopathological outcome despite the lack of apparent effect on the cortical blood flow, and an adverse effect on the systemic blood pressure. This is the first report that describes data on a reproducible model of reperfusion after temporary occlusion of the MCA in a cat, extending the findings of the Glasgow group, who observed similar neuroprotection in models of permanent MCA occlusion.

Topics: Animals; Blood Pressure; Brain Ischemia; Cats; Cerebral Arteries; Cerebral Cortex; Dizocilpine Maleate; Electroencephalography; Male; Regional Blood Flow; Reperfusion; Synaptic Transmission; Tissue Survival

[3H]dopamine ([3H]DA) release was measured from rat striatal slices under normoxic and hypoxic conditions. In some experiments hypoxia was combined with glucose withdrawal. Hypoxia increased the evoked release of dopamine without affecting resting release. Hypoglycemia itself increased only the resting release of [3H]DA. In the absence of glucose hypoxia provoked a dramatic rise in both resting and stimulation-evoked release of dopamine. This effect was partly reduced by Ca2+ withdrawal, and was abolished in the presence of tetrodotoxin (1 microM). The NMDA-receptor antagonist MK-801 (3 microM) attenuated the effect of hypoxia and hypoglycemia on [3H]DA release. It was suggested that activation of NMDA receptors is involved in dopamine release during hypoxia and energy deprivation.

Topics: Animals; Brain Ischemia; Cell Hypoxia; Corpus Striatum; Disease Models, Animal; Dizocilpine Maleate; Dopamine; Glucose; Male; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Tetrodotoxin

Excitatory neurotransmitters appear to cause cell death during ischemia by inducing depolarization, influx of ions, and metabolic failure in the postsynaptic neuron. If this hypothesis is correct, then postsynaptic membrane hyperpolarization and inhibition of metabolism may be protective. Antagonists of the excitotoxic amino acid glutamate protect neurons in culture and in animal models of stroke but appear to cause unacceptable side effects in humans. We propose an alternative strategy of protection using agonists of the inhibitory neurotransmitter gamma-aminobutyric acid.. We caused multifocal cerebral ischemia in rats and rabbits by injecting microspheres into the carotid circulation. We administered saline, muscimol, or MK-801 within 5 minutes of stroke onset. We used a bioassay to measure outcome. In rats, we also used learning to assess cortical function, and we performed detailed quantitative brain morphometry 3 months after infarction.. Using the bioassay, we found that muscimol exerted a protective effect in rats (p less than 0.01). There was a dose-response effect seen in muscimol-treated rabbits. Rats treated with muscimol or MK-801 exhibited significantly better visual-spatial learning compared with saline-treated subjects (p less than 0.001). Hemisphere volume after ischemia was comparable in all groups.. Agonists of gamma-aminobutyric acid and antagonists of glutamate appear to protect brain during ischemia. Since agonists of gamma-aminobutyric acid are known to have fewer side effects in humans, they may prove more useful in the clinical setting as neuroprotective agents.

Topics: Animals; Brain Ischemia; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Glutamic Acid; Learning; Male; Muscimol; Rabbits; Rats; Rats, Sprague-Dawley; Reperfusion Injury

Excitatory amino acids are implicated in the development of neuronal cell damage following periods of reversible cerebral ischemia or insulin-induced hypoglycemic coma. To explore the importance of glutamate receptor activation in the posthypoglycemic phase, we exposed rats to 20 min of insulin-induced severe hypoglycemia. The rats were treated immediately after the hypoglycemic insult with four regimes of glutamate receptor antagonists: (1) the AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazole propriate)-receptor antagonist NBQX [2.3-dihydroxy-6-nitro-7-sulfamoyl-benzo (F) quinoxaline] given as a bolus dose of 30 mg.kg-1 i.p., followed by an i.v. infusion of 225 micrograms.kg-1.min-1 for 6 h; (2) the non-competitive NMDA-receptor antagonist, dizocilpine (MK-801) 1 mg.kg-1 given i.v.; (3) a combined NBQX treatment, (a bolus dose of 10 mg.kg-1 i.p., followed by an i.v. infusion of 225 micrograms.kg-1.min-1 for 6 h), with dizocilpine 0.33 mg.kg-1 given twice i.p. at 0 and 15 min after recovery and (4) the competitive NMDA-receptor blocker CGP 40,116 [D-(E)-2-amino-4-methyl-5-phosphono-3- pentenoic acid] 10 mg.kg-1 given i.p. In the striatum, all glutamate receptor blockers significantly decreased neuronal damage by approximately 30%. An approximately 50% decrease in neuronal damage was demonstrated in neocortex and hippocampus following the combined treatment with NBQX and dizocilpine, while protection was variable following the treatment with a single glutamate-receptor antagonist.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 2-Amino-5-phosphonovalerate; Animals; Blood Pressure; Brain Ischemia; Cell Death; Dizocilpine Maleate; Electroencephalography; Excitatory Amino Acid Antagonists; Insulin Coma; Male; Necrosis; Neurons; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate

We examined whether MK-801, an N-methyl-D-aspartate (NMDA)-receptor antagonist, or anisomycin, a reversible protein synthesis inhibitor, inhibits the induction of ischemic tolerance following preconditioning with sublethal ischemia in gerbil hippocampus. Preconditioning with 2 min of ischemia, which induced heat shock protein-72 immunoreactivity, prevented hippocampal CA1 neuronal damage following 3 min of ischemia produced 3 days later. MK-801, but not anisomycin, inhibited the induction of tolerance although the heat shock protein synthesis was reduced in both groups. The present result suggests that NMDA receptor activation, causing stress response, induces the ischemic tolerance.

Topics: Animals; Anisomycin; Body Temperature; Brain Ischemia; Dizocilpine Maleate; Electroencephalography; Gerbillinae; Hippocampus; Immunohistochemistry; Male; Receptors, N-Methyl-D-Aspartate

Pretreatment with MK-801, a non-competitive N-methyl-D-aspartate (NMDA) antagonist, failed to protect the vagal component of reflex bradycardia from 5-min global cerebral ischemia in dogs under pentobarbital anesthesia. On the other hand, MK-801 completely prevented the development of the post-ischemic cerebral hypoperfusion without affecting the cerebral blood flow in sham-operated animals. The results suggest that NMDA receptors may participate in the development of the secondary disturbance of the cerebral circulation, but are not involved in the post-ischemic dysfunction of the baroreflex system.

Topics: Animals; Brain Ischemia; Cerebrovascular Circulation; Dizocilpine Maleate; Dogs; Female; Male; Pressoreceptors; Receptors, N-Methyl-D-Aspartate; Vagus Nerve

To determine the effect of suppression of epileptiform activity that develops after hypoxic-ischemic injury in the immature brain, chronically instrumented near-term fetal sheep (119-133 days) were subjected to 30 minutes of complete cerebral ischemia: 6 were given a 0.3-mg/kg bolus of MK-801 at 6 hours after the insult followed by continuous infusion of 1 mg/kg over the next 36 hours, and were compared to 6 control sheep. Electrocorticographic activity and edema within the parasagittal region of the cortex were quantified with real-time spectral analysis and impedance measurements, respectively. Histological outcome was assessed 72 hours later. The intense epileptiform activity seen from 9 +/- 2 to 30 +/- 3 hours in the control group was completely suppressed in the MK-801-treated group. The onset of secondary cortical edema was delayed from 9.4 +/- 1.1 hours to 14.8 +/- 0.7 hours (p < 0.01). Neuronal damage was reduced, particularly in the lateral cortex and hippocampus (p < 0.05). Infarction of the parasagittal cortex was not prevented. These results suggest that N-methyl-D-aspartate-mediated epileptiform activity that develops after a global hypoxic-ischemic insult worsens neuronal outcome in the immature brain.

Topics: Animals; Brain Ischemia; Cell Death; Cerebral Cortex; Dizocilpine Maleate; Electric Impedance; Electroencephalography; Epilepsy; Fetal Diseases; Hippocampus; Sheep

The Protective effects of dizocilpine maleate (DM) against anoxia in mice and ischemic damage in rats of 4-vessel occlusion (4-VO) were studied. DM 0.5 or 1.0 mg.kg-1 ip significantly prolonged the survival time of mice in closed containers. DM 0.5 and 1.0 mg.kg-1 ip 30 min prior to 4-VO obviously accelerated the electroencephalographic recovery, reduced the neuronal loss in the hippocampus, and increased the survival rate after 72-h reperfusion. These effects followed a dose-dependent manner. Our results indicate that selective non-competitive N-methyl-D-aspartate receptor blocker DM protects against anoxic and ischemic cerebral damage.

Topics: Animals; Brain Ischemia; Dizocilpine Maleate; Dose-Response Relationship, Drug; Electroencephalography; Hippocampus; Hypoxia; Male; Mice; Rats; Rats, Sprague-Dawley; Reperfusion Injury

FOS and JUN proteins are transcription factors thought to be involved in coupling neuronal excitation to target gene expression. Cortical infarction of consistent size and location was produced by irradiating the rat brain with Xenon light through the intact skull for 20 min following systemic injection of the photo-sensitizing dye, rose bengal. To investigate the time course and distribution pattern of five cellular immediate early gene (IEG)-encoded proteins after focal ischemia, the expression of c-FOS, FOS B, c-JUN, JUN B and JUN D was studied immunocytochemically in sham-operated control animals and at different postischemic time intervals up to 24 h. A separate group of animals was pretreated with the non-competitive N-methyl-D-aspartate (NMDA) antagonist MK-801. Photochemically induced focal ischemia caused a rapid induction of FOS and JUN proteins in the entire ipsilateral cortex apart from the ischemic focus. Immunoreactivity in the ipsilateral subcortical gray and white matter and in the entire contralateral hemisphere was indistinguishable from control animals. Individual IEG-encoded proteins were sequentially induced with increased levels of immunoreactivity persisting for different time periods up to 24 h. c-FOS, FOS B, c-JUN and JUN B exhibited a characteristic distribution pattern as reflected by different staining intensities in individual cortical layers. The rapid IEG induction in the entire ipsilateral sensorimotor and limbic structure-associated cortices after photochemically induced infarction most likely reflects spreading depression caused by ischemia and mediated by NMDA receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Antibody Formation; Brain Ischemia; Cerebral Cortex; Cerebral Infarction; Dizocilpine Maleate; Immunohistochemistry; Male; Photochemistry; Proto-Oncogene Proteins c-fos; Proto-Oncogene Proteins c-jun; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate

The protective effect of vinconate, a vinca alkaloid derivative, on ischemia-induced neuronal damage was investigated using a model of rat forebrain ischemia caused by occlusion of four vessels. Hippocampal cell loss was observed histologically and neurochemically 5 days after 10 min of ischemia. Treatment with vinconate (50 and 200 mg/kg i.p.) before cerebral ischemia significantly suppressed neuronal cell loss in the hippocampal CA1 region and the decrease in the content of neuroactive amino acids in the hippocampus. The release of neuroactive amino acids in the hippocampus was significantly increased by cerebral ischemia. Pretreatment with vinconate (50 and 200 mg/kg i.p.) significantly attenuated the increased release of glutamic acid and aspartic acid, but not the release of gamma-aminobutyric acid (GABA), taurine and glycine. This suppressive effect of vinconate was antagonized by scopolamine (10(-5) M). The addition of vinconate (10(-11)-10(-4) M) had no effect on the binding of [3H]MK-801. These results indicate that pretreatment with vinconate attenuates the ischemia-induced release of excitatory amino acids into the extracellular space of the hippocampus via the stimulation of presynaptic muscarinic acetylcholine receptors. The present results also suggest that this suppressive effect of vinconate on the release of excitatory amino acids (glutamic acid and aspartic acid) may play a crucial role in the protective action of this agent against ischemia-induced neuronal damage in the hippocampus.

Topics: Amino Acids; Animals; Brain Ischemia; Dizocilpine Maleate; Drug Interactions; Hippocampus; Male; Neurons; Rats; Rats, Wistar; Scopolamine; Synaptic Membranes; Vinca Alkaloids

Following permanent occlusion of the left middle cerebral artery (MCA) in rats, electrophysiological and hemodynamic characteristics of the periinfarct border zone were investigated in sham-operated (n = 6), untreated (n = 6), and MK-801-treated (3.0 mg/kg; n = 6) animals. For this purpose, direct current potential (DC), EEG, and blood flow (laser-Doppler flowmetry) were recorded from the cortex in the periphery of the MCA territory. In sham-operated rats, a single negative cortical DC deflection was observed after electrocoagulation of the cortex, whereas in untreated MCA-occluded animals, three to eight transient DC deflections were monitored during the initial 3 h of ischemia. The duration of these cortical DC shifts gradually increased from 1.2 +/- 0.3 to 3.7 +/- 2.7 min (mean +/- SD; p less than 0.05) during this time. In animals treated intraperitoneally with MK-801 (3.0 mg/kg) immediately after MCA occlusion, the number of cortical DC shifts significantly declined to one to three deflections (p less than 0.005). The EEG of the treated animals revealed low-amplitude burst-suppression activity. In the untreated and treated experimental group, the reduction of cortical blood flow amounted to 69 +/- 25 and 49 +/- 13% of control, respectively. Despite the more pronounced cortical oligemia, MK-801 treatment resulted in a significant decrease of the volume of the ischemically injured tissue from 108 +/- 38.5 (untreated group) to 58 +/- 11.5 (p less than 0.05) mm3. Our results suggest that repetitive cortical DC deflections in the periinfarct border zone contribute to the expansion of ischemic brain infarcts.

Topics: Action Potentials; Animals; Arterial Occlusive Diseases; Brain Ischemia; Cerebral Arterial Diseases; Cerebral Cortex; Cerebrovascular Circulation; Dizocilpine Maleate; Electroencephalography; Male; Rats

The levels of brain free fatty acids rapidly increase after the onset of ischemia. The purpose of this study was to investigate the action of phospholipases A2 and C during complete ischemia based on the effects of a phospholipase C inhibitor (phenylmethylsulfonyl fluoride) and the N-methyl-D-aspartate antagonist MK-801 on the release of free fatty acids in rat neocortex. Complete brain ischemia was induced in rats with cardiac arrest by intracardiac injection of KCl. Free fatty acid levels in the neocortex were measured 0, 2, 4, and 8 minutes after cardiac arrest. Phenylmethylsulfonyl fluoride inhibited the release of free fatty acids primarily from phosphatidylinositol during the first 2 minutes of ischemia and from phosphatidylcholine and phosphatidylethanolamine at 4 to 8 minutes of ischemia. Conversely, MK-801 inhibited free fatty acid release mainly from phosphatidylcholine and phosphatidylethanolamine at 2 to 4 minutes of ischemia. These results indicate that the release of free fatty acids during the first 2 minutes of ischemia can be attributed mostly to the action of phospholipase C, and that the activation of phospholipase C further influences the activation of phospholipase A2 in the subsequent course, while phospholipase A2 predominantly acts after 2 minutes of ischemia.

Topics: Animals; Brain Ischemia; Cerebral Cortex; Dizocilpine Maleate; Fatty Acids, Nonesterified; Male; Phenylmethylsulfonyl Fluoride; Phospholipases; Phospholipases A; Phospholipases A2; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate; Type C Phospholipases

Excitatory amino acids and their receptors are involved in mediating ischemic neuronal damage. The sigma-agonists are believed to interact with the N-methyl-D-aspartate receptor. Therefore, we studied the neuroprotective, hypothermic, and motor deficit effects of the sigma-agonist SKF 10,047 and the N-methyl-D-aspartate antagonist MK-801.. Neuroprotective effects were compared using an in vitro ischemia model of cultured rat cerebellar granule cells and the gerbil model of global brain ischemia induced by 5 minutes of bilateral carotid artery occlusion followed by 7 days of reperfusion.. In vitro, (+)MK-801 protected against 100 microM glutamate with a 50% protective concentration of 30 nM, followed by (-)MK-801 (150 nM), cyclazocine (0.5 microM), (+)SKF 10,047 (3.3 microM), pentazocine (5 microM), and (-)SKF 10,047 (10 microM). In vivo, (+)SKF 10,047 pretreatment (60 mg/kg) or multiple postischemic treatments provided neuroprotection comparable with MK-801 pretreatment (10 mg/kg). When ischemic animals were administered the multiple dosing regimen of (+)SKF 10,047, no hypothermic effect was noted in the temporalis muscle over 4 hours' postischemia. Motor deficits monitored by a swing grid test showed that 50% recovery from (+)SKF 10,047 was 5.5 times faster than recovery from MK-801.. These results are the first to report a hypothermia-free, in vivo neuroprotective effect of (+)SKF 10,047, a prototypical drug of the sigma-agonist class.

Topics: Animals; Body Temperature; Brain Ischemia; Cells, Cultured; Cerebellum; Cyclazocine; Dizocilpine Maleate; Gerbillinae; Pentazocine; Phenazocine; Rats; Rats, Inbred Strains; Reference Values; Stereoisomerism

Excessive activation of the N-methyl-D-aspartate receptor by glutamate produces an influx of Ca2+, which in turn is thought to lead to ischemic cell death. In this study we evaluated the combined treatment of the N-methyl-D-aspartate antagonist dizocilpine (MK-801) and the dihydropyridine Ca2+ channel blocker nicardipine for the reduction of hippocampal CA1 neuronal loss.. Global ischemia was induced by bilateral carotid artery occlusion in the gerbil. Body temperature was maintained between 36.5 degrees C and 37.5 degrees C during surgery. MK-801 (5.0 mg/kg) was injected 15 minutes after occlusion whereas nicardipine was given by injection and via a micro-osmotic pump (1.0 mg/kg/day) for 3 days.. Postischemic treatment with MK-801 reduced CA1 cell loss by 27.0%, whereas nicardipine reduced CA1 cell loss by 13.3%. Combined postischemic treatment with these drugs yielded an additive, protective effect (44.5% reduction of CA1 loss) that did not appear to result from postischemic hypothermia as assessed by skull and rectal temperature recordings.. Our results demonstrate that MK-801 plus nicardipine significantly attenuates CA1 cell death after forebrain ischemia in the gerbil. Excitatory amino acid antagonists in combination with Ca2+ channel antagonists may be an effective therapy in patients exposed to global ischemic insult.

Topics: Animals; Body Temperature; Brain Ischemia; Cell Death; Dizocilpine Maleate; Drug Combinations; Female; Gerbillinae; Hippocampus; Neurons; Nicardipine; Rectum; Skull

1. The effect of a novel, highly potent and selective kappa-opioid receptor agonist, GR89696, has been evaluated in two animal models of cerebral ischaemia: transient bilateral carotid artery occlusion in the Mongolian gerbil and permanent, unilateral middle cerebral artery occlusion in the mouse. 2. In the Mongolian gerbil model, administration of GR89696 (3 to 30 micrograms kg-1, s.c.), immediately before and at 4 h after insult, produced a dose-dependent reduction in the hippocampal CA1 neuronal cell loss resulting from a 7-min bilateral carotid occlusion. Similar effects were obtained with two other kappa-agonists, GR86014 (1 mgkg-1, s.c.) and GR91272 (1 mgkg-1, s.c.). The neuroprotective effect of GR89696 was completely blocked by prior administration of the opioid receptor antagonist, naltrexone, at 10 mgkg-1, s.c. Repeated post-treatment with GR89696 (100 micrograms kg-1, s.c.) or GR44821 (10 mgkg-1, s.c.) was also effective in protecting completely the hippocampal CA1 neurones from ischaemia-induced neurodegeneration. 3. In the permanent, unilateral middle cerebral artery occlusion model in the mouse, repeated administration of GR89696 at 300 micrograms kg-1, s.c. produced a 50% reduction in cerebrocortical infarct volume. In these experiments GR89696 was dosed 5 min, 4, 8, 12, 16, 20 and 24 h after occlusion on the first day and then three times daily for the next three days. GR89696 (300 micrograms kg-1) also produced a significant 35% reduction in infarct volume in this model when the initiation of dosing was delayed for 6 h after the insult. 4. The results indicate that the potent kappa-opioid receptor agonist, GR89696, is neuroprotective in both global and focal cerebral ischaemia models and suggest that, with this class of compound, there may be a considerable time window for pharmacological intervention.

Topics: Animals; Brain Ischemia; Carotid Artery Diseases; Dizocilpine Maleate; Dose-Response Relationship, Drug; Female; Gerbillinae; Male; Mice; Nervous System Diseases; Piperazines; Pyrrolidines; Receptors, Opioid; Receptors, Opioid, kappa

Evidence from animal stroke models suggests that the proximate cause of neuronal degeneration after ischemia is massive release of glutamate and activation of NMDA receptors. However, in the physiologic presence of oxygen and glucose in the rat hippocampal slice preparation, the neurotoxicity of glutamate, as measured by inhibition of protein synthesis, requires high concentrations and is not prevented by glutamate receptor antagonists. Thus, the NMDA receptor-mediated neurotoxic effects of extracellular glutamate accumulation during ischemia might depend on additional factors, such as neuronal depolarization. In the experiments reported here, slices were exposed to glutamate in a medium intended to mimic the ionic conditions found during ischemia, high potassium (128 mM) and low sodium (26 mM). This depolarizing medium itself inhibited protein synthesis in a manner which was partially mediated by NMDA receptor activation, since it was significantly reversed by the noncompetitive NMDA antagonist, MK-801. Furthermore, the effect of glutamate under depolarizing conditions was also significantly decreased by MK-801, suggesting that glutamate was acting at NMDA receptors. Thus, depolarization appears to enhance the sensitivity of neurons to toxic NMDA receptor activation by glutamate. Under conditions that mimic ischemia, hypoxia plus hypoglycemia, a similar protective effect of NMDA receptor antagonists was observed. Depolarization and ischemia both appeared to attenuate the neurotoxicity of non-NMDA receptor agonists. It appears that under conditions of normal glucose and oxygen, high concentrations of bath applied glutamate inhibit protein synthesis at sites other than the NMDA receptor. However, when the Na+ gradient is decreased, as occurs during ischemia, glutamate's NMDA effects predominate. These findings suggest that ionic shifts may play a central role in permitting NMDA receptor-mediated ischemic neuronal damage.

Topics: Animals; Brain Ischemia; Dizocilpine Maleate; Glucose; Glutamates; Hippocampus; In Vitro Techniques; Membrane Potentials; Nervous System Diseases; Neuromuscular Depolarizing Agents; Neurons; Oxygen; Potassium; Rats; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter; Sodium; Time Factors

The effects of a competitive, N-methyl-D-aspartate (NMDA) receptor antagonist, D(-)E-4-(3-phosphonoprop-2-enyl)-piperazine-2-carboxylic acid (D-CPPene), on the volume of ischemic brain damage was assessed by quantitative histological study in 35 chloralose-anesthetized cats. Focal cerebral ischemia was produced by permanent occlusion of one middle cerebral artery and the animals were killed by transcardiac perfusion fixation 6 hours later. Pretreatment with D-CPPene (1.5, 4.5, or 15 mg/kg, administered intravenously 15 minutes prior to occlusion, with subsequent drug infusions to maintain a plateau in the plasma drug concentrations) effected dose-dependent reductions in the volume of ischemic brain damage. At the highest dose studied (15 mg/kg, plus an infusion of 170 micrograms/kg/min), D-CPPene reduced the volume of ischemic damage in the cerebral cortex by more than 75% compared to vehicle-treated control animals. The plasma concentration of D-CPPene, which is associated with a half maximal reduction in the volume of ischemic damage, was estimated to be 24 micrograms/ml during the initial 120 minutes after the middle cerebral artery occlusion. Treatment with D-CPPene (15 mg/kg, plus an infusion of 170 micrograms/kg/min) initiated 1 hour after occlusion reduced the volume of ischemic brain damage in the cerebral cortex by 30%, but this response did not achieve statistical significance. Precise definition of dose dependency for the anti-ischemic effects of NMDA antagonists and the therapeutic time window are influenced greatly by brain pharmacokinetics of the agents.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Binding, Competitive; Brain; Brain Ischemia; Cats; Caudate Nucleus; Cerebral Cortex; Dizocilpine Maleate; Drug Administration Schedule; Half-Life; N-Methylaspartate; Piperazines; Receptors, N-Methyl-D-Aspartate

Ischemia may increase glutamate release, which can lead to neuronal damage. The therapeutic value of drugs that antagonize glutamate's effects are being investigated in CNS ischemia. This study examined the efficacy of a noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist, MK-801 [(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten- 5,10-imine hydrogen maleate], in reducing ischemic injury. We explored the limits of this therapy and different properties of MK-801 that might be involved in its neuroprotective actions. Two focal CNS ischemia models were used, a multiple cerebral embolic model (MCEM) and a rabbit spinal cord ischemia model (RSCIM). When animals were treated 5 minutes after the onset of injury, MK-801 was effective in reducing ischemic damage in both models. However, when treatment was delayed 10 minutes after the ischemic insult in the MCEM, no neuroprotection was observed even when the MK-801 dose was increased eightfold. We also did not find a beneficial effect of MK-801 pretreatment with a dose that was one tenth of the effective dose in the RSCIM. Studies using the (-) MK-801 isomer showed that MK-801 neuroprotection exhibited stereoselectivity. The contribution of anticonvulsant activity and sedation to MK-801's neuroprotective actions was examined indirectly using phenytoin and midazolam, respectively. Neither drug was effective in reducing ischemic injury in the MCEM. This suggests that MK-801's neuroprotective efficacy in ischemia is mediated through its NMDA receptor antagonist activity independent of its anticonvulsant or sedative properties. These results support the hypothesis that excessive NMDA receptor excitation may be involved in ischemic neuronal damage.

Topics: Animals; Blood Pressure; Body Temperature; Brain Ischemia; Dizocilpine Maleate; Heart Rate; Ischemia; Male; Rabbits; Receptors, N-Methyl-D-Aspartate; Regional Blood Flow; Spinal Cord

This investigation tested in rats whether MK-801, an N-methyl-D-aspartate receptor antagonist, would improve the balance of oxygen supply and consumption in the focal ischemic area of the brain induced by occlusion of the middle cerebral artery. Fifteen minutes after middle cerebral artery occlusion, 5 mg/kg MK-801 was administered intravenously to the MK-801 group (n = 12), and normal saline was given to the control group (n = 12). One hour after the occlusion in each group, regional cerebral blood flow was determined in six rats using [14C]iodoantipyrine, and regional arterial and venous oxygen saturations were determined using a microspectrophotometric technique in the other six rats. In both groups of animals, the cerebral blood flow of the ischemic cortex was significantly lower than that of the contralateral cortex (36 +/- 16 [SD] and 67 +/- 14 ml/min/100 g for the control group; 33 +/- 10 and 58 +/- 11 ml/min/100 g for the MK-801 group, respectively). Oxygen extraction was significantly higher in the ischemic cortex (8.8 +/- 2.1 ml O2/100 ml blood) than in the contralateral cortex (5.6 +/- 0.3) for the control group. However, for the MK-801 group, there was no significant difference between the ischemic cortex (6.1 +/- 1.0) and the contralateral cortex (5.7 +/- 1.1). Oxygen extraction in the ischemic cortex of the MK-801 group was significantly lower than that of the control group. Calculated ischemic regional oxygen consumption was similar to the nonischemic values in the control group, whereas the ischemic value was reduced to 61% of the value of the contralateral cortex in the MK-801 group.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Brain; Brain Ischemia; Cerebrovascular Circulation; Dizocilpine Maleate; Male; Oxygen; Oxygen Consumption; Rats; Spectrophotometry

The effect of NB-818, a new dihydropyridine calcium entry blocker, on delayed neuronal death (DND) in the hippocampal CA1 subfield of gerbils after 5 minutes of forebrain ischemia induced by bilateral carotid artery occlusion was examined. Gerbils were treated intraperitoneally with NB-818 (0.1-3 mg/kg) just after release of the occlusion. Four days after the ischemia, they were fixed by perfusing 10% buffered-formalin, and the neuronal cell density (NCD, cell/mm) in the CA1 subfield was estimated under microscopy. The average NCD in the ischemic control group was 43 +/- 10.8 cells/mm, whereas NB-818 (3 mg/kg) significantly ameliorated DND with an average NCD of 143 +/- 24.2 cells/mm (P less than 0.01). In addition, NB-818 (3 mg/kg) significantly inhibited DND at 1, 2 and 4 weeks after transient ischemia: the average NCD of the NB-818 and ischemic control groups were 80 +/- 9.4 (P less than 0.01) and 43 +/- 7.7 cells/mm, 92 +/- 13.7 (P less than 0.05) and 52 +/- 9.3 cells/mm, and 57 +/- 5.0 (P less than 0.01) and 43 +/- 12.4 cells/mm, respectively. In this experiment, NB-818 exhibited a protective effect on DND in the hippocampal CA1 subfield after transient forebrain ischemia, and its effect persisted for up to 4 weeks. These findings suggest that NB-818 may be useful for clinical treatment of neurological deficit after an ischemic insult.

Topics: Animals; Brain Ischemia; Calcium Channel Blockers; Cell Survival; Dihydropyridines; Dizocilpine Maleate; Gerbillinae; Hippocampus; Histocytochemistry; Injections, Intraperitoneal; Male; Neurons

Both oxygen free radicals and excitatory amino acids have been implicated as important cellular toxins in ischemic brain. Recent in vitro studies suggest that there may be a mutual interaction between these two mediators. We explored the relation between oxygen free radicals and excitatory amino acids in the development of ischemic brain edema in vivo. Male Sprague-Dawley rats were treated with the free radical scavenger dimethylthiourea 1 hour before ischemia or with the excitotoxin antagonist MK-801 30 minutes before ischemia produced by occlusion of the middle cerebral artery. Groups of seven or eight animals were treated with vehicle, low-dose (375 mg/kg) dimethylthiourea, high-dose (750 mg/kg) dimethylthiourea, low-dose (0.5 mg/kg) MK-801, high-dose (2.0 mg/kg) MK-801, or both high-dose dimethylthiourea and low-dose MK-801. After 4 hours of ischemia, brain water content was determined. In eight vehicle-treated controls, mean +/- SEM water content of tissue in the center of the ischemic zone was 83.29 +/- 0.18%. A significant reduction of brain edema was observed in all drug-treated groups: for example, 50.2% (p less than 0.001) in the high-dose dimethylthiourea group, 53.7% (p less than 0.001) in the low-dose MK-801 group, and 66.4% (p less than 0.001) in the combined dimethylthiourea and MK-801 group. Combined treatment with dimethylthiourea and MK-801 provided no significant additive effect over that resulting from treatment with MK-801 alone.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Amino Acids; Animals; Brain; Brain Edema; Brain Ischemia; Dizocilpine Maleate; Drug Interactions; Free Radicals; Male; Neurotoxins; Oxygen; Potassium; Rats; Rats, Inbred Strains; Sodium; Thiourea

1. An acute model of focal ischaemia, which involves permanent occlusion of the middle cerebral artery of the rat with 4 h survival, was used to find the minimum effective plasma concentration of dizocilpine (MK-801) and to determine its dose-effect relationship. 2. MK-801 was administered at the time of occlusion and was given as an i.v. bolus followed by an infusion for 4 h to maintain a steady state plasma concentration of the drug throughout the study. MK-801 was given at 3 dose levels; 0.04 mg kg-1 i.v. bolus + 0.6 micrograms kg-1 min-1 infusion; 0.12 mg kg-1 i.v. bolus + 1.8 micrograms kg-1 min-1 infusion; 0.4 mg kg-1 i.v. bolus + 6 micrograms kg-1 min-1 infusion, which gave mean plasma levels over the 4 h of 8.0 ng ml-1, 18.9 ng ml-1 and 113.2 ng ml-1 respectively. 3. MK-801 at 8.0 ng ml-1 gave 10% reduction in the volume of ischaemic brain damage in the cerebral cortex which just reached significance. The middle dose of MK-801 (18.9 ng ml-1) gave a highly significant reduction in the volume of ischaemic brain damage in the cerebral cortex and hemisphere, volumes of ischaemic tissue being reduced by 60% and 50% compared to saline-treated animals, respectively. The highest plasma concentration of MK-801 (113.2 ng ml-1) resulted in a 35% reduction in the volume of hemispheric damage and a 40% reduction in the volume of cortical damage, which were significant.4. The reduction in the amount of protection afforded by the highest dose of MK-801 may be due to the hypotensive effect of this dose. There was no protection against the volume of damage in the caudate nucleus for any of the doses of MK-801 tested.5. Therefore the minimum effective plasma concentration of MK-801 was 8.0 ngml1, although the greatest protection was seen with a plasma level of 18.9 ng ml- 1. This correlates well with the concentration of MK-801 required to block N-methyl-D-aspartate (NMDA) receptors and prevent NMDA receptor mediated neurotoxicity in vitro.

Topics: Animals; Brain Ischemia; Cerebral Arterial Diseases; Dizocilpine Maleate; Dose-Response Relationship, Drug; Male; Rats; Rats, Inbred Strains

The excitatory amino acid inhibitor MK-801 has been shown in many animals species to protect against hypoxic-ischemic brain injury. We sought to determine whether hypoxic-ischemic injury to the newborn pig's brain could be prevented by the use of MK-801.. Hypoxic-ischemic injury to the brain was induced in forty 0-3-day-old piglets. They were randomized to receive either 3 mg/kg MK-801 (MK-801 group, n = 20) or vehicle (control group, n = 19) prior to insult. At time 0, the carotid arteries were ligated and the blood pressure was reduced by one third by hemorrhage. At 15 minutes, inspired oxygen was reduced from 50% to 6%. At 30 minutes, inspired oxygen was changed to 100%, carotid ligatures were released, and the withdrawn blood was reinfused. An additional 14 piglets received 3 mg/kg MK-801 but not hypoxic-ischemic injury (drug-only group), and a final group of 11 piglets were subjected to only a sham operation (sham group).. Neurological examination scores at 24, 48, and 72 hours showed that MK-801 and drug-only piglets were significantly worse than the controls. Pathological examination of the brains at 72 hours showed significantly greater damage in the brains of the MK-801 and control pigs relative to the sham and drug-only groups. No differences were found between the control and the MK-801 groups. No differences were found between the sham and drug-only groups.. MK-801, at a dose of 3 mg/kg, causes neurological dysfunction in piglets lasting at least 72 hours, but neither causes brain damage nor ameliorates the effects of hypoxic-ischemic injury to the brain of the newborn pig.

Topics: Animals; Animals, Newborn; Blood Pressure; Brain Ischemia; Carotid Arteries; Disease Models, Animal; Dizocilpine Maleate; Hypoxia, Brain; Ligation; Oxygen; Swine

Calcium ion can enter ischemic neurons through both receptor-operated and voltage-sensitive Ca2+ channels. To attenuate this Ca2+ entry and Ca2(+)-induced neuronal injury, we tried a combined treatment with the noncompetitive N-methyl-D-aspartate (NMDA) antagonist, MK-801, and the dihydropyridine calcium antagonist, nimodipine, in a cat middle cerebral artery occlusion (1 hour) and reperfusion (3 hours) model. We measured changes in cytosolic free calcium, nicotinamide adenine dinucleotide/reduced nicotinamide adenine dinucleotide redox state, and blood flow in the cat cortex using a newly developed fluorometric technique with indo-1, a fluorescent intracellular Ca2+ indicator. The combined treatment, starting 5 minutes into ischemia, was effective in reducing both Ca2+ entry and histologic damage and in enhancing recovery of the electroencephalogram following reperfusion. MK-801 alone was also effective, but to a lesser extent. These data suggest that the dual blockade of Ca2+ entry using MK-801 and nimodipine may be a useful tool for protection against ischemic brain damage.

Topics: Animals; Brain; Brain Ischemia; Calcium; Cats; Cerebrovascular Circulation; Cytosol; Dizocilpine Maleate; Drug Therapy, Combination; Electroencephalography; Male; NAD; Nimodipine; Oxidation-Reduction; Reperfusion

Topics: Brain Ischemia; Dizocilpine Maleate; Drug Therapy, Combination; Humans; Nimodipine

The phencyclidine analogue [3H](1-[2-thienyl]cyclohexyl)piperidine (3H-TCP) binds to the ion channel associated with the N-methyl-D-aspartate receptor channel complex. In vitro autoradiography indicates that the distribution of 3H-TCP binding in brain closely parallels that of [3H]glutamate binding to the N-methyl-D-aspartate receptor. In nine 7-day-old rats, an acute focal hypoxic-ischemic insult produced by unilateral carotid artery ligation and subsequent exposure to 8% oxygen acutely reduced 3H-TCP binding ipsilateral to the ligation by 30% in the CA1, by 27% in the CA3, by 26% in the dentate gyrus, and by 17% in the striatum compared with values from the contralateral hemisphere. In 10 littermates that received 1 mg/kg of the neuroprotective noncompetitive N-methyl-D-aspartate antagonist MK-801 immediately before hypoxic exposure, the regional distribution of 3H-TCP binding in hypoxic-ischemic brain was relatively preserved and there were no interhemispheric asymmetries in 3H-TCP binding densities. In addition, in three unoperated rats decapitated 24 hours after MK-801 treatment, 3H-TCP binding was reduced by 15-35%; similar bilateral suppression of 3H-TCP binding was detected in MK-801-treated ligates. Our data indicate that 3H-TCP autoradiography can be used to assay the efficacy of neuroprotective agents in this experimental model of perinatal stroke.

Topics: Animals; Animals, Newborn; Aspartic Acid; Autoradiography; Binding Sites; Brain; Brain Ischemia; Dibenzocycloheptenes; Dizocilpine Maleate; Glutamates; Glutamic Acid; Hypoxia; N-Methylaspartate; Phencyclidine; Rats; Rats, Inbred Strains; Tritium

Ischemic damage to the brain, whether induced experimentally or observed clinically, often produces a pattern of delayed selective cell death in subfield CA1 of hippocampus which has been associated with significant neurologic deficits. The present study demonstrates that this selective vulnerability of CA1 neurons to ischemia, with relative preservation of their neighbors, is expressed in organotypic tissue culture and is prevented by the N-methyl-D-aspartate (NMDA) receptor blocker, MK-801. These data provide conclusive evidence that this selective cell death does not have a vascular etiology but is mediated by factors intrinsic to the hippocampal neurons and/or local circuitry. This model system provides an opportunity both to examine mechanisms of ischemic cell death in an avascular environment and to study methods of prevention in the absence of systemic variables.

Topics: Brain Ischemia; Cell Survival; Culture Techniques; Dizocilpine Maleate; Glutamates; Glutamic Acid; Hippocampus; Osmolar Concentration; Time Factors

We studied the effects of iontophoretically administered MK-801 (50-150 nA) on ischemic changes on the CA1 hippocampal field potential. Twenty rats under urethane anesthesia, of which the hippocampal field response was depressed or lost upon ligation of the carotid arteries, were used. MK-801 applications starting before carotid ligation, decreased the depression of the field response in 8 of 11 trials. MK-801 was applied after the appearance of ischemic changes and partly restored the deteriorated hippocampal field potential in 16 of 34 penetrations. MK-801 was ineffective in preventing or restoring the severely depressed or lost evoked activity. During ischemia a DC potential shift of -32.6 +/- 3.7 mV (n = 10) was recorded. MK-801 reduced the amplitude of the DC potential shift by 50% when applied before (n = 6) or after (n = 4) the initiation of ischemia. Activation of N-methyl-D-aspartate (NMDA) receptors by glutamate or N-methyl-DL-aspartate (NMDLA) induces a slow negative wave on the field response. During ischemia a similar negative wave spontaneously appeared in 9 trials and was also induced with low currents (5-10 nA) of NMDLA which were insufficient to evoke the NMDA-mediated wave before ischemia. These data provide electrophysiological evidence that NMDA receptors are activated during ischemia and MK-801 reduces ischemia neuronal dysfunction.

Topics: Animals; Brain Ischemia; Dizocilpine Maleate; Electric Stimulation; Evoked Potentials; Glutamates; Glutamic Acid; Male; Neurons; Rats; Receptors, N-Methyl-D-Aspartate

The effects of glutamate and the excitatory amino acid antagonist, MK-801, were investigated on the time course of the shift of water from extracellular to intracellular space (progression or cytotoxic oedema) after total brain ischaemia in rats. Administration of sodium glutamate intravenously before ischaemia accelerated the shift of water dose-dependently. On the contrary, preischaemic administration of MK-801, an NMDA antagonist, delayed the progression of cytotoxic oedema due to brain ischaemia. We consider that glutamate and NMDA antagonists may have important roles in the development and prevention of cytotoxic oedema in the ischaemic state.

Topics: Animals; Body Water; Brain; Brain Ischemia; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Extracellular Space; Glutamates; Glutamic Acid; Intracellular Fluid; Magnetic Resonance Spectroscopy; Male; Rats; Rats, Inbred Strains

We studied the time course and molecular mechanisms of changes in brain polyamines and their rate-regulatory synthetic enzyme ornithine decarboxylase during reversible forebrain ischemia and recirculation in the gerbil. Bilateral carotid occlusion induced an acute (less than 2 minutes), transient increase in ornithine decarboxylase activity and putrescine level. After 15 minutes of ischemia, recirculation evoked an immediate (less than 1 minute) increase in ornithine decarboxylase activity and putrescine concentration that progressed over a 15-minute period. A small rise in spermidine and spermine also was observed. A secondary increase in ornithine decarboxylase activity and the levels of putrescine and spermidine commenced after 6 hours of recirculation. Pretreatment with a-difluoromethylornithine, a specific suicide inhibitor of ornithine decarboxylase, or MK-801, a noncompetitive N-methyl-D-aspartate receptor antagonist, abolished all early and delayed increases in ornithine decarboxylase activity and polyamine levels. This is the first demonstration that both ischemia and postischemic recirculation evoke rapid, transient increases in the activity of ornithine decarboxylase and the levels of polyamines, most notably the ornithine decarboxylase product, putrescine. Our results indicate that N-methyl-D-aspartate receptor activation (by an ischemically induced elevation of extracellular glutamate) is responsible for initiating the early and the delayed stimulation of ornithine decarboxylase activity. Ornithine decarboxylase activation causes the rapid rise in the levels of putrescine and higher polyamines observed in the acute response to ischemia and the acute and delayed response to postischemic recirculation. These polyamine changes may be involved in the pathophysiology of Ca2+ entry and neuronal death after brain ischemia.

Topics: Animals; Brain; Brain Ischemia; Cerebrovascular Circulation; Dizocilpine Maleate; Female; Gerbillinae; Ornithine Decarboxylase; Polyamines; Putrescine; Receptors, N-Methyl-D-Aspartate

Excessive Ca2+ influx through NMDA receptor-coupled channels has been linked to neuronal cell death. Using an in vitro model of transient brain ischemia, we investigated possible protective effects of NMDA receptor antagonists ketamine or MK-801 and of calmidazolium, an inhibitor of intracellular Ca2(+)-activated proteins. Brain ischemia/recovery was simulated in isolated hippocampal slices and injury monitored by measurement of ATP levels. Omission of both glucose and oxygen (but not oxygen alone) for 20 min led to persistent ATP deficits after 4 h recovery. Addition of ketamine or MK-801 at 1 microM permitted ATP to recover within 1 h, as did addition of calmidazolium at 10 microM. Our findings are consistent with other reports that NMDA receptor antagonists can protect neuronal tissue from ischemic damage. The role of inappropriately activated Ca2(+)-mediated signaling processes in the mechanism(s) of such injury is suggested by the protection also seen with calmidazolium, an inhibitor of calmodulin and other structurally related proteins such as calpain(s) and protein kinase C. The inhibition of intracellular Ca2+ target proteins may be an alternative for protection of the brain against injury due to insults that activate NMDA receptors.

Topics: Adenosine Triphosphate; Animals; Brain Ischemia; Dizocilpine Maleate; Hippocampus; Imidazoles; In Vitro Techniques; Ketamine; Rats; Receptors, N-Methyl-D-Aspartate; Time Factors

The blockade of postsynaptic receptors for excitatory amino acids is a promising new field for the possible treatment of cerebral ischaemia. The most important receptor seems to be the N-methyl-D-aspartate (NMDA) subtype of the glutamate receptors and MK-801 is a potent non-competitive antagonist to the NMDA receptor. 31P NMR Spectroscopy was used to measure the recovery of intracellular pH and the high energy phosphates Phosphocreatine (PCr) and ATP after ten minutes of temporary global cerebral ischaemia in the rat. Cerebral ischaemia was obtained by combining bilateral carotid ligation and systemic hypotension (2 vessel occlusion model). Two intervention groups with intravenous injection of MK-801 in doses of 0.25 mg/kg and 0.5 mg/kg 15 minutes before onset of ischaemia were compared to a control group. Both intervention groups showed a more rapid recovery of PCr and ATP than the control group, but there were no significant differences for intracellular pH.

Topics: Adenosine Triphosphate; Animals; Brain; Brain Ischemia; Dizocilpine Maleate; Energy Metabolism; Magnetic Resonance Spectroscopy; Male; Phosphocreatine; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate

The dose-dependent effects of MK-801, a glutamate receptor antagonist, on changes in CBF, CBF-PaCO2 responsiveness (133Xe clearance), EEG, and blood-brain barrier (methylene blue) were examined after a 15-min period of reversible complete global ischemia induced in halothane-anesthetized cats by occlusion of the vertebral and carotid arteries. Pretreatment with doses of MK-801 of greater than or equal to 0.5 mg/kg had no effect on resting CBF measures and produced a dose-dependent slowing of the dominant EEG frequency. In animals receiving this agent, there was an almost immediate return of baseline EEG patterns upon reinstitution of flow, no hypoperfusion after 2 h of reflow, preservation of CBF and CBF-PaCO2 responsiveness, and maintenance of blood-brain barrier integrity. In contrast, parallel control animals and animals treated with MK-801 at a dose of 0.1 mg/kg exhibited poor recovery based on the above parameters. MK-801 also diminished in a dose-dependent manner the CSF levels of 6-keto-prostaglandin (PG) F1 alpha (stable metabolite of PGI2) and thromboxane (Tx) B2 (stable metabolite of TxA2), which were otherwise elevated in vehicle-treated animals 2 h after reflow. Of particular interest, the CSF TxB2/6-keto-PGF1 alpha ratio in vehicle-treated animals was near 2. In animals pretreated with MK-801, at doses of greater than or equal to 0.5 mg/kg, this ratio was nearly 1. These observations are consistent with a possible triggering role of glutamate release in initiating at least part of the acute sequelae of ischemia. Such release in an electrically silent cell would increase Ca2+ influx and activate free fatty acid metabolism, leading to probable changes in vascular function and changes in blood-brain barrier permeability.

Topics: Animals; Arteries; Blood Pressure; Blood-Brain Barrier; Brain; Brain Ischemia; Carbon Dioxide; Cats; Cerebrovascular Circulation; Dibenzocycloheptenes; Dizocilpine Maleate; Electroencephalography; Female; Hydrogen-Ion Concentration; Male; Partial Pressure; Prostaglandins; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter

We have administered antagonists acting competitively or noncompetitively at the N-methyl-D-aspartate receptor after a short period of incomplete ischaemia and evaluated selective neuronal loss in the CA1 region of the rat hippocampus. The competitive antagonists D-(-)-2-amino-7-phosphonoheptanoate (2APH); 100 or 330 mg/kg; 3-(2-carboxypiperazin-4-yl)propyl-1-phosphonate (CPP); 3.3 or 10 mg/kg; and CGS 19755 (cis-4-phosphonomethyl-2-piperidine carboxylate) 3.3 or 10 mg/kg; and the noncompetitive antagonists MK801 [+)5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate), 0.3, 1, or 3 mg/kg, and dextrorphan, 2, 6, 18, or 54 mg/kg, were administered intraperitoneally 15 min and 5 h after a 10-min incomplete ischaemia period; additionally MK801 (1 or 3 mg/kg) and CGS 19755 (10 or 30 mg/kg) were administered 5 and 10 h postischaemia. Seven days after ischaemia, the brains were fixed by perfusion. CA1 pyramidal cell counts were performed on Nissl-stained sections using an ocular grid piece. Ventilated (no ischaemia) control animals had a mean of 406 +/- 13 CA1 neurones/3 grid lengths. Ischaemia reduced this mean to 157 +/- 23. A significant protective effect against this cell loss was seen after two injections (at 15 min and 5 h postischaemia) of 2APH, CPP (10 mg/kg), CGS 19755 (10 mg/kg), MK801 (1 mg/kg), and dextrophan (54 mg/kg). Delayed injection (5 and 10 h postischaemia) of CGS 19755 (10 and 30 mg/kg) and MK801 (1 and 3 mg/kg) did not provide any protection against pyramidal cell loss.

Topics: 2-Amino-5-phosphonovalerate; Amino Acids; Animals; Anticonvulsants; Aspartic Acid; Brain; Brain Ischemia; Dibenzocycloheptenes; Dizocilpine Maleate; Male; N-Methylaspartate; Neurons; Pipecolic Acids; Piperazines; Piperidines; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter

Changes in intra- and extracellular free calcium concentration were evaluated with ion-selective microelectrodes during periods of anoxia and ischemia in three different regions of intact rat brain. Recordings stable for at least 2 min and in most cases for 4-6 min were chosen for analysis. Under normoxic conditions neuronal [Ca2+]i varied between less than 10(-8) and 10(-7) M from cell to cell but no systematic regional differences were observed. Elimination of O2 or interruption in blood flow caused, within 30-60 s, slight intracellular alkalinization followed by a small rise in [Ca2+]i, a mild degree of hyperpolarization, and disappearance of electrical activity in the cortex, in that order. It is postulated that a decline in cellular energy levels, as manifested by H+ uptake associated with creatine phosphate hydrolysis, leads to an increase in [Ca2+]i, which activates Ca2(+)-dependent K+ channels and consequently enhances gK. 2-4 min later there was a sudden, large rise in [K+]e, a fall in [Ca2+]e and a rapid elevation of [Ca2+]i. The magnitude of the latter was greatest in a high proportion of hippocampal neurons in area CA1 and some cortical cells, while it was smallest and relatively delayed in thalamic neurons. In the hippocampus area CA1 increases in [Ca2+]i to as much as 6-8 x 10(-4) were observed; some of these could be reversed when O2 or blood flow were restored to normal. Pretreatment of animals with ketamine and MK-801, antagonists of excitatory amino acid transmitters, markedly slowed and decreased the rises in [Ca2+]i. The effects of the two agents were most pronounced in the hippocampus. It is concluded that the receptor-operated channels are largely responsible for Ca2+ entry into certain cells during hypoxia/ischemia. This pathway may be of primary importance in parts of the hippocampus and cortex, regions of the brain that are particularly vulnerable to O2 deprivation and which receive high glutamatergic input and have an abundance of excitatory amino acid receptors.

Topics: Animals; Brain Ischemia; Calcium; Calcium Channels; Cerebral Cortex; Dibenzocycloheptenes; Dizocilpine Maleate; Female; Hippocampus; Ketamine; Male; Membrane Potentials; Microelectrodes; Nifedipine; Oxygen Consumption; Rats; Rats, Inbred Strains; Thalamus

Global cerebral ischemia is well known to cause neuronal necrosis in selectively vulnerable sectors of the hippocampus. Since the hippocampus of the rat is involved in spatial navigation, learning, and memory, selective deficits in these abilities may arise from ischemic brain damage. Previous studies have shown (a) a detectable neurobehavioural deficit due to ischemic brain damage limited to half of the CA1 sector of the hippocampus and (b) a reduction of ischemic neuronal necrosis with the noncompetitive N-methyl-D-Aspartate (NMDA) antagonist MK-801. This study was designed to determine the relationship between the improvement in structural brain damage in postischemically treated rats and any improvement in neurobehavioural performance, using a learning-set water task. Seventeen male Wistar rats received 10.5 min of forebrain ischemia induced by carotid clamping and hypotension. Brain temperature was estimated with probes in the temporalis muscle. Ten of these animals received no therapy (controls), and seven animals received 5 mg/kg MK-801 iv, 20 min postischemia. Six additional rats underwent a sham operation. Postischemic hypothermia was prevented with heating lamps. Four controls and one MK-801 treated animal died. The survivors were then tested on a place learning-set task in a swimming pool paradigm, and quantitative histopathologic analysis of their entire brains was done. The learning-set task revealed defects in spatial navigation, reflected as increased errors and latency in the performance of the untreated control rats. The performance of the MK-801 treated group progressively approached that of sham-operated rats over the course of testing and was significantly better than controls. Importantly, no long-term detrimental effect of MK-801 on the learning-set task performance was seen. Quantitative neuropathology revealed significantly less damage in the MK-801 treated group in all major brain regions. In the hippocampus, MK-801 treated animals showed hippocampal damage limited to the vulnerable portion of the pyramidal cell band comprising 48.8% of the CA1 pyramidal cells, as opposed to 72.4% in untreated controls. Extra-hippocampal damage was evident only in untreated control animals. MK-801 totally prevented neuronal necrosis in both the cerebral cortex and striatum and also prevented infarction in the neocortex and thalamus. Three conclusions emerge from the study. First, postischemic MK-801 mitigates structural brain damage in several bra

Topics: Animals; Anticonvulsants; Brain Ischemia; Dibenzocycloheptenes; Discrimination Learning; Dizocilpine Maleate; Hippocampus; Male; Memory; Mental Recall; Neuronal Plasticity; Orientation; Rats; Rats, Inbred Strains

The effects of the N-methyl-D-aspartate (NMDA) antagonist MK-801 on capillary beds and CBF following 1 h of transient incomplete focal cerebral ischemia were studied by examining 133Xe CBF, capillary diameter, and area of perfused vasculature. Capillary diameter increased from a control of 5.24 +/- 0.37 to 8.62 +/- 0.57 microns (p less than 0.001) and area of perfused vasculature from 20,943 +/- 1,151 to 30,442 +/- 1,691 microns2/x 10 magnification field (p less than 0.001) with MK-801 1.0 mg/kg administered 30 min prior to ischemia. After flow restoration in control animals, there was a relative hypoperfusion with eventual normalization of CBF over 60 min. Alternatively, in MK-801 1.0 mg/kg animals, there was rapid normalization of CBF upon flow restoration without the postischemic hypoperfusion observed in controls. On histological analysis, there was consistently less neuronal edema in MK-801-treated animals. These results support the hypothesis that hypoperfusion following incomplete focal cerebral ischemia may be due in part to NMDA-mediated cellular edema with subsequent extravascular capillary bed compression.

Topics: Animals; Brain Ischemia; Capillaries; Cerebrovascular Circulation; Dizocilpine Maleate; Rabbits

The purpose of the present study was to test whether the anticonvulsant, memantine (1-amino-3,5-dimethyladamantane), can protect neurons against hypoxic or ischemic damage. To this end, we used a rat model of transient forebrain ischemia and cultured neurons from chick embryo cerebral hemispheres. Ischemia was induced for 10 min by clamping both carotid arteries and lowering the mean arterial blood pressure to 40 mm Hg; the rats were allowed to recover for 7 days. Cultured neurons were made hypoxic with 1 mmol/l NaCN added to the incubation medium for 30 min followed by a recovery period of 3 days. The possible effects of memantine were compared with those produced by a typical non-competitive NMDA antagonist, dizocilpine. Similar effects were obtained with both drugs. The drugs reduced the damage caused by transient ischemia to neurons of the hippocampal CA1 subfield. Memantine (10 and 20 mg/kg) had a dose-dependent effect when administered intraperitoneally to the rats 1 h before ischemia. Dizocilpine was active in this model at a dosage of 1 mg/kg. When administered after ischemia, 10 mg/kg memantine significantly protected CA1 neurons against ischemic damage. Furthermore, the drugs protected cultured neurons against hypoxic damage. The lowest effective concentration was 0.1 mumol/l for dizocilpine and 1 mumol/l for memantine. Thus, memantine possesses neuroprotective activity but is less potent than dizocilpine.

Topics: Adenosine Triphosphate; Animals; Brain Ischemia; Cell Hypoxia; Cells, Cultured; Chick Embryo; Dizocilpine Maleate; Memantine; Nerve Tissue Proteins; Nervous System Diseases; Neurons; Potassium Cyanide; Rats; Rats, Inbred Strains

Recent in vitro and in vivo experiments have suggested that excitatory amino acid antagonists, particularly those active at the N-methyl-D-aspartate receptor subtype, are effective in ameliorating ischemic injury due to their antiexcitotoxic activity. However, these drugs are also potent and effective in vivo anticonvulsants. The present experiments compared the noncompetitive N-methyl-D-aspartate antagonists phencyclidine and MK-801 with the anticonvulsant phenytoin in a model of focal brain ischemia. Fisher F-344 rats were subjected to tandem occlusion of the middle cerebral and ipsilateral common carotid arteries under halothane anesthesia. Compounds were administered intravenously 30 minutes and 24 hours after arterial occlusion; infarct size was assessed at 48 hours after occlusion. Phencyclidine had no effect on infarct volume at 1 mg/kg, significantly reduced (by 36%) infarct volume at 3 mg/kg, and produced a nonsignificant 26% decrease at 10 mg/kg. The more potent and selective noncompetitive antagonist MK-801 reduced (by 32%) infarct volume significantly at 0.1 mg/kg, produced a nonsignificant 23% decrease at 0.3 mg/kg, and had no effect at 0.5 mg/kg. Phenytoin, which is not a glutamate antagonist, reduced the infarct volume by 45% at 28 mg/kg. A single dose of phenytoin (28 mg/kg) administered 30 minutes after occlusion was neuroprotective, but delaying drug administration for more than 2 hours was ineffective. These data suggest that blockade of the N-methyl-D-aspartate receptor is effective in reducing the infarct size after focal cerebral ischemia. The neuroprotective activity of phenytoin suggests that this may be related to the common anticonvulsant action.

Topics: Animals; Brain Ischemia; Disease Models, Animal; Dizocilpine Maleate; Male; N-Methylaspartate; Phencyclidine; Phenytoin; Rats; Rats, Inbred F344

It is well known that levels of free fatty acids (FFA) in the brain rapidly increase during ischemia. FFA release during ischemia is generally due to the disturbance of reacylation by ATP depletion and deacylation from membrane phospholipids by the action of phospholipase. The present study examined the regional difference in brain FFA levels and also the action of phospholipase from the effect of NMDA antagonist (MK-801) and phospholipase C inhibitor (PMSF) on FFA release during complete ischemia in rat brain. Complete brain ischemia was induced with cardiac arrest by intracardiac injection of KCI. A focused microwave was irradiated to the head of rats 0, 2, 4 and 8 minutes after cardiac arrest. Samples of the neocortex, striatum, hippocampus and thalamus were dissected. FFA were measured in each sample. In the vulnerable regions such as neocortex, hippocampus and striatum, arachidonic acid and other FFA levels rapidly increased from the onset of ischemia. All FFA levels in the thalamus were significantly lower than those in the other regions during ischemia. The regional difference of FFA levels during ischemia seemed to be responsible for the regional difference of the vulnerability to ischemia. MK-801 inhibited the FFA release mainly from phosphatidylcholine and phosphatidylethanolamine between 2 and 4 minutes of ischemia. On the other hand, PMSF inhibited the FFA release mainly from phosphatidylinositol during the first 2 minutes of ischemia and from phosphatidylcholine and phosphatidylethanolamine until 8 minutes of ischemia.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Brain; Brain Ischemia; Dizocilpine Maleate; Fatty Acids, Nonesterified; Male; Phenylmethylsulfonyl Fluoride; Phospholipases; Rats; Rats, Inbred Strains; Type C Phospholipases

8319, ((+-)-2-Amino-N-ethyl-alpha-(3-methyl-2-thienyl)benzeneethanamine 2HCl), is a novel compound with the profile of a non-competitive NMDA antagonist. The compound displaced [3H] TCP with high affinity (IC50 = 43 nM), but was inactive at the NMDA, benzodiazepine and GABA sites; in vivo, 8319 showed good efficacy as an anticonvulsant and potential neuroprotective agent. It blocked seizures induced by NMDLA, supramaximal electroshock, pentylenetetrazol (PTZ), picrotoxin, and thiosemicarbazide with ED50's of 1-20 mg/kg ip. As a neuroprotective agent, 8319 (30-100 mg/kg sc) prevented the death of dorsal hippocampal pyramidal cells induced by direct injection of 20 nmol NMDA. At 15 mg/kg ip, the compound was also effective against hippocampal neuronal necrosis induced via bilateral occlusion of the carotid arteries in gerbils. In summary, 8319 is a noncompetitive NMDA antagonist with good anticonvulsant activity and may possess neuroprotective properties useful in the treatment of brain ischemia.

Topics: Aniline Compounds; Animals; Anticonvulsants; Binding, Competitive; Brain Diseases; Brain Ischemia; Dizocilpine Maleate; Gerbillinae; Hippocampus; Male; Mice; Mice, Inbred Strains; N-Methylaspartate; Rats; Rats, Inbred Strains; Thiophenes; Tritium

Electrophysiological recordings were made from hippocampal slices of gerbils which had undergone a 5 min period of bilateral carotid occlusion. Normal CA1 population spikes, field potentials and synaptic responses were obtained from slices of gerbils pretreated with the N-methyl-D-aspartate antagonist, MK-801 (10 mg/kg, i.p., 1 h before the occlusion). In contrast, no CA1 population spike could be obtained from slices of untreated gerbils, although field potentials and population spikes were obtained from the dentate granule cell region of these slices in response to perforant path stimulation. These studies indicate that the CA1 pyramidal cells identified histologically as protected by MK-801 from selective neuronal degeneration produced by a 5 min period of global ischaemia, are also functionally protected and retain their normal synaptic responsiveness.

Topics: Animals; Brain Ischemia; Dizocilpine Maleate; Electric Stimulation; Evoked Potentials; Female; Gerbillinae; Hippocampus; In Vitro Techniques; Male; Pyramidal Tracts; Staining and Labeling; Synapses

In cerebral asphyxia, enhanced postsynaptic stimulation of N-methyl-D-aspartate (NMDA) receptor by excessive glutamate may mediate neuronal injury and death. The neuroprotective potential of the novel, potent NMDA receptor antagonist MK-801 was assessed by evaluating hippocampal behavioral and histologic outcomes in an experimental rat model of neonatal hypoxia/ischemia. Seven-day-old rats with and without MK-801 pretreatment were subjected to unilateral carotid ligation followed by 2 hours of hypoxia. At age 30 days, spontaneous alternation behavior was measured using a conventional wooden T maze. Hypoxic-ischemic animals pretreated with saline demonstrated a significant impairment in spontaneous alternation behavior compared with that of normal control rats and the hypoxic-ischemic rats pretreated with MK-801. Hippocampal neuronal damage in the CA1 and CA3 regions was prevented in animals pretreated with MK-801 vs saline-treated controls. Therefore, while saline-treated rats with hippocampal lesions showed defective memory and hippocampal neuronal destruction, pretreatment with MK-801 protected rats. Thus, MK-801 appears to protect hippocampal neurons from hypoxia/ischemia and may be potentially beneficial in preventing neonatal asphyxial brain damage.

Topics: Animals; Brain Ischemia; Dibenzocycloheptenes; Dizocilpine Maleate; Female; Hippocampus; Hypoxia, Brain; Male; Neurons; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter

The functional integrity of the N-methyl-D-aspartate receptor complex following focal cerebral ischemia in the rat has been examined at a time when brain tissue is irreversibly damaged. Twelve hours after unilateral permanent middle cerebral artery occlusion, [3H]-MK-801 binding was not significantly altered in the ischemic cerebral cortex compared to sham-operated animals. Moreover, the enhancement of [3H]MK-801 binding by exogenous glutamate was preserved in an area of the brain that was permanently damaged by the ischemic insult.

Topics: Animals; Arterial Occlusive Diseases; Brain; Brain Ischemia; Cerebral Arteries; Dibenzocycloheptenes; Dizocilpine Maleate; Glutamates; Glutamic Acid; Male; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter

We evaluated the neuroprotective effect of MK-801, a noncompetitive, selective N-methyl-D-aspartate receptor antagonist, in a neonatal hypoxic-ischemic animal model. Seven-day-old rats underwent bilateral ligation of the carotid arteries followed by exposure to an 8% oxygen atmosphere for 1 hr. We sacrificed the animals 72 hrs later and assessed the hypoxic-ischemic brain damage histologically. MK-801 (10 mg/kg), administered IP 0.5 hr before the hypoxia, completely prevented hypoxic-ischemic infarction in cerebral cortex, while treatment immediately and 1 hr after the end of the hypoxia resulted in 76% and 52% reduction in the infarcted area, respectively. MK-801, given 0.5 hr before and immediately after the insult, reduced striatal damage and, given 0.5 hr before, attenuated neuronal necrosis in hippocampal regions. These results show that in neonates MK-801 is neuroprotective even when administered up to 1 hr after the end of a hypoxic-ischemic insult.

Topics: Animals; Animals, Newborn; Brain; Brain Damage, Chronic; Brain Ischemia; Dibenzocycloheptenes; Dizocilpine Maleate; Hypoxia; Rats; Rats, Inbred Strains

This study was designed to determine if dizocilipine maleate (MK-801), administered following 11 min of complete ischemia in dogs, could favorably alter neurologic outcome and hippocampal damage. Eighteen dogs were anesthetized and subjected to complete cerebral ischemia by temporary occlusion of the ascending aorta and the venae cavae via a thoracotomy. Five min postischemia, 9 dogs were given dizocilipine 150 micrograms/kg, followed by an infusion of 1.25 microgram/kg/min for 8 h. Control dogs were given equal volumes of placebo. Dogs were evaluated neurologically at 24, 48, and 72 h; thereafter, the brains were perfused, fixed and harvested. There was no significant difference in outcome between dizocilipine- and placebo-treated dogs: 5 of 9 given dizocilipine were normal, 1 was mildly injured and 3 were severely injured or dead. In the control animals given placebo, 3 of 9 were normal, 2 were mildly injured and 4 were moderately to severely injured. Histopathologic examination was limited to the hippocampus. CA1 and CA2,3,4 pyramidal neurons were graded according to degree of injury on a 5-point scale. There were no differences in histopathologic grades between the two groups. However, in both groups combined there was a significant correlation between neurologic outcome grade and histopathologic grade. The only notable systemic effect of dizocilipine appeared to be prolonged sedation which extended beyond 24 h postischemia but was not evident at 48 h postischemia. The authors conclude that more outcome studies in more sensitive models are needed.

Topics: Animals; Brain Ischemia; Dibenzocycloheptenes; Disease Models, Animal; Dizocilpine Maleate; Dogs; Female; Hippocampus; Male

Recent evidence implicates the endogenous excitatory amino acids, glutamate (Glu) and aspartate, in hypoxic/ischemic neuronal degeneration. In a preceding article (Ikonomidou et al., 1989) we described a new model for studying hypoxic/ischemic neuronal degeneration in the infant rat brain that entails unilateral common carotid artery ligation followed by exposure to a partial vacuum for 75 min. Promising features of this model include a low mortality rate and high incidence of acute brain damage disseminated over numerous brain regions. In addition, there is a striking similarity between the type of cytopathology characterizing this model of hypoxic/ischemic neuronal degeneration and that which has been described in infant animals treated with Glu. MK-801 is a powerful antagonist of the N-methyl-D-aspartate (NMDA) receptor ionophore complex (a subtype of Glu receptor). In the present study, after unilateral carotid artery ligation was performed on 10-d-old rat pups, they were treated either with MK-801 (1 mg/kg i.p.) or saline 15 min before exposure to the hypobaric condition. MK-801 exerted a strong neuroprotective effect without serious side effects; the majority of saline control animals sustained severe brain damage, whereas the majority of MK-801-treated pups had no brain damage. These and other recent findings suggest that the NMDA receptor may play an important role in hypoxic/ischemic neuronal degeneration in the immature brain and provide hope that NMDA antagonists such as MK-801 may be effective in preventing such degeneration.

Topics: Animals; Animals, Newborn; Atmospheric Pressure; Body Temperature; Brain; Brain Ischemia; Dibenzocycloheptenes; Dizocilpine Maleate; Female; Male; Nerve Degeneration; Neurons; Rats

The purpose of this study was to determine the effectiveness of the non-competitive N-methyl-D-aspartate receptor antagonist dizocilpine, or (+)-5-methyl-10,11-dihydro-5H-dibenzo(a,d)cyclohepten-5,10-imine (MK-801) in mitigating ischemic neuronal necrosis in the rat. Ten minutes of transient forebrain ischemia was induced by a combination of bilateral carotid clamping and hypotension to 50 mm Hg. Control animals received intravenous saline, whereas treated animals received dizocilpine, either 1 mg/kg iv 20 min. pre ischemia, 1 mg/kg iv 20 min. post ischemia, 10 mg/kg iv 20 min. post ischemia, 10 mg/kg ip 2 hrs. post ischemia, 10 mg/kg ip 24 hrs. post ischemia. The groups receiving dizocilpine before or up to 20 min. after ischemia all showed a significant reduction in the number of dead neurons as assessed by quantitative histopathology in hippocampus, caudate nucleus and cerebral cortex after one week of recovery. However, dizocilpine administered either 2 or 24 hrs. after ischemia afforded no protection. These results suggest that the potent non-competitive NMDA antagonist dizocilpine may have some value in protecting the brain from hippocampal and cortical neuronal necrosis after a short insult consisting of dense transient cerebral ischemia. Noteworthy is the fact that pharmacologic intervention in the post-ischemic period was successful in preventing neuronal death, provided that drug administration occurred within dizocilpine's "therapeutic window".

Topics: Animals; Brain; Brain Ischemia; Dibenzocycloheptenes; Dizocilpine Maleate; Drug Evaluation; Male; Necrosis; Rats; Rats, Inbred Strains; Time Factors

Cultured neurons of chick cerebral embryo hemispheres were used to study drug effects against neuronal damage caused by hypoxia during long-term recovery. Sodium cyanide (NaCN, 1 mmol/l) induces hypoxia-like conditions by inhibiting oxydative phosphorylation. The sensitivity of the cultured neurons against this type of hypoxia was determined after 3, 4, 5 and 6 days of cultivation followed by 4, 3, 2 days and 1 day of recovery, respectively. The ATP level and the viability of cells as well as the total cell number and the protein content of the cultures were used to characterize the extent of posthypoxic neuronal damage. A hypoxic period of 30 min after 4 days of cultivation followed by 3 days of recovery seemed to be appropriate for determining protective drug effects. The drug effects obtained were comparable to those from in vivo models of cerebral ischemia or hypoxia. The results suggest that cultured neurons exposed to hypoxia and to long-term recovery could be suitable for studying post-hypoxic neuronal damage as well as neuroprotective drug effects.

Topics: Adenosine Triphosphate; Animals; Anticonvulsants; Brain Ischemia; Cell Count; Cell Survival; Cells, Cultured; Chick Embryo; Clonazepam; Diazepam; Dibenzocycloheptenes; Dizocilpine Maleate; Flunarizine; Haloperidol; Imipramine; Ketamine; Lidocaine; Nafronyl; Neurons; Pyrithioxin; Telencephalon

The effects of MK-801 upon local CBF after permanent middle cerebral artery (MCA) occlusion have been examined using [14C]iodoantipyrine autoradiography in halothane-anaesthetised rats. MK-801 (0.5 mg kg-1 i.v.) or saline was administered 30 min before MCA occlusion and CBF measured approximately 40 min after occlusion. In the hemisphere contralateral to the occluded MCA, MK-801 significantly reduced local CBF in 19 of the 22 regions examined from the levels in saline-treated rats. In the contralateral hemisphere, after treatment with MK-801, blood flow was reduced by an average of 37% with little variation in the magnitude of the reductions in different regions. In the hemisphere ipsilateral to MCA occlusion, MK-801 reduced CBF in almost every region located outside the territory of the occluded MCA. Within the territory of the occluded MCA, blood flow in the MK-801-treated rat did not significantly differ from values in vehicle-treated rats in any of the five cortical areas examined, although in the caudate nucleus there was a tendency for CBF to be lower in rats pretreated with MK-801. MK-801 had no effect on the amount of hypoperfused cerebral tissue (CBF less than 30 ml 100 g-1 min-1) in the ipsilateral hemisphere at any coronal plane examined; e.g., at coronal plane anterior 7.2 mm, 51 +/- 5% of the hemisphere displayed CBF of less than 30 ml 100 g-1 min-1 in saline-treated rats with MCA occlusion compared with 52 +/- 8% of the hemisphere in rats treated with MK-801 prior to MCA occlusion.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Aspartic Acid; Blood Pressure; Brain Ischemia; Cerebrovascular Circulation; Dibenzocycloheptenes; Dizocilpine Maleate; Male; N-Methylaspartate; Rats; Rats, Inbred Strains

The effects of the glutamate N-methyl-D-aspartate receptor antagonist MK-801 in reducing ischaemic brain damage have been examined in anaesthetised cats, with drug treatment being initiated 2 h after the induction of cerebral ischaemia. Focal cerebral ischaemia was produced by permanent occlusion of one middle cerebral artery, and the animals were killed 6 h later. The amount of early irreversible ischaemic damage was assessed at 16 predetermined stereotactic planes. Treatment with MK-801 (5 mg/kg, i.v.) 2 h after middle cerebral artery occlusion reduced significantly the volume of ischaemic damage (from 1,625 +/- 384 mm3 of the cerebral hemisphere in vehicle-treated cats to 792 +/- 385 mm3 in MK-801-treated cats). The demonstration of reduced ischaemic brain damage with MK-801, when the agent is administered after the induction of ischaemia, extends the therapeutic potential of such agents in the treatment of focal cerebral ischaemia in humans.

Topics: Animals; Anticonvulsants; Aspartic Acid; Brain Ischemia; Cats; Dibenzocycloheptenes; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; N-Methylaspartate

1. An in vitro receptor autoradiography procedure is described for visualizing binding sites for the excitatory amino acid antagonist radiolabelled MK-801, in rat and gerbil brain sections. 2. Ten micron sections were labelled by incubation at room temperature for 20 min in 30 nM [3H]-MK-801. This was followed by 2 rinses for 20 s in fresh buffer solution. Specifically bound ligand determined with 100 microM unlabelled MK-801 amounted to 55-60% of total. 3. Phencyclidine, (+/-)-SKF 10047, ketamine and 2-aminophosphonovaleric acid (APV) (all 100 microM) prevented the specific binding of [3H]-MK-801. L-Glutamate and N-methyl D-aspartate (NMDA) (100 microM) had no effect. However, L-glutamate prevented the inhibition by APV. 4. The highest concentrations of [3H]-MK-801 binding sites occurred in the hippocampal formation, cerebral cortex, olfactory bulb and thalamus. Very low levels were detected in the brain stem and cerebellum. 5. The distribution of [3H]-MK-801 binding sites was comparable to that of NMDA sites and phencyclidine sites (labelled with [3H]-TCP) but not with high-affinity sigma sites labelled with [3H]-3-PPP. 6. The density of [3H]-MK-801 binding sites in the gerbil hippocampus was examined 1, 2, 6 and 22 days after unilateral carotid artery occlusion for 10 min. Only at 6 and 22 days was the binding reduced (by 36% and 46% respectively) in the CA1 region whereas a significant neuronal loss was apparent at day 2. In CA2 a decrease in binding was only evident at day 22. 7. These results indicate that binding sites for [3H]-MK-801 can be detected in mammalian brain sections by receptor autoradiography. Their distribution supports an association with the NMDA receptor complex and the loss in the hippocampus after carotid artery occlusion indicates their presence on pyramidal cells is vulnerable to ischaemic insult.

Topics: Animals; Aspartic Acid; Autoradiography; Binding Sites; Brain; Brain Ischemia; Densitometry; Dibenzocycloheptenes; Dizocilpine Maleate; Gerbillinae; Glutamates; In Vitro Techniques; Muscle, Smooth, Vascular; N-Methylaspartate; Rats; Rats, Inbred Strains; Synaptic Membranes

Topics: Animals; Animals, Newborn; Anticonvulsants; Brain Ischemia; Dibenzocycloheptenes; Dizocilpine Maleate; Hypoxia, Brain; Organ Size; Rats; Receptors, Neurotransmitter; Receptors, Phencyclidine

The neuroprotective effects of MK-801, a noncompetitive antagonist of N-methyl-D-aspartate (NMDA) receptors, were evaluated in models of cerebral ischemia using Mongolian gerbils. Bilateral occlusion of the carotid arteries for a period of 5 min resulted in a consistent pattern of degeneration of hippocampal CA1 and CA2 pyramidal neurons, which was quantified using an image analyzer. Systemic administration of MK-801 (0.01-10 mg/kg, i.p.) 1 hr prior to the occlusion caused a dose-dependent protection of the CA1 and CA2 neurons. The ED50 value for neuroprotection by MK-801 was calculated to be 0.3 mg/kg, and at doses greater than or equal to 3 mg/kg the majority of animals were completely protected against the ischemic insult. Systemic administration of MK-801 (1 or 10 mg/kg, i.p.) 1 hr prior to unilateral occlusion of the right carotid artery resulted in significant protection against hippocampal neurodegeneration following 10 min of occlusion, and increased the survival rate after 30 min of occlusion. The potent neuroprotective effects of MK-801 in these cerebral ischemia models add further weight to the evidence that NMDA receptors are involved in the mechanism of ischemia-induced neuronal degeneration.

Topics: Animals; Arterial Occlusive Diseases; Brain Ischemia; Carotid Artery Diseases; Dibenzocycloheptenes; Dizocilpine Maleate; Female; Gerbillinae; Hippocampus; Male; Nerve Degeneration