dizocilpine-maleate and Nerve-Degeneration

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

Convincing evidence has accumulated that indicates neuroplastic changes within the spinal cord in response to repeated exposure to opioids. Such neuroplastic changes occur at both cellular and intracellular levels. It has been generally acknowledged that the activation of N-methyl-D-aspartate (NMDA) receptors plays a pivotal role in the development of neuroplastic changes following repeated opioid exposure. Intracellular cascades can also be activated subsequent to NMDA receptor activation. In particular, protein kinase C has been shown to be a key intracellular element that contributes to the behavioral manifestation of neuroplastic changes. Moreover, interactions between NMDA and opioid receptors can lead to potentially irreversible degenerative neuronal changes in the spinal cord in association with the development of opioid tolerance. Interestingly, similar cellular and intracellular changes occur in the spinal cord following peripheral nerve injury. These findings indicate that interactions exist in the spinal cord neural structures between two seemingly unrelated conditions-chronic opioid exposure and a pathological pain state. These observations may help understand mechanisms of chemical intolerance and multiple chemical sensitivity as well as have significant clinical implications in pain management with opioid analgesics.

Topics: Analgesics; Analgesics, Opioid; Animals; Benzamides; Dizocilpine Maleate; Drug Tolerance; Hot Temperature; Humans; Hyperalgesia; Morphine; Multiple Chemical Sensitivity; N-Methylaspartate; Narcotics; Nerve Degeneration; Nerve Tissue Proteins; Neuralgia; Neuronal Plasticity; Nitric Oxide; Phosphorylation; Poly(ADP-ribose) Polymerases; Posterior Horn Cells; Protein Kinase C; Protein Processing, Post-Translational; Rats; Receptors, N-Methyl-D-Aspartate; Receptors, Opioid; Receptors, Opioid, mu; Sciatic Nerve; Spinal Cord

Excitotoxic Ca

Topics: Animals; Brain; Calcium Channels; Dizocilpine Maleate; Mice; Mice, Knockout; Mitochondria; Mitochondrial Proteins; Nerve Degeneration; Neurons; Neuroprotective Agents; Zinc

Thiamine/vitamin B1 deficiency can lead to behavioral changes and neurotoxicity in humans. This may due in part to vascular damage, neuroinflammation and neuronal degeneration in the diencephalon, which is seen in animal models of pyrithiamine-enhanced thiamine deficiency. However, the time course of the progression of these changes in the animal models has been poorly characterized. Therefore, in this study, the progression of: 1) activated microglial association with vasculature; 2) neurodegeneration; and 3) any vascular leakage in the forebrain during the progress of thiamine deficiency were determined. A thiamine deficient diet along with 0.25 mg/kg/d of pyrithiamine was used as the mouse model. Vasculature was identified with Cd31 and microglia with Cd11b and Iba1 immunoreactivity. Neurodegeneration was determined by FJc labeling. The first sign of activated microglia within the thalamic nuclei were detected after 8 d of thiamine deficiency, and by 9 d activated microglia associated primarily with vasculature were clearly present but only in thalamus. At the 8 d time point neurodegeneration was not present in thalamus. However at 9 d, the first signs of neurodegeneration (FJc + neurons) were seen in most animals. Over 80% of the microglia were activated at 10 d but almost exclusively in the thalamus and the number of degenerating neurons was less than 10% of the activated microglia. At 10 d, there were sporadic minor changes in IgG presence in thalamus indicating minor vascular leakage. Dizocilpine (0.2-0.4 mg/kg) or phenobarbital (10-20 mg/kg) was administered to groups of mice from day 8 through day 10 to block neurodegeneration but neither did. In summary, activated microglia start to surround vasculature 1-2 d prior to the start of neurodegeneration. This response may be a means of controlling or repairing vascular damage and leakage. Glutamate excitotoxicity and vascular leakage likely only play a minor role in the early neurodegeneration resulting from thiamine deficiency. However, failure of dysfunctional vasculature endothelium to supply sufficient nutrients to neurons could be contributing to the neurodegeneration.

Topics: Animals; Blood Vessels; Calcium-Binding Proteins; CD11b Antigen; Diet; Dizocilpine Maleate; Female; Mice; Microfilament Proteins; Microglia; Nerve Degeneration; Phenobarbital; Pyrithiamine; Thalamus; Thiamine Deficiency; Time Factors

Inhibitory GABAergic and glycinergic neurotransmission in the spinal cord play a central role in the regulation of neuronal excitability, by maintaining a balance with the glutamate-mediated excitatory transmission. Glutamatergic agonists infusion in the spinal cord induce motor neuron death by excitotoxicity, leading to motor deficits and paralysis, but little is known on the effect of the blockade of inhibitory transmission. In this work we studied the effects of GABAergic and glycinergic blockade, by means of microdialysis perfusion (acute administration) and osmotic minipumps infusion (chronic administration) of GABA and glycine receptors antagonists directly in the lumbar spinal cord. We show that acute glycinergic blockade with strychnine or GABAergic blockade with bicuculline had no significant effects on motor activity and on motor neuron survival. However, chronic bicuculline infusion, but not strychnine, induced ipsilateral gait alterations, phalange flaccidity and significant motor neuron loss, and these effects were prevented by AMPA receptor blockade with CNQX but not by NMDA receptor blockade with MK801. In addition, we demonstrate that the chronic infusion of bicuculline enhanced the excitotoxic effect of AMPA, causing faster bilateral paralysis and increasing motor neuron loss. These findings indicate a relevant role of GABAergic inhibitory circuits in the regulation of motor neuron excitability and suggest that their alterations may be involved in the neurodegeneration processes characteristic of motor neuron diseases such as amyotrophic lateral sclerosis.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Atrophy; Bicuculline; Dizocilpine Maleate; Drug Interactions; GABA Antagonists; Gait; Male; Motor Activity; Motor Neurons; Muscle Hypotonia; Nerve Degeneration; Rats; Receptors, Glycine; Spinal Cord; Strychnine

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

MK-801 is a use-dependent NMDA receptor open channel blocker with a very slow off-rate. These properties can be exploited to 'pre-block' a population of NMDARs, such as synaptic ones, enabling the selective activation of a different population, such as extrasynaptic NMDARs. However, the usefulness of this approach is dependent on the stability of MK-801 blockade after washout. We have revisited this issue, and confirm that recovery of NMDAR currents from MK-801 blockade is enhanced by channel opening by NMDA, and find that it is further increased when Mg(2+) is also present. In the presence of Mg(2+), 50% recovery from MK-801 blockade is achieved after 10' of 100 μM NMDA, or 30' of 15 μM NMDA exposure. In Mg(2+)-free medium, NMDA-induced MK-801 dissociation was found to be much slower. Memantine, another PCP-site antagonist, could substitute for Mg(2+) in accelerating the unblock of MK-801 in the presence of NMDA. This suggests a model whereby, upon dissociation from its binding site in the pore, MK-801 is able to re-bind in a process antagonized by Mg(2+) or another PCP-site antagonist. Finally we show that even when all NMDARs are pre-blocked by MK-801, incubation of neurons with 100 μM NMDA in the presence of Mg(2+) for 2.5 h triggers sufficient unblocking to kill >80% of neurons. We conclude that while synaptic MK-801 'pre-block' protocols are useful for pharmacologically assessing synaptic vs. extrasynaptic contributions to NMDAR currents, or studying short-term effects, it is problematic to use this technique to attempt to study the effects of long-term selective extrasynaptic NMDAR activation. This article is part of the Special Issue entitled 'Glutamate Receptor-Dependent Synaptic Plasticity'.

Topics: Animals; Cations; Cells, Cultured; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Interactions; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Magnesium; Memantine; Membrane Potentials; N-Methylaspartate; Nerve Degeneration; Neurons; Rats; Receptors, N-Methyl-D-Aspartate; Time Factors

NMDA receptor (NMDAR) antagonists like ketamine and MK-801 possess remarkable antidepressant effects with fast onset. However, they over-stimulate the retrosplenial cortex, evoking psychosis-like effects and neuronal injury, revealed by de novo induction of the heat shock protein 70 (Hsp70). Moreover, early in the development MK-801 triggers widespread cortical apoptosis, inducing extensive caspase-3 expression. Altogether these data raise strong concerns on the clinical applicability of NMDAR antagonist therapies. Therefore, the development of novel therapeutics targeting more specifically NMDAR to avoid psychotomimetic effects is necessary. Here we investigated a GluN2B (NR2B) antagonist in behavioral and neurotoxicity paradigms in rats to assess its potential as possible alternative to unspecific NMDA receptor antagonists. We found that treatment with the GluN2B specific antagonist Ro 25-6981 evoked robust antidepressant-like effects. Moreover, Ro 25-6981 did not cause hyperactivity as displayed after treatment with unspecific NMDAR antagonists, a correlate of psychosis-like effects in rodents. Additionally, Ro 25-6981, unlike MK-801, did not induce caspase-3 and HSP70 expression, markers of neurotoxicity in the perinatal and adult brain, respectively. Moreover, unexpectedly, in the adult retrosplenial cortex Ro 25-6981 pretreatment significantly reduced MK-801-triggered neurotoxicity. Our results suggest that GluN2B antagonists may represent valuable alternatives to unspecific NMDAR antagonists with robust antidepressant efficacy and a more favorable side-effect profile.

Topics: Age Factors; Animals; Antidepressive Agents; Caspase 3; Dizocilpine Maleate; Drug Interactions; Female; Gyrus Cinguli; Hallucinogens; HSP70 Heat-Shock Proteins; Male; Mice; Motor Activity; Nerve Degeneration; Phenols; Piperidines; Rats; Receptors, N-Methyl-D-Aspartate

NMDA glutamate receptors (NMDARs) have critical functional roles in the nervous system but NMDAR over-activity can contribute to neuronal damage. The open channel NMDAR blocker, memantine is used to treat certain neurodegenerative diseases, including Parkinson's disease (PD) and is well tolerated clinically. We have investigated memantine block of NMDARs in substantia nigra pars compacta (SNc) dopamine neurones, which show severe pathology in PD. Memantine (10 μM) caused robust inhibition of whole-cell (synaptic and extrasynaptic) NMDARs activated by NMDA at a high concentration or a long duration, low concentration. Less memantine block of NMDAR-EPSCs was seen in response to low frequency synaptic stimulation, while responses to high frequency synaptic stimulation were robustly inhibited by memantine; thus memantine inhibition of NMDAR-EPSCs showed frequency-dependence. By contrast, MK-801 (10 μM) inhibition of NMDAR-EPSCs was not significantly different at low versus high frequencies of synaptic stimulation. Using immunohistochemistry, confocal imaging and stereological analysis, NMDA was found to reduce the density of cells expressing tyrosine hydroxylase, a marker of viable dopamine neurones; memantine prevented the NMDA-evoked decrease. In conclusion, memantine blocked NMDAR populations in different subcellular locations in SNc dopamine neurones but the degree of block depended on the intensity of agonist presentation at the NMDAR. This profile may contribute to the beneficial effects of memantine in PD, as glutamatergic activity is reported to increase, and memantine could preferentially reduce over-activity while leaving some physiological signalling intact.

Topics: Animals; Cell Count; Dizocilpine Maleate; Dopaminergic Neurons; Dose-Response Relationship, Drug; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Male; Memantine; N-Methylaspartate; Nerve Degeneration; Rats; Receptors, N-Methyl-D-Aspartate; Substantia Nigra; Tyrosine 3-Monooxygenase

Mercury is a well-known neurotoxin implicated in a wide range of neurological or psychiatric disorders including autism spectrum disorders, Alzheimer's disease, Parkinson's disease, epilepsy, depression, mood disorders and tremor. Mercury-induced neuronal degeneration is thought to invoke glutamate-mediated excitotoxicity, however, the underlying mechanisms remain poorly understood. Here, we examine the effects of various mercury concentrations (including pathological levels present in human plasma or cerebrospinal fluid) on cultured, rat cortical neurons.. We found that inorganic mercuric chloride (HgCl₂--at 0.025 to 25 μM) not only caused neuronal degeneration but also perturbed neuronal excitability. Whole-cell patch-clamp recordings of pyramidal neurons revealed that HgCl₂ not only enhanced the amplitude and frequency of synaptic, inward currents, but also increased spontaneous synaptic potentials followed by sustained membrane depolarization. HgCl₂ also triggered sustained, 2-5 fold rises in intracellular calcium concentration ([Ca²⁺]i). The observed increases in neuronal activity and [Ca²⁺]i were substantially reduced by the application of MK 801, a non-competitive antagonist of N-Methyl-D-Aspartate (NMDA) receptors. Importantly, our study further shows that a pre incubation or co-application of MK 801 prevents HgCl₂-induced reduction of cell viability and a disruption of β-tubulin.. Collectively, our data show that HgCl₂-induced toxic effects on central neurons are triggered by an over-activation of NMDA receptors, leading to cytoskeleton instability.

Topics: Animals; Calcium; Cell Survival; Cells, Cultured; Cerebral Cortex; Cytoskeleton; Dizocilpine Maleate; Humans; Intracellular Space; Membrane Potentials; Mercuric Chloride; Nerve Degeneration; Nerve Net; Neurites; Neurons; Pyramidal Cells; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Synapses

Developing central white matter is subject to ischemic-type injury during the period that precedes myelination. At this stage in maturation, central axons initiate a program of radial expansion and ion channel redistribution. Here we test the hypothesis that during radial expansion axons display heightened ischemic sensitivity, when clusters of Ca(2+) channels decorate future node of Ranvier sites.. Functionality and morphology of central axons and glia were examined during and after a period of modeled ischemia. Pathological changes in axons undergoing radial expansion were probed using electrophysiological, quantitative ultrastructural, and morphometric analysis in neonatal rodent optic nerve and periventricular white matter axons studied under modeled ischemia in vitro or after hypoxia-ischemia in vivo.. Acute ischemic injury of central axons undergoing initial radial expansion was mediated by Ca(2+) influx through Ca(2+) channels expressed in axolemma clusters. This form of injury operated only in this axon population, which was more sensitive to injury than neighboring myelinated axons, smaller axons yet to initiate radial expansion, astrocytes, or oligodendroglia. A pharmacological strategy designed to protect both small and large diameter premyelinated axons proved 100% protective against acute ischemia studied under modeled ischemia in vitro or after hypoxia-ischemia in vivo.. Recent clinical data highlight the importance of axon pathology in developing white matter injury. The elevated susceptibility of early maturing axons to ischemic injury described here may significantly contribute to selective white matter pathology and places these axons alongside preoligodendrocytes as a potential primary target of both injury and therapeutics.

Topics: Age Factors; Animals; Animals, Newborn; Apoptosis; Astrocytes; Axons; Disease Models, Animal; Dizocilpine Maleate; Glucose; Green Fluorescent Proteins; Hypoxia; Hypoxia-Ischemia, Brain; Mice; Mice, Transgenic; Myelin Sheath; Nerve Degeneration; Nerve Fibers, Myelinated; Neuroprotective Agents; Oligodendroglia; omega-Agatoxin IVA; Optic Nerve; Organ Culture Techniques; Rats; Rats, Sprague-Dawley; Recovery of Function; Thy-1 Antigens

Cannabinoid CB1 receptors (CB1Rs) regulate the neurodegenerative damage of experimental autoimmune encephalomyelitis (EAE) and of multiple sclerosis (MS). The mechanism by which CB1R stimulation exerts protective effects is still unclear. Here we show that pharmacological activation of CB1Rs dampens the tumor necrosis factor α (TNFα)-mediated potentiation of striatal spontaneous glutamate-mediated excitatory postsynaptic currents (EPSCs), which is believed to cogently contribute to the inflammation-induced neurodegenerative damage observed in EAE mice. Furthermore, mice lacking CB1Rs showed a more severe clinical course and, in parallel, exacerbated alterations of sEPSC duration after induction of EAE, indicating that endogenous cannabinoids activate CB1Rs and mitigate the synaptotoxic action of TNFα in EAE. Consistently, we found that mice lacking the fatty acid amide hydrolase (FAAH), and thus expressing abnormally high brain levels of the endocannabinoid anandamide, developed a less severe EAE associated with preserved TNFα-induced sEPSC alterations. CB1Rs are important modulators of EAE pathophysiology, and might play a mechanistic role in the neurodegenerative damage of MS patients.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Amidohydrolases; Animals; Arachidonic Acids; Cannabinoid Receptor Modulators; Corpus Striatum; Dizocilpine Maleate; Dronabinol; Encephalomyelitis, Autoimmune, Experimental; Endocannabinoids; Etanercept; Excitatory Postsynaptic Potentials; Female; Glutamic Acid; Immunoglobulin G; Mice; Mice, Inbred C57BL; Mice, Knockout; Nerve Degeneration; Neurons; Polyunsaturated Alkamides; Receptor, Cannabinoid, CB1; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Receptors, Tumor Necrosis Factor; Tumor Necrosis Factor-alpha

In addition to their original applications to lowering cholesterol, statins display multiple neuroprotective effects. N-methyl-D-aspartate (NMDA) receptors interact closely with the dopaminergic system and are strongly implicated in therapeutic paradigms of Parkinson's disease (PD). This study aims to investigate how simvastatin impacts on experimental parkinsonian models via regulating NMDA receptors.. Regional changes in NMDA receptors in the rat brain and anxiolytic-like activity were examined after unilateral medial forebrain bundle lesion by 6-hydroxydopamine via a 3-week administration of simvastatin. NMDA receptor alterations in the post-mortem rat brain were detected by [³H]MK-801(Dizocilpine) binding autoradiography. 6-hydroxydopamine treated PC12 was applied to investigate the neuroprotection of simvastatin, the association with NMDA receptors, and the anti-inflammation. 6-hydroxydopamine induced anxiety and the downregulation of NMDA receptors in the hippocampus, CA1(Cornu Ammonis 1 Area), amygdala and caudate putamen was observed in 6-OHDA(6-hydroxydopamine) lesioned rats whereas simvastatin significantly ameliorated the anxiety-like activity and restored the expression of NMDA receptors in examined brain regions. Significant positive correlations were identified between anxiolytic-like activity and the restoration of expression of NMDA receptors in the hippocampus, amygdala and CA1 following simvastatin administration. Simvastatin exerted neuroprotection in 6-hydroxydopamine-lesioned rat brain and 6-hydroxydopamine treated PC12, partially by regulating NMDA receptors, MMP9 (matrix metalloproteinase-9), and TNF-a (tumour necrosis factor-alpha).. Our results provide strong evidence that NMDA receptor modulation after simvastatin treatment could partially explain its anxiolytic-like activity and anti-inflammatory mechanisms in experimental parkinsonian models. These findings contribute to a better understanding of the critical roles of simvastatin in treating PD via NMDA receptors.

Topics: Animals; Anti-Inflammatory Agents; Anxiety; Apoptosis; Autoradiography; Cell Survival; Disease Models, Animal; Dizocilpine Maleate; Dopamine; Glutamic Acid; Immunohistochemistry; L-Lactate Dehydrogenase; Male; Matrix Metalloproteinase 9; Nerve Degeneration; Oxidopamine; Parkinson Disease; PC12 Cells; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Simvastatin; Substantia Nigra; Tritium; Tumor Necrosis Factor-alpha; Tyrosine 3-Monooxygenase

Excess glutamate release and stimulation of post-synaptic glutamatergic receptors have been implicated in the pathophysiology of many neurological diseases. The hippocampus, and the pyramidal cell layer of the cornu ammonus 1 (CA1) region in particular, has been noted for its selective sensitivity to excitotoxic insults. The current studies examined the role of N-methyl-D-aspartate (NMDA) receptor subunit composition and sensitivity to stimulatory effects of the polyamine spermidine, an allosteric modulator of NMDA NR2 subunit activity, in hippocampal CA1 region sensitivity to excitotoxic insult. Organotypic hippocampal slice cultures of 8 day-old neonatal rat were obtained and maintained in vitro for 5 days. At this time, immunohistochemical analysis of mature neuron density (NeuN); microtubule associated protein-2(a,b) density (MAP-2); and NMDA receptor NR1 and NR2B subunit density in the primary cell layers of the dentate gyrus (DG), CA3, and CA1 regions, was conducted. Further, autoradiographic analysis of NMDA receptor distribution and density (i.e. [(125)I]MK-801 binding) and spermidine (100 microM)-potentiated [(125)I]MK-801 binding in the primary cell layers of these regions was examined. A final series of studies examined effects of prolonged exposure to NMDA (0.1-10 microM) on neurodegeneration in the primary cell layers of the DG, CA3, and CA1 regions, in the absence and presence of spermidine (100 microM) or ifenprodil (100 microM), an allosteric inhibitor of NR2B polypeptide subunit activity. The pyramidal cell layer of the CA1 region demonstrated significantly greater density of mature neurons, MAP-2, NR1 and NR2B subunits, and [(125)I]MK-801 binding than the CA3 region or DG. Twenty-four hour NMDA (10 microM) exposure produced marked neurodegeneration (approximately 350% of control cultures) in the CA1 pyramidal cell region that was significantly reduced by co-exposure to ifenprodil or DL-2-Amino-5-phosphonopentanoic acid (APV). The addition of spermidine significantly potentiated [(125)I]MK-801 binding and neurodegeneration induced by exposure to a non-toxic concentration of NMDA, exclusively in the CA1 region. This neurodegeneration was markedly reduced with co-exposure to ifenprodil. These data suggest that selective sensitivity of the CA1 region to excitotoxic stimuli may be attributable to the density of mature neurons expressing polyamine-sensitive NR2B polypeptide subunits.

Topics: Animals; CA1 Region, Hippocampal; CA3 Region, Hippocampal; Dentate Gyrus; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Hippocampus; In Vitro Techniques; Male; N-Methylaspartate; Nerve Degeneration; Neuroprotective Agents; Neurotoxins; Piperidines; Pyramidal Cells; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Spermidine; Valine

Brain injury during the last trimester to the first 1-4 years in humans is now thought to trigger an array of intellectual and emotional problems later in life, including disorders such as schizophrenia. In adult schizophrenic brains, there is a specific loss of neurons that co-express glutamic acid decarboxylase-parvalbumin (GAD67-PV). Loss of this phenotype is thought to occur in mature animals previously exposed to N-methyl-D: -aspartate receptor (NMDAR) antagonists during late gestation or at postnatal day 7 (P7). However, in similarly treated animals, we have previously shown that GAD67 and PV are unaltered in the first 24 h. To more precisely define when changes in these markers first occur, we exposed rat pups (P7 or P6-P10) to the NMDAR antagonist MK801 and at P11 co-stained brain sections for GAD67 or PV. In the cingulate cortex, we found evidence for a reduction in PV (GAD67 levels were very low to undetectable). In contrast, in the somatosensory cortex, we found that expression of GAD67 was reduced, but PV remained stable. Further, repeated but not single doses of MK801 were necessary to see such changes. Thus, depending on the region, NMDAR antagonism appears to influence expression of PV or GAD67, but not both. These observations could not have been predicted by previous studies and raise important questions as to how the GAD67-PV phenotype is lost once animals reach maturity. More importantly, such differential effects may be of great clinical importance, given that cognitive deficits are seen in children exposed to anesthetics that act by blocking the NMDAR.

Topics: Aging; Animals; Animals, Newborn; Cell Count; Cell Differentiation; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; gamma-Aminobutyric Acid; Glutamate Decarboxylase; Gyrus Cinguli; Immunohistochemistry; Interneurons; Nerve Degeneration; Parvalbumins; Phenotype; Rats; Receptors, N-Methyl-D-Aspartate; Schizophrenia; Somatosensory Cortex

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

Prenatal supplementation of rat dams with dietary choline has been shown to provide their offspring with neuroprotection against N-methyl-d-aspartate (NMDA) antagonist-mediated neurotoxicity. This study investigated whether postnatal dietary choline supplementation exposure for 30 and 60 days of rats starting in a pre-puberty age would also induce neuroprotection (without prenatal exposure). Male and female Sprague-Dawley rats (postnatal day 30 of age) were reared for 30 or 60 concurrent days on one of the four dietary levels of choline: 1) fully deficient choline, 2) 1/3 the normal level, 3) the normal level, or 4) seven times the normal level. After diet treatment, the rats received one injection of MK-801 (dizocilpine 3mg/kg) or saline control. Seventy-two hours later, the rats were anesthetized and transcardially perfused. Their brains were then postfixed for histology with Fluorojade-C (FJ-C) staining. Serial coronal sections were prepared from a rostrocaudal direction from 1.80 to 4.2mm posterior to the bregma to examine cell degeneration in the retrosplenial and piriform regions. MK-801, but not control saline, produced significant numbers of FJ-C positive neurons, indicating considerable neuronal degeneration. Dietary choline supplementation or deprivation in young animals reared for 30-60days did not alter NMDA antagonist-induced neurodegeneration in the retrosplenial region. An interesting finding is the absence of the piriform cortex involvement in young male rats and the complete absence of neurotoxicity in both hippocampus regions and DG. However, neurotoxicity in the piriform cortex of immature females treated for 60days appeared to be suppressed by low levels of dietary choline.

Topics: Animals; Animals, Newborn; Brain; Choline; Cytoprotection; Dietary Supplements; Disease Models, Animal; Dizocilpine Maleate; Female; Male; Nerve Degeneration; Neuroprotective Agents; Neurotoxins; Rats; Rats, Sprague-Dawley

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

There is accumulating evidence that excitotoxicity and oxidative stress resulting from excessive activation of glutamate (N-methyl-D-aspartate) NMDA receptors are major participants in striatal degeneration associated with 3-nitropropionic acid (3NP) administration. Although excitotoxic and oxidative mechanisms are implicated in 3NP toxicity, there are conflicting reports as to whether NMDA receptor antagonists attenuate or exacerbate the 3NP-induced neurodegeneration. In the present study, we investigated the involvement of NMDA receptors in striatal degeneration, protein oxidation and motor impairment following systemic 3NP administration. We examined whether NMDA receptor antagonists, memantine and ifenprodil, influence the neurotoxicity of 3NP. The development of striatal lesion and protein oxidation following 3NP administration is delayed by memantine but not affected by ifenprodil. However, in behavioral experiments, memantine failed to improve and ifenprodil exacerbated the motor deficits associated with 3NP toxicity. Together, these findings suggest caution in the application of NMDA receptor antagonists as a neuroprotective agent in neurodegenerative disorders associated with metabolic impairment.

Topics: Adenosine Diphosphate; Animals; Corpus Striatum; Dizocilpine Maleate; Drug Interactions; Male; Memantine; Motor Activity; Nerve Degeneration; Neuroprotective Agents; Nitro Compounds; Piperidines; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerases; Propionates; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Tyrosine

In addition to protective effects within the adult central nervous system (CNS), in vivo application of N-methyl-d-aspartate inhibitors such as (+) MK-801 have been shown to induce neurodegeneration in neonatal rats over a specific developmental period. We have systematically mapped the nature and extent of MK-801-induced neurodegeneration throughout the neonatal murine brain in order to genetically dissect the mechanism of these effects. Highest levels of MK-801-induced neurodegeneration are seen in the cerebellar external germinal layer; while mature neurons of the internal granule layer are unaffected by MK-801 treatment. Examination of external germinal layer neurons by electron microscopy, terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling (TUNEL) and bromodeoxyuridine (BrdU) labeling, and caspase-3 activation demonstrate that these neurons die through the process of programmed cell death soon after they exit from the cell cycle. Significantly, ablation of caspase-3 activity completely inhibited the MK-801-induced (and developmental) programmed cell death of external germinal layer neurons. Similar to caspase-3, inactivation of muscarinic acetylcholine receptors in vivo using scopolamine inhibited MK-801-induced programmed cell death. By contrast, the GABAergic agonist diazepam, either alone or in combination with MK-801, enhanced programmed cell death within external germinal layer neurons. These data demonstrate that, in vivo, cerebellar granule neurons undergo a dramatic change in intracellular signaling in response to molecules present in the local cellular milieu during their first 24 h following exit from the cell cycle.

Topics: Analysis of Variance; Animals; Animals, Newborn; Bromodeoxyuridine; Caspase 3; Cell Death; Cerebellum; Cholinergic Antagonists; Diazepam; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; GABA Modulators; In Situ Nick-End Labeling; Mice; Mice, Inbred C57BL; Mice, Knockout; Microscopy, Electron, Transmission; Nerve Degeneration; Neurons; Scopolamine; Stem Cells

We have previously shown that P7 rat pups injected with the N-methyl-d-aspartate receptor (NMDAR) blocker MK801 displayed robust apoptotic injury within hours after injection. Further studies from our lab suggest that loss of calcium cannot be compensated for when vulnerable neurons lack calcium buffering capabilities. Thus, to elevate calcium in these neurons prior to MK801 exposure, we injected P7 rats with the calcium channel agonist BayK 8644. Whereas BayK 8644 did not induce apoptosis by itself, it was found to block MK801-induced injury in a dose-dependent manner. Reversal of MK801 toxicity was complete in the caudate-putamen, partial in the somatosensory cortex but was not observed in the retrosplenial cortex. These results suggest that postnatal brain injury resulting from agents that block the NMDAR, which include commonly used anesthetics as well as drugs of abuse, may be prevented in vulnerable neurons by compensatory increases in calcium prior to exposure to these antagonists.

Topics: 3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; Animals; Animals, Newborn; Apoptosis; Brain; Brain Damage, Chronic; Calcium Channel Agonists; Calcium Channels; Calcium Signaling; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Neostriatum; Nerve Degeneration; Neurons; Rats; Receptors, N-Methyl-D-Aspartate; Somatosensory Cortex

Intraventricular (i.c.v.) kainic acid (KA) causes an acute excitotoxic lesion to the CA3 region of rodent hippocampus. Recent evidence implicated c-fos gene in regulating neuron survival and death following an excitotoxic insult. In this study we attempted to prevent KA-induced damage in CA3 neurons with NMDA preconditioning, which produced a marked expression of c-fos in the hippocampus. NMDA (0.6-6 microg, i.c.v.) was injected to anesthetized rats alone or 1 h before KA (0.15 microg, i.c.v.). Following KA injection, vibratome sections were processed for immunohistochemistry/electron microscopy. c-Fos and Nissl staining were used to estimate the extent of neuronal excitation and damage, respectively. Quantitative evaluation of c-Fos-labeled cells showed significantly less c-Fos in CA3a than in neighboring CA3b and CA2 from 1 to 4 h after KA alone. Attenuation of expressed c-Fos in CA3a was accompanied by damage of neurons with more apoptotic than necrotic signs. NMDA preconditioning elevated CA3a c-Fos expression and at 1 and 2 h exceeded markedly that after KA alone. However, at 4 h after KA, NMDA-preconditioned c-Fos induction in CA3a diminished to the same level as that seen after KA alone. The onset of neuronal degeneration was delayed in similar way. While NMDA-induced c-Fos expression in CA3a could be blocked by MK-801 completely, MK-801 and CNQX were both without significant effect on KA-induced c-Fos expression and neuronal damage. In conclusion, inhibition of c-Fos expression and onset of neuronal damage in CA3a following icv KA injection might be transiently delayed by i.c.v. NMDA preconditioning.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Cell Death; Dizocilpine Maleate; Female; Gene Expression; Genes, fos; Hippocampus; Kainic Acid; N-Methylaspartate; Nerve Degeneration; Neurons; Neuroprotective Agents; Proto-Oncogene Proteins c-fos; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate

The organotin trimethyltin (TMT) is known to cause neuronal degeneration in the central nervous system. A systemic injection of TMT produced neuronal damage in the cerebral frontal cortex of mice. To elucidate the mechanism(s) underlying the toxicity of TMT toward neurons, we prepared primary cultures of neurons from the cerebral cortex of mouse embryos for use in this study. Microscopic observations revealed that a continuous exposure to TMT produced neuronal damage with nuclear condensation in an incubation time-dependent manner up to 48 h. The neuronal damage induced by TMT was not blocked by N-methyl-D-aspartate receptor channel-blocker MK-801. The exposure to TMT produced an elevation of the phosphorylation level of c-Jun N-terminal kinase (JNK)(p46), but not JNK(p54), prior to neuronal death. Under the same conditions, a significant elevation was seen in the phosphorylation level of stress-activated protein kinase 1, which activates JNKs. Furthermore, TMT enhanced the expression and phosphorylation of c-Jun during a continuous exposure. The JNK inhibitor SP600125 was effective in significantly but only partially attenuating the TMT-induced nuclear condensation and accumulation of lactate dehydrogenase in the culture medium. Taken together, our data suggest that the neuronal damage induced by TMT was independent of excitotoxicity but that at least some of it was dependent on the JNK cascades in primary cultures of cortical neurons.

Topics: Animals; Anthracenes; Cell Survival; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Enzyme Activation; Fungicides, Industrial; Immunoblotting; Injections, Intraperitoneal; JNK Mitogen-Activated Protein Kinases; Male; MAP Kinase Kinase 4; Mice; Mice, Inbred Strains; N-Methylaspartate; Nerve Degeneration; Neurons; Phosphorylation; Time Factors; Trimethyltin Compounds

The acoustic startle reflex in rats can be inhibited if a prepulse stimulus is presented just before the startle stimulus (prepulse inhibition; PPI). When postnatal day 7 (P7) rats are exposed to agents that block the NMDA receptor (NMDAR), robust apoptosis is observed within hours and is thought to be followed at later ages by a significant loss of PPI. To understand these observations further, we exposed rat pups to vehicle or the NMDAR antagonist MK801 (1 mg/kg) at P6, P8, and P10. We then examined animals for PPI at P28 and P56. Compared to vehicle controls, we found no evidence for PPI deficits in the MK801-treated group, although we did observe prepulse-induced delay in response time at P56 (but not at P28). In a parallel study, we also performed histological analysis of brain sections for evidence of the pro-apoptotic marker activated caspase-3, 8 h after vehicle or MK801 injection into P6 animals. We found that there was a robust increase in this marker of cell death in the inferior colliculus of MK801 compared to vehicle-treated animals. Thus, transient blockade of the NMDAR during the postnatal period not only promotes early apoptosis in a brain region critical for acoustic processing but also leads to auditory deficits at a later age, suggesting that injury-induced loss of collicular neurons leads to network reorganization in the auditory system that is progressive in nature.

Topics: Aging; Animals; Animals, Newborn; Apoptosis; Auditory Pathways; Biomarkers; Caspase 3; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Glutamic Acid; Inferior Colliculi; Male; Nerve Degeneration; Nerve Net; Neuronal Plasticity; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Sensory Gating; Time

N-methyl-d-aspartate receptor antagonist drugs (NMDA-A), such as dizocilpine (MK801), induce long-lasting behavioral disturbances reminiscent to psychotic disorders in humans. To identify cortical structures affected by NMDA-A, we used a single dose of MK801 (10 mg/kg) that caused low and high neurodegeneration in intact and orchiectomized male rats, respectively. Degenerating somas (neuronal death) and axonal/synaptic endings (terminal degeneration) were depicted by a silver technique, and functionally affected cortical neuronal subpopulations by Egr-1, c-Fos, and FosB/DeltaFosB-immunolabeling. In intact males, MK801 triggered a c-Fos induction that remained high for more than 24 h in selected layers of the retrosplenial, somatosensory and entorhinal cortices. MK801-induced neurodegeneration reached its peak at 72 h. Degenerating somas were restricted to layer IV of the granular subdivision of the retrosplenial cortex, and were accompanied by suppression of Egr-1 immunolabeling. Terminal degeneration extended to selected layers of the retrosplenial, somatosensory and parahippocampal cortices, which are target areas of retrosplenial cortex. Induction of FosB/DeltaFosB by MK801 also extended to the same cortical layers affected by terminal degeneration, likely reflecting the damage of synaptic connectivity. In orchiectomized males, the neurodegenerative and functional effects of MK801 were exacerbated. Degenerative somas in layer IV of the retrosplenial cortex significantly increased, with a parallel enhancement of terminal degeneration and FosB/DeltaFosB-expression in the mentioned cortical structures, but no additional areas were affected. These observations reveal that synaptic dysfunction/degeneration in the retrosplenial, somatosensory and parahippocampal cortices might underlie the long-lasting impairments induced by NMDA-A.

Topics: Animals; Cerebral Cortex; Dizocilpine Maleate; Early Growth Response Protein 1; Excitatory Amino Acid Antagonists; Gene Expression Regulation; Genes, Immediate-Early; Immunohistochemistry; Male; Nerve Degeneration; Neurons; Parahippocampal Gyrus; Presynaptic Terminals; Proto-Oncogene Proteins c-fos; Rats; Rats, Wistar; Somatosensory Cortex; Synaptic Transmission; Time

MK801 (dizocilpine) induces selective neurotoxic effects in the retrosplenial cortex, ranging from neuronal vacuolization to irreversible neurodegeneration depending on the dose administered. Although lamotrigine prevents MK801-induced neuronal vacuolization in the retrosplenial cortex 4 h after injection, it is not clear whether lamotrigine attenuates the subsequent neurodegeneration that occurs 3-4 days later. Because early growth response factor-1 (egr-1) plays a key role in neurodegeneration and its expression is induced in the retrosplenial cortex following MK801 treatment, it is possible that lamotrigine may attenuate MK801-induced neurodegeneration via inhibition of egr-1 expression in the retrosplenial cortex. To address this issue, we treated rats with lamotrigine (10 or 20 mg/kg) followed by MK801 (2 mg/kg) and measured changes in the levels of egr-1 mRNA and immunoreactivity in the retrosplenial cortex and other brain regions 3 h later. We also evaluated the effects of these treatments on neurodegeneration 4 days following treatment using Fluoro-Jade B staining. MK801 treatment increased egr-1 mRNA and immunoreactivity in the restrosplenial, cingulate, entorhinal and piriform cortices, but decreased levels in hippocampal subfields. These MK801-induced changes in egr-1 expression were significantly inhibited by lamotrigine pretreatment. In addition, MK801-induced neurodegeneration in the retrosplenial cortex was partially blocked by lamotrigine pretreatment in a dose dependent manner. These results demonstrate that lamotrigine pretreatment prevents the MK801-induced upregulation of egr-1 expression in a region-selective manner, and suggest that this effect may contribute, in part, to the attenuation of MK801-induced neurodegeneration in the retrosplenial cortex.

Topics: Animals; Cerebral Cortex; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Early Growth Response Protein 1; Hippocampus; Immunohistochemistry; Lamotrigine; Male; Nerve Degeneration; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; RNA, Messenger; Time Factors; Triazines

MK801, PCP, and ketamine are non-competitive NMDA receptor-antagonists drugs that in humans produce psychomimetic effects and neurocognitive disturbances reminiscent to those of schizophrenia. The administration of these drugs in animals has been used as a pharmacological model to study the NMDA receptor hypofunction-hypothesis of schizophrenia. In animals, the biological effect of MK801 is dose-dependent. Low doses induce behavioral disturbances and higher doses, in addition, promote neurotoxicity in many brain regions, particularly the granular retrosplenial cortex (RSG). The neurotoxic effect of MK801 is sexually dimorphic, being females markedly more sensitive than males; however, the involvement of gonadal hormones is elusive. Here we show that a single intraperitoneal injection of 5 mg/kg of MK801 induced overt neurodegeneration in RSG of female rats, including abundant somatic degeneration in layer 4, and somatodendritic and terminal degeneration in layers 1, 4, and 5. MK801-degeneration in males was scarce and mainly evidenced by the presence of few argirophilic somas in layer 4. Ovariectomized rats were not significantly different than intact females, while orchiectomized rats showed robust MK801-toxicity. Testosterone and dihydrotestosterone (DHT) inhibit MK801-toxicity in orchiectomized rats. In ovariectomized rats only DHT, but not testosterone, prevented MK801-induced degeneration, while in intact females, DHT was only partially protective. Treatment of intact males with estradiol benzoate significantly enhanced MK801-toxicity. Altogether, our experiments indicate that non-aromatizable androgens protect RSG from MK801-toxicity, while estrogens counteract this protection. Thus, the balance of androgens and estrogens delineate the sexual dimorphism of the RSG to the toxic effect of MK801.

Topics: Androgens; Animals; Cell Differentiation; Cerebral Cortex; Contraceptive Agents; Dihydrotestosterone; Dizocilpine Maleate; Estradiol; Excitatory Amino Acid Antagonists; Female; Gonadal Hormones; Injections, Intraperitoneal; Injections, Subcutaneous; Male; Nerve Degeneration; Neurons; Orchiectomy; Ovariectomy; Rats; Rats, Wistar; Sex Factors; Testosterone Propionate

About 30% of patients with epilepsy do not respond adequately to drug therapy, making pharmacoresistance a major problem in the treatment of this common brain disorder. Mechanisms of intractability are not well understood, but may include limitation of antiepileptic drug access to the seizure focus by overexpression of the drug efflux transporter P-glycoprotein (Pgp) at the blood-brain barrier. Increased expression of Pgp has been determined both in epileptogenic brain tissue of patients with intractable epilepsy and in rodent models of temporal lobe epilepsy, including the pilocarpine model. The mechanisms underlying the increase of Pgp after seizures are unclear. We have recently suggested that the excitatory neurotransmitter glutamate, which is excessively released by seizures, is involved in the seizure-induced overexpression of Pgp in the brain. This hypothesis was evaluated in the present study in the pilocarpine model in rats. After 90 min of status epilepticus (SE), diazepam was administered, followed by either vehicle or the glutamate receptor antagonist MK-801 (dizocilpine). Following SE in vehicle treated rats, Pgp expression in brain capillary endothelial cells increased about twofold in the hippocampus, which was completely prevented by MK-801. Furthermore, neurodegeneration developing in the hippocampus and parahippocampal regions was reduced by the glutamate antagonist. In contrast, the Pgp inhibitor tariquidar did not affect the SE-induced overexpression of Pgp or neurodegeneration in most regions examined. The data indicate that seizure-induced glutamate release is involved in the regulation of Pgp expression, which can be blocked by MK-801. The finding that MK-801 counteracts both Pgp overexpression and neuronal damage when administered after SE may offer a clinically useful therapeutic option in patients with refractory SE.

Topics: Animals; Apoptosis; ATP Binding Cassette Transporter, Subfamily B, Member 1; Brain Chemistry; Capillaries; Dizocilpine Maleate; Epilepsy, Temporal Lobe; Excitatory Amino Acid Antagonists; Female; Glutamic Acid; Image Processing, Computer-Assisted; Immunohistochemistry; Muscarinic Agonists; Nerve Degeneration; Pilocarpine; Quinolines; Rats; Rats, Wistar; Seizures; Status Epilepticus

PNQX (9-methyl-amino-6-nitro-hexahydro-benzo(F)quinoxalinedione) is a selective AMPA antagonist with demonstrated neuroprotective effects in focal ischemia in rats. Here we report corresponding effects in mouse hippocampal slice cultures subjected to oxygen and glucose deprivation (OGD) and in transient global cerebral ischemia in gerbils. For in vitro studies, hippocampal slice cultures derived from 7-day-old mice and grown for 14 days, were submersed in oxygen-glucose deprived medium for 30 min and exposed to PNQX for 24 h, starting together with OGD, immediately after OGD, or 2 h after OGD. For comparison, other cultures were exposed to the NMDA antagonist MK-801 using the same protocol. Both PNQX and MK-801 displayed significant neuroprotective effects in all hippocampal subfields when present during and after OGD. When added just after OGD, only PNQX retained some neuroprotective effect. When added 2 h after OGD neither PNQX nor MK-801 had an effect. Transient global cerebral ischemia was induced in Mongolian gerbils by occlusion of both common carotid arteries for 4.5 min, with PNQX (10 mg/kg) being injected i.p. 30, 60 and 90 min after the insult. Subsequent analysis of brain sections stained for the neurodegeneration marker Fluoro-Jade B and immunostained for the astroglial marker glial fibrillary acidic protein revealed a significant PNQX-induced decrease in neuronal cell death and astroglial activation. We conclude that, PNQX provided neuroprotection against both global cerebral ischemia in gerbils in vivo and oxygen-glucose deprivation in mouse hippocampal slice cultures.

Topics: Animals; Body Temperature; Cell Death; Coloring Agents; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Fluoresceins; Gerbillinae; Glucose; Hippocampus; Hypoxia, Brain; Ischemic Attack, Transient; Mice; Mice, Inbred C57BL; Microtubule-Associated Proteins; Nerve Degeneration; Neuroprotective Agents; Organ Culture Techniques; Organic Chemicals; Quinoxalines; Receptors, AMPA; Tolonium Chloride

We compared the neuroprotective and metabolic effects of chronic treatment with ionotropic or metabotropic glutamate receptor antagonists, in rats bearing a unilateral nigrostriatal lesion induced by 6-hydroxydopamine (6-OHDA). The ionotropic, N-methyl-D-aspartate receptor antagonist MK-801 increased cell survival in the substantia nigra pars compacta (SNc) and corrected the metabolic hyperactivity (increased cytochrome oxidase activity) of the ipsilateral substantia nigra pars reticulata (SNr) associated with the lesion, but showed no effects on the 6-OHDA-induced hyperactivity of the subthalamic nucleus (STN). Significant-although less pronounced-protection of SNc neurons was also observed following treatment with the metabotropic glutamate receptor (mGluR5) antagonist 2-methyl-6-(phenylehtynyl)-pyridine (MPEP). As opposed to MK-801, MPEP abolished the STN metabolic hyperactivity associated with the nigrostriatal lesion, without affecting SNr activity. Specific modulation of STN hyperactivity obtained with mGluR5 blockade may, therefore, open interesting perspectives for the use of this class of compounds in the treatment of Parkinson's disease.

Topics: Animals; Cell Survival; Corpus Striatum; Disease Models, Animal; Dizocilpine Maleate; Electron Transport Complex IV; Energy Metabolism; Excitatory Amino Acid Antagonists; Male; Nerve Degeneration; Neural Pathways; Neuroprotective Agents; Oxidopamine; Parkinsonian Disorders; Pyridines; Rats; Rats, Sprague-Dawley; Receptor, Metabotropic Glutamate 5; Receptors, Glutamate; Receptors, Metabotropic Glutamate; Receptors, N-Methyl-D-Aspartate; Substantia Nigra; Subthalamic Nucleus

Alzheimer's disease is characterized by extracellular beta-amyloid plaques, intraneuronal Tau-inclusions and cell death of cholinergic neurons. Recent evidence indicates that the vascular system may play an important role in the development of this progressive neurodegenerative disease. The aim of this study was to observe, if brain capillary endothelial cells (BCEC) may produce and secrete factors which induce cell death of cholinergic neurons, and if this effect is counteracted by (1) NGF, MK-801 or vitamin C, (2) modulated by experimentally-induced inflammation with interleukin-1beta and lipopolysaccharide (IL-1beta and LPS) or (3) by blocking of different intracellular signalling pathways. Cholinergic neurons were cultivated in organotypic brain slices of the nucleus basalis of Meynert and treated with conditioned medium derived from BCEC, supplemented with different protective factors. BCEC were stimulated with IL-1beta and LPS or different intracellular pathway inhibitors before collection of conditioned medium. Cholinergic neurons were detected by immunohistochemistry for choline-acetyltransferase. Possible effects on BCEC viability and proliferation were determined by nuclear staining, BrdU incorporation and release of nitrite and lactate-dehydrogenase. BCEC released factors that can kill cholinergic neurons. This neurotoxic effect was blocked by NGF and MK-801 (a NMDA-antagonist), but not by vitamin C. Pretreatment of BCEC with intracellular pathway inhibitors did not change the neurotoxicity, but pretreatment with IL-1beta and LPS abolished the neurotoxic effect. In summary, BCEC produce and secrete molecules which induce excitotoxic cell death of cholinergic neurons. It is concluded that excitotoxic factors secreted by vascular cells may contribute to the development of cholinergic neurodegeneration as it occurs in Alzheimer's disease.

Topics: Alzheimer Disease; Animals; Antioxidants; Ascorbic Acid; Brain; Cell Death; Cells, Cultured; Cholinergic Fibers; Culture Media, Conditioned; Dizocilpine Maleate; Endothelial Cells; Inflammation; Interleukin-1beta; Lipopolysaccharides; Nerve Degeneration; Nerve Growth Factor; Neuroprotective Agents; Rats; Rats, Sprague-Dawley

Alzheimer's disease (AD) is the most common form of neurodegenerative disease in the elderly. Anti-inflammatory agents have been shown to be beneficial in preventing neurodegenerative disorders such as AD. In this study we investigated the possible antioxidant and neuroprotective properties of two non-steroidal anti-inflammatory drugs (NSAIDS), tolmetin and sulindac, using quinolinic acid (QA)-induced neurotoxicity as a model. We used the thiobarbituric acid assay to measure the extent of lipid peroxidation and the nitroblue tetrazolium assay to measure the superoxide anion generated in rat brain homogenate. QA (1 mM) induced lipid peroxidation in rat brain homogenate was significantly curtailed by co-treatment of the homogenate with tolmetin and/or sulindac. Tolmetin and sulindac both reduced the generation of superoxide anions by the known neurotoxin, potassium cyanide (KCN). Intrahippocampal injections of QA induced neurotoxicity in rat hippocampus. N-Methyl-D-Aspartate (NMDA) receptor counts were conducted do give an indication of the amount protection offered by the NSAIDS. QA drastically reduced the number of NMDA binding sites by approximately 37%. This sharp decrease was considerably attenuated by the pre-treatment of the rats with tolmetin and sulindac (5 mg/kg/bd for five days). This study shows the antioxidant and neuroprotective properties of tolmetin and sulindac and hereby postulates that these drugs have important implications in the prevention or treatment of neurodegenerative diseases such as AD.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Brain Chemistry; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hippocampus; Kinetics; Lipid Peroxidation; Male; Nerve Degeneration; Neurons; Neuroprotective Agents; Oxidative Stress; Potassium Chloride; Quinolinic Acid; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Sulindac; Superoxides; Thermodynamics; Tolmetin

The TGF-beta signaling pathway is a key organizer of injury and immune responses, and recent studies suggest it fulfills critical roles in CNS function and maintenance. TGF-beta receptor activation results in phosphorylation of Smad proteins, which subsequently translocate to the nucleus to regulate gene transcription by binding to Smad binding elements (SBE). Using SBE-luciferase reporter mice, we recently discovered that the brain has the highest Smad baseline activity of any major organ in the mouse, and we now demonstrate that this signal is primarily localized to pyramidal neurons of the hippocampus. In vivo excitatory stimulation with kainic acid (KA) resulted in an increase in luciferase activity and phosphorylated Smad2 (Smad2P), and nuclear translocation of Smad2P in hippocampal CA3 neurons correlated significantly with luciferase activity. Although this activation was most prominent at 24 h after KA administration in neurons, Smad2P immunoreactivity gradually increased in astrocytes and microglial cells at 3 and 5 days, consistent with reactive gliosis. Bioluminescence measured over the skull in living mice peaked at 12-72 h and correlated with the extent of microglial activation and pathological markers of neurodegeneration 5 days after injury. Treatment with the glutamate receptor antagonist MK-801 strongly reduced bioluminescence and pathology. These results show that Smad2 signaling is a sensitive marker of neuronal activation and CNS injury that can be used to monitor KA-induced neuronal degeneration. This and related mouse models may provide valuable tools to study mechanisms and treatments for neurodegeneration.

Topics: Animals; Biomarkers; Cells, Cultured; Central Nervous System; Dizocilpine Maleate; Genes, Reporter; Kainic Acid; Mice; Microscopy, Confocal; Nerve Degeneration; Signal Transduction; Smad Proteins

We investigated the relationship between the degeneration of spinal motor neurons and activation of N-methyl-d-aspartate (NMDA) receptors after neuraxial morphine following a noninjurious interval of aortic occlusion in rats. Spinal cord ischemia was induced by aortic occlusion for 6 min with a balloon catheter. In a microdialysis study, 10 muL of saline (group C; n = 8) or 30 mug of morphine (group M; n = 8) was injected intrathecally (IT) 0.5 h after reflow, and 30 mug of morphine (group SM; n = 8) or 10 muL of saline (group SC; n = 8) was injected IT 0.5 h after sham operation. Microdialysis samples were collected preischemia, before IT injection, and at 2, 4, 8, 24, and 48 h of reperfusion (after IT injection). Second, we investigated the effect of IT MK-801 (30 mug) on the histopathologic changes in the spinal cord after morphine-induced spastic paraparesis. After IT morphine, the cerebrospinal fluid (CSF) glutamate concentration was increased in group M relative to both baseline and group C (P < 0.05). This increase persisted for 8 hrs. IT MK-801 significantly reduced the number of dark-stained alpha-motoneurons after morphine-induced spastic paraparesis compared with the saline group. These data indicate that IT morphine induces spastic paraparesis with a concomitant increase in CSF glutamate, which is involved in NMDA receptor activation. We suggest that opioids may be neurotoxic in the setting of spinal cord ischemia via NMDA receptor activation.

Topics: Analgesics, Opioid; Animals; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Injections, Spinal; Male; Morphine; Motor Neurons; Nerve Degeneration; Neuroprotective Agents; Paraparesis; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Spinal Cord; Spinal Cord Ischemia

The regulatory mechanisms of neuropeptide-metabolizing enzymes often play a critical role in the pathogenesis of neuronal damage. A systemic administration of pentylenetetrazol (PTZ), an antagonist of GABA(A) receptor ion channel binding site, causes generalized epilepsy in an animal model. In the present study, we examined the involvement of prolyl oligopeptidase (POP), thimet oligopeptidase/neurolysin (EP 24.15/16) and glial proteins in PTZ-treated rat brain regions, and the suppressive effect of MK-801, a non-competitive NMDA receptor antagonist, pretreatment for their proteins. The activity of POP significantly decreased in the hippocampus at 30min and 3h, and in the frontal cortex at 3h after PTZ treatment, and pretreatment with MK-801 recovered the activity in the cortex at 3h. The activity of EP 24.15/16 significantly decreased in the hippocampus at 3h and 1 day, and in the cortex at 3h after the PTZ administration, whereas pretreatment with MK-801 recovered the change of the activity. The Western blot analysis of EP 24.15 showed significant decrease of the protein level in the hippocampus 3h after the PTZ treatment, whereas pretreatment with MK-801 recovered. The expression of GFAP and CD11b immunohistochemically increased in the hippocampus of the PTZ-treated rat as compared with controls. Pretreatment with MK-801 also recovered the GFAP and CD11b expression. These data suggest that PTZ-induced seizures of the rats cause indirect activation of glutamate NMDA receptors, then decrease POP and EP 24.15/16 enzyme activities and EP 24.15 immunoreactivity in the neuronal cells of the hippocampal formation. We speculate that changes of those peptidases in the brain may be related to the levels of the neuropeptides regulating PTZ-induced seizures.

Topics: Animals; Brain; CD11b Antigen; Cerebral Cortex; Convulsants; Disease Models, Animal; Dizocilpine Maleate; Down-Regulation; Epilepsy; Excitatory Amino Acid Antagonists; GABA Antagonists; Glial Fibrillary Acidic Protein; Hippocampus; Male; Metalloendopeptidases; Nerve Degeneration; Neuroglia; Neurons; Pentylenetetrazole; Prolyl Oligopeptidases; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Serine Endopeptidases

NMDA antagonists are of potential therapeutic benefit for several conditions. However, their ability to produce neurotoxicity and psychosis has hampered their clinical use. A better understanding of these side effects and the mechanism underlying them could result in their safer use and in improving our understanding of psychotic illnesses. By disinhibiting certain multisynaptic circuits, moderate doses of NMDA antagonists produce reversible neurotoxicity in the retrosplenial cortex in rats older than 1 month. Higher doses of these same agents result in the death of neurons in the retrosplenial cortex and several other brain regions. It is unknown whether susceptibility to this irreversible neurodegeneration has a similar age dependency profile. We, therefore, examined the sensitivity of rats of various ages (PND20-60) to the irreversible neurodegenerative effect of the selective NMDA antagonist, MK-801. Quantification of the severity of neurodegeneration with stereology revealed that the retrosplenial cortex, induseum griseum, and dentate gyrus had decreasing amounts of damage with decreasing age and onset of sensitivity around PND30. The piriform cortex also displayed a decreased amount of degeneration in younger age groups. However, a low level of degeneration continued to occur in the posterior piriform cortex in the PND20-25 animals. The stage of degeneration appeared to be more advanced, suggesting that these neurons were dying by a different mechanism. We conclude that for most neuronal populations, susceptibility to the irreversible and reversible neurodegenerative effects of NMDA antagonists has a similar age dependency profile, consistent with the proposal that the same disinhibitory mechanism underlies both neurotoxicities.

Topics: Age Factors; Aging; Animals; Brain; Cell Count; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Nerve Degeneration; Rats; Rats, Sprague-Dawley; Silver Staining

Altered cholinergic function is considered as a potential contributing factor in the pathogenesis of schizophrenia. We hypothesize that cortical cholinergic denervation may result in changes in glutamatergic activity. Therefore, we lesioned the cholinergic corticopetal projections by local infusion of 192 IgG-saporin into the nucleus basalis magnocellularis of rats. Possible effects of this lesion on glutamatergic systems were examined by phencyclidine-induced locomotor activity, and also by N-methyl-D-aspartate receptor binding. We find that cholinergic lesioning of neocortex leads to enhanced sensitivity to phencyclidine in the form of a dramatic increase in horizontal activity. Further, N-methyl-D-aspartate receptor binding is unaffected in denervated rats. These results suggest that aberrations in cholinergic function might lead to glutamatergic dysfunctions, which might be of relevance for the pathophysiology for schizophrenia.

Topics: Acetylcholine; Acetylcholinesterase; Analysis of Variance; Animals; Antibodies, Monoclonal; Basal Nucleus of Meynert; Behavior, Animal; Cerebral Cortex; Denervation; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Inhibitors; Hyperkinesis; Immunohistochemistry; Immunotoxins; Male; Motor Activity; N-Glycosyl Hydrolases; Nerve Degeneration; Phencyclidine; Protein Binding; Radioligand Assay; Rats; Rats, Sprague-Dawley; Ribosome Inactivating Proteins, Type 1; Saporins; Tritium

The intrahippocampal perfusion of 4-aminopyridine (4-AP) in the rat produces immediate seizures and delayed neuronal death, due to the overactivation of N-methyl-D-aspartate (NMDA) receptors by endogenous glutamate released from nerve endings. With the same time course, 4-AP also induces the expression of the cell stress marker heat shock protein 70 (HSP70) in the contralateral non-damaged hippocampus. We have used this experimental model to study the mechanisms of the delayed neuronal stress and death. The NMDA receptor antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo(a,d)cyclohepten-5,10-imine maleate (MK-801), administered intraperitoneally 30 or 60 but not 120 min after 4-AP perfusion, when animals show intense electroencephalography epileptiform activity, prevented the delayed neurodegeneration whereas the seizures continued for about 3 h as in the control animals. With an identical time window, MK-801 treatment also modified the pattern of HSP70 expression; the protein was expressed in the protected perfused hippocampus but no longer in the undamaged contralateral hippocampus. The possible role of Ca2+ in the delayed cell death and HSP70 expression was also studied by coperfusing the intracellular Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetra-acetic acid tetrakis(acetoxymethyl ester) with 4-AP. This treatment resulted in protective and HSP70 effects very similar to those of MK-801. These results suggest that the seizures are not linked to neurodegeneration and that NMDA receptors need to be continuously overactivated by endogenous glutamate for at least 60 min in order to induce delayed neuronal stress and death, which are dependent on Ca2+ entry through the NMDA receptor channel.

Topics: 4-Aminopyridine; Animals; Cell Count; Cell Death; Chelating Agents; Disease Models, Animal; Dizocilpine Maleate; Drug Interactions; Egtazic Acid; Electroencephalography; Epilepsy; Excitatory Amino Acid Antagonists; Functional Laterality; Glutamic Acid; Hippocampus; Male; Microdialysis; Nerve Degeneration; Neurons; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Stress, Physiological; Time Factors

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

The purpose of this study was to examine the optimal dose and therapeutic window of opportunity of the nonsteroidal anti-inflammatory drug naproxen in an animal model of excitotoxic neuronal injury. Injection of N-methyl-D-aspartate (NMDA; 18-20 nmol) into the CA1 region of the left hippocampus resulted in significant brain edema as measured by the percentage of total forebrain water content that occurred 24 h after intrahippocampal microinjection of NMDA with approximately 50% loss of CA1 neurons assessed 72 h later. Naproxen pretreatment (20 mg/kg) resulted in significantly less brain edema. Ten, 15, or 20 mg/kg naproxen, administered systemically 1 day (b.i.d.) before and for 3 days after (b.i.d.) NMDA injection, attenuated the neuronal damage by 27.2 +/- 7.8, 39.6 +/- 11.1, and 57.0 +/- 5.2%, respectively. By comparison, a single dose of MK-801 (2 mg/kg i.p.) given 20 min before NMDA injection inhibited subsequent hippocampal injury by 65.6 +/- 8.8%. Most importantly, neuroprotection was still evident when naproxen treatment (20 mg/kg i.p.) was initiated 6 h after NMDA microinjection. Protection was lost if administration of naproxen was delayed for 20 h. These findings demonstrate that naproxen can prevent excitotoxic neuronal injury in vivo, that it is nearly as effective as direct NMDA receptor antagonism, and that it has an extended therapeutic time window. As such, naproxen may be a particularly promising pharmaceutical for the treatment of neurological diseases associated with overactivation of NMDA receptors.

Topics: Animals; Dizocilpine Maleate; Male; Mice; N-Methylaspartate; Naproxen; Nerve Degeneration; Neuroprotective Agents; Time Factors

Pharmacological blockade of NMDA receptor function induces apoptotic neurodegeneration in the developing rat brain. However, the use of NMDA receptor antagonists as anesthetics and sedatives represents a difficult-to-avoid clinical practice in pediatrics. This warrants the search for adjunctive neuroprotective measures that will prevent or ameliorate neurotoxicity of NMDA receptor antagonists. The NMDA receptor antagonist MK801 triggered apoptosis in the neonatal rat forebrain, most notably in cortex and thalamus. MK801 exposure reduced mRNA levels of erythropoietin (EPO) and the EPO receptor, suggesting that loss of endogenous EPO activity may contribute to MK801-induced apoptosis. Coadministration of recombinant EPO (rEPO) conferred 50% neuroprotection, partially restored MK801-induced reduction of brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) mRNA, and prevented decreased phosphorylation levels of extracellular signal-regulated protein kinase-1/2 (ERK1/2) and Akt. These observations indicate that rEPO partly rescues newborn rats from MK801-mediated brain damage by enhancing neurotrophin-associated signaling pathways.

Topics: Animals; Animals, Newborn; Apoptosis; Brain; Brain-Derived Neurotrophic Factor; Dizocilpine Maleate; Erythropoietin; Excitatory Amino Acid Antagonists; Glial Cell Line-Derived Neurotrophic Factor; Mice; Mice, Transgenic; Mitogen-Activated Protein Kinases; Nerve Degeneration; Nerve Growth Factors; Neuroprotective Agents; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Rats; Rats, Wistar; Receptors, Erythropoietin; Receptors, N-Methyl-D-Aspartate; Recombinant Fusion Proteins; RNA, Messenger; Signal Transduction

The developing rodent brain is vulnerable to pharmacological blockade of N-methyl-d-aspartate (NMDA) receptors which can lead to severe and disseminated apoptotic neurodegeneration. Here, we show that systemic administration of the NMDA receptor antagonist MK801 to 7-day-old rats leads to impaired activity of extracellular signal-regulated kinase 1/2 (ERK1/2) and reduces levels of phosphorylated cAMP-responsive element binding protein (CREB) in brain regions which display severe apoptotic neurodegeneration. Impaired ERK1/2 and CREB activity were temporally paralleled by sustained depletion of neurotrophin expression, particularly brain-derived neurotrophic factor (BDNF). BDNF supplementation fully prevented MK801-induced neurotoxicity in immature neuronal cultures and transgenic constitutive activation of Ras was associated with marked protection against MK801-induced apoptotic neuronal death. These data indicate that uncoupling of NMDA receptors from the ERK1/2-CREB signaling pathway in vivo results in massive apoptotic deletion of neurons in the developing rodent brain.

Topics: Age Factors; Animals; Apoptosis; Cells, Cultured; Cyclic AMP Response Element-Binding Protein; Dizocilpine Maleate; Down-Regulation; Excitatory Amino Acid Antagonists; Gene Expression; Gyrus Cinguli; MAP Kinase Signaling System; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases; Nerve Degeneration; Nerve Growth Factors; Neurons; Proto-Oncogene Proteins c-bcl-2; ras Proteins; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Transcription, Genetic

We used primary cultures of cortical neurons to examine the relationship between beta-amyloid toxicity and hyperphosphorylation of the tau protein, the biochemical substrate for neurofibrillary tangles of Alzheimer's brain. Exposure of the cultures to beta-amyloid peptide (betaAP) induced the expression of the secreted glycoprotein Dickkopf-1 (DKK1). DKK1 negatively modulates the canonical Wnt signaling pathway, thus activating the tau-phosphorylating enzyme glycogen synthase kinase-3beta. DKK1 was induced at late times after betaAP exposure, and its expression was dependent on the tumor suppressing protein p53. The antisense induced knock-down of DKK1 attenuated neuronal apoptosis but nearly abolished the increase in tau phosphorylation in betaAP-treated neurons. DKK1 was also expressed by degenerating neurons in the brain from Alzheimer's patients, where it colocalized with neurofibrillary tangles and distrophic neurites. We conclude that induction of DKK1 contributes to the pathological cascade triggered by beta-amyloid and is critically involved in the process of tau phosphorylation.

Topics: Aged; Aged, 80 and over; Alzheimer Disease; Amyloid beta-Peptides; Animals; Apoptosis; bcl-2-Associated X Protein; Cells, Cultured; Dizocilpine Maleate; Gene Expression Regulation; Glutamic Acid; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Humans; Immunoenzyme Techniques; Intercellular Signaling Peptides and Proteins; Nerve Degeneration; Nerve Tissue Proteins; Neurofibrillary Tangles; Neurons; Oligodeoxyribonucleotides; Peptide Fragments; Phosphorylation; Protein Processing, Post-Translational; Proteins; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Quinoxalines; Rats; RNA, Messenger; Signal Transduction; tau Proteins; Tumor Suppressor Protein p53; Wnt Proteins

Recent experimental evidence indicates that erythropoietin (Epo), in addition to its hormonal role in regulating red cell production, operates as a neuroprotective agent. So far, the neuroprotective effect of human recombinant Epo (rhEpo) has been mainly demonstrated in models of cerebral ischemia/hypoxia and in selected in vivo studies of traumatic neuronal injury. To further investigate the potential role of this multifunctional trophic factor in post-traumatic cell death, we examined the protective effects of rhEpo in a newly developed model of mechanical trauma in organotypic hippocampal slices. Organotypic rat hippocampal slices were subjected to traumatic injury by allowing a stylus to impact on the CA1 area with an energy of 6 microJ. Hippocampal damage was identified and measured 24 and 48 h later with the fluorescent dye propidium iodide (PI). In untreated slices, the impact induced a significant increase in the mean hippocampal PI fluorescence, co-localized with the area of impact at 24 h (primary post-traumatic injury) and progressively spread to the whole slice between 24 and 48 h (secondary post-traumatic injury). Addition of rhEpo (1-100 UI/mL) or of the NMDA antagonist MK-801 (30 microM) immediately after the traumatic injury reduced hippocampal damage by approximately 30% when observed 24 h later. At 48 h after trauma, the protective effect of rhEpo was greater (by about 47%) and significantly more pronounced than that of MK-801 (28%). Our results suggest that the neuroprotective activity of rhEpo is particularly effective against delayed, secondary post-traumatic damage. This well tolerated agent could provide a therapeutic benefit in pathologies involving post-traumatic neurodegeneration.

Topics: Animals; Animals, Newborn; Brain Injuries; Dizocilpine Maleate; Down-Regulation; Erythropoietin; Excitatory Amino Acid Antagonists; Hippocampus; In Vitro Techniques; Models, Biological; Nerve Degeneration; Neurons; Neuroprotective Agents; Propidium; Rats; Rats, Wistar; Recombinant Fusion Proteins; Recombinant Proteins; Treatment Outcome

Single doses of an NMDA antagonist cause an adult or a prepubertal form of neurodegeneration, depending on the age of the animal. Single doses of ethanol (EtOH) by blocking NMDA receptors produce apoptotic neurodegeneration in young animals. This capability could account, in part, for the ability of EtOH to produce the fetal alcohol syndrome. We investigated whether EtOH could produce NMDA antagonist-induced neurotoxicity (NAN), a different neurotoxicity that is seen only in adult animals. In spite of producing blood EtOH levels (30 to 600 mg/dl) known to block NMDA receptors, EtOH was unable to produce neurotoxicity in the adult central nervous system (CNS). Moreover, EtOH in a dose-dependent fashion (ED(50)=138 mg/dl) prevented the selective and powerful NMDA antagonist, MK-801, from producing NAN in adult animals, suggesting that activity at another site might be negating the neurotoxic effect of EtOH's inherent NMDA antagonistic activity. Because GABA(A) agonism and non-NMDA glutamate antagonism, properties which EtOH possesses, can prevent NAN, we proceeded to study whether GABA(A) antagonists (or agents capable of reversing EtOH's GABAergic effects) and non-NMDA agonists could reverse EtOH's protective effect. Bicuculline, Ro15-4513, finasteride, kainic acid or AMPA, alone or in combination, did not significantly reverse EtOH's protective effect. Given that EtOH has effects on a wide range of ion channels and receptors, determining the precise mechanism of EtOH's protective effect will take additional effort. The inability of EtOH to acutely produce NAN in the adult CNS indicates that, in contrast to fetuses, brief exposure of the adult CNS to EtOH is non-toxic for neurons.

Topics: Age Factors; Animals; Apoptosis; Cerebral Cortex; Dizocilpine Maleate; Dose-Response Relationship, Drug; Ethanol; Excitatory Amino Acid Antagonists; Female; Nerve Degeneration; Neuroprotective Agents; Pyramidal Cells; Rats; Receptors, N-Methyl-D-Aspartate

The roles that glutamate N-methyl-D-aspartate (NMDA) and dopamine D1-like and D2-like receptors play in the cortical neurotoxicity occurring in rats exposed to multiple doses of amphetamine (AMPH) for 2 days was evaluated. Neurodegeneration in rats that did not become hyperthermic during AMPH exposure was quantified by counting isolectin B4-labeled phagocytic microglia and Fluoro-Jade (F-J)-labeled neurons in the somatosensory parietal cortex, piriform cortex and posterolateral cortical amygdaloid nucleus (PLCo). The NMDA receptor antagonist, dizocilpine (0.63 mg/kg day) blocked AMPH-induced neurodegeneration in the somatosensory cortex. However, it did not affect degeneration in the piriform cortex and PLCo indicating that limbic degeneration was not NMDA-mediated. The dopamine antagonists, eticlopride (D2/3, 0.25 mg/kg day) and SCH-23390 (D1, 0.25 mg/kg day), blocked the stereotypic behavior and neurodegeneration in the somatosensory cortex. However, eticlopride had a lesser protective effect in the limbic regions. As well, the dopamine D2/D3 agonist quinpirole (1.5 mg/kg day) protected against cortical neurodegeneration when it was given during AMPH exposure and continued until sacrifice. The dopamine D1 agonist (SKF-38393, 12.5 mg/kg day) had no significant effect on neurodegeneration. These data indicate that there are significant differences in NMDA and dopamine D2 modulation of AMPH-induced neurodegeneration in the somatosensory cortex compared to the limbic cortices, and limbic cortical degeneration is not necessarily dependent on excessive stimulation of NMDA receptors as it is in the somatosensory cortex. Although excessive dopamine receptor stimulation during amphetamine exposure may trigger the neurodegenerative processes, continued D2 stimulation after AMPH exposure is neuroprotective in the cortex.

Topics: Amphetamine; Animals; Dizocilpine Maleate; Dopamine Agents; Excitatory Amino Acid Antagonists; Limbic System; Male; Nerve Degeneration; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Receptors, Dopamine; Receptors, N-Methyl-D-Aspartate; Somatosensory Cortex

Several studies involving postnatal administration of the N-methyl-D-aspartate (NMDA) antagonists, dizocilpine (MK-801; 3 x 0.5 mg/kg, at 08.00, 16.00 and 24.00 h) on Postnatal day 11, or Ketamine (1 x 50 mg/kg) or Ethanol (1 x 2.5 g/kg, Ethanol-Low, or 2 x 2.5 g/kg, 2-h interval, Ethanol-High) on Postnatal day 10, are described. Some mice from each treatment/vehicle group were sacrificed 24 h after NMDA antagonist treatment and brain regions were taken for fluoro-jade staining analysis. Functional analysis was initiated at 60 days of age. All three treatments inducing an antagonistic action at NMDA receptors, MK-801, Ketamine and Ethanol-High induced a similar pattern of initial hypoactivity followed by marked and lasting hyperactivity in the motor activity test chambers. In each case, the basal hyperactivity level was abolished by acute treatment with a low dose of D-amphetamine (0.25 mg/kg). All three treatments, MK-801, Ketamine and Ethanol-High, induced a deficit in acquisitive performance in the radial arm maze test of instrumental learning. The deficit induced by postnatal MK-801 was abolished by acute treatment with the low dose of D-amphetamine. All three treatments, MK-801, Ketamine and Ethanol-High, resulted in normal acquisitive performance during the first three test days in the circular swim with the submerged platform maintained in a constant position, but on the fourth test day, with the platform position shifted to a different "quadrant", induced marked deficits. Fluoro-jade staining analyses indicated a devastating cell degeneration in several brain regions of mice administered NMDA antagonists postnatally, including the hippocampus, frontal cortex, parietal cortex, and cerebellum. Severe cell degeneration in the laterodorsal thalamus due to Ethanol or diazepam (5 mg/kg) appeared not to affect the different aspects of function. The pattern of dysfunctional outcome and apoptotic cell loss following postnatal NMDA antagonist treatment offers a plausible similarity to the major aspects of 'syndromatic continuity' in ADHD, hyperactivity, inattention and impulsivity, thereby providing an interesting animal model of the disorder.

Topics: Animals; Animals, Newborn; Apoptosis; Attention Deficit Disorder with Hyperactivity; Central Nervous System Depressants; Central Nervous System Stimulants; Dextroamphetamine; Diazepam; Dizocilpine Maleate; Ethanol; Excitatory Amino Acid Antagonists; Female; Fluoresceins; Fluorescent Dyes; Hypnotics and Sedatives; Ketamine; Maze Learning; Mental Disorders; Mice; Motor Activity; Nerve Degeneration; Nervous System Diseases; Organic Chemicals; Pregnancy; Receptors, N-Methyl-D-Aspartate; Swimming; Weight Gain

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

In order to examine the possible involvement of mu-calpain in methylmercury (MeHg)-induced neurotoxicity in developing cortical neurons, we performed biochemical and immunohistochemical studies utilizing two antibodies which specifically recognize the 150-kDa mu-calpain-specific alpha-spectrin breakdown product (SBDP) and the active form of mu-calpain in rats on postnatal day 16. Soluble fractions of the cerebral cortex from control rats exhibited slight immunoreactivity for SBDP. Although the amount of SBDP in the cerebral cortex was only slightly increased the day after the final treatment of MeHg (10 mg/kg) for 3 or 7 consecutive days, there was a prominent accumulation of SBDP 3 days after the final treatment of MeHg for 7 consecutive days. On the other hand, the 76-kDa isoform of mu-calpain gradually increased after chronic treatment of MeHg, but markedly decreased 3 days after the final treatment of MeHg for 7 consecutive days. At this stage, many cortical neurons were densely stained with anti-SBDP antibody. The delayed increase in SBDP corresponded well with the delayed nature of the MeHg-induced neurotoxicity. When MK-801 (0.1 mg/kg), a non-competitive antagonist of N-methyl-D-aspartate (NMDA), was administered intraperitoneally with MeHg for 7 consecutive days, both neuronal damage and accumulation of SBDP were markedly depressed in the cerebral cortex 3 days after the final treatment. Our results indicate that mu-calpain activation and mu-calpain-mediated proteolysis of alpha-spectrin preceded neuronal damage in the developing cerebral cortex induced by chronic treatment of MeHg.

Topics: Animals; Calpain; Cerebral Cortex; Dizocilpine Maleate; Enzyme Activation; Immunoblotting; Immunohistochemistry; Mercury Poisoning, Nervous System; Methylmercury Compounds; N-Methylaspartate; Nerve Degeneration; Neurons; Neuroprotective Agents; Rats; Rats, Wistar; Spectrin

The neurotoxic effect following a single intraperitoneal injection of MK-801 (10 mg/kg) in adult female Wistar rats at different survival times was studied with the 1994 version of de Olmos' Amino-Cupric-Silver (A-Cu-Ag) technique for detection of neural degeneration. In addition to the well documented somatodendritic degeneration observable in cortical olfactory structures, dentate gyrus, retrosplenial and sensory cortices, we detected this type of neuronal degeneration also in the main olfactory bulb, motor and anterior cingulate cortices, thalamus and cerebellum. Terminal degeneration, not reported by previous authors, was detected in cortical olfactory structures, hippocampal formation, sensory, infralimbic, prelimbic, agranular insular, ectorhinal, perirhinal and lateral orbital cortices. These results demonstrate that the A-Cu-Ag procedure is more efficient than other silver methods for detecting the degeneration induced by MK-801. In fact, the use of the A-Cu-Ag method has made it possible to infer the connectional relations between the damaged cell bodies and corresponding terminal degeneration. Our results also indicate that the A-Cu-Ag technique may be a suitable method for the staining of neurons undergoing apoptotic-like degeneration. The probable degenerative mechanism of MK-801 in the main olfactory system is discussed.

Topics: Animals; Apoptosis; Brain; Brain Diseases; Copper; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Nerve Degeneration; Rats; Rats, Wistar; Silver; Staining and Labeling; Time Factors

Most patients with temporal lobe epilepsy (TLE), the most common type of epilepsy, show pronounced loss of neurons in limbic brain regions, including the hippocampus. The massive neurodegeneration in the hippocampus is known as hippocampal sclerosis, and is considered one of the hallmarks of this type of difficult-to-treat epilepsy. There is a long and ongoing debate on whether this sclerosis is the result of an initial pathological event, such as a status epilepticus (S.E.), stroke or head trauma, which often precedes the development of TLE, or is caused by the spontaneous recurrent seizures (SRS) once epilepsy has developed. At present, pharmacological prevention of limbic sclerosis is not available. In a clinical situation, such prevention would only be possible if delayed cell death developing after an initial pathological event is involved. Assuming that sclerotic brain lesions provoke epileptogenesis and that delayed cell death is involved in these lesions, it should be possible to prevent both the lesions and the epilepsy by a prophylactic treatment after an initial insult such as an S.E. In order to test this hypothesis, we used a rat model of TLE in which limbic brain lesions and epilepsy with SRS develop after a kainate-induced S.E. A single low dose of the N-methyl-D-aspartate (NMDA) receptor blocker dizocilpine (MK-801) significantly reduced the damage in limbic regions, including the hippocampus and piriform cortex, and completely protected several rats from such damage when given after an S.E. of 90 min induced by kainate, strongly suggesting that delayed cell death is involved in the damage. This was substantiated by the use of molecular and immunohistochemical markers of delayed active ("programmed") cell death. However, the neuroprotection by dizocilpine did not prevent the development of SRS after the S.E., suggesting that structures not protected by dizocilpine may play a role in the genesis of SRS or that epileptogenesis is not the consequence of structural lesions in the limbic system. The only brain regions that exhibited neuronal damage in all rats with SRS were the hilus of the dentate gyrus and the mediodorsal thalamus, although treatment with dizocilpine reduced the severity of damage in the latter region. The data indicate that NMDA receptor blockade immediately after a prolonged S.E. is an effective means to reduce the damage produced by a sustained S.E. in several brain regions, including the hippocampus, but show that this pa

Topics: Animals; Cell Death; Dentate Gyrus; Disease Models, Animal; Dizocilpine Maleate; DNA Fragmentation; Epilepsy, Temporal Lobe; Excitatory Amino Acid Antagonists; Female; Hippocampus; Kainic Acid; Limbic System; Mediodorsal Thalamic Nucleus; Nerve Degeneration; Neurons; Neuroprotective Agents; Olfactory Pathways; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Status Epilepticus; Treatment Outcome

Multiple sclerosis is increasingly being recognized as a neurodegenerative disease that is triggered by inflammatory attack of the CNS. As yet there is no satisfactory treatment. Using experimental allergic encephalo myelitis (EAE), an animal model of multiple sclerosis, we demonstrate that the cannabinoid system is neuroprotective during EAE. Mice deficient in the cannabinoid receptor CB1 tolerate inflammatory and excitotoxic insults poorly and develop substantial neurodegeneration following immune attack in EAE. In addition, exogenous CB1 agonists can provide significant neuroprotection from the consequences of inflammatory CNS disease in an experimental allergic uveitis model. Therefore, in addition to symptom management, cannabis may also slow the neurodegenerative processes that ultimately lead to chronic disability in multiple sclerosis and probably other diseases.

Topics: Animals; Aspartic Acid; Axons; Benzoxazines; Cannabinoids; Cyclohexanols; Dizocilpine Maleate; Encephalomyelitis, Autoimmune, Experimental; Excitatory Amino Acid Agonists; Gene Deletion; Humans; Mice; Mice, Transgenic; Monomeric GTP-Binding Proteins; Morpholines; Multiple Sclerosis; N-Methylaspartate; Naphthalenes; Nerve Degeneration; Nuclear Proteins; Receptors, Cannabinoid; Receptors, Drug; Receptors, N-Methyl-D-Aspartate; Retina; Saccharomyces cerevisiae Proteins; Spinal Cord; Uveitis

Acetylcholinesterase (AChE) was analyzed in organotypic hippocampal slice cultures (OHSCs) during recovery from a brief period (20 min) of combined hypoxia and hypoglycemia. Simulated ischemia transiently increased AChE activity in OHSCs in a time-dependent manner reaching a 1.5 fold increase at 6 h post-ischemia. The ischemia-induced AChE increase was totally abolished by incubation with 10 microM dizocilpine (MK-801), a neuroprotective NMDA receptor blocker.

Topics: Acetylcholine; Acetylcholinesterase; Animals; Animals, Newborn; Cell Death; Cerebral Infarction; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Hippocampus; Hypoxia-Ischemia, Brain; Nerve Degeneration; Organ Culture Techniques; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Up-Regulation

The intrahippocampal administration of 4-aminopyridine (4-AP) induces epileptic seizures and neurodegeneration, due probably to stimulation of glutamate release from synaptic terminals. We have studied the time course of the neurodegenerative changes produced by 4-AP, perfused through microdialysis cannulas in rat hippocampus, and correlated them with the expression of the inducible heat shock protein 70 (HSP70), detected immunocytochemically. Electroencephalographic seizure activity appeared immediately after the beginning of 4-AP perfusion. The first signs of histological neuronal damage were observed in CA1 and CA3 subfields of the perfused hippocampus 3 h after treatment and progressed until reaching a maximal neuronal loss at 24 h. In 4-AP-treated rats HSP70 was expressed mainly in neurons of the contralateral hippocampus, with a time course and cellular distribution very similar to the neurodegeneration observed in the perfused hippocampus, but no neuronal damage was observed. The N-methyl-D-aspartate (NMDA) receptor antagonists MK-801 and (3-phosphonopropyl)-piperazine-2-carboxylic acid prevented the seizures, the neurodegeneration and the expression of HSP70. These data demonstrate that the 4-AP-induced release of endogenous glutamate overactivates NMDA receptors in the perfused hippocampus and that the resulting neuronal hyperexcitability propagates to the contralateral hippocampus, generating a glutamate-mediated neuronal stress sufficient to induce the expression of HSP70 but not to produce neurodegeneration. These findings provide a useful model for investigating the relationships between neuronal hyperexcitation, neurodegeneration and the role of HSP expression.

Topics: 4-Aminopyridine; Animals; Anticonvulsants; Cell Count; Chromatography, High Pressure Liquid; Dizocilpine Maleate; Drug Interactions; Electroencephalography; Epilepsy; Extracellular Space; Functional Laterality; Gene Expression; Glutamic Acid; Hippocampus; HSP70 Heat-Shock Proteins; Immunohistochemistry; Male; Microdialysis; Nerve Degeneration; Neuroprotective Agents; Piperazines; Potassium Channel Blockers; Quinoxalines; Rats; Rats, Wistar; Somatosensory Cortex; Stress, Physiological; Time Factors

The strain and sex of a species under investigation may influence the animal's physiological response to a variety of stimuli. Strain and sex differences are important considerations when evaluating animal models. In the rodent MK-801 model of schizophrenia, degenerative changes occur widely in the main olfactory system and in a number of cortical brain regions. In the present report, we compare the effects of MK-801 neurotoxicity in two strains of female rats and also two lines within each strain. The magnitude and regional extent of the neurodegeneration detected with the amino-cupric-silver method varied markedly both between the Sprague-Dawley and Wistar rat strains and also between two lines derived from each strain. For example, terminal degeneration occurred in layer VI of somatosensory cortex and the central extended amygdala in Sprague-Dawley but not Wistar rats. Moreover, MK-801 treatment led to somatodendritic degeneration in the dentate gyrus of the dorsal hippocampus and basolateral amygdala in Wistar rats from Charles River Laboratories but not those from Ferreyra Institute. There are thus both strain and intrastrain differences in the magnitude of the neurodegenerative response to MK-801 treatment. The differing neurotoxicity of MK-801 between rat strains and between lines within a strain may reflect genetic variation and/or differences in hepatic biotransformation and thus the bioavailability of the drug between strains and lines within a strain.

Topics: Animals; Brain; Brain Diseases; Copper; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Genetic Predisposition to Disease; Nerve Degeneration; Rats; Rats, Sprague-Dawley; Rats, Wistar; Silver; Species Specificity; Staining and Labeling

Staurosporine-induced apoptosis was associated with a 20% cellular survival rate in primary rat forebrain cultures. Treatment with the NR2B subunit-selective NMDA receptor antagonist conantokin-G (0.1-1 microM) increased the survival rate up to 78%. No protection was provided by the nonselective NMDA receptor antagonist dizocilpine (0.01-10 microM) but 34-64% cellular survival was provided by ifenprodil (0.01-10 microM), another NR2B subunit-selective antagonist. These results suggest a novel anti-apoptotic mechanism linked to the NR2B receptor subunit.

Topics: Animals; Apoptosis; Cell Survival; Cells, Cultured; Conotoxins; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Nerve Degeneration; Piperidines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Staurosporine

We have recently shown that focal administration of dizocilpine hydrogen maleate (MK-801, a non-competitive N-methyl-D-aspartate antagonist) within the nucleus accumbens increases locomotor activity in a dopamine-independent manner. The purpose of this study was to investigate the neural network underlying locomotor stimulation induced by N-methyl-D-aspartate receptor blockade in the accumbens. In the first experiment, we examined the effect of different doses (1, 5 and 25 nmol) of the active and inactive enantiomers of the N-methyl-D-aspartate antagonist, (+)- and (-)-MK-801, respectively, focally administered in the nucleus accumbens. Only the active enantiomer induced a significant increase in locomotor activity; furthermore, the effect induced by the two highest doses of (+)-MK-801 was significantly different from that induced by (-)-MK-801. In the second part of the study, we performed ibotenic acid lesions to the major output nuclei of the accumbens, the ventral pallidum, mediodorsal thalamus, ventrolateral/ventromedial thalamus and pedunculopontine tegmental nucleus, to observe their effect on locomotor activity induced by focal (+)-MK-801 (25 nmol) administration into the accumbens. None of the lesions had any effect on spontaneous locomotor activity. Hyperactivity induced by accumbens MK-801 administrations was unaffected by ibotenic acid lesions of the pedunculopontine tegmental nucleus, while lesions of the mediodorsal thalamus induced only a partial inhibition. In contrast, ibotenic acid lesions of the ventral pallidum and ventrolateral/ventromedial thalamus completely blocked the motor response induced by accumbens MK-801. These data indicate that the intact mediodorsal thalamus, which has been proposed as a part of the loop that relays accumbens information to the prefrontal cortex, does not seem to be a structure of primary importance in MK-801 locomotor activity. On the contrary, the motor nuclei of the thalamus appear to play a more relevant role, suggesting that different neural substrates may mediate dopamine and glutamate functional output from the nucleus accumbens.

Topics: Animals; Denervation; Dizocilpine Maleate; Dose-Response Relationship, Drug; Efferent Pathways; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Globus Pallidus; Ibotenic Acid; Male; Mediodorsal Thalamic Nucleus; Motor Activity; Nerve Degeneration; Nerve Net; Neurons; Nucleus Accumbens; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Reticular Formation; Ventral Thalamic Nuclei

The developing cortical neurons have been well documented to be extremely vulnerable to the toxic effect of methylmercury (MeHg). In the present study, a possible involvement of N-methyl-D-aspartate (NMDA) receptors in MeHg neurotoxicity was examined because the sensitivity of cortical neurons to NMDA neurotoxicity has a similar developmental profile. Rats on postnatal day 2 (P2), P16, and P60 were orally administered MeHg (10 mg/kg) for 7 consecutive days. The most severe neuronal damage was observed in the occipital cortex of P16 rats. When MK-801 (0.1 mg/kg), a non-competitive antagonist of NMDA, was administered intraperitoneally with MeHg, MeHg-induced neurodegeneration was markedly ameliorated. Furthermore, there was a marked accumulation of nitrotyrosine, a reaction product of peroxynitrite and L-tyrosine, after chronic treatment of MeHg in the occipital cortex of P16 rats. The accumulation of nitrotyrosine was also significantly suppressed by MK-801. In the present electrophysiological study, the amplitude of synaptic responses mediated by NMDA receptors recorded in cortical neurons of P16 rats was significantly larger than those from P2 and P60 rats. These observations strongly suggest that a generation of peroxynitrite through activation of NMDA receptors is a major causal factor for MeHg neurotoxicity in the developing cortical neurons. Furthermore, enhanced sensitivity of NMDA receptors may make the cortical neurons of P16 rats most susceptible to MeHg neurotoxicity.

Topics: Age Factors; Animals; Animals, Newborn; Antigens, CD; Antigens, Neoplasm; Antigens, Surface; Avian Proteins; Basigin; Blood Proteins; Cerebral Cortex; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Glutamic Acid; Membrane Glycoproteins; Mercury Poisoning, Nervous System; Methylmercury Compounds; Nerve Degeneration; Neurons; Nitrates; Quinoxalines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Tyrosine

Proinflammatory cytokines are supposed to be involved in the pathophysiology of neuronal damage following excitotoxic lesions. We examined the effect of rolipram, a TNF-alpha-inhibitor, on excitotoxic neuronal damage. Quinolinic acid (240 nmol in 1 microl) was injected stereotactically into the striatum of male Wistar rats. Four groups of QA rats were treated i.p. with solvent, MK-801 (4 mg/kg) or rolipram (0.3 mg/kg) which was started either 6 or 24 h after QA injection and continued with daily applications for 14 days. QA injection induced neuronal damage which affected 93% of the striatal area. MK-801 reduced this damage to 12% of the striatal area. Treatment with rolipram when started at 6 h after QA injection resulted in neuronal damage amounting to 60%; the result after starting at 24 h was not different from solvent (91%). The present results demonstrate that rolipram reduces neuronal damage induced by intrastriatal QA application.

Topics: Animals; Antidepressive Agents; Cell Count; Dizocilpine Maleate; Drug Interactions; Excitatory Amino Acid Antagonists; Immunohistochemistry; Male; Neostriatum; Nerve Degeneration; Neurons; Neuroprotective Agents; Neurotoxins; Quinolinic Acid; Rats; Rats, Wistar; Rolipram; Tumor Necrosis Factor-alpha

The neurochemical alterations associated with neurodegeneration in prion diseases are not well defined. It is therefore of interest to study the influence of prion infection on messenger molecules and their receptors. In the present study we have analyzed the possible involvement of NMDA receptors in prion-infected mice using ligand binding autoradiography and iodinated MK-801, a noncompetitive NMDA antagonist. The results show a reduced binding of MK-801 in discrete regions of hippocampus at 110 days after infection, that is before the appearance of behavioral symptoms. In addition, early transient increases in MK-801 binding were observed in several layers. The exact neuroanatomical correlate of these changes in MK-801 binding, as well as its functional significance in relation to prion symptomatology, remain to be analyzed.

Topics: Animals; Autoradiography; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Hippocampus; Male; Mice; Mice, Inbred Strains; Nerve Degeneration; Prions; Receptors, N-Methyl-D-Aspartate; Scrapie

Retinal cell death induced by over-stimulation of glutamate receptors is related to the programmed cell death or apoptosis. However, little is known about the intracellular events that lead to this cell death process in the retina. In this study, we asked if caspase-3 family cysteine proteases regulate cell death in an explant culture of adult rat retina after exposure to excessive glutamate. Cells with DNA fragmentation were first detected in the ganglion cell layer 3 h after a brief exposure to 20 mM glutamate; whilst those in the inner nuclear layer were first observed 6 h after the glutamate lesion. Caspase-3-like activity, as indicated by immunostaining of the fractin antibody that recognizes actin fragments generated by caspase-3 family proteases, was seen 40 min after glutamate treatment. Staining was first detected in the ganglion cell layer and then in the inner nuclear layer, preceding the appearance of cells with DNA fragmentation in these layers. Colocalization study showed that all cells with DNA breaks were fractin positive, indicating that caspase-3 family activity was involved in the glutamate-induced cell death in the adult rat retina. Furthermore, DEVD-CHO, a tetrapeptide inhibitor for caspase-3 family members, reduced dramatically the fractin staining and significantly alleviated glutamate-induced cell death and DNA fragmentation in the ganglion cell layer and inner nuclear layer. Inhibitor for caspase-1-like activity, YVAD-CHO, neither reduced the fractin staining nor showed comparable neuroprotective effects to the retina. We conclude that glutamate-induced apoptotic cell death in adult rat retina is mediated by a specific activation of cysteine proteases related to the caspase-3 family, and an intervention to the caspase-3 proteases provides effective protection to retinal neurons against glutamate excitotoxicity.

Topics: Animals; Apoptosis; Carbocyanines; Caspase 1; Caspase 3; Caspase Inhibitors; Caspases; Cells, Cultured; Cysteine Proteinase Inhibitors; Dizocilpine Maleate; DNA Fragmentation; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Female; Fluorescent Dyes; Glutamic Acid; In Situ Nick-End Labeling; Nerve Degeneration; Oligopeptides; Rats; Rats, Sprague-Dawley; Retina; Retinal Ganglion Cells

N-Methyl-D-aspartate (NMDA) antagonists act by an anti-excitotoxic action to provide neuroprotection against acute brain injury, but these agents can also cause toxic effects. In low doses they induce reversible neuronal injury, but in higher doses they cause irreversible degeneration of cerebrocortical neurons. GABAmimetic drugs protect against the reversible neurotoxic changes in rat brain. Here we show that two GABAmimetic anesthetic agents--propofol and sodium thiopental--protect against the irreversible neurodegenerative reaction induced by the powerful NMDA antagonist, MK-801.

Topics: Anesthetics, Intravenous; Animals; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Interactions; Excitatory Amino Acid Antagonists; Female; GABA Agonists; gamma-Aminobutyric Acid; Glutamic Acid; Gyrus Cinguli; Necrosis; Nerve Degeneration; Neurons; Neuroprotective Agents; Neurotoxins; Propofol; Rats; Rats, Sprague-Dawley; Receptors, GABA-A; Thiopental

Endogenous cannabinoid receptor ligands (endocannabinoids) may rescue neurons from glutamate excitotoxicity. As these substances also accumulate in cultured immature neurons following neuronal damage, elevated endocannabinoid concentrations may be interpreted as a putative neuroprotective response. However, it is not known how glutamatergic insults affect in vivo endocannabinoid homeostasis, i.e. N-arachidonoylethanolamine (anandamide) and 2-arachidonoylglycerol (2-AG), as well as other constituents of their lipid families, N-acylethanolamines (NAEs) and 2-monoacylglycerols (2-MAGs), respectively. Here we employed three in vivo neonatal rat models characterized by widespread neurodegeneration as a consequence of altered glutamatergic neurotransmission and assessed changes in endocannabinoid homeostasis. A 46-fold increase of cortical NAE concentrations (anandamide, 13-fold) was noted 24 h after intracerebral NMDA injection, while less severe insults triggered by mild concussive head trauma or NMDA receptor blockade produced a less pronounced NAE accumulation. By contrast, levels of 2-AG and other 2-MAGs were virtually unaffected by the insults employed, rendering it likely that key enzymes in biosynthetic pathways of the two different endocannabinoid structures are not equally associated to intracellular events that cause neuronal damage in vivo. Analysis of cannabinoid CB(1) receptor mRNA expression and binding capacity revealed that cortical subfields exhibited an up-regulation of these parameters following mild concussive head trauma and exposure to NMDA receptor blockade. This may suggest that mild to moderate brain injury may trigger elevated endocannabinoid activity via concomitant increase of anandamide levels, but not 2-AG, and CB(1) receptor density.

Topics: Animals; Arachidonic Acids; Brain Concussion; Cannabinoid Receptor Modulators; Cerebral Cortex; Corpus Striatum; Craniocerebral Trauma; Dizocilpine Maleate; Endocannabinoids; Ethanolamines; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Glycerides; Male; N-Methylaspartate; Nerve Degeneration; Polyunsaturated Alkamides; Rats; Rats, Sprague-Dawley; Receptors, Cannabinoid; Receptors, Drug; RNA, Messenger

The selective 5-HT(1A) receptor agonist Repinotan HCl (BAY x3702) has been reported to attenuate cortical damage and improve functional performance in experimental models of cerebral ischemia and acute subdural hematoma. Using a clinically relevant contusion model of traumatic brain injury, we tested the hypothesis that a 4-h continuous infusion of Repinotan HCl (10 microg/kg/h i.v.) commencing 5 min post-injury would ameliorate functional outcome and attenuate histopathology. Forty isoflurane-anesthetized male adult rats were randomly assigned to receive either a controlled cortical impact (2.7 mm tissue deformation, 4 m/s) or sham injury (Injury/Vehicle=10, Injury/MK-801=10, Injury/Repinotan HCl=10, Sham/Vehicle=10), then tested for vestibulomotor function on post-operative days 1-5 and for spatial learning on days 14-18. Neither Repinotan HCl nor the non-competitive N-methyl-D-aspartate receptor antagonist MK-801, which served as a positive control, improved vestibulomotor function on beam balance and beam walk tasks relative to the Injury/Vehicle group, but both did significantly attenuate spatial learning and memory deficits on a water maze task. Repinotan HCl also reduced hippocampal CA(1) and CA(3) neuronal loss, as well as cortical tissue damage, compared to the Injury/Vehicle group at 4 weeks post-trauma. No significant difference in histological outcome was revealed between the Repinotan HCl- and MK-801-treated groups.These findings extend the therapeutic efficacy of Repinotan HCl to a contusion model of experimental brain injury and demonstrate for the first time that 5-HT(1A) receptor agonists confer neuroprotection and attenuate spatial learning deficits following controlled cortical impact injury. This treatment strategy may be beneficial in a clinical context where memory impairments are common following human traumatic brain injury.

Topics: Animals; Benzopyrans; Body Temperature; Brain; Brain Injuries; Cognition; Cognition Disorders; Dizocilpine Maleate; Drug Administration Schedule; Excitatory Amino Acid Antagonists; Hippocampus; Male; Maze Learning; Nerve Degeneration; Neurons; Neuroprotective Agents; Postural Balance; Rats; Rats, Sprague-Dawley; Receptors, Serotonin; Receptors, Serotonin, 5-HT1; Serotonin Receptor Agonists; Thiazoles; Vestibular Nuclei

1. Administration of 3,4-methylenedioxymethamphetamine (MDMA, 'ecstasy') to mice produces acute hyperthermia and long-term degeneration of striatal dopamine nerve terminals. Attenuation of the hyperthermia decreases the neurodegeneration. We have investigated the mechanisms involved in producing the neurotoxic loss of striatal dopamine. 2. MDMA produced a dose-dependent loss in striatal dopamine concentration 7 days later with 3 doses of 25 mg kg(-1) (3 h apart) producing a 70% loss. 3. Pretreatment 30 min before each MDMA dose with either of the N-methyl-D-aspartate antagonists AR-R15896AR (20, 5, 5 mg kg(-1)) or MK-801 (0.5 mg kg(-1)x3) failed to provide neuroprotection. 4. Pretreatment with clomethiazole (50 mg kg(-1)x3) was similarly ineffective in protecting against MDMA-induced dopamine loss. 5. The free radical trapping compound PBN (150 mg kg(-1)x3) was neuroprotective, but it proved impossible to separate neuroprotection from a hypothermic effect on body temperature. 6. Pretreatment with the nitric oxide synthase (NOS) inhibitor 7-NI (50 mg kg(-1)x3) produced neuroprotection, but also significant hypothermia. Two other NOS inhibitors, S-methyl-L-thiocitrulline (10 mg kg(-1)x3) and AR-R17477AR (5 mg kg(-1)x3), provided significant neuroprotection and had little effect on MDMA-induced hyperthermia. 7. MDMA (20 mg kg(-1)) increased 2,3-dihydroxybenzoic acid formation from salicylic acid perfused through a microdialysis tube implanted in the striatum, indicating increased free radical formation. This increase was prevented by AR-R17477AR administration. Since AR-R17477AR was also found to have no radical trapping activity this result suggests that MDMA-induced neurotoxicity results from MDMA or dopamine metabolites producing radicals that combine with NO to form tissue-damaging peroxynitrites.

Topics: Animals; Chlormethiazole; Corpus Striatum; Cyclic N-Oxides; Dizocilpine Maleate; Dopamine; Dose-Response Relationship, Drug; Enzyme Inhibitors; Fever; Free Radicals; Hydroxybenzoates; Injections, Intraperitoneal; Male; Mice; Microdialysis; N-Methyl-3,4-methylenedioxyamphetamine; Nerve Degeneration; Neurons; Neuroprotective Agents; Nitric Oxide Synthase; Nitrogen Oxides; Pyridines; Salicylic Acid

The neuroactive steroids pregnenolone (3beta-hydroxy-5-pregnen-20-one) and dehydroepiandrosterone (DHEA, 3alpha-hydroxy-5-androstene-17-one) are negative allosteric modulators of the GABA(A) receptors and positive modulators of acetylcholine, NMDA and sigma(1) receptors. Pregnenolone was recently shown to potentiate the neuronal damage induced by excessive glutamate in cell culture models, whereas dehydroepiandrosterone was reported to present some neuroprotective activity. The in vivo relevance of these effects was investigated in mice submitted to an hypoxic insult, the repeated exposure to carbon monoxide (CO) gas, a model that leads to neurodegeneration in the CA(1) hippocampal area and learning deficits. Recording spontaneous alternation behaviour in the Y-maze assessed short-term memory and long-term memory was examined using a passive avoidance task. After exposure to CO, mice showed a progressive deterioration of their learning ability, reaching significance after 3 days and being maximal after 7 days. Pregnenolone administered before CO significantly facilitated the hypoxia-related deficits, which could be measured 1 day after CO and appeared maximal after 3 days. Dizocilpine blocked the deficits in vehicle- and pregnenolone-treated CO-exposed animals, showing that pregnenolone selectively facilitated the NMDA receptor-dependent excitotoxicity. Dehydroepiandrosterone blocked the appearance of the CO-induced deficits, even after 7 days. Interestingly, the sigma(1) receptor antagonist N, N-dipropyl-2-(4-methoxy-3-(2-phenylethoxy)phenyl)ethylamine (NE-100) failed to affect the dehydroepiandrosterone-induced protection, showing the lack of involvement of sigma(1) receptors. Cresyl violet-stained sections of the mouse hippocampal formation showed that the neurodegeneration observed in the CA(1) area after exposure to CO was augmented by pregnenolone and blocked by dehydroepiandrosterone. These results show that pregnenolone and dehydroepiandrosterone, although being similarly involved in modulating the excitatory/inhibitory balance in the brain, do not equally affect the extent of excitotoxic insults.

Topics: Animals; Anisoles; Antipsychotic Agents; Behavior, Animal; Carbon Monoxide Poisoning; Dehydroepiandrosterone; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hippocampus; Learning; Male; Memory; Memory, Short-Term; Mice; Nerve Degeneration; Pregnenolone; Propylamines

Non-competitive N-methyl-D-aspartate (NMDA) receptor antagonists, ketamine, phencyclidine (PCP) and dizocilpine (MK-801), produce psychosis in people. In rodents they produce cytoplasmic vacuoles in injured retrosplenial cortical neurons that express HSP70 heat shock protein. This study examined possible circuits and receptors that mediate this neuronal injury. Bilateral, but not unilateral, injection of dizocilpine (5, 10, 15, 20 microg/microL per side) into the anterior thalamus induced HSP70 protein in pyramidal neurons in deep layer III of rat retrosplenial cortex 24 h later. In contrast, bilateral dizocilpine injections (5, 10, 15, 20 microg/microL per side) into the retrosplenial cortex or into the diagonal band of Broca did not induce HSP70. Bilateral injections of muscimol (0.1, 1, 10 microg/microL per side), a GABAA (gamma-aminobutyric acid) agonist, into the anterior thalamus blocked HSP70 induction in the retrosplenial cortex produced by systemic dizocilpine (1 mg/kg). Bilateral thalamic injections of baclofen (0.1, 1, 10 microg/microL per side), a GABAB agonist, were ineffective. Anterograde tracer studies confirmed that neurons in the anterior thalamus project to superficial layer III of the retrosplenial cortex where the dendrites of HSP70-immunostained neurons in deep layer III reside. Bilateral blockade of NMDA receptors on GABA neurons in the reticular nuclei of the thalamus is proposed to decrease GABA neuronal firing, decrease GABA release and decrease activation of GABAA receptors. This activates thalamic projection neurons that damage retrosplenial cortical neurons presumably via unblocked cortical glutamate alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionate (AMPA) and kainate receptors. The increases of blood flow that occur in the thalamus and retrosplenial cortex of people that have psychosis produced by NMDA antagonists could be related to thalamic excitation of the retrosplenial cortex produced by these drugs.

Topics: Animals; Anterior Thalamic Nuclei; Baclofen; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; GABA Agonists; gamma-Aminobutyric Acid; Glutamic Acid; Gyrus Cinguli; HSP70 Heat-Shock Proteins; Muscimol; Nerve Degeneration; Neural Pathways; Phytohemagglutinins; Pyramidal Cells; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Schizophrenia

Whereas a cardinal role for beta-amyloid protein (Abeta) has been postulated as a major trigger of neuronal injury in Alzheimer's disease, the pathogenic mechanism by which Abeta deranges nerve cells remains largely elusive. Here we report correlative in vitro and in vivo evidence that an excitotoxic cascade mediates Abeta neurotoxicity in the rat magnocellular nucleus basalis (MBN). In vitro application of Abeta to astrocytes elicits rapid depolarization of astroglial membranes with a concomitant inhibition of glutamate uptake. In vivo Abeta infusion by way of microdialysis in the MBN revealed peak extracellular concentrations of excitatory amino acid neurotransmitters within 20-30 min. Abeta-triggered extracellular elevation of excitatory amino acids coincided with a significantly enhanced intracellular accumulation of Ca2+ in the Abeta injection area, as was demonstrated by 45Ca2+ autoradiography. In consequence of these acute processes delayed cell death in the MBN and persistent loss of cholinergic fibre projections to the neocortex appear as early as 3 days following the Abeta-induced toxic insult. Such a sequence of Abeta toxicity was effectively antagonized by the N-methyl-D-aspartate (NMDA) receptor ligand dizocilpine maleate (MK-801). Moreover, Abeta toxicity in the MBN decreases with advancing age that may be associated with the age-related loss of NMDA receptor expression in rats. In summary, the present results indicate that Abeta compromises neurons of the rat MBN via an excitotoxic pathway including astroglial depolarization, extracellular glutamate accumulation, NMDA receptor activation and an intracellular Ca2+ overload leading to cell death.

Topics: Alzheimer Disease; Amyloid beta-Peptides; Animals; Aspartic Acid; Astrocytes; Basal Nucleus of Meynert; Calcium; Calcium Radioisotopes; Cells, Cultured; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Microdialysis; Nerve Degeneration; Neuroprotective Agents; Neurotoxins; Rats; Receptors, N-Methyl-D-Aspartate; Taurine

Previous studies have utilized a lesion model of cortical injury that produces transient behavioral impairments to investigate the recovery of function process. To better understand the recovery process, it would be beneficial to use a lesion model that produces more severe, enduring, behavioral impairments. The purpose of experiment 1 was to validate whether large lesions of the sensorimotor cortex (SMC), which included the rostral forelimb and caudal forelimb regions, produced enduring behavioral deficits. Rats were given large unilateral electrolytic lesions of the SMC, administered either the N-methyl-D-aspartate (NMDA) antagonist, MK-801 or saline 16 h after injury, and tested on a battery of behavioral tests. Enduring behavioral deficits were observed, for at least 6 months, on two tests of forelimb placing while transient deficits were observed on the foot-fault and somatosensory neutralization tests. Administration of MK-801 facilitated recovery on the somatosensory neutralization test; however, it did not induce recovery on either forelimb placing test. A second experiment was performed to determine if earlier administration of MK-801, the NMDA antagonist magnesium chloride (MgCl(2)), or the anti-oxidant N-tert-butyl-alpha-phenylnitrone (PBN) could induce behavioral recovery in this chronic model. Treatment with these drugs induced behavioral recovery on the forelimb placing tests, whereas, the saline-treated rats did not show any signs of behavioral recovery for at least 3 months. Anatomical analysis of the striatum showed that MK-801 and MgCl(2) but not PBN reduced the extent of lesion-induced striatal atrophy. These results suggest that administration of MK-801, MgCl(2), or PBN shortly after cortical injury can induce recovery of function when recovery is otherwise not expected in un-treated rats.

Topics: Animals; Antioxidants; Brain Injuries; Cyclic N-Oxides; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Forelimb; Lameness, Animal; Magnesium Chloride; Male; Motor Activity; Motor Cortex; Nerve Degeneration; Neuroprotective Agents; Nitrogen Oxides; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Touch; Vibrissae

The cytokines interleukin-1 beta (IL-1 beta) and IL-1 receptor antagonist (IL-1ra) are rapidly induced in response to excitotoxic and ischemic brain damage. The aim of the present study was to investigate the influence of a non-competitive (dizocilpine maleate, MK-801) and a competitive ((R)-CPP) NMDA receptor antagonist on the transient cytokine expression in the rat brain induced by systemic kainic acid administration. Peripheral administration of kainic acid (10 mg/kg, i.p.) results in a transient expression of IL-1 beta and IL-1ra mRNA, mainly in microglia, in regions showing neurodegeneration such as the hippocampus, thalamus, amygdala, and certain cortical regions. In addition, a few neurons expressing IL-1ra mRNA were observed in the piriform cortex and amygdala following kainic acid injection. Administration of MK-801 (i.p.) 1 h prior to kainic acid injection reduced cytokine expression in all of these regions. MK-801 at 3.0 mg/kg decreased the IL-1 beta mRNA expression, blocked or decreased the IL-1ra mRNA expression, depending on the brain region. MK-801 at 5.0 mg/kg abolished IL-1ra mRNA expression in all of the regions, whereas the IL-1 beta mRNA expression was decreased or blocked, depending on the brain region, or the time point investigated. Peripheral administration of (R)-CPP (15 mg/kg, i.p.) 15 min prior to the kainic acid injection abolished the IL-1 beta mRNA expression. The IL-1ra mRNA expression was abolished in all regions except for a few neurons in the piriform cortex. The finding that NMDA receptor antagonists inhibit the IL-1 beta and IL-1ra mRNA synthesis induced by kainic acid suggests that NMDA receptor activation may be involved in triggering cytokine synthesis following excitotoxic brain damage.

Topics: Animals; Behavior, Animal; Dizocilpine Maleate; Epilepsies, Myoclonic; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Gene Expression; In Situ Hybridization; Interleukin 1 Receptor Antagonist Protein; Interleukin-1; Kainic Acid; Male; Microglia; Nerve Degeneration; Piperazines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Sialoglycoproteins

An experimental study was conducted to evaluate the effects of methylprednisolone and MK-801 after the compressive injury of spinal cord in rats.. To investigate the effect of methylprednisolone and non-competitive NMDA antagonist MK-801 in long-term functional outcome after spinal cord injury (SCI).. A randomized group A of Sprague-Dawley rats were treated with MK-801 (1.0 mg/kg, n=10; Group A) after a compression injury. A group of methylprednisolone (MP)-treated (30 mg/kg, n=10; Group B) and non-treated animals (n=9; Group C) were included for comparison. The functional motor outcome such as inclined plane (IP), toe spreading reflex (TSR), and modified Tarlov scale (TS) were measured in each animal at regular time points up to 8 weeks post-treatment. Histologically the injury site was scored in four groups and immunohistochemically Wallerian Degeneration (WD), astrocytosis and expression of beta-amyloid protein was identified.. In examining the IP data, no significant difference was recognized between the group means (P-value>0.5). For the TSR, there were no differences in the group responses. For the TS, the differences were not statistically significant. Only group B showed significance in cavitation scores compared to group A (P>0.0094), WD was significantly different than group C (P>0.03), astrocytosis was significantly higher than group A (P>0.001) and modest presence of beta-amyloid protein.. Our data indicate that one time bolus administration of MK-801 lacks any significant effect on axonal function in chronically injured rats. Daily bolus administration of MP at 30 mg/kg also did not ensure a better functional outcome. Immunohistochemically we have been able to show significant differences in WD, astrocytosis and small insignificant changes in beta-amyloid protein.

Topics: Amyloid beta-Peptides; Animals; Anti-Inflammatory Agents; Axons; Chronic Disease; Dizocilpine Maleate; Drug Therapy, Combination; Gliosis; Male; Methylprednisolone; Nerve Degeneration; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Recovery of Function; Spinal Cord; Spinal Cord Injuries

Programmed cell death (apoptosis) occurs during normal development of the central nervous system. However, the mechanisms that determine which neurons will succumb to apoptosis are poorly understood. Blockade of N-methyl-D-aspartate (NMDA) glutamate receptors for only a few hours during late fetal or early neonatal life triggered widespread apoptotic neurodegeneration in the developing rat brain, suggesting that the excitatory neurotransmitter glutamate, acting at NMDA receptors, controls neuronal survival. These findings may have relevance to human neurodevelopmental disorders involving prenatal (drug-abusing mothers) or postnatal (pediatric anesthesia) exposure to drugs that block NMDA receptors.

Topics: Animals; Apoptosis; Brain; Calcium Channel Blockers; Dizocilpine Maleate; Dopamine Antagonists; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Fetus; Haloperidol; Immunohistochemistry; In Situ Nick-End Labeling; Microscopy, Electron; Muscarinic Antagonists; Nerve Degeneration; Neurons; Quinoxalines; Rats; Receptors, N-Methyl-D-Aspartate; Scopolamine

Evidence has accumulated that Zn2+ plays a central role in neurodegenerative processes following brain injuries including ischaemia or epilepsy. In the present study, we examined patterns and possible mechanisms of Zn2+ neurotoxicity. Inclusion of 30-300 microM Zn2+ for 30 min caused neuronal necrosis apparent by cell body and mitochondrial swelling in cortical cell cultures. This Zn2+ neurotoxicity was not attenuated by antiapoptosis agents, inhibitors of protein synthesis or caspase. Blockade of glutamate receptors or nitric oxide synthase showed no beneficial effect against Zn2+ neurotoxicity. Interestingly, antioxidants, trolox or SKF38393, attenuated Zn(2+)-induced neuronal necrosis. Pretreatment with insulin or brain-derived neurotrophic factor increased the Zn(2+)-induced free radical injury. Kainate or AMPA facilitated Zn2+ entry and potentiated Zn2+ neurotoxicity in a way sensitive to trolox. Reactive oxygen species and lipid peroxidation were generated in the early phase of Zn2+ neurotoxicity. These findings indicate that entry and accumulation of Zn2+ result in generation of toxic free radicals and then cause necrotic neuronal degeneration under certain pathological conditions in the brain.

Topics: 2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine; 6-Cyano-7-nitroquinoxaline-2,3-dione; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Amino Acid Chloromethyl Ketones; Animals; Antioxidants; Apoptosis; Brain-Derived Neurotrophic Factor; Cells, Cultured; Cerebral Cortex; Chromans; Cysteine Proteinase Inhibitors; Dizocilpine Maleate; Dopamine Agonists; Drug Synergism; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Female; Free Radicals; Hypoglycemic Agents; Insulin; Kainic Acid; Lipid Peroxidation; Mice; Microscopy, Electron; Mitochondrial Swelling; Necrosis; Nerve Degeneration; Neurons; Neurotoxins; Oligopeptides; Oxidative Stress; Pregnancy; Zinc

Morbidity and mortality from head trauma is highest among children. No animal model mimicking traumatic brain injury in children has yet been established, and the mechanisms of neuronal degeneration after traumatic injury to the developing brain are not understood. In infant rats subjected to percussion head trauma, two types of brain damage could be characterized. The first type or primary damage evolved within 4 hr and occurred by an excitotoxic mechanism. The second type or secondary damage evolved within 6-24 hr and occurred by an apoptotic mechanism. Primary damage remained localized to the parietal cortex at the site of impact. Secondary damage affected distant sites such as the cingulate/retrosplenial cortex, subiculum, frontal cortex, thalamus and striatum. Secondary apoptotic damage was more severe than primary excitotoxic damage. Morphometric analysis demonstrated that the N-methyl-D-aspartate receptor antagonists 3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonate and dizocilpine protected against primary excitotoxic damage but increased severity of secondary apoptotic damage. 2-Sulfo-alpha-phenyl-N-tert-butyl-nitrone, a free radical scavenger, did not affect primary excitotoxic damage but mitigated apoptotic damage. These observations demonstrate that apoptosis and not excitotoxicity determine neuropathologic outcome after traumatic injury to the developing brain. Whereas free radical scavengers may prove useful in therapy of head trauma in children, N-methyl-D-aspartate antagonists should be avoided because of their propensity to increase severity of apoptotic damage.

Topics: Animals; Apoptosis; Brain; Brain Injuries; Caudate Nucleus; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Frontal Lobe; Gyrus Cinguli; In Situ Nick-End Labeling; N-Methylaspartate; Nerve Degeneration; Parietal Lobe; Piperazines; Rats; Rats, Wistar; Thalamus; Time Factors

Accumulated evidence has shown that apoptosis and necrosis contribute to neuronal death after ischemia. The present study was performed to study the temporal and spatial patterns of neuronal necrosis and apoptosis after ischemia in retina and to outline mechanisms underlying necrosis and apoptosis.. Retinal ischemia was induced by increasing intraocular pressure to a range of 160 mm Hg to 180 mm Hg for 90 minutes in adult rats. The patterns of neuronal cell death were determined using light and electron microscopy and were visualized by TdT-dUTP nick-end labeling (TUNEL). The mRNA expression profile of p53 was examined using reverse transcription-polymerase chain reaction (RT-PCR) and in situ hybridization histochemistry. Immunohistochemistry was performed using anti-p53, anti-microtubule associated protein-2, and anti-glial fibrillary acidic protein antibodies.. Within 4 hours after ischemia, neurons in the inner nuclear cell layer (INL) and ganglion cell layer (GCL) underwent marked necrosis, made apparent by swelling of the cell body and mitochondria, early fenestration of the plasma membrane, and irregularly scattered condensation of nuclear chromatin. After 3 days, the INL and GCL neurons showed further degeneration through apoptosis marked by cell body shrinkage, aggregation, and condensation of nuclear chromatin. Apoptotic neurons were also observed sparsely in the outer nuclear cell layer. Intravitreal injections of MK-801 prevented early neuronal degeneration after ischemia. Of note, mRNA and protein levels of p53, the tumor suppressor gene known to induce apoptosis, were increased in the retinal areas undergoing apoptosis 1 to 3 days after ischemic injury.. Ischemia produces the N-methyl-D-aspartate-mediated necrosis and slowly evolving apoptosis of neurons in the retina. The latter may depend on the expression of the p53 proapoptosis gene.

Topics: Animals; Apoptosis; Dizocilpine Maleate; DNA Primers; Excitatory Amino Acid Antagonists; Glial Fibrillary Acidic Protein; In Situ Hybridization; In Situ Nick-End Labeling; Ischemia; Male; Microtubule-Associated Proteins; Necrosis; Nerve Degeneration; Neurons; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Retina; Retinal Diseases; Retinal Vessels; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tumor Suppressor Protein p53

1. Spinal cord ischemia evoked a biphasic increase in CSF-Glu during 20 min of ischemia (40%) and at 2 hr after reperfusion (70%) in the nontreated group that was attenuated by all treated groups. But MK-801 (15 micrograms i.t.) did not affect the increased Glu at 2 hr (80%). 2. The argyrophilia observed in laminae II-V at 8 hr after reperfusion was attenuated by hypothermia (33 degrees C) and combination with MK-801, but the attenuation was less with MK-801. 3. Mild hypothermia attenuated the biphasic increase in CSF-Glu and corresponding development of neuronal damage after spinal cord ischemia. 4. Mild hypothermia with NMDA antagonism did not yield any further effects, suggesting that hypothermia itself plays a pivotal role in the protection.

Topics: Animals; Combined Modality Therapy; Denervation; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Hypothermia, Induced; Injections, Spinal; Male; Microdialysis; Nerve Degeneration; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Reperfusion Injury; Spinal Cord

Unilateral neurotoxin lesion of rat caudate-putamen and globus pallidus resulted in delayed, transneuronal degeneration of GABAergic substantia nigra pars reticulata neurons. To explore whether the disinhibition of endogenous glutamate excitatory input played a role in the degeneration of substantia nigra pars reticulata neurons, animals with unilateral striatal-pallidal lesions received three daily intraperitoneal injections of either dizocilpine maleate (MK-801, 1 or 10 mg/kg), an N-methyl-D-aspartate glutamate receptor blocker, or 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(f)quinoxaline (NBQX, 30 mg/kg), an alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptor blocker, that began 24 h after the striatal-pallidal neurotoxin lesion. Drug treatment affected neither the volume of the initial lesion nor the volume of striatal-pallidal glial fibrillary acidic protein immunoreactivity. Neuron number in the substantia nigra pars reticulata ipsilateral to the lesioned striatopallidum was reduced on average by 37% in untreated control rats, in low dose MK-801, and NBQX-treated rats (P<0.0001). However, in animals treated with high doses of MK-801 there was no difference in the number of neurons in the substantia nigra pars reticulata ipsilateral or contralateral to the neurotoxin lesion. These data demonstrate that dose-related treatment with N-methyl-D-aspartate glutamate receptor blockers protects substantia nigra pars reticulata neurons, and suggests that glutamatergic mechanisms play a role in delayed transneuronal degeneration.

Topics: Animals; Corpus Striatum; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; Excitatory Amino Acid Antagonists; Globus Pallidus; Ibotenic Acid; Male; Nerve Degeneration; Neurons; Neuroprotective Agents; Neurotoxins; Putamen; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Glutamate; Substantia Nigra

Administration of dopamine agonists to 6-hydroxydopamine (6-OHDA) lesioned rats enhances the rotational response to subsequent administration of dopamine agonist, an effect called 'priming'. Previously, we have shown that 6-OHDA rats primed with three injections of the D1/D2 dopamine agonist apomorphine (0.5 mg/kg) permitted a challenge with an otherwise inactive dose of the D2 agonist quinpirole (0.25 mg/kg) to elicit robust rotational behavior and to induce Fos expression in striatoentopeduncular neurons. In this study, the time-course and role of N-methyl-d-aspartate (NMDA) glutamate receptors on apomorphine-priming of these D2 responses were investigated. The enhanced rotational behavior and striatal Fos expression observed following challenge with quinpirole (0.25 mg/kg) peaked 1 day following the third apomorphine priming injection and persisted, in reduced form, for at least 4 months. Pretreatment with the NMDA antagonists MK-801 or 3-[(+)-2-carboxypiperazin-4-yl]-propyl-1-phosphonate (CPP) dose-dependently attenuated apomorphine-priming of quinpirole-mediated rotational behavior and striatal Fos induction compared to 6-OHDA rats primed with apomorphine alone. Taken together, these data suggest that priming of these D2-mediated responses in 6-OHDA rats develops rapidly, persists for several months, and is dependent on concomitant NMDA receptor stimulation. Since this priming effect resembles response fluctuations observed in patients with Parkinson's disease receiving long-term l-dihydroxyphenylalanine therapy, the results of the present study suggest that interventions that prevent the development of this enhanced response, such as NMDA antagonists, could prove useful in reducing the incidence these response fluctuations.

Topics: Animals; Apomorphine; Behavior, Animal; Brain Chemistry; Corpus Striatum; Dizocilpine Maleate; Dopamine Agonists; Excitatory Amino Acid Antagonists; Male; Nerve Degeneration; Oxidopamine; Piperazines; Proto-Oncogene Proteins c-fos; Quinpirole; Rats; Rats, Sprague-Dawley; Receptors, Dopamine D2; Receptors, N-Methyl-D-Aspartate; Rotation; Sympatholytics; Time Factors

Fimbria-fornix transection produces neuronal injury in the septum. This cellular pathology is characterized by somatodendritic vacuolar abnormalities in neurons. Because these cellular changes are reminiscent of some of the morphological abnormalities seen with glutamate receptor-mediated excitoxicity, we tested whether excitotoxic injury to the septal complex of adult rats mimics the degeneration observed within the dorsolateral septal nucleus and medial septal nucleus following fimbria-fornix transection. The septal complex was evaluated at various time-points (6 h to 14 days) by light and electron microscopy following unilateral injection of the N-methyl-D-aspartate receptor agonist quinolinate or the non-N-methyl-D-aspartate receptor agonist kainate, and the morphological changes observed were compared to those abnormalities in the medial septal nucleus and dorsolateral septal nucleus at three to 14 days after fimbria-fornix transection. The patterns of cytoplasmic abnormalities and vacuolar injury were morphologically similar in the somatodendritic compartment of neurons following excitotoxicity and axotomy paradigms. These similarities were most evident when comparing the persistently injured neurons in the penumbral regions of the excitotoxic lesions at one to 14 days recovery to neurons in the medial septal nucleus and dorsolateral septal nucleus at seven and 14 days after fimbria-fornix transection. Pretreatment with the N-methyl-D-aspartate receptor antagonist dizocilpine maleate prior to unilateral fimbria-fornix transection attenuated the somatodentritic vacuolar damage found within the ipsilateral dorsolateral and medial septal nuclei at 14 days recovery. Because glutamate is the principal transmitter of hippocampal efferents within the fimbria-fornix, we conclude that postsynaptic glutamate receptor activation participates in the evolution of septal neuron injury following fimbria-fornix transection. Thus, excitotoxicity is a possible mechanism for transneuronal degeneration following central nervous system axotomy.

Topics: Animals; Denervation; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Hippocampus; Kainic Acid; Male; Nerve Degeneration; Neuroprotective Agents; Neurotoxins; Quinolinic Acid; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Septum Pellucidum

Administration of the excitotoxin kainate produces seizure activity and selective neuronal death in various brain areas. We examined the degeneration pattern of hippocampal neurons following systemic injections of kainate in the hamster and the rat. As reported, treatment with kainate resulted in severe neuronal loss in the hilus and CA3 in the rat. While the hilar neurons were also highly vulnerable to kainate in the hamster, neurons in the CA1 area, but not CA3, were highly sensitive to kainate. In both animals, immunoreactivity to anti-p50 nuclear factor kappa B antibody was increased in nuclei of the hilar neurons within 4 h following administration of kainate. Kainate treatment also increased the nuclear factor kappa B immunoreactivity in hamster CA1 neurons and rat CA3 neurons 24 h later. Neurons showing intense nuclear factor kappa B signal were stained with acid fuchsin. Kainate also increased DNA binding activity of p50 and p65 nuclear factor kappa B in the nuclear extract of the hippocampal formation as analysed by electrophoretic mobility shift assay in the hamster, suggesting that activation of nuclear factor kappa B may contribute to kainate-induced hippocampal degeneration. Administration of 100 nmol dizocilpine maleate 3 h prior to kainate attenuated kainate-induced activation of nuclear factor kappa B and neuronal death in CA1 in the hamster. The present study provides evidence that the differential vulnerability of neurons in the rat and the hamster hippocampus to kainate is partly mediated by mechanisms involving N-methyl-D-aspartate-dependent activation of nuclear factor kappa B.

Topics: Animals; Cell Death; Cell Nucleus; Cricetinae; Dizocilpine Maleate; Epilepsy; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Female; Hippocampus; Kainic Acid; Mesocricetus; N-Methylaspartate; Nerve Degeneration; Neurons; Neurotoxins; NF-kappa B; Rats; Rats, Sprague-Dawley

Adult rats intubated with a single dose of ethanol (alcohol; approximately 5 g/kg) for 5 to 10 successive days incur neurodegeneration in the entorhinal cortex, dentate gyrus, and olfactory bulbs accompanied by cerebrocortical edema and electrolyte (Na+, K+) accumulation. The brain damage is not lessened by cotreatment with the NMDA receptor antagonist MK-801; also, as reported elsewhere, MK-801 as well as non-NMDA receptor and Ca2+ channel antagonists are not neuroprotective in a similar, but more compressed, intoxication protocol. However, cotreatment with the electrolyte transport inhibitor/diuretic furosemide reduces alcohol-dependent cerebrocortical damage by 75-85% while preventing brain hydration and electrolyte elevations; olfactory bulb neurodegeneration is not attenuated. In parallel in vitro studies, rat organotypic entorhinal/hippocampal slice cultures exposed to alcohol (50-200 mM) 15 h/day for 6 days, mirroring episodic intoxication in vivo, demonstrate concentration-related release of the cytotoxic indicator, lactate dehydrogenase. Analogous to the in vivo findings, furosemide blocks this alcohol-induced in vitro cytotoxicity. Our results showing neuroprotection by furosemide indicate that brain edema and swelling are essential events in the brain damage induced by episodic alcohol exposure. Furosemide and related agents might be useful as neuroprotective agents in alcohol abuse. We suggest that the neurodegeneration is elicited in part by edema-dependent oxidative stress, but the regional selectivity of the damage may be best explained by physical (mechanical) compression of the limbic cortex against the adjacent tympanic bulla and subsequent neuronal cytoskeletal collapse. A scheme for these apparently nonexcitotoxic metabolic and mechanical pathways initiated by repeated alcohol exposure is proposed.

Topics: Alcoholic Intoxication; Animals; Brain; Brain Edema; Dentate Gyrus; Diuretics; Dizocilpine Maleate; Entorhinal Cortex; Ethanol; Excitatory Amino Acid Antagonists; Furosemide; Male; Nerve Degeneration; Neuroprotective Agents; Olfactory Bulb; Organ Culture Techniques; Organ Specificity; Potassium; Rats; Rats, Sprague-Dawley; Sodium

N-methyl-D-aspartate (NMDA) glutamate receptors mediate critical components of cardiorespiratory control in anesthetized animals. The role of NMDA receptors in the ventilatory responses to peripheral and central chemoreceptor stimulation was investigated in conscious, freely behaving rats. Minute ventilation (VE) responses to 10% O2, 5% CO2, and increasing intravenous doses of sodium cyanide were measured in intact rats before and after intravenous administration of the NMDA receptor antagonist MK-801 (3 mg/kg). After MK-801, eupcapnic tidal volume (VT) decreased while frequency increased, resulting in a modest reduction in VE. Inspiratory time (TI) decreased, whereas expiratory time remained unchanged. The VE responses to hypercapnia were qualitatively similar in control and MK-801 conditions, with slight reductions in respiratory drive (VT/TI) after MK-801. In contrast, responses to hypoxia were markedly attenuated after MK-801 and were primarily due to reduced frequency changes, whereas VT was unaffected. Sodium cyanide doses associated with significant VE increases were 5 and 50 microg/kg before and after MK-801, respectively. Thus 1-log shift to the right of individual dose-response curves occurred with MK-801. Selective carotid body denervation reduced VE during hypoxia by 70%, and residual hypoxic ventilatory responses were abolished after MK-801. These findings suggest that, in conscious rats, carotid and other peripheral chemoreceptor-mediated hypoxic ventilatory responses are critically dependent on NMDA receptor activation and that NMDA receptor mechanisms are only modestly involved during hypercapnia.

Topics: Animals; Blood Gas Analysis; Carotid Body; Chemoreceptor Cells; Denervation; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hemodynamics; Hypercapnia; Male; Nerve Degeneration; Neurons, Afferent; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Respiratory Mechanics

Rats repeatedly intoxicated with alcohol (ethanol, three times daily) over a 4-day period display neuronal degeneration in the dentate gyrus; entorhinal, piriform, insular, orbital, and perirhinal cortices; and in the olfactory nerve fibers and terminals in the olfactory bulb. Postulating a role for excitotoxicity, we have attempted to prevent the degeneration using antagonists that are neuroprotective in this type of brain damage. In an initial study, continuous subcutaneous infusion of a high dose of the glutamate/NMDA receptor antagonist MK-801 (2 mg/kg/day) by itself caused extensive neuronal degeneration in several brain regions and severe behavioral intoxication that precluded survival if combined with high blood alcohol levels (approximately 300 mg/dl). Moreover, the lower, nonneurotoxic blood alcohol levels (approximately 150 mg/dl) that were compatible with survival worsened the MK-801-induced brain damage. In a subsequent experiment, daily intraperitoneal injections of a lower dose of MK-801 (1 mg/kg/day) resulted in no MK-801 toxicity and, when combined with neurotoxic levels of alcohol, no reduction in alcohol-induced neurotoxicity. Nimodipine, a voltage-gated Ca2+ channel blocker, reduced the neuronal damage in the dentate gyrus, but greatly increased it in the piriform cortex when administered intragastrically at 600 mg/kg/day; it provided no protection from alcohol-dependent degeneration when given intragastrically at 100 mg/kg/day. Continuous intracerebroventricular delivery of 0.24 to 0.29 mg/day of 6,7-dinitro-quinoxaline-2,3-dione, a glutamate/alpha-amino-3-hydroxy-5-methyl-4-isoxazole receptor antagonist, failed to diminish alcohol-dependent neuronal damage in any brain region. We conclude that brain damage from episodic "binge" alcohol intoxication is not primarily mediated by excitotoxic mechanisms, implying that other, nonexcitotoxic pathophysiological mechanisms, are involved. Furthermore, MK-801, far from protecting from the alcohol-induced damage, at high doses causes widespread neuropathology that is significantly potentiated by alcohol.

Topics: Alcoholic Intoxication; Animals; Brain; Calcium Channel Blockers; Dentate Gyrus; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Administration Schedule; Excitatory Amino Acid Antagonists; Male; Nerve Degeneration; Neurons; Neuroprotective Agents; Nimodipine; Quinoxalines; Rats; Rats, Sprague-Dawley

We examined the effects of nerve growth factor (NGF) on free radical neurotoxicity in striatal cell cultures. Following exposure to 30 microM Fe2+ or 1 mM L-buthionine-[S,R]-sulfoximine (BSO), an inhibitor of gamma-glutamylcysteine synthetase, striatal neurons underwent cell body swelling and then widespread death over the next 24 h. The degeneration was prevented by addition of 100 microM trolox, an antioxidant. Addition of 100 ng/ml BDNF beginning 12 h before Fe2+ or BSO potentiated necrosis of most striatal neurons after exposure to 10 microM Fe2+ or 1 mM BSO. In contrast, treatment with 100 ng/ml NGF selectively potentiated the oxidative degeneration of striatal cholinergic neurons. The present findings provide additional evidence that NGF, like other neurotrophins, can potentiate oxidative neuronal cell necrosis.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Acetylcholinesterase; Animals; Antioxidants; Brain-Derived Neurotrophic Factor; Buthionine Sulfoximine; Cells, Cultured; Chromans; Corpus Striatum; Cycloheximide; Dizocilpine Maleate; Drug Synergism; Fetus; Free Radicals; Iron; Necrosis; Nerve Degeneration; Nerve Growth Factors; Neuroglia; Neurons; Neurotoxins; Rats; Rats, Sprague-Dawley

The neuroprotective effects of lowering body temperature have been well documented in various models of neuronal injury. The present study investigated the effects a lower ambient or core body temperature would have on damage to striatal dopamine (DA) neurons produced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Mice received systemic MPTP treatment at two different temperatures, 4 degrees C and 22 degrees C. MPTP-treated mice maintained at 4 degrees C demonstrated (1) a greater hypothermic response, (2) a significant reduction in striatal DA content and tyrosine hydroxylase (TH) activity, and (3) significantly greater striatal 1-methyl-4-phenylpyridinium (MPP+) levels, as compared to mice dosed with MPTP at room temperature. Parallel studies with methamphetamine (METH) were conducted since temperature appears to play a pivotal role in the mediation of damage to DA neurons by this CNS stimulant in rodents. As previously reported, METH-induced hyperthermia and the subsequent loss of striatal DA content were attenuated in animals dosed at 4 degrees C. We also evaluated the effects a hypothermic state induced by pharmacological agents would have on striatal neurochemistry and MPP+ levels following MPTP treatment. Concurrent administration of MK-801 or 8-OHDPAT increased the striatal MPP+ levels following MPTP treatment. However, only 8-OHDPAT potentiated the MPTP-induced decrements of striatal DA content and TH activity; MK-801 did not affect MPTP decreases in these striatal markers of dopaminergic damage. Altogether, these findings indicate that temperature has a profound effect on striatal MPP+ levels and MPTP-induced damage to DA neurons in mice.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; 1-Methyl-4-phenylpyridinium; 8-Hydroxy-2-(di-n-propylamino)tetralin; Animals; Body Temperature; Corpus Striatum; Dizocilpine Maleate; Dopamine; Dopamine Agents; Excitatory Amino Acid Antagonists; Methamphetamine; Mice; MPTP Poisoning; Nerve Degeneration; Neurons; Neurotoxins; Serotonin Receptor Agonists; Tyrosine 3-Monooxygenase

We have recently established a rat substantia nigra (SN) slice preparation in which a sensitive index of excitatory amino acid (EAA) toxicity was degeneration of the dendritic arbor of catecholamine neurons labelled by immunostaining for tyrosine hydroxylase (TH). The present study examined the pharmacological characteristics of EAA-induced neurotoxicity. Rats were anesthetised by halothane inhalation and killed, the brain was rapidly removed, and 400-microm-thick SN slices cut in the horizontal plane on a vibratome. Slices were incubated in saline buffer at 35 degreesC for 15 min to 6 h in the presence or absence or absence of kainic acid (KA) or N-methyl-D-aspartate (NMDA) in concentrations ranging from 10 to 500 microM. The slices were then fixed and resectioned into 40-microm sections that were coplanar with the parent slice. Dopaminergic SN neurons were labeled using antibody to tyrosine hydroxylase (TH) coupled to diaminobenzidine. A feature of the immunostaining was that it labeled not only the cell body but also the prolific dendritic arborization of SN neurons. Dendritic damage was quantified by counting the proportion of neurons with intact dendrites after treatment with EAA. KA and NMDA caused loss of dendrites that was prevented by CNQX (20 microM) and MK-801 (20 microM), respectively, indicating that activation of either NMDA or non-NMDA receptors produces neurotoxicity. EAA-induced dendritic damage was observed within 2 h of treatment with a low concentration (10 microM) of KA and within 15 min if the concentration was increased to 500 microM. Thus the loss of dendrites occurs rapidly and precedes disintegration of the cell bodies. Furthermore, brief (15 min) exposure to EAA initiated damage in the dendrites which progressed after the EAA was removed from its receptor. The observations are consistent with the postulated role of EAAs in neurodegenerative diseases. Labeling the dendritic arbor provides a sensitive approach to investigating the cellular mechanisms of neurodegeneration of catecholamine neurons.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Catecholamines; Dendrites; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Kainic Acid; Male; N-Methylaspartate; Nerve Degeneration; Neurotoxins; Organ Culture Techniques; Rats; Rats, Sprague-Dawley; Substantia Nigra

Injury of a peripheral nerve gives rise to adaptive functional and structural alterations in spinal neurons. We report that the rearrangement of the spinal circuitry in response to sciatic nerve transection in adult rats involves a delayed mode of degeneration of lumbar spinal cord neurons. Nuclear fragmentation was detected by the TUNEL technique 7 days after sciatic neurectomy but not after 3 or 14 days. Dying cells were preferentially located in the ipsilateral superficial dorsal horn and expressed the neuronal cytoskeletal marker SMI-31. Degeneration was prevented by continuous systemic treatment with the NMDA receptor-antagonist MK-801. These data are supportive that apoptosis is induced in spinal neurons in a transsynaptic manner by an early signal from injured afferent fibres via activation of spinal NMDA receptors.

Topics: Animals; Denervation; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Genetic Techniques; Immunohistochemistry; Male; Nerve Degeneration; Neurofilament Proteins; Neurons; Phosphoproteins; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Sciatic Nerve; Spinal Cord; Time Factors; Wounds, Penetrating

The tumor promoter okadaic acid is a potent and specific inhibitor of protein phosphatases 1 and 2A and therefore it is a useful tool for studying the participation of protein phosphorylation in cellular processes. Since it has been shown that in cultured neurons OKA behaves as a potent neurotoxin, in the present work we have administered different doses of this compound into the dorsal rat hippocampus, in order to assess its neurotoxicity in vivo. Cresyl violet staining of hippocampal sections revealed that as early as 3 h after injection of 300 ng OKA a notable neurodegeneration occurred in the CA1 subfield, the dentate gyrus, and the hilus, particularly in the former. Neuronal death was more evident at 24 h and at this time the extent of damage was dose-dependent. The process of neuronal death was accompanied by a loss of the microtubule-associated protein MAP2, as assessed by immunocytochemistry. Moreover, OKA treatment resulted in a notable expression of the inducible heat shock protein 72 in the surviving neurons of the injected hippocampus and in the corresponding CA1 and hilus of the apparently normal contralateral hippocampus. The expression of the heat shock protein was partially prevented in the injected hippocampus and completely blocked in the contralateral CA1 region, by the systemic previous administration of the NMDA receptor antagonist MK-801. These results suggest that protein hyperphosphorylation due to inhibition of phosphatases in vivo induces neuronal stress and subsequent neurodegeneration.

Topics: Animals; Dentate Gyrus; Dizocilpine Maleate; Enzyme Inhibitors; Functional Laterality; Gene Expression Regulation; Heat-Shock Proteins; Hippocampus; Immunohistochemistry; Male; Nerve Degeneration; Neurons; Neuroprotective Agents; Neurotoxins; Okadaic Acid; Phosphoprotein Phosphatases; Rats; Rats, Wistar

Blockade of N-methyl-D-aspartate (NMDA) glutamate receptors by MK-801 induces neuronal degeneration in the posterior cingulate/retrosplenial cortex and other corticolimbic regions although damage in the latter has not been adequately characterized. This disseminated corticolimbic damage is of interest since NMDA hypofunction, the mechanism that triggers this neurodegenerative syndrome, has been postulated to play a role in the pathophysiology of Alzheimer's disease (AD). Several histological methods, including electron microscopy, were used to evaluate the neurotoxic changes in various corticolimbic regions of rat brain following MK-801 or a combination of MK-801 plus pilocarpine. We found that MK-801 triggers neuronal degeneration in a widespread pattern similar to that induced by phencyclidine and that females showed more damage than males. The neurotoxic reaction involved additional brain regions when muscarinic receptors were hyperactivated by administering pilocarpine with MK-801. Ultrastructural evaluation revealed that a major feature of the neurotoxic action involves degeneration of dendritic spines which entails loss of synaptic complexes. The ultrastructural appearance of degenerating neurons was generally inconsistent with an apoptotic mechanism, although evidence equivocally consistent with apoptosis was observed in some instances. The cell death process evolved relatively slowly and was still ongoing 7 days posttreatment. Relevance of these results to AD is discussed.

Topics: Alzheimer Disease; Animals; Cerebral Cortex; Dizocilpine Maleate; Drug Combinations; Excitatory Amino Acid Antagonists; Female; Limbic System; Male; Nerve Degeneration; Pilocarpine; Rats; Rats, Sprague-Dawley; Sex Characteristics; Time Factors

In acute brain injury syndromes, the potent N-methyl-D-aspartate (NMDA) antagonist, MK-801, can prevent neuronal degeneration, and the general anesthetics, isoflurane and propofol, may also provide neuroprotective benefits. An obstacle to the use of NMDA antagonists for neuroprotective purposes is that they can cause a neurotoxic vacuole reaction in cerebrocortical neurons. This study demonstrates the ability of isoflurane and propofol to prevent the neurotoxic vacuole reaction induced by MK-801. Low sedative doses of inhaled isoflurane (1%) or intravenous (i.v.) propofol (7.5 mg/kg/h) were as effective as higher general anesthetic doses. Thus, in the clinical management of acute brain injury conditions such as stroke and brain trauma, administration of one of these anesthetic agents together with an NMDA antagonist may be an excellent formula for obtaining optimal neuroprotection while eliminating serious side effects.

Topics: Anesthetics, Inhalation; Animals; Brain Damage, Chronic; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Gyrus Cinguli; Hallucinogens; Isoflurane; Nerve Degeneration; Neuroprotective Agents; Propofol; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

Previously, we have reported that intranigral infusions of malonate, an inhibitor of mitochondrial function, lead to the degeneration of the dopaminergic neurons of the nigrostriatal pathway that is mediated, at least in part, through NMDA receptor activation and nitric oxide formation. In the present study, unilateral focal infusions of malonate into the nucleus basalis magnocellularis (nbM) of male Sprague-Dawley rats (weighing 250-300 g) resulted in a dose-related depletion in ipsilateral cortical and amygdaloid choline acetyltransferase (ChAT) activity. Infusion of a 3 mumol dose of malonate into the nbM of vehicle-treated animals resulted in a 41 and 54% decrease in cortical and amygdaloid ChAT activity, respectively. Systemic pretreatment with lamotrigine (16 mg/kg, i.p.) and MK-801 (5 mg/kg, i.p.) attenuated the depletions in cortical and amygdaloid ChAT activity that resulted from an infusion of this dose of malonate into the nbM. Acetylcholinesterase (AChE) histochemistry of the nbM following focal infusion of malonate (3 mumol) showed a marked decrease in the number of AChE-positive neurons that was partially prevented by MK-801 pretreatment. Before examining the role of nitric oxide formation in malonate-induced toxicity, the ability of systemic administration of N omega-nitro-L-arginine (L-NA) to inhibit nitric oxide synthase (NOS) activity in the nbM and cerebellum was investigated. L-NA (2, 10, and 20 mg/kg, i.p.) produced a dose-related inhibition of nbM and cerebellar NOS activity that was maximal following a dose of 10 mg/kg L-NA. This level of NOS inhibition persisted for at least 13 h following L-NA (10 mg/kg) administration. Subsequently, the effect of L-NA pretreatment on malonate toxicity was evaluated. Following pretreatment with L-NA (2 and 10 mg/kg, i.p.), the toxic action of malonate on cortical and amygdaloid ChAT activity was not altered. In addition, infusion of a lower dose of malonate (2 mumol) into the nbM resulted in decreases in cortical and amygdaloid ChAT activity that were not altered by pretreatment with L-NA (2 and 10 mg/kg, i.p.). In 7-nitroindazole (7-NI; 25 and 50 mg/kg, i.p.)-pretreated animals, malonate (3 mumol) produced decreases in cortical and amygdaloid ChAT activity that were attenuated by both doses of 7-NI. Thus, malonate-induced destruction of the basal forebrain cholinergic neurons was attenuated by systemic pretreatment with lamotrigine, MK-801, and 7-NI but not by L-NA.

Topics: Animals; Dizocilpine Maleate; Enzyme Inhibitors; Indazoles; Lamotrigine; Male; Malonates; Nerve Degeneration; Neurons; Neuroprotective Agents; Nitric Oxide Synthase; Nitroarginine; Parasympathetic Nervous System; Prosencephalon; Rats; Rats, Sprague-Dawley; Triazines

Fluoro-Jade is an anionic fluorochrome capable of selectively staining degenerating neurons in brain slices. The histochemical application of Fluoro-Jade results in a simple, sensitive and reliable method for staining degenerating neurons and their processes. The technique will detect neuronal degeneration resulting from exposure to a variety of neurotoxic insults. Fluoro-Jade can be combined with other fluorescent methodologies including immunofluorescence, fluorescent axonal tract tracing, and fluorescent Nissl counterstaining. Compared to conventional methodologies, Fluoro-Jade is a more sensitive and definitive marker of neuronal degeneration than hematoxylin and eosin (H&E) or Nissl type stains, while being comparably sensitive yet considerably simpler and more reliable than suppressed silver techniques.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Animals; Antihypertensive Agents; Axons; Dizocilpine Maleate; Dopamine Agents; Eosine Yellowish-(YS); Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Eye Enucleation; Fluorescent Antibody Technique; Fluorescent Dyes; Hallucinogens; Hematoxylin; Ibogaine; Kainic Acid; Male; Metals; Nerve Degeneration; Nissl Bodies; Nitro Compounds; Phencyclidine; Propionates; Rats; Rats, Sprague-Dawley; Sensitivity and Specificity; Silver Staining; Stereotaxic Techniques

Apoptosis and necrosis are generally recognized as two distinct pathways of cell death, based on biochemical and morphological characteristics. Despite rapid advances in elucidating molecular mechanisms of cell death, little is known about the morphological progression of death in neurons and the relationship between different mechanisms of neuronal death and the resulting subcellular alterations. With excitotoxicity, a clinically relevant model of neuronal death, apoptotic DNA laddering and morphologic evidence of necrosis can occur simultaneously in the same region of adult brain. Here, we tested the hypothesis that activation of N-methyl-D-aspartic acid (NMDA) and non-NMDA glutamate receptors (GluR) results in a spectrum of morphologically distinct phenotypes of neuronal death, with apoptosis and necrosis as its endpoints. The ultrastructural morphologies of newborn and adult neurons at different times following intrastriatal injections of non-NMDA and NMDA GluR agonists were compared to apoptosis, as established during naturally occurring neuronal death in the developing rat brain. Excitotoxic neuronal death in newborn striatum was morphologically indistinguishable from developmental apoptosis. In the adult, non-NMDA receptor agonist-induced neuronal death was characterized by extensive chromatin condensation that was reminiscent of, but not identical to, apoptosis during normal development. In contrast, quinolinate, an NMDA receptor agonist, produced only minor chromatin clumping and rapid cytoplasmic disintegration, which is suggestive of necrosis. These findings support the concept that degenerative phenotypes of excitotoxically injured neurons are influenced by the degree of brain maturity and GluR subtype stimulation, independent of the severity of excitotoxic insult, along a morphological continuum or gradient ranging from apoptosis to necrosis.

Topics: Animals; Apoptosis; Brain; Cell Death; Dizocilpine Maleate; Kainic Acid; Male; Necrosis; Nerve Degeneration; Neurons; Neurotoxins; Quinolinic Acid; Rats; Rats, Sprague-Dawley; Receptors, Amino Acid; Receptors, N-Methyl-D-Aspartate

Neuronal calcium loading attributable to hypoxic/ischemic injury is believed to trigger neurotoxicity. We examined in organotypic hippocampal slice cultures whether artificially and reversibly enhancing the Ca2+ buffering capacity of neurons reduces the neurotoxic sequelae of oxygen-glucose deprivation (OGD), whether such manipulation has neurotoxic potential, and whether the mechanism underlying these effects is pre- or postsynaptic. Neurodegeneration caused over 24 hr by 60 min of OGD was triggered largely by NMDA receptor activation and was attenuated temporarily by pretreating the slices with cell-permeant Ca2+ buffers such as 1, 2 bis(2-aminophenoxy)ethane-N,N,N',N'-tetra-acetic acid acetoxymethyl ester (BAPTA-AM). This pretreatment produced a transient, reversible increase in intracellular buffer content as demonstrated autoradiographically using slices loaded with 14C-BAPTA-AM and by confocal imaging of slices loaded with the BAPTA-AM analog calcium green-acetoxymethyl ester (AM). The time courses of 14C-BAPTA retention and of neuronal survival after OGD were identical, indicating that increased buffer content is necessary for the observed protective effect. Protection by Ca2+ buffering originated presynaptically because BAPTA-AM was ineffective when endogenous transmitter release was bypassed by directly applying NMDA to the cultures, and because pretreatment with the low Ca2+ affinity buffer 2-aminophenol-N,N,O-triacetic acid acetoxymethyl ester, which attenuates excitatory transmitter release, attenuated neurodegeneration. Thus, in cultured hippocampal slices, enhancing neuronal Ca2+ buffering unequivocally attenuates or delays the onset of anoxic neurodegeneration, likely by attenuating the synaptic release of endogenous excitatory neurotransmitters (excitotoxicity).

Topics: Animals; Buffers; Calcium; Carbon Radioisotopes; Cell Death; Cell Hypoxia; Cell Survival; Cells, Cultured; Chelating Agents; Dizocilpine Maleate; Egtazic Acid; Excitatory Amino Acid Antagonists; Glucose; Hippocampus; Nerve Degeneration; Neurons; Neuroprotective Agents; Neurotoxins; Neurotransmitter Agents; Oxygen; Presynaptic Terminals; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate

Exposure of the rodent striatum to quinolinic acid (QA, N-methyl-D-aspartate receptor agonist) induces immediate early gene (IEG; c-fos, c-jun, jun-B, zif/268) expression that may extend 12-24 h after injection. In order to determine the specificity of the prolonged IEG response to the QA injection, the temporal pattern of c-fos mRNA expression was examined during the first 4 h after administration of saline or QA (40 micrograms). As early as 30 min after intrastriatal injection, both saline and QA increased c-fos mRNA levels. In the saline group, this increase in IEG expression was only transient and returned to baseline by 1 h. In contrast, c-fos mRNA levels within QA-injected animals continued to rise significantly at 1 and 4 h. In a second experiment, rats received 4 ng to 40-micrograms injections of QA followed by sacrifice at 6 h to determine if increasing QA doses caused the appearance of the prolonged IEG response phase. The prolonged IEG response was evident at 6 h only in animal groups that received higher dose ranges (4-40 micrograms) of QA. A final experiment was undertaken to determine if blockage of NMDA receptor stimulation would also inhibit the prolonged IEG response at 6 h in relationship to neuronal sparing evidenced at 24 h post-QA injection. The NMDA receptor antagonist, MK-801, blocked the prolonged IEG response at 6 h following QA (40 micrograms) injection while also preventing striatal neuropeptide mRNA decline by 24 h. Delaying the MK-801 administration for 1-2 h post-QA injection revealed that the intensity of the prolonged IEG mRNA response may be predictive of neuronal demise within the QA lesion site. These results suggest that prolonged IEG expression is associated with QA excitotoxicity of the rodent striatum and subsequent neuronal degeneration.

Topics: Animals; Calcium; Corpus Striatum; Dizocilpine Maleate; DNA-Binding Proteins; Early Growth Response Protein 1; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Gene Expression Regulation; Genes, fos; Genes, Immediate-Early; Genes, jun; Immediate-Early Proteins; Male; Nerve Degeneration; Nerve Tissue Proteins; Neurons; Neuropeptides; Neurotoxins; Proto-Oncogene Proteins c-fos; Proto-Oncogene Proteins c-jun; Quinolinic Acid; Rats; Rats, Inbred F344; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Signal Transduction; Transcription Factors; Transcription, Genetic

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

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a known neurotoxicant primarily selective for catecholaminergic neurons, including those of the nigrostriatal dopaminergic system, thereby mimicking the pathology of Parkinson's disease (PD). In this study, serial transbrain sectioning, followed by staining with a newly developed fluorochrome (Fluoro-Jade) specific for degenerating neurons, was used to detect additional sites of MPTP-induced neuronal degeneration in mice. Male CD-1 mice received a single 50 mg/kg dose of MPTP intraperitoneally at room temperature or at a reduced temperature (6 degrees C), which has been shown to potentiate striatal dopamine depletion. Neuronal degeneration was observed in the substantia nigra pars compacta (SN), ventral tegmental area (VTA) and retrorubral field (RRF) of only animals dosed in the low temperature environment. Neuronal degeneration was also observed in other catecholaminergic nuclei in both treatment groups. In addition, degenerating cell bodies and fibers were detected in the midline and intralaminar thalamic nuclei of all dosed animals, regardless of the dosing environment. Pharmacological manipulations which prevented nigral degeneration (deprenyl and nomifensine pretreatment) also prevented the degeneration of thalamic neurons. MK-801 pretreatment, however, resulted in a disproportionate protection of the thalamic neurons. These findings confirm and extend our previous observations regarding the protective effect of hyperthermia in CD-1 mice and also suggest that regions of the thalamus may be relevant to the pathophysiology of PD.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Animals; Dizocilpine Maleate; Dopamine Agents; Male; Mice; Mice, Inbred Strains; Nerve Degeneration; Neurons; Neuroprotective Agents; Nomifensine; Selegiline; Substantia Nigra; Temperature; Thalamus

Damage resulting from a partial acute lesion of white matter in the central nervous system (CNS) gradually spreads also to neurons that escaped the primary injury, resulting in their degeneration. Such spreading has been referred to as secondary degeneration. In order to demonstrate that this degeneration is indeed secondary to that caused by the acute insult, as well as to investigate the mechanism underlying the spread of damage and ways in which to protect neurons from such damage, we have proposed the use of partial lesion of the rodent optic nerve as a model. In this model we examined whether an antagonist of a receptor-mediated channel, shown to be beneficial in gray matter lesions, can protect neurons from undergoing secondary degeneration following white matter lesion. A well-calibrated partial crush lesion inflicted on the optic nerve of adult rats was immediately followed by a single intraperitoneal injection of the N-methyl-D-aspartate receptor antagonist, MK-801 (1 mg/kg). Protection of neurons from secondary degeneration was assessed by retrograde labeling and by measurement of the visual evoked potential (VEP) response to light. Two weeks after the injury, the mean number of neurons that were still intact was about threefold higher in the MK-801-treated group than in the saline-treated control group, indicating a treatment-induced protection of neurons that had escaped primary injury. A positive VEP response to light was obtained in 90% of the MK-801 treated animals and in only 50% of injured controls. The questions regarding whether the secondary degeneration of initially spared neurons starts in their cell bodies or in their axons, and consequently the identity of the primary site of their protection by MK-801, are discussed in relation to the absence of N-methyl-D-aspartate receptors on nerve fibers. The present findings may have implications for both acute and chronic injuries of the CNS.

Topics: Analysis of Variance; Animals; Body Temperature; Brain Injuries; Case-Control Studies; Disease Models, Animal; Dizocilpine Maleate; Evoked Potentials, Visual; Excitatory Amino Acid Antagonists; Male; Nerve Degeneration; Neural Pathways; Neuroprotective Agents; Optic Nerve; Optic Nerve Injuries; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

Single administration of a high dose of an uncompetitive NMDA receptor antagonist-dizocilpine maleate (MK-801)-results in transient neuronal vacuolization and cell death in retrosplenial cortex in rodents. In this study expression of cathepsin D (CatD), a major lysosomal aspartic protease, was investigated in brains of female rats treated with 1, 5, or 10 mg/kg of MK-801. Northern blot analysis demonstrated that the CatD mRNA level was moderately increased in retrosplenial cortex 24 h-7 days after the treatment. Concomitantly, increased CatD immunoreactivity was observed, predominantly in the degenerating neurons in layer III of retrosplenial cortex. Neuronal response was spatially distinguished from glial reactivation marked by increased mRNA and protein levels of glial fibrillary acidic protein, as demonstrated by Northern blot and immunohistochemistry in retrosplenial cortex 24 h-7 days after MK-801 treatment. These data suggest that activation of the lysosomal proteolytic system of neurons may play a role in MK-801-evoked neurodegeneration.

Topics: Animals; Astrocytes; Cathepsin D; Dizocilpine Maleate; Enzyme Induction; Excitatory Amino Acid Antagonists; Female; Glial Fibrillary Acidic Protein; Nerve Degeneration; Nerve Tissue Proteins; Neurons; Parietal Lobe; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Vacuoles

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

This study examined the effects of chronic intrastriatal infusion of L-trans-pyrrolidine-2,4-dicarboxylate (PDC), a selective competitive inhibitor of high affinity glutamate transport systems, via osmotic minipumps in rats. Injection of PDC at the rate of 25 nmol/h for 14 days caused striatal lesion. Histological evaluation on frontal striatal sections showed that the lesion was circumscribed to a circular area showing a dramatic neuronal loss accompanied by gliosis and representing 30% of the whole striatal surface at the level of the injection site. A total loss of neurons expressing glutamate decarboxylase (GAD67), enkephalin or substance P mRNA was observed on a similar circular area, suggesting degeneration of the two populations of striatal efferent neurons. In the whole striatum outside the region devoided of hybridization signal, a selective 27% decrease in enkephalin mRNA expression occurred, suggesting a higher sensitivity of enkephalin neurons versus substance P neurons to glutamate uptake-mediated alterations. Injection of PDC at the rate of 25 nmol/h for 3 days produced striatal lesion of similar extent. In contrast, PDC at the rate of 5 nmol/h did not produce neuronal damage when administered over 14 days. This study provides new in vivo evidence that defective glutamate transport is one of the critical conditions that may give rise to toxicity of an endogenous transmitter system in the striatum, and may underlie neuronal death in neurodegenerative diseases.

Topics: Animals; Biological Transport; Corpus Striatum; Dicarboxylic Acids; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Glutamic Acid; In Situ Hybridization; Infusion Pumps, Implantable; Molecular Weight; Nerve Degeneration; Neurotransmitter Uptake Inhibitors; Osmotic Pressure; Pyrrolidines; Rats; Rats, Wistar

The mechanisms of neuronal degeneration following traumatic head injury are not well understood and no adequate treatment is currently available for the prevention of traumatic brain damage in humans. Seven day old rat pups were subjected to mechanical percussion of the head. Cortical damage in infant rats was reduced by pre-treatment with the N-methyl-D-aspartate (NMDA) antagonists dizocilpine (MK-801) or 3-((+/-)-2-carboxypiperazin-4-yl)-propyl-I-phosphonate (CPP). The AMPA antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo (f) quinoxaline (NBQX) did not significantly suppress cortical damage in infant rats. In adult rats, traumatic head injury leads to primary (at impact-cortex) and secondary (distant-hippocampus) damage to the brain. Morphometric analysis demonstrated that both cortical and hippocampal damage was mitigated by pre-treatment with either the NMDA antagonist CPP or the non-NMDA antagonist NBQX. Neither treatment prevented primary damage in the cortex when therapy was started after trauma. Delayed treatment of rats with NBQX, but not with CPP, beginning between 1 and 7 h after trauma prevented the hippocampal damage. No protection was seen when therapy with NBQX was started 10 h after trauma. These data indicate that NMDA antagonists may possess better neuroprotective properties against excitotoxic processes triggered by traumatic brain injury in young individuals whereas AMPA antagonists may be more beneficial in adults.

Topics: Animals; Animals, Newborn; Brain Injuries; Cerebral Cortex; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hippocampus; Nerve Degeneration; Parietal Lobe; Rats

Neuroprotection against excitotoxicity by a combined therapy with the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 and the L-type Ca2+ channel blocker nimodipine was examined using an in vivo rat model of NMDA-induced neurodegeneration. Attention was focused on the neuroprotective potential of this combined drug treatment before and after NMDA-exposure. NMDA was unilaterally injected in the magnocellular nucleus basalis (MBN). Neuronal damage was assessed 12 days after the NMDA-injection by measuring the reduction of cholinergic cortical fibres that originate from the MBN neurons. In controls that received no drug treatment, NMDA-exposure damaged MBN neurons such that 66% of the cholinergic terminals were lost in the ipsilateral parietal cortex. Pretreatment with a nimodipine diet (860 ppm) combined with application of MK-801 (5 mg/kg i.p.) before NMDA-exposure reduced fibre loss by 89% thereby providing a near complete neuroprotection. Combined therapy of MK-801 (5 mg/kg i.p.) and nimodipine (15 mg/kg i.p.) 8 min after NMDA-infusion reduced neuronal injury by 82%, while the same combination given 2 h after the excitotoxic treatment still yielded a 66% protection against neurotoxic damage invoked by NMDA. In conclusion, the present data show that a dual blockade of NMDA-channels and voltage-dependent calcium channels (VDCC's) up to 2 h after NMDA-exposure is able to provide a significant protection against NMDA-neurotoxicity.

Topics: Aging; Analysis of Variance; Animals; Brain; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Interactions; Male; N-Methylaspartate; Nerve Degeneration; Nerve Fibers; Neurons; Nimodipine; Rats; Rats, Wistar

We investigated the involvement of glutamatergic neurotransmission in the modulation of D1 receptor-mediated stimulation of acetylcholine outflow in dorsal striatum in freely moving rats, and the relative roles of the thalamostriatal and corticostriatal pathways in this regulation using in vivo microdialysis. The selective N-methyl-D-aspartate non-competitive antagonist dizocilpine maleate (0.1 mg/kg i.p.), but not the kainate/quisqualate receptor antagonist, 6,7-dinitroquinoxaline-2,3-dione (3 micrograms per side i.c.v.), completely prevented the rise in striatal extracellular acetylcholine elicited by maximal effective doses of the full D1 agonist SKF 82958 (3 mg/kg s.c.) and of the dopamine releaser d-amphetamine (2 mg/kg s.c.). Acute bilateral electrolytic lesions of the parafascicular nucleus of the thalamus prevented the stimulation of striatal acetylcholine output by SKF 82958 and d-amphetamine but only slightly reduced basal acetylcholine release. In contrast acute interruption of the corticostriatal pathway did not alter the effect of the two dopaminergic drugs although it markedly reduced basal striatal acetylcholine release. Lesions of the parafascicular thalamic nucleus, or a low dose of dizocilpine maleate (0.1 mg/kg i.p.), also prevented the acetylcholine-increasing effect of the neuroleptic remoxipride (10 mg/kg s.c.), an effect known to be D1 receptor dependent. The results suggest that striatal projections arising from the parafascicular thalamic nucleus and utilizing N-methyl-D-aspartate receptors play a critical role in the D1-mediated stimulation of acetylcholine release in dorsal striata.

Topics: Acetylcholine; Animals; Benzazepines; Dextroamphetamine; Dizocilpine Maleate; Dopamine Agonists; Dopamine Antagonists; Dopamine Uptake Inhibitors; Excitatory Amino Acid Antagonists; Male; Microdialysis; N-Methylaspartate; Neostriatum; Nerve Degeneration; Neurons, Afferent; Prefrontal Cortex; Rats; Rats, Inbred Strains; Receptors, Dopamine D1; Receptors, N-Methyl-D-Aspartate; Remoxipride; Synaptic Transmission; Thalamic Nuclei

After partial transection of one optic nerve in adult cats the majority of beta retinal ganglion cells degenerate and die 1 week after axotomy, whilst other cell classes degenerate slowly and survive for a long period after the lesion. We have investigated the effects of intravitreal and intraperitoneal injections of MK-801, a NMDA-glutamate receptor antagonist, on the early degeneration of retinal ganglion cells after partial optic nerve section. Control animals received saline intravitreal injections. Retinal flat mounts were retrogradely labelled with horseradish peroxidase and counterstained with Cresyl Violet. We evaluated the ganglion cell loss in the three experimental groups 1 week after lesion and compared them with normal uninjured controls and injured untreated retinae. In untreated retinae 49% of ganglion cells die 1 week after the lesion. Systemic MK-801 or saline prolonged survival of 41% of retinal ganglion cells that would die without treatment. Intravitreal MK-801 or saline prolonged survival of 71% of retinal ganglion cells that would die without treatment, but the results of saline administration had a larger range of variability. In untreated retinae many pyknotic cells were observed. They decreased in number after systemic MK-801 treatment and in some retinae treated with intravitreal injections of saline solution. There were no pyknotic cells after local, intravitreal MK-801 treatment. These results support the hypothesis that NMDA-receptor mediated neurotoxicity plays an important role in the early retinal ganglion cell death after retrobulbar axotomy. They also support the existence of an endogenous source of neurotrophins whose release is triggered by eyeball injury. We conclude that the early death of beta retinal ganglion cells after axotomy occurs by a mechanism that can be controlled by neurotrophins and antagonists to NMDA-glutamate receptors.

Topics: Animals; Cats; Cell Survival; Dizocilpine Maleate; Female; Histocytochemistry; Horseradish Peroxidase; Male; Nerve Degeneration; Optic Nerve Injuries; Receptors, N-Methyl-D-Aspartate; Retinal Ganglion Cells

Several histological and behavioral experiments were conducted to investigate the neurotoxic effects of MK-801 in male mice. Moderate subcutaneous (s.c.) doses of MK-801 (0.5 and 1.0 mg/kg) induced the formation of intracytoplasmic vacuoles in pyramidal neurons in layers III and IV of the posterior cingulate/retrosplenial (PC/RS) cortex in 50% and 100% of the mice from the two respective treatment groups. Electron microscopic analysis of the vacuoles indicated that mitochondria and endoplasmic reticulum are the cellular organelles most prominently involved in this pathomorphological change. Treating mice with a high systemic dose of MK-801 (10 mg/kg s.c. or intraperitoneal (i.p.)) caused selective, irreversible degeneration of a small number of PC/RS cortical neurons. Compared to saline controls, the acquisition performance of mice treated i.p. with 10 mg/kg MK-801 was chronically impaired on a spatial learning task (modified hole board food search task) when tested at several posttreatment intervals (up to at least 5 months), although the groups did not differ on activity or sensorimotor tests conducted 2 weeks posttreatment. In summary, MK-801 caused histopathological changes in the mouse brain similar to those observed in the rat. Furthermore, high dose MK-801 treatment that killed a small number of mouse PC/RS cortical neurons resulted in a chronic acquisition impairment in spatial learning, an effect not previously demonstrated in any species.

Topics: Animals; Behavior, Animal; Brain; Dizocilpine Maleate; Dose-Response Relationship, Drug; Injections, Intraperitoneal; Injections, Subcutaneous; Learning; Male; Memory; Mice; Mice, Inbred ICR; Microscopy, Electron; Motor Activity; Nerve Degeneration; Paraffin Embedding; Rats; Space Perception

To study the sequence of degenerative events possibly associated with cholinergic cell death in Alzheimer's disease, septal cholinergic neurons derived from rat embryonic brains were exposed to chronic excitotoxic stress by glutamate. Counts of choline acetyltransferase (ChAT)-immunopositive neurons and measurement of ChAT activity revealed that concentrations of glutamate on the order of 70 microM killed 50% of cholinergic neurons after 24 hr of treatment. Neurotoxic effects were not aimed at cholinergic neurons specifically, since other populations of cells present in these cultures were also affected at similar concentrations. The noncompetitive N-methyl-D-aspartate (NMDA) receptor channel antagonist MK-801 (10 microM) abolished acute neuronal swelling and rescued from late degeneration both cholinergic and noncholinergic cells when concentrations of glutamate up to 500 microM were added to the cultures. Protective effects declined progressively with increasing concentrations of the amino acid, even when MK-801 was raised to its highest nontoxic levels, e.g., 50 microM. the kainate/quisqualate receptor antagonist CNQX provided no protection alone or in combination with MK-801. Nerve growth factor (NGF), used in standard culture conditions to stimulate the expression of the cholinergic phenotype, was shown not to influence excitotoxic neurodegenerative changes. Several observations suggested that nitric oxide (NO) may act as an intercellular messenger of NMDA-mediated cell death in septal cultures: 1) Most of the cholinergic neurons contained the NO synthase enzyme as characterized by NADPH-diaphorase (NADPH-d) staining; 2) sodium nitroprusside (SNP) [a chemical with the ability of generating NO] was capable of mimicking some of the aspects of the glutamate-induced degenerative process; 3) the rise in cyclic GMP which was observed in the presence of toxic levels of glutamate and which is usually taken as an index of NO production, was antagonized by MK-801 and by the inhibitor of the NO synthase enzyme, L-NOARG. Yet, the fact that L-NOARG and its congener, L-NAME, were ineffective in preventing glutamate-induced neurodegenerative changes in our culture system did not substantiate our working hypothesis. Altogether these results suggest that glutamate-induced cholinergic neuronal death is the consequence of an overstimulation of NMDA receptors and that neither NGF nor NO plays a key role in the degenerative process.

Topics: Animals; Cell Death; Cells, Cultured; Dizocilpine Maleate; Excitatory Amino Acids; Glutamic Acid; NADPH Dehydrogenase; Nerve Degeneration; Nerve Growth Factors; Neurons; Nitric Oxide; Parasympathetic Nervous System; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Septum Pellucidum

Systemic administration of N-methyl-D-aspartate (NMDA) receptor antagonists induces a well defined behaviour in rodents characterized by, for example increased locomotion and ataxia. It is not clear in what brain region(s) NMDA antagonists induce this behaviour. We have studied the possible involvement of olfactory pathways by making adult mice anosmic via intranasal injection of zinc sulphate, a procedure that is known to destroy the olfactory epithelium. The NMDA antagonist MK-801 was given intraperitoneally (0.1-1.0 mg/kg) and the animals were scored for locomotion and ataxia 60-90 min later. Before MK-801 administration, olfactory-lesioned mice did not differ from non-lesioned controls with regard to locomotion or ataxia. MK-801 caused locomotor activation (> or = 0.2 mg/kg) and ataxia (> or = 0.5 mg/kg) in both groups. In general, olfactory-lesioned animals showed more locomotion and less ataxia after MK-801 administration than non-lesioned animals. Lesioned animals displayed 2.0- (P < 0.05) and 3.7-fold (P < 0.05) more extensive locomotor activation than non-lesioned animals after 0.5 and 1.0 mg/kg of MK-801, respectively. No difference in the degree of ataxia was seen between the two groups at 0.5 mg/kg, whereas non-lesioned animals showed a 2.1-fold higher degree of ataxia after 1.0 mg/kg of MK-801, indicating that the enhanced MK-801-induced locomotor activation in olfactory-lesioned mice was not simply due to less ataxia. These results suggest that olfactory input is involved in NMDA antagonist-induced behaviour.

Topics: Animals; Ataxia; Behavior, Animal; Body Weight; Denervation; Dizocilpine Maleate; Excitatory Amino Acids; Male; Mice; Motor Activity; Nerve Degeneration; Neurons, Afferent; Olfaction Disorders; Olfactory Bulb; Organ Size; Sulfates; Zinc Compounds; Zinc Sulfate

It is well documented that neurons exposed to high concentrations of excitatory amino acids, such as glutamate and aspartate, degenerate and die. The clearance of these amino acids from the synaptic cleft depends mainly on their transport by high-affinity sodium-dependent carriers. Using microdialysis in vivo and HPLC analysis, we have studied the effect of the administration of inhibitors of the glutamate transporter (L-trans-pyrrolidine-2,4-dicarboxylate and dihydrokainate) on the extracellular concentration of endogenous amino acids in the rat striatum. In addition, we have analyzed whether the changes observed in the concentration of glutamate and aspartate were injurious to striatal cells. Neuronal damage was assessed by biochemical determination of choline acetyltransferase and glutamate decarboxylase activities, 7 days after the microdialysis procedure. In other experiments, pyrrolidine dicarboxylate and dihydrokainate, as well as two other inhibitors of the glutamate carrier, DL-threo-beta-hydroxyaspartate and L-aspartate-beta-hydroxamate, were microinjected into the striatum, and neuronal damage was assessed, both biochemically and histologically, 7 or 14 days after the injection. Dihydrokainate and pyrrolidine dicarboxylate produced a similar remarkable increase in the concentration of extracellular aspartate and glutamate. However, the former induced also notable elevations in the concentration of other amino acids. Clear neuronal damage was observed only after dihydrokainate administration, which was partially prevented by intraperitoneal injection of (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate or by intrastriatal coinjection of 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(f)quinoxaline. No cell damage was observed with the other three glutamate carrier inhibitors used. It is concluded that an increased extracellular glutamate level in vivo due to dysfunction of its transporter is not sufficient for inducing neuronal damage. The neurotoxic effects of dihydrokainate could be explained by direct activation of glutamate postsynaptic receptors, an effect not shared by the other inhibitors used.

Topics: Animals; Cell Death; Choline; Choline O-Acetyltransferase; Corpus Striatum; Dicarboxylic Acids; Dizocilpine Maleate; Extracellular Space; gamma-Aminobutyric Acid; Glutamate Decarboxylase; Glutamic Acid; Kainic Acid; Male; Microdialysis; Nerve Degeneration; Neurons; Neurotransmitter Uptake Inhibitors; Pyrrolidines; Quinoxalines; Rats; Rats, Wistar

The effects of neurotrophins on several forms of neuronal degeneration in murine cortical cell cultures were examined. Consistent with other studies, brain-derived neurotrophic factor, neurotrophin-3, and neurotrophin-4/5 all attenuated the apoptotic death induced by serum deprivation or exposure to the calcium channel antagonist nimodipine. Unexpectedly, however, 24-hour pretreatment with these same neurotrophins markedly potentiated the necrotic death induced by exposure to oxygen-glucose deprivation or N-methyl-D-aspartate. Thus, certain neurotrophins may have opposing effects on different types of death in the same neurons.

Topics: Animals; Apoptosis; Brain-Derived Neurotrophic Factor; Calcium; Cell Death; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Mice; N-Methylaspartate; Necrosis; Nerve Degeneration; Nerve Growth Factors; Nerve Tissue Proteins; Neurons; Neurotrophin 3; Quinoxalines; Receptors, AMPA

The effects of nicotine on glutamate-induced cytotoxicity were examined using primary cultures of rat cortical neurons. The cell viability was significantly reduced when cultures were briefly exposed to glutamate or N-methyl-D-aspartate (NMDA) then incubated with normal medium for 1 h. A 1-h exposure of the cultures to kainate or alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) reduced cell viability. Incubating cultures with nicotine for 1-24 h protected cortical neurons against glutamate cytotoxicity. Maximum protection against glutamate cytotoxicity was induced with a 2-h nicotine incubation. Exposure to nicotine for up to 2 h did not affect cell viability by itself although cell viability was reduced in a time-dependent manner when the exposure exceeded 4 h. Neuroprotection by nicotine was dependent on both the concentration and incubation period. Nicotine reduced the NMDA cytotoxicity but did not attenuate that of kainate and AMPA. The neuroprotective effects of nicotine against glutamate cytotoxicity were antagonized by mecamylamine and hexamethonium but not by atropine. These results indicate that nicotinic receptor stimulation induces neuroprotection against glutamate cytotoxicity mediated by NMDA receptors.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Glutamates; Glutamic Acid; Nerve Degeneration; Neurons; Nicotine; Parasympathetic Nervous System; Parasympathomimetics; Piperazines; Rats; Receptors, N-Methyl-D-Aspartate

An investigation has been made into the effect of 3,4-methylenedioxymethamphetamine (MDMA or 'Ecstasy') administration on the concentration of 5-hydroxytryptamine (5-HT), uptake of [3H]5-HT and [3H]paroxetine binding in rat cerebral cortex tissue. Four days after 2 injections of MDMA (20 mg/kg i.p., 6 hr apart) the concentrations of 5-HT and its metabolite 5-HIAA were reduced by 60%. The binding of [3H]paroxetine to the presynaptic 5-HT transporter was decreased and high affinity uptake of [3H]5-HT was reduced by a similar amount, indicating neurodegeneration of 5-HT terminals. Pretreatment with chlormethiazole (100 mg/kg i.p.), 10 min before each MDMA injection prevented the decrease in both [3H]parotextine binding and uptake of [3H]5-HT. The loss in 5-HT and 5-HIAA content was also attenuated. Pretreatment with dizocilpine (1 mg/kg i.p.) or haloperidol (2 mg/kg i.p.) also prevented the MDMA-induced loss of [3H]paroxetine binding and attenuated the loss of 5-HT and 5-HIAA content. All three compounds also decreased the degree of hyperthermia that follows MDMA administration, although previous studies suggest that the long term neurodegeneration is not associated with the acute hyperthermic response. These data support the findings of others that MDMA injection produces degeneration of 5-HT nerve terminals in the cortex, confirm that chlormethiazole, dizocilpine and haloperidol attenuate MDMA-induced neurotoxic loss of 5-HT and demonstrate for the first time that these compounds prevent the neurodegeneration of 5-HT nerve terminals that follows MDMA administration.

Topics: Animals; Chlormethiazole; Dizocilpine Maleate; Haloperidol; Hydroxyindoleacetic Acid; Male; N-Methyl-3,4-methylenedioxyamphetamine; Nerve Degeneration; Nerve Endings; Neuroprotective Agents; Radioligand Assay; Rats; Selective Serotonin Reuptake Inhibitors; Serotonin

Neural degeneration was observed in a similar set of limbic structures following the continuous administration of several NMDA antagonists (phencyclidine, dizocilpine, and LY235959). The earliest signs involved terminals and processes, followed by cell bodies. In retrosplenial cortex the predominant staining showed a distribution very similar to that observed for cholinergic innervations. Considerable degeneration was also observed in entorhinal cortex and its principal output, dentate gyrus of hippocampus, and in olfactory regions such as olfactory tubercle and tenia tecta, and in piriform cortex. These results, when considered together with those from studies of glucose metabolism following NMDA antagonists, suggest that a hypermetabolic circuit was involved, and indicate that both competitive and non-competitive NMDA antagonists can induce these effects.

Topics: Animals; Dizocilpine Maleate; Dose-Response Relationship, Drug; Isoquinolines; Limbic System; N-Methylaspartate; Nerve Degeneration; Phencyclidine; Rats; Time Factors

The novel free radical scavenger and electron-trapping agent, idebenone, protects cultured cortical neurons against necrotic degeneration induced by either a brief exposure to N-methyl-D-aspartate (NMDA) or a prolonged exposure to kainate. As opposed to the NMDA receptor antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]- cyclohepten-5,10-imine hydrogen maleate (MK801), idebenone rescued cortical neurons even when applied 30 min after the NMDA pulse, suggesting that the drug interferes with the chain of toxic reactions triggered by an excessive stimulation of excitatory amino acid receptors.

Topics: Animals; Benzoquinones; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Mice; N-Methylaspartate; Nerve Degeneration; Neurons; Neuroprotective Agents; Neurotoxins; Time Factors; Ubiquinone

Succinic dehydrogenase in mouse cortical explant cultures was inhibited by 3-nitropropionic acid (3-NPA). ATP concentrations declined upon application of 3-NPA. At 4 h, ATP levels of cultures treated with 3-NPA alone were no different from those in cultures treated additionally with MK-801 (20 microM), 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 10 microM) or a combination thereof. However, MK-801 and MK-801 plus CNQX mitigated morphological lesions caused by 3-NPA. CNQX alone did not influence the extent of morphological damage. In conclusion, MK-801, at concentrations which were neuroprotective against 3-NPA lesions in cortical explant cultures, did not modify 3-NPA dependent decreases in cellular ATP levels. These data indicate that the neuroprotective effects of glutamate receptor antagonists in this model are probably receptor mediated and do not involve effects on cellular metabolism.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Adenosine Triphosphate; Animals; Culture Techniques; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Mice; Nerve Degeneration; Neurotoxins; Nitro Compounds; Oxidative Phosphorylation; Plant Extracts; Propionates

Rotation-mediated aggregating cell cultures from fetal rat telencephalons containing glial and neuronal cells mature in a fashion comparable to that known to occur in brain in vivo. Large aggregates of 300-500 microM diameters can now reproducibly be cultivated and maintained for more than 40 days in a well defined serum free medium. Validity of the use of such cultures for in vitro studies of various physiological, pharmacological and toxicological phenomenon has already been demonstrated. In this communication some observations suggesting the usefulness of such cultures for pharmacological studies clarifying the possible effects of drugs and other agents on excitatory amino acid induced pathological processes will be presented. The advantages and limitations of the use of aggregated brain cell culture based models for the development of agents potentially useful for the treatment of aging and dementia will also be discussed.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Acetylcholinesterase; Animals; Biomarkers; Cell Aggregation; Choline O-Acetyltransferase; Dizocilpine Maleate; DNA; Fetus; Gestational Age; Glutamate-Ammonia Ligase; Glutamic Acid; L-Lactate Dehydrogenase; N-Methylaspartate; Nerve Degeneration; Nerve Regeneration; Neuroglia; Neurons; Rats; Reproducibility of Results; Telencephalon

Excitatory input regulates cell birth and survival in many systems. The granule cell population of the rat dentate gyrus is formed primarily during the postnatal period. Excitatory afferents enter the dentate gyrus and begin to form synapses with granule cells during the first postnatal week, the time of maximal cell birth and death. In order to determine whether excitatory input plays a role in the regulation of cell birth and survival in the developing granule cell layers and their germinal regions, the subependymal layer and hilus, we treated rat pups with the N-methyl D-aspartate (NMDA) receptor antagonists MK-801, CGP 37849, or CGP 43487 during the first postnatal week and examined the numbers of 3H-thymidine-labeled cells, pyknotic cells, and healthy cells in these regions. In order to determine the cell type that was affected, sections from brains of MK-801-treated rats were processed for 3H-thymidine autoradiography combined with immunohistochemistry for the marker of radial glia, vimentin, and the marker of mature astrocytes, glial fibrillary acidic protein (GFAP). Within the dentate gyrus, NMDA receptor blockade resulted in the following changes: (1) the density of 3H-thymidine-labeled cells was increased, (2) the density of pyknotic cells was increased, (3) the density of 3H-thymidine-labeled pyknotic cells was increased, and (4) the density of healthy cells was decreased. The infrapyramidal blade/hilus showed changes throughout its extent, whereas the suprapyramidal blade showed changes only at the rostral level. No change in the numbers of 3H-thymidine-labeled vimentin-immunoreactive or GFAP-immunoreactive cells was observed in the dentate gyrus with MK-801 treatment, indicating that glia are not primarily affected by NMDA receptor blockade. Blockade of NMDA receptors resulted in gross morphologic changes in the dentate gyrus; in most cases, the infrapyramidal blade was indistinguishable from the hilus. Moreover, in several brains of animals treated with CGP 37849 or CGP 43487 on postnatal day (P)5, an abnormal aggregation of cells was observed ventral to the normal location of the infrapyramidal blade. This cellular cluster contained many pyknotic and 3H-thymidine-labeled cells and may represent cells that normally comprise the infrapyramidal blade. Dramatic changes to the subependymal layer were also seen following NMDA receptor blockade. The cross-sectional area of this region was significantly increased with MK-801, CGP 37849, or CGP 4348

Topics: 2-Amino-5-phosphonovalerate; Animals; Autoradiography; Brain; Dizocilpine Maleate; Female; Glial Fibrillary Acidic Protein; Hippocampus; Immunohistochemistry; Nerve Degeneration; Nerve Regeneration; Pregnancy; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Thymidine; Vimentin

We have used the laser-induced photochemical thrombosis model in adult rats to evaluate the significance of the non-N-methyl-D-aspartate (non-NMDA) subtype of glutamate receptors in situations of focal spinal cord ischemia. The animals were pretreated with the selective non-NMDA antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(f)-quinoxaline (NBQX) or, for comparison, the NMDA antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d] cyclohepten-5-10-imine (MK-801). Neurological function was quantified using evaluations of motor score and inclined plane. The MK-801-treated rats had higher motor scores during the 3-week observation period while NBQX-treated rats only performed significantly better at 1 week. Both treatments caused significantly better performance in the inclined plane test. NBQX and MK-801 reduced the volume of necrosis by approximately 47% at 3 weeks postlesion. We conclude that blockade of both NMDA and non-NMDA subtypes of glutamate receptors reduces ischemic necrosis, possibly by preventing excessive stimulation of these receptors by released excitatory amino acids in the lesion area.

Topics: Analysis of Variance; Animals; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Ischemia; Lasers; Nerve Degeneration; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Reference Values; Spinal Cord; Time Factors

HIV-1-associated cognitive/motor complex is a frequent neurological complication of the acquired immunodeficiency syndrome (AIDS). The pathogenesis of this syndrome implicates immunopathological and toxic events such as the production of cytokines. The HIV envelope glycoprotein gp120 seems also to play a major role in this process. Gp120 could produce a slow neuronal death probably via the release of neurotoxic factors by CNS macrophages/monocytes. NMDA antagonists and Ca2+ channel blockers in vitro have a powerful neuroprotective effect against gp120 neurotoxicity. The purpose of the present work is to determine whether gp120-induced neurotoxicity is associated with an abnormal neuronal depolarization induced by putative neurotoxins. We have compared in vitro the neuroprotective effects of Tetrodotoxin a Na+ channel blocker, the Ca2+ channel blocker nifedipine and the NMDA antagonist MK-801 in primary cortical neurons taken from embryonic rat and intoxicated with gp120. We observed comparable neuroprotective effects with the 3 precited compounds suggesting that gp120-induced neurotoxic factors act on Na+ channels, NMDA receptors and Ca2+ channels in a cascade of cellular events. We confirmed that the presence of macrophages is needed to trigger a marked gp120-induced neurotoxicity. These results underline the fact that depolarization is an important component of gp120 neurotoxicity in primary neuronal cultures.

Topics: Animals; Cell Survival; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; HIV Envelope Protein gp120; Macrophages, Alveolar; Nerve Degeneration; Neurons; Nifedipine; Rats; Receptors, N-Methyl-D-Aspartate; Tetrodotoxin

The neurotoxin beta-N-oxalylamino-L-alanine (BOAA), found in Lathyrus sativus seeds, is thought to be the causative agent of neurolathyrism. We have investigated the in vivo mechanism of action of BOAA by focal injection (1 microliter) in the dorsal hippocampus of male Wistar rats and comparing the pathological outcome with the effects of injections (1 microliter) of alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionate (AMPA), kainate (KA) or N-methyl-D-aspartate (NMDA). Cellular damage induced by the excitatory amino acids in the pyramidal (CA1-CA4) and dentate granule neurones (DG) was assessed histologically 24 h after the injection. The study shows that BOAA (50 nmol) induces hippocampal toxicity with a highly selective pattern of regional cellular damage. The CA1, CA4 and DG subfields show 70-90% neuronal injury whereas CA2 and CA3 show only minimal damage. This pattern of cellular damage is similar to that induced by AMPA (1 nmol) and NMDA (25 nmol) but not KA (0.5 nmol). BOAA-induced neurotoxicity is prevented in a dose-dependent manner by focal co-injection of the non-NMDA receptor antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX) (1-25 nmol) but not by a dose of MK-801 (3 mg/kg i.p.) which is neuroprotective against an injection of NMDA. Delayed focal injections of NBQX (25 nmol) up to 2 h after the BOAA injection result in a significant protection of all pyramidal and granular cell regions. These results indicate that the in vivo hippocampal toxicity of BOAA is mediated by AMPA receptors rather than by KA or NMDA receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Amino Acids, Diamino; Animals; beta-Alanine; Dizocilpine Maleate; Dose-Response Relationship, Drug; Hippocampus; Kainic Acid; Male; N-Methylaspartate; Nerve Degeneration; Neurons; Quinoxalines; Rats; Rats, Wistar

Injection of the N-methyl-D-aspartate receptor agonist, quinolinic acid, into the rat striatum in vivo results in the degeneration of cholinergic and GABAergic neurons, as determined seven days later using the marker enzymes, choline acetyltransferase and glutamate decarboxylase, respectively. Such damage was dose-dependently prevented by CGP 37849 or MK-801 (competitive and uncompetitive N-methyl-D-aspartate receptor antagonists, respectively) administered systemically or intrastriatally at the same time as quinolinic acid. The neuroprotective activity of CGP 37849 was associated with the D-enantiomer, CGP 40116 (ED50 7.5 mg/kg i.p.), which was approximately 1.5-fold and 3.5-fold more potent than the related compounds, D-CPPene and CGS 19755, respectively. CGP 37849 was a weaker neuroprotectant than MK-801 (ED50 0.8 mg/kg i.p) when administered systemically, but was dramatically more potent following coinjection with quinolinic acid (ED50's 0.2 and 117 nmol, respectively). When injected intrastriatally 0.5-2 h post-quinolinic acid, CGP 37849 was protective over the entire period studied, whereas MK-801 was less effective at all post-quinolinic acid injection times. The finding that CGP 37849 is neuroprotective when administered intrastriatally 1-2 h post-quinolinic acid supports the hypothesis that a period exists following excitotoxic insult in which neurons are not committed to die, and can be rescued by blockade of ongoing N-methyl-D-aspartate receptor activation. Competition studies indicated that, when coinjected with 100-400 nmol quinolinic acid into the striatum, CGP 37849 exhibited kinetics predicted of a competitive N-methyl-D-aspartate receptor antagonist (declining neuroprotective potency with increasing doses of agonist), whereas MK-801 displayed a complex picture, with weak protective activity at low doses of quinolinic acid. Following systemic administration, neither antagonist was markedly affected by the dose of excitotoxin. When given i.p. at up to 6 h post-quinolinic acid, CGP 37849 and MK-801 showed essentially identical profiles of post-insult protection; degeneration of cholinergic neurons was reduced significantly throughout the entire post-insult period, whereas GABAergic neurons were protected only when drugs were administered 2 h or earlier post-quinolinic acid. The data indicate that competitive and uncompetitive N-methyl-D-aspartate receptor antagonists are effective neuroprotectants in vivo, and that parameters such as drug li

Topics: 2-Amino-5-phosphonovalerate; Acetylcholine; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Binding, Competitive; Biomarkers; Cell Death; Choline O-Acetyltransferase; Corpus Striatum; Dizocilpine Maleate; Drug Administration Schedule; gamma-Aminobutyric Acid; Glutamate Decarboxylase; Injections; Injections, Intraperitoneal; Kainic Acid; Male; Nerve Degeneration; Nerve Tissue Proteins; Neurons; Neurotoxins; Pipecolic Acids; Piperazines; Quinolinic Acid; Rats; Receptors, N-Methyl-D-Aspartate

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

Excitotoxic amino acids contain two acidic groups, but cysteine represents an exception to this rule. The hypothesis that cysteine toxicity is mediated by the oxidized and diacidic metabolites cysteine sulphinate and/or cysteate was tested in the present study. The issue was approached in three different ways. Firstly, the distribution of brain injury after subcutaneous administration of cysteine (1 mg/g) to 4-day-old rats was compared with that caused by cysteine sulphinate (3 mg/g). Secondly, the effects of excitatory amino acid receptor antagonists on cysteine and cysteine sulphinate toxicity were investigated. Thirdly, the cerebral concentrations of cysteine sulphinate were determined after cysteine administration and compared with those obtained after cysteine sulphinate injection. The cerebral cortex was the region most vulnerable to cysteine toxicity, followed by the hippocampus (especially the medial subicular neurons), amygdala, caudoputamen, cerebellum and septum. Pronounced extravasation of red blood cells was observed in lesioned areas. One day after cysteine administration, the injury was infarction-like and sharply demarcated. Cysteine sulphinate-induced damage resembled cysteine-induced lesions in some respects: the anterior cingulate and retrosplenial cortices, as well as medial subicular cells, were quite vulnerable. However, the differences prevailed. Cysteine sulphinate, but not cysteine, killed neurons of the superficial part of the tectum, the medial habenula, the ventromedial hypothalamus and the arcuate nucleus. Further, while cysteine toxicity was prominent in deep cortical layers, cysteine sulphinate preferentially damaged superficial cortical neurons. Cysteine toxicity was abolished by pretreatment with MK-801, a selective NMDA antagonist, but not by 2,3-dihydroxy-6-nitro-7-sulphamoyl-benzo(F)quinoxaline, a selective AMPA receptor blocker. In contrast, the considerably smaller lesion seen after cysteine sulphinate administration was only partially prevented by MK-801. Large (19-fold) increases in cortical cysteine sulphinate concentration were noted after injection of a toxic dose of cysteine. This corresponds to 90 nmol cysteine sulphinate/g protein. The cysteate concentration was not increased above the detection limit. Injection of a toxic dose of cysteine sulphinate elevated cysteine sulphinate concentration in the frontomedial cortex (a region consistently injured by cysteine sulphinate) almost three orders of magnitude more

Topics: Amino Acids; Animals; Animals, Newborn; Brain; Cerebral Cortex; Cysteic Acid; Cysteine; Dizocilpine Maleate; Female; Male; Nerve Degeneration; Neurons; Neurotoxins; Neurotransmitter Agents; Organ Specificity; Quinoxalines; Rats; Rats, Sprague-Dawley

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder affecting motor neurons. Glutamate, a potent central-nervous-system toxin, has been proposed as one possible factor in this motoneuron disease. Serum from patients with ALS is known to be toxic when added to neurons in culture. We report on the toxicity to rat neurons in culture of cerebrospinal fluid (CSF) from patients with ALS. CSF were obtained from 10 ALS patients, 10 neurological controls, and 10 other controls. ALS CSF was added at dilutions of 50%, 20%, or 10% and neuron survival was assessed after 24 h. The neuroprotective effects of antagonists to two glutamate receptors were also assessed. ALS CSF was significantly neurotoxic, with a neuronal survival rate of only 47% compared with 80% or so for control CSF. This neurotoxicity was blocked by CNQX, an antagonist to the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)/kainate receptor but not by two N-methyl-D-aspartate (NMDA) antagonists. ALS CSF contains a specific neurotoxic factor which is AMPA/kainate-like which could have a role in the neuronal degeneration of this disease.

Topics: 2-Amino-5-phosphonovalerate; 6-Cyano-7-nitroquinoxaline-2,3-dione; Aged; Amino Acids; Amyotrophic Lateral Sclerosis; Animals; Cell Survival; Cells, Cultured; Dizocilpine Maleate; Dose-Response Relationship, Drug; Humans; Middle Aged; Motor Neurons; Nerve Degeneration; Quinoxalines; Rats; Receptors, AMPA; Receptors, Glutamate; Receptors, Kainic Acid

The effect of the N-methyl-D-aspartate receptor agonist quinolinic acid on extracellular levels of striatal amino acids, following its injection directly into the rat striatum, has been investigated using intracerebral dialysis in the attempt to elucidate the cellular mechanisms underlying delayed neurodegeneration. A neurotoxic dose (200 nmol) of quinolinic acid caused an elevation in the levels of aspartate (x 6), glutamate (x 2), asparagine (x 2), serine (x 2.5), glycine (x 3), and threonine (x 2) which peaked in the fractions 20-40 min after the injection and achieved statistical significance for aspartate and asparagine. The dialysate content of these amino acids returned to basal values within 1 h and no further changes were observed in the following 4 h. Injection of an equivalent dose of nicotinic acid did not mimic the effect of quinolinate, indicating that osmotic and/or mechanical damage was not responsible for the observed phenomena. Pretreatment with the N-methyl-D-aspartate receptor channel blocker dizocilpine (MK-801) completely blocked the quinolinate-induced increase of the amino acids, thus confirming that N-methyl-D-aspartate receptor activation is required for this effect to occur. Seven days after the injection of quinolinate, histological analysis showed an extensive loss of neuronal elements in the injected striatum, which was completely prevented in the dizocilpine-treated animals. Sections from striata of animals injected with nicotinic acid showed normal-appearing neurons and no differences were detectable from controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Amino Acids; Animals; Asparagine; Aspartic Acid; Corpus Striatum; Dialysis; Dizocilpine Maleate; Glutamates; Glutamic Acid; Glycine; Male; Microinjections; Nerve Degeneration; Niacin; Quinolinic Acid; Rats; Rats, Sprague-Dawley; Serine; Threonine

In exploring the recently discovered phenomenon of indirect excitotoxicity, we noted that intrahippocampal injections of the nonspecific aminotransferase inhibitor gamma-acetylenic GABA (GAG; 60-240 nmol) caused excitotoxic lesions in rats. When assessed 3 days following the injection, GAG was shown to be approximately equally toxic to CA3/hilar neurons and CA1 pyramids, while CA2 neurons and granule cells were clearly less vulnerable. Choline acetyltransferase activity, a marker of extrinsic afferents, remained unchanged in the GAG-lesioned hippocampus, indicating the axon-sparing nature of the insult. In contrast, a lesion caused by 240 nmol of GAG resulted in a significant reduction in 3H-MK-801 binding, which was used as a marker for NMDA receptor-bearing hippocampal neurons. GAG-induced lesions were blocked by the NMDA receptor antagonists MK-801 and AP7 but were not influenced by the nature of the anesthetic used during surgery. Iontophoretic application of GAG did not excite CA1/CA3 cells in the rat hippocampus. In vitro, GAG proved to be a relatively potent inhibitor (IC50: 43 microM) of kynurenine aminotransferase, the biosynthetic enzyme of the endogenous neuroprotectant kynurenic acid. GAG also inhibited the neosynthesis of kynurenic acid in hippocampal slices (IC50: 790 microM). Thus, GAG shares several characteristics of the recently described indirect excitotoxin aminooxyacetic acid (AOAA; Exp. Neurol. 113: 378, 1991). GAG and AOAA appear to belong to a new family of excitotoxic agents which produce lesions indirectly by metabolic derangement and/or inhibition of kynurenate production.

Topics: 2-Amino-5-phosphonovalerate; 4-Aminobutyrate Transaminase; Alkynes; Amino Acids; Aminocaproates; Animals; Anticonvulsants; Axons; Choline O-Acetyltransferase; Dizocilpine Maleate; Hippocampus; In Vitro Techniques; Kynurenic Acid; Lyases; Male; Nerve Degeneration; Neurons; Pyramidal Cells; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Stereotaxic Techniques; Transaminases

The hamster neurotropic (HNT) strain of measles virus causes non-inflammatory encephalopathy in Balb/c mice, associated with neurodegeneration in hippocampal CA1 and CA3 regions. This loss of pyramidal cells can be prevented by twice daily systemic treatment with 1 mg/kg dizocilpine (5-methyl-10,11-dihydro-5H-dibenzo(a,d)cyclo-hepten-5,10-imine maleate; MK-801) for 7 days. By varying the MK-801 treatment protocol, we now found that drug administration during the last 4 days prior to sacrifice (i.e. days 4-7 post inoculation, p.i.) is essential for neuroprotection. In contrast, MK-801 treatment during the first days (days 0-4 p.i.) did not prevent the neuronal necrosis. These data suggest that the concentration of an excitotoxic factor in the mouse brain increases after virus inoculation, reaching toxic levels by days 4-5 p.i. This novel 'subacute' mouse model of neurodegeneration therefore constitutes an attractive tool for mechanistic and interventional studies in excitotoxicity research.

Topics: Animals; Disease Models, Animal; Dizocilpine Maleate; Female; Hippocampus; Measles; Measles virus; Mice; Mice, Inbred BALB C; Nerve Degeneration; Neurotoxins; Receptors, N-Methyl-D-Aspartate

The role of nitric oxide in N-methyl-D-aspartate (NMDA) neurotoxicity was investigated in murine cortical cell cultures. Exposure of cultures to 300 microM NMDA for 5 min resulted in death of 50-80% of neurons over the subsequent 24 h. This injury was not attenuated by hemoglobin, the nitric oxide synthase (NOS) inhibitors NG monomethyl-L-arginine (MMA) or N omega-nitro-L-arginine (NA), or L-arginine depletion. Hemoglobin and NOS inhibitors consistently prevented the increase in cyclic guanosine monophosphate (cGMP) seen after NMDA exposure. These results suggest that NMDA neurotoxicity in this cell culture system is mediated, at least in part, by mechanisms other than NOS activation.

Topics: Animals; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Hemoglobins; Mice; N-Methylaspartate; Nerve Degeneration; Neurons; Neurotoxins; Nitric Oxide

alpha-Dendrotoxin (Dtx), a snake polypeptide, increases neuronal excitability by blocking certain fast-activating, voltage-dependent K+ channels. Thus, the behavioural, electrocortical (ECoG) and neuropathological effects of Dtx, injected into rat brain areas, were studied. A unilateral injection of 35 pmol of Dtx into the CA1 hippocampal area or the dendate gyrus (DG; upper blade) immediately produced motor and ECoG seizures, followed at 24 h by multi-focal brain damage and significant neuronal loss. Whilst brain damage was seen bilaterally, significant neuronal loss occurred only in regions (CA1, CA3, CA4 and DG) ipsilateral to the site of injection. A lower dose (3.5 pmol) of toxin elicited motor and ECoG seizures but failed to produce brain damage. Seizures were observed 50 min after injecting Dtx (35 pmol) into the amygdala, though significant neuronal loss was not evident. 4-Aminopyridine (100 nmol), given into the CA1 area elicited a similar motor and ECoG pattern to that of Dtx except no brain damage could be seen at 24 h. Systemic pretreatment with antagonists of N-methyl-D-aspartate receptors (MK-801 or CGP 37849) did not protect against the effects typically evoked by injecting Dtx into the CA1 area.

Topics: 2-Amino-5-phosphonovalerate; Animals; Brain Diseases; Dizocilpine Maleate; Elapid Venoms; Electroencephalography; Hippocampus; Injections; Male; Nerve Degeneration; Neurotoxins; Potassium Channels; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Seizures

Focal brain ischemia was induced by middle cerebral artery occlusion in the rat. The volume of cerebral damage was determined 2 days later by MRI in vivo and in the same animals histologically. The edema volume as measured by MRI and the histologically determined infarction was highly correlated. As a consequence, the neuroprotective effect of the N-methyl-D-aspartate (NMDA) receptor antagonists CGP 40116 and MK 801 were similar with both methods. Excitotoxic neurodegeneration in the rat striatum was induced by direct injection of quinolinic acid. The degree of damage was evaluated in vivo 1 day later by quantitative MRI, and 7 days later by measuring the activities of neuronal marker enzymes choline acetyltransferase and glutamic acid decarboxylase. Striatal damage assessed using the three approaches was highly correlated. Cerebroprotective efficacy of the NMDA receptor antagonist CGP 40116 was indistinguishable based on all methods. MRI was more reproducible than the enzymatic methods and was faster and simpler than histologic examination for routine analysis of excitotoxic damage and cerebroprotection in vivo in a pharmaceutical research environment.

Topics: 2-Amino-5-phosphonovalerate; Animals; Brain; Cerebral Arteries; Choline O-Acetyltransferase; Clinical Enzyme Tests; Constriction; Dizocilpine Maleate; Glutamate Decarboxylase; Ischemic Attack, Transient; Magnetic Resonance Imaging; Nerve Degeneration; Quinolinic Acid; Rats; Rats, Inbred F344

Tetanus toxin (TT) blocks GABA-mediated inhibitory neurotransmission in the mammalian CNS via selective inhibition of transmitter release. The loss of central inhibition produces an excitatory focus resembling human limbic epilepsy. We now report that the net excitation caused by an unopposed action of glutamic acid may also produce neuronal degeneration in the rat brain. Anaesthetized rats were placed in a stereotaxic frame and TT (1 microliter dissolved in phosphate buffer, pH 7.0) was injected unilaterally into the dorsal hippocampus. Injection of TT (1000 mouse minimum lethal doses, MLDs; n = 3-6 rats per group) produced time-dependent neuronal loss in the CA1 pyramidal cell layer which was significant (p < 0.05) 7 and 10 days, but not 1 day, after the injection. Systemic treatment with competitive (CGP 37849, 3 mg/kg i.p) or non-competitive (MK801, 0.3 mg/kg i.p.) antagonists at the N-methyl D-aspartate (NMDA) receptor complex 1 h before and 1 h after TT and then once daily for 10 days protected rats from the hippocampal damage produced by TT (1000 MLDs). In addition, in rats bearing a monolateral surgical lesion of the Schaffer collaterals, through which CA1 neurones receive a robust excitatory input from CA3 pyramids, the bilateral injection of TT (1000 MLDs/side) produced significant neuronal loss in the unlesioned hippocampus whereas the contralateral appeared to be preserved. In conclusion, these results demonstrate that excitatory neurotransmission may be involved in the neuropathology elicited by intrahippocampal TT in rats.

Topics: 2-Amino-5-phosphonovalerate; Animals; Dizocilpine Maleate; gamma-Aminobutyric Acid; Hippocampus; Male; Nerve Degeneration; Neurons; Rats; Rats, Wistar; Receptors, GABA-A; Receptors, N-Methyl-D-Aspartate; Synaptic Transmission; Tetanus Toxin

The effect of a single injection of 3-acetylpyridine (3-AP), which led to a degeneration of the excitatory cerebellar climbing fibers, was studied on the binding of [3H]MK-801, a non-competitive NMDA antagonist, in the rat cerebellar cortex. The same treatment increased also the binding of [3H]CGP 39653, a new NMDA competitive antagonist. Saturation isotherms showed a significant increase of the maximal number of binding sites (Bmax) for [3H]CGP 39653 and [3H]MK-801 (+48 and 36% respectively) with no change in the affinity 4-9 days after the administration of 3-AP. Our data demonstrate that in the cerebellar cortex both NMDA recognition site labelled by [3H]CGP 39653 and its modulatory site labelled by [3H]MK-801 may undergo plastic changes when the glutamatergic receptors and transmission are denervated.

Topics: 2-Amino-5-phosphonovalerate; Animals; Cerebellar Cortex; Dizocilpine Maleate; Kinetics; Male; Nerve Degeneration; Nerve Fibers; Pyridines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Synaptic Transmission

Topics: 2-Amino-5-phosphonovalerate; Animals; Anticonvulsants; Carbamazepine; Diazepam; Dizocilpine Maleate; Epilepsy, Generalized; gamma-Aminobutyric Acid; Glutamates; Glutamic Acid; Hippocampus; Male; Nerve Degeneration; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Tetanus Toxin

Lipocortin-1 (annexin-1), an endogenous phospholipid and calcium binding protein, has been shown to significantly attenuate the damage produced by focal cerebral ischaemia in the rat. In the present study we have therefore investigated its effect on N-methyl-D-aspartate (NMDA) induced neuronal damage. Unilateral intrastriatal infusion of a potent and selective NMDA agonist, cis-2,4-methanoglutamate (MGlu), induced an extensive lesion of the striatum in the rat, which was inhibited (greater than 80%) by prior injection of MK801 (4 mg/kg, i.p.). Infusion of 1.2 micrograms of an active fragment of lipocortin-1 (N-terminal 1-188 aa) immediately after MGlu significantly reduced the extent of damage by 44.2 +/- 8.0%. In contrast, infusion of 3 microliters of neutralizing anti-lipocortin-1 antibody with MGlu increased lesion size by 158.9 +/- 22.0%. These findings indicate that the damage produced by intrastriatal infusion of MGlu is mediated by the NMDA receptor. Lipocortin-1 fragment markedly attenuated, and the neutralizing antibody increased, this NMDA mediated neuronal damage. These observations may explain the neuroprotective action of lipocortin following cerebral ischaemia.

Topics: Animals; Annexins; Calcium-Binding Proteins; Corpus Striatum; Dizocilpine Maleate; Dose-Response Relationship, Drug; Glutamates; Male; Nerve Degeneration; Neurons; Peptide Fragments; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate

Photochemically induced ischemic lesions in the rat spinal cord were studied using neurological tests and morphological evaluation in order to investigate ischemia-mediated pathophysiological mechanisms in traumatic spinal cord injury. One week after ischemic lesioning, animals were severely impaired with 85% decrease of performance in neurological tests. During the next 2 weeks considerable recovery occurred. Pretreatment with the noncompetitive N-methyl-D-aspartate antagonist MK-801 at a dose of 0.5-1.0 mg/kg significantly improved the recovery of function after spinal ischemia while lower doses exerted no protection. Morphologically, no dose-response effect on the extent of tissue necrosis was found, but a significant difference between groups with severe neurological deficit versus mildly affected groups was observed. Immunohistochemical staining for glial fibrillary acidic protein in the area close to the lesion revealed extensive gliosis, while neurofilament immunohistochemistry showed an irregular pattern of fiber loss with large variability between animals. The degree of gliosis or loss of neurofilament immunoreactivity in nonnecrotic tissue was not affected by MK-801. These results suggest that excessive stimulation of N-methyl-D-aspartate receptors participates in the development of spinal cord ischemia and possibly also participates after traumatic spinal cord injury.

Topics: Animals; Behavior, Animal; Dizocilpine Maleate; Female; Glial Fibrillary Acidic Protein; Immunohistochemistry; Ischemia; Lasers; N-Methylaspartate; Nerve Degeneration; Neurofilament Proteins; Radiation Injuries, Experimental; Rats; Rats, Sprague-Dawley; Spinal Cord

In cynomologus monkeys, systemic administration of MK-801, a noncompetitive antagonist for the N-methyl-D-aspartate receptor, prevented the development of the parkinsonian syndrome induced by the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). MK-801 also attenuated dopamine depletion in the caudate and putamen and protected dopaminergic neurons in the substantia nigra from the degeneration induced by the neurotoxin. Nevertheless, 7 days after MPTP administration in the caudate and putamen of monkeys also receiving MK-801, the levels of toxic 1-methyl-4-phenylpyridinium were even higher than those measured in monkeys receiving MPTP alone. This indicates that the protective action of MK-801 is not related to MPTP metabolism and strongly suggests that, in primates, the excitatory amino acids could play a crucial role in the mechanism of the selective neuronal death induced by MPTP.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; 1-Methyl-4-phenylpyridinium; Animals; Caudate Nucleus; Disease Models, Animal; Dizocilpine Maleate; Dopamine; Dose-Response Relationship, Drug; Macaca fascicularis; Male; Nerve Degeneration; Neurons; Parkinson Disease, Secondary; Putamen; Substantia Nigra

Injection of the N-methyl-D-aspartate receptor agonist quinolinate, or N-methyl-D-aspartate itself, into the rat brain produces neurodegeneration which can be prevented by N-methyl-D-aspartate receptor antagonists administered up to 5 h after excitotoxin injection. The present study was designed to investigate aspects of the mechanisms involved in this delayed form of neurodegeneration. Following its injection into the rat striatum, extracellular levels of [3H]quinolinate were monitored using a microdialysis probe located 1 mm from the site of injection. Peak concentrations were observed 10-20 min after injection and [3H]quinolinate levels decayed in a biexponential fashion, the initial component having an apparent t1/2 of 13.7 +/- 5.2 min (n = 3). Estimations of the extracellular concentrations of quinolinate after an injection of 200 nmol indicated a peak level of 13.7 +/- 6.0 mM (n = 3) at 10-20 min which declined to 1.2 +/- 0.13 mM (n = 3) by 2 h and substantial levels were present up to 5 h, the period over which N-methyl-D-aspartate receptor antagonists are effective in this model. Administration of dizocilpine at 1, 2, 3 or 5 h after injection of 100, 200 or 400 nmol quinolinate resulted in a similar temporal profile of neuroprotection, as assessed by measuring the activities of choline acetyltransferase and glutamate decarboxylase in striatal homogenates, which was independent of the degree of neurodegeneration produced by the different excitotoxin doses. Overall, these results suggest that the neuronal degeneration caused by quinolinate in vivo is critically dependent upon events occurring after the initial peak of excitoxin levels in the extracellular space.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adrenergic alpha-Antagonists; Animals; Anticonvulsants; Convulsants; Corpus Striatum; Dialysis; Diazepam; Dizocilpine Maleate; Glutamate Decarboxylase; Haloperidol; Ibotenic Acid; Injections; Male; Nerve Degeneration; Pipecolic Acids; Piperazines; Piperidines; Quinolinic Acid; Quinolinic Acids; Rats; Rats, Inbred Strains; Stereotaxic Techniques

Recent evidence implicates excitatory amino acids (EAAs), acting as excitotoxic agents, in the pathogenesis of neurological disorders involving the spinal cord. In this study, we used the chick embryo spinal cord as an in vitro model for studying the sensitivity of spinal neurons to the excitotoxic effects of EAA agonists. Compounds tested include the prototypic receptor-specific agonists, N-methyl-D-aspartate (NMDA), quisqualic acid (Quis), and kainic acid (KA), and the plant-derived excitotoxic food poisons, beta-N-oxalylamino-L-alanine, beta-N-methylamino-L-alanine, and domoic acid. Each agonist induced concentration-dependent acute degeneration of neurons distributed throughout the spinal cord. These cytopathological changes consisted of acute edematous degeneration of dendrosomal structures in the dorsal horn and intermediate zone, and dark cell changes with intracytoplasmic vacuolization of motor neurons; this damage is identical to that induced by excitotoxin agonists in other regions of the central nervous system. The NMDA receptor-specific antagonist MK-801 completely blocked toxicity of NMDA, and the nonNMDA antagonist CNQX preferentially blocked the toxicity of Quis- and KA-type agonists in the spinal cord. Our findings suggest that (1) the majority of spinal neurons have all three subtypes of EAA receptors, making them acutely vulnerable to excitotoxin exposure; and (2) EAA antagonists are effective in preventing excitotoxin-induced damage of the spinal cord.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Amino Acids, Diamino; Animals; beta-Alanine; Chick Embryo; Cyanobacteria Toxins; Dizocilpine Maleate; Glutamates; Glutamic Acid; Ibotenic Acid; Kainic Acid; Motor Neurons; N-Methylaspartate; Nerve Degeneration; Neurotoxins; Quinoxalines; Quisqualic Acid; Spinal Cord

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Aging; Alzheimer Disease; Animals; Dizocilpine Maleate; Haplorhini; Humans; Huntington Disease; Nerve Degeneration; Neurotoxins; Parkinson Disease; Quinolinic Acid; Quinolinic Acids; Receptors, Dopamine

The ability of the N-methyl-D-aspartate receptor antagonists, MK-801, ketamine and alaptide [a newly synthesized cyclo(1-amino-1-cyclopentane-carbonyl-L-alanyl) with protective properties in models of hypoxia], to prevent neuronal degeneration caused by intracerebroventricular application of quinolinic acid was investigated. Neurodegenerative effects of quinolinate in the hippocampal formation were found to increase with the degree of maturity of glutamatergic target structures. A protective potency of the N-methyl-D-aspartate receptor antagonists was observed at all developmental stages studied (12- and 30-day-old and adult rats). MK-801 showed the highest efficacy, alaptide the lowest. These findings suggest a parallelism in maturity of glutamatergic transmission processes as one prerequisite of quinolinate vulnerability and postnatal increases of target fields of the protectives. Application of MK-801 or ketamine after quinolinate injection intensified their protective effects when compared to simultaneous or preadministration. This observation is interpreted as indicating that quinolinate is a prompter of a delayed neurodegenerative process rather than acting immediately as a toxicant.

Topics: Animals; Cell Hypoxia; Dizocilpine Maleate; Hippocampus; Injections, Intraventricular; Ketamine; Male; Nerve Degeneration; Neuropeptides; Peptides, Cyclic; Quinolinic Acid; Quinolinic Acids; Rats; Rats, Inbred Strains

The purpose of this study was to determine whether prior transient cerebral ischemia, in conscious mice, would alter the biological responses resulting from excessive activation of N-methyl-D-aspartate (NMDA) receptors, in an early stage. The responses to the activation of NMDA receptors by an intracerebroventricular injection of NMDA, such as wild running, tonic and clonic convulsions, absence of the visual placing reflex, loss of the righting reflex, impaired motor function and a high mortality rate, were to a large extent prevented if 30 min before treatment, either a 10-min period of global cerebral ischemia was induced or a 1 nmol intraventricular injection of NMDA was given but not if either of the above procedures was done one day before the test dose of NMDA. In contrast, behavioral symptoms, in response to activation of non-NMDA-type glutamate receptors elicited by intraventricular injection of either kainic acid or AMPA, were not clearly affected. Transient systemic hypercapnic anoxia (22-sec exposure to 100% CO2 gas), before treatment with NMDA did not significantly reduce the NMDA-induced behavior. The severity of these behavioral responses and high mortality rate observed after intraventricular injection of pentylenetetrazole (PTZ, 30 mumol) were not altered by either prior global ischemic insult or by a preexposure to NMDA given intraventricularly. The NMDA antagonist, MK801 (0.1 and 0.3 mg/kg i.p.) greatly reduced the behavioral effects and mortality rate, resulting from the intraventricular injection of NMDA and somewhat reduced the effects of the intraventricular injection of PTZ.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Behavior, Animal; Carbon Dioxide; Dizocilpine Maleate; Female; Hypoxia; Injections, Intraventricular; Ischemic Attack, Transient; Mice; Mice, Inbred Strains; N-Methylaspartate; Nerve Degeneration; Pentylenetetrazole; Receptors, N-Methyl-D-Aspartate; Survival Rate

Using the excitotoxic animal model of Huntington's disease, two experimental treatments were evaluated. The first experiment explored the effect of MK801 (a systemically active anticonvulsant, and noncompetitive NMDA antagonist) pretreatment on quinolinic acid (QA)-induced striatal degeneration and behavioral deficits. MK801 prevented QA-induced neuropathological changes in the striatum and the anatomical protection was correlated with the absence of deficits in the cataleptic response to haloperidol. The second experiment tested the ability of three types of fetal grafts to reverse behavioral deficits induced by kainic acid (KA) lesion. Fetal (E15-16) striatal, cortical and tectal grafts were delivered into the KA-lesioned striatum one week or one month after lesion. Animals in this experiment were evaluated on a motor coordination task, haloperidol-induced catalepsy and amphetamine-induced locomotor activity. Striatal grafts attenuated the deficits induced by KA in all of the tasks observed, and no effect of time of grafting was detected. Tectal grafts had a partial beneficial effect, attenuating the decrease in the cataleptic response to haloperidol observed after KA lesions. No effect of time of grafting was detected for these grafts. In contrast, a clear effect of time of grafting was detected for the cortical grafts. Early cortical grafts reversed the exaggerated response to amphetamine observed after KA lesions whereas late cortical grafts resulted in sham-like scores on the catalepsy test. Histochemical analysis showed that most of the grafts survived, had acetylcholinesterase (AChE) positive fibers and cell bodies, and were metabolically active as indicated by cytochrome oxidase (CO) positive staining. It is suggested that striatal grafts may have restored to some extent the striatal GABAergic control over output structures, and that trophic factors play a role in behavioral recovery as is evident from the beneficial effects of the tectal grafts. Although the mechanisms underlying the differential effects observed after early or late cortical grafts are unknown, the interaction between the cellular components and trophic factors present in the cortical grafts and the condition of the lesioned host at the time of grafting may yield a host-graft complex with a unique profile.

Topics: Acetylcholinesterase; Amphetamine; Animals; Brain Tissue Transplantation; Catalepsy; Cerebral Cortex; Corpus Striatum; Dizocilpine Maleate; Electron Transport Complex IV; Female; Fetal Tissue Transplantation; Huntington Disease; Immunohistochemistry; Kainic Acid; Motor Activity; Nerve Degeneration; Pregnancy; Quinolinic Acid; Quinolinic Acids; Rats; Rats, Inbred Strains

After electrical stimulation of the optic layer (OL) of superior colliculus (SC) slices, the postsynaptic potential (PSP) was recorded in the superficial gray layer (SGL) of the SC. The degeneration studies of retinotectal or corticotectal inputs to the SGL of the SC indicated that this PSP evoked in the SGL of the SC slices was retinotectal in origin. Neurotransmission in this pathway may be mediated by glutamate, because the PSP amplitude was reduced and blocked by application of kynurenate or quinoxaline dione (DNQX) to the medium. Furthermore, the concentration of glutamate in the right SGL was significantly reduced by 32% after left optic denervation and by 30% after ablation of the right visual cortex, compared with that in the left SGL. Long-term potentiation (LTP) in the SGL was induced by tetanic stimulation (50 Hz, 20 s) to the OL. The LTP formation was facilitated by the removal of Mg2+ from the medium. The effects of glutamate antagonists D-amino-5-phosphonovalerate (D-APV), gamma-D-glutamylglycine (gamma-DGG), and (+)-5-methyl-10,11-dihydro-5H-dibenzo, a,d-cycloheptene-5,10-imine maleate (MK-801) on the induction of LTP were investigated. D-APV (100 microM) or gamma-DGG (1 mM) masked the expression of LTP by tetanic stimulation, however LTP was induced after removal of the agents. LTP formation was observed without further tetanic stimulation following the removal of D-APV from the medium even 80 min after the tetanic stimulation. LTP once formed was not influenced by application of D-APV.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 2-Amino-5-phosphonovalerate; Animals; Aspartic Acid; Denervation; Dibenzocycloheptenes; Dipeptides; Dizocilpine Maleate; Evoked Potentials; Excitatory Amino Acid Antagonists; gamma-Aminobutyric Acid; Glutamates; Glutamic Acid; In Vitro Techniques; Kynurenic Acid; Magnesium; Mice; Nerve Degeneration; Optic Nerve; Quinoxalines; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter; Superior Colliculi; Visual Cortex

The neuroprotective potential of an antagonist (7-chlorokynurenic acid (7-CIKYNA)) and a low efficacy partial agonist (HA-966) for the glycine modulatory site on the N-methyl-D-aspartate (NMDA) receptor complex has been examined using a neuronal cell culture/hypoxia model of neurodegeneration. Their effects were compared to those of the potent uncompetitive NMDA antagonist, MK-801. Hypoxic cell injury was assessed visually and quantified by measuring the appearance of two cytosolic enzymes, lactate dehydrogenase (LDH) and neurone specific enolase (NSE), in the culture medium. MK-801 prevented the hypoxia-induced cell mortality in a concentration-related manner with an IC50 of 15 nM against increases in LDH levels. HA-966 and 7-CIKYNA also produced concentration-related protective effects with IC50s of 175 and 18 microM, respectively. Although both glycine antagonists were considerably weaker than MK-801 their maximum neuroprotective effects were comparable to that produced by MK-801, i.e. complete protection. This indicates that the level of NMDA receptor activation which can take place in the presence of the partial agonist HA-966 is insufficient to cause permanent neuronal damage. Concentration-effect curves were similar when NSE was used as the marker enzyme, supporting previous observations that the increases in LDH levels accurately and specifically reflect neuronal cell death. These results provide further evidence that hypoxia-induced injury to cortical neuronal cultures is mediated by an excessive stimulation of NMDA receptors and that glycine-site antagonists and partial agonists may have therapeutic potential in conditions where pathologically high levels of NMDA receptor activation are thought to occur.

Topics: Animals; Binding Sites; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Fetus; Glycine; Hypoxia; Kinetics; Kynurenic Acid; L-Lactate Dehydrogenase; Nerve Degeneration; Neurons; Phosphopyruvate Hydratase; Pyrrolidinones; Rats; Receptors, N-Methyl-D-Aspartate

The in vivo efficacies and potencies of various excitatory amino acid agonists in inducing cholinergic neuronal degeneration were compared following unilateral injections into the rat striatum. Kainic acid (KA), alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), ibotenic acid (IBO), and N-methyl-D-aspartic acid (NMDA) all produced dose-related decreases in choline acetyltransferase (ChAT) activity. The relative order of potency was KA greater than AMPA greater than IBO greater than NMDA. Quisqualic acid (QUIS) was about as potent as NMDA, but the maximal decrease in ChAT activity was less (36%). N-acetylaspartyl-L-glutamate (NAAG) did not significantly decrease ChAT activity when up to 1,000 nmoles was injected. Approximate equitoxic doses of agonists were then used to examine the ability of i.p. administered CGS-19755 and MK-801 to prevent in vivo excitatory amino acid-induced cholinergic and GABAergic neuronal degeneration. NMDA-induced decreases in ChAT and glutamic acid decarboxylase (GAD) activities were prevented by CGS-19755 (10-40 mg/kg) and MK-801 (1-10 mg/kg). CGS-19755 (40 mg/kg) and MK-801 (10 mg/kg) did not prevent loss of ChAT or GAD induced by KA or AMPA, but did prevent the degenerative effects of IBO. This study shows that CGS-19755 and MK-801, two NMDA receptor antagonists that act by different mechanisms, are completely selective following systemic administration. Moreover, the in vivo excitotoxic effects of IBO are mediated at NMDA receptor sites that are blocked by these compounds.

Topics: Animals; Aspartic Acid; Choline O-Acetyltransferase; Cholinergic Fibers; Corpus Striatum; Dibenzocycloheptenes; Dizocilpine Maleate; gamma-Aminobutyric Acid; Ibotenic Acid; Male; N-Methylaspartate; Nerve Degeneration; Oxazoles; Pipecolic Acids; Piperidines; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter

Topics: 3,4-Methylenedioxyamphetamine; Animals; Anticonvulsants; Brain; Corpus Striatum; Dibenzocycloheptenes; Dizocilpine Maleate; Male; Methamphetamine; N-Methyl-3,4-methylenedioxyamphetamine; Nerve Degeneration; Rats; Rats, Inbred Strains; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter; Receptors, Serotonin; Tryptophan Hydroxylase

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

Ketamine and MK-801 act at phencyclidine receptors to block transmitter activity through the N-methyl-D-aspartate (NMDA) subtype of glutamate (GLU) receptor. These agents also block the potent excitotoxic actions of NMDA and are of interest for their potential ability to protect against neurodegenerative processes mediated by the excitotoxic action of endogenous Glu at NMDA receptors. Here we show that degeneration of thalamic neurons caused by persistent seizure activity in the corticothalamic tract (putative glutamergic transmitter pathway) is prevented by systemic administration of ketamine or MK-801, despite the failure of these agents to eliminate persistent electrographic seizure activity recorded from cortex and thalamus.

Topics: Animals; Anticonvulsants; Bicuculline; Cerebral Cortex; Dibenzocycloheptenes; Dizocilpine Maleate; Electroencephalography; Ketamine; Nerve Degeneration; Neurons; Rats; Rats, Inbred Strains; Seizures; Thalamus

The ability of the noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 to prevent neuronal degeneration in the rat striatum and hippocampus caused by intracerebral injection of excitotoxins has been examined. Excitotoxic damage was assessed after 7 d, using histological and biochemical [choline acetyltransferase (ChAT) glutamate decarboxylase (GAD)] measurements. Systemically administered MK-801 was found to protect against neurodegeneration caused by NMDA (200 nmol) and the naturally occurring NMDA receptor agonist quinolinate (120-600 nmol) but not against that induced by kainate (5 nmol) or alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA; 50 nmol), indicating a selectivity for NMDA receptor-mediated neuronal loss. Neurotoxicity caused by NMDA (200 nmol) or quinolinate (200 nmol) was prevented by MK-801 (1-10 mg/kg, i.p.) administered in a single dose after excitotoxin injection. In the striatum, significant protection of cholinergic neurons (assessed by ChAT measurements) was observed when MK-801 was given up to 5 hr after injection of NMDA or quinolinate, whereas protection of GABAergic neurons (assessed by GAD measurements) was obtained up to 2 hr. The results suggest that GABAergic neurons degenerate more rapidly than cholinergic neurons. The competitive NMDA receptor antagonist 3-[(+/-)-2-carboxypiperazin-4-yl]-propyl-1-phosphonate (100 mg/kg, i.p.) gave partial protection of striatal neurons when administered 1 hr after quinolinate injection. In the rat hippocampus, administration of 10 mg/kg MK-801 i.p. 1 hr after quinolinate injection caused almost complete protection of pyramidal and granule neurons, whereas the degeneration of CA3/CA4 pyramidal neurons caused by kainate injection was unaffected. These observations indicate that neurons in rat striatum and hippocampus do not die as an immediate consequence of exposure to high concentrations of NMDA agonists but that a delayed process is involved that requires NMDA receptor activation. In this respect, intracerebral injections of NMDA agonists may mimic the pathological changes that are thought to occur in the brain following periods of cerebral ischemia, where delayed neuronal degeneration occurs.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Anesthesia; Animals; Aspartic Acid; Brain; Dibenzocycloheptenes; Dizocilpine Maleate; Ibotenic Acid; Isoflurane; Kainic Acid; Male; N-Methylaspartate; Nerve Degeneration; Piperazines; Quinolinic Acid; Quinolinic Acids; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter; Time Factors

The neuroprotective effects of the non-competitive N-methyl-D-aspartate receptor antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801) have been evaluated in the gerbil hippocampus when the drug was administered i.p. at various times during and after a 5 min period of transient forebrain ischaemia, induced by bilateral common carotid artery occlusion. A single dose of 1, 3 or 10 mg/kg of MK-801 gave significant protection of hippocampal CA1 and CA2 pyramidal neurons when administered during the occlusion and up to 24 h following the period of ischaemia. A dose of 0.3 mg/kg was effective when administered during the occlusion period but gave no protection at 30 min or 2 h post-ischaemia. Experiments in which MK-801 was administered in repeated doses indicated that significant protection was achieved with 1 mg/kg of MK-801 repeated post-ischaemically and with 1 mg/kg MK-801 supplemented with repeated doses of 0.3 mg/kg of MK-801. However 0.3 mg/kg of MK-801 followed by repeated doses of 0.03 mg/kg administered post-ischaemically was not neuroprotective. These results indicate that MK-801 can protect hippocampal neurons from ischaemia-induced neuronal degeneration when it is administered up to 24 h after the insult. These data provide further evidence that therapeutic intervention in the post-ischaemic period can successfully prevent neurodegenerative events, and that the delayed degeneration of hippocampal neurons following an ischaemic insult occurs by an N-methyl-D-aspartate receptor-mediated process.

Topics: Animals; Dibenzocycloheptenes; Dizocilpine Maleate; Dose-Response Relationship, Drug; Female; Gerbillinae; Hippocampus; Ischemic Attack, Transient; Male; Nerve Degeneration; Time Factors

MK-801 is a novel, potent and selective non-competitive antagonist of the N-methyl-D-aspartate (NMDA) subtype of excitatory amino acid receptors. Pretreatment of rats with MK-801 (1-10 mg/kg, i.p.) prevented neuronal degeneration in the hippocampus and striatum caused by direct intracranial injections of NMDA (20-120 nmol), but did not protect against the loss of neurones induced by kainate (2.5 nmol) injected into the striatum. Thus, MK-801 is a selective antagonist of neuronal degeneration caused by excessive stimulation of NMDA receptors in vivo.

Topics: Animals; Aspartic Acid; Choline O-Acetyltransferase; Corpus Striatum; Dibenzocycloheptenes; Dizocilpine Maleate; Hippocampus; Kainic Acid; Male; N-Methylaspartate; Nerve Degeneration; Rats; Rats, Inbred Strains

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