dizocilpine-maleate and 2-3-dioxo-6-nitro-7-sulfamoylbenzo(f)quinoxaline

Hypoxia is the most common cause of neonatal seizures and encephalopathy. We have previously developed an in vivo experimental model of perinatal hypoxia which exhibits age-dependent acute and chronic epileptogenic effects. Between postnatal day (P) 10-12, the rat exhibits acute seizure activity during global hypoxia, while no seizures are induced at earlier (P5) or older (P60) ages. Rats exposed to hypoxia between P10-12 have reduced seizure thresholds to chemical convulsants in adulthood. The nonNMDA antagonists NBQX appears to suppress both the acute and long term epileptogenic effects of hypoxia. The age-dependency of the hyperexcitable response to hypoxia in vivo can be reproduced in vitro using hippocampal slices. In Mg(2+)-free media, hypoxia induced ictal discharges within 60 s of onset in 79% of slices from normal P10 rat pups compared to 11% of adult slices (p < 0.001). Model systems such as that described here allow for correlation of in vitro and in vivo electrophysiology and should provide data regarding the pharmacological and physiological characteristics of hypoxia-induced seizure activity in the immature brain which could ultimately be applied to therapeutic strategies.

Topics: Age Factors; Animals; Disease Models, Animal; Dizocilpine Maleate; Electroencephalography; Electrophysiology; Epilepsy, Temporal Lobe; Hippocampus; Hypoxia; In Vitro Techniques; Lorazepam; Quinoxalines; Rats

Current antiparkinsonian therapies focus on either replacing dopamine via precursor (L-DOPA) administration, or directly stimulating post-synaptic dopamine receptors with dopamine agonists. Unfortunately, this approach is associated with numerous side effects and these drugs lose efficacy with disease progression. This article reviews recent evidence which suggests that negative modulation of glutamatergic neurotransmission has antiparkinsonian effects in a variety of rodent and primate models of parkinsonism. The pronounced synergism between dopaminergic agents and glutamate receptor antagonists may provide a means of using very low doses of the two drug classes in concert to treat Parkinson's disease effectively and minimize dose-related drug side effects.

Topics: Animals; Antiparkinson Agents; Basal Ganglia; Disease Models, Animal; Dizocilpine Maleate; Dopamine; Dopamine Agents; Drug Evaluation, Preclinical; Drug Synergism; Drug Tolerance; Excitatory Amino Acid Antagonists; Glutamates; Glutamic Acid; Haplorhini; Humans; Mice; MPTP Poisoning; Oxidopamine; Parkinson Disease; Parkinson Disease, Secondary; Quinoxalines; Rats; Receptors, Dopamine; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Synaptic Transmission

d-Serine plays an important role in modulating N-methyl-d-aspartate receptor (NMDAR) neurotransmission in the mammalian brain by binding to the receptor's glycine modulatory site (GMS). The cytosolic enzyme serine racemase (SR) converts L-serine to d-serine, while the peroxisomal enzyme d-amino acid oxidase (DAAO) catalyzes the breakdown of d-serine. Although it is important to understand how the activities of SR and DAAO regulate d-serine levels, very little is known about the mechanisms that regulate the expression of SR and DAAO. In this study, we investigated whether the different centrally active drugs affect the expression of SR and DAAO in adult mouse brain. We found that the NMDAR antagonist, MK801, and cocaine, psychotropic drugs that both augment glutamate release, reduce the expression of SR and DAAO. This regulation is brain region selective, and in the case of cocaine, is reversed in part byα-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo[f]quinoxaline-2,3-dione (NBQX). However, d-serine and antipsychotics do not regulate SR and DAAO protein levels. In a genetic model of SR disruption, we found that DAAO expression was unaltered in SR conditional knockout mice, in which tissue d-serine content remains fairly stable despite marked reduction in SR expression. This study reveals a new mechanism by which AMPAR activity could regulate NMDAR function via d-serine availability.

Topics: Animals; Brain; Cocaine; Corpus Striatum; D-Amino-Acid Oxidase; Dizocilpine Maleate; Female; Gene Expression; Gene Expression Regulation; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Quinoxalines; Racemases and Epimerases; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Serine

Mechanism of ictogenesis of D- and L-stereroisomers of homocysteic acid was studied in 12-day-old rats by means of antagonists of N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors. There was no qualitative difference between the two stereoisomers in generation of emprosthotonic (flexion) as well as generalized tonic-clonic seizures. Moderate differences were observed in the first, nonconvulsive effects of the two isomers. As generation of the two types of seizures is concerned, NMDA and AMPA participate in generalized tonic-clonic seizures whereas NMDA receptors play a dominant role in generation of flexion seizures.

Topics: 2-Amino-5-phosphonovalerate; Animals; Benzodiazepines; Dizocilpine Maleate; Homocysteine; Male; Quinoxalines; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Seizures; Stereoisomerism

Topics: Animals; Apoptosis; Cells, Cultured; Dizocilpine Maleate; Dopaminergic Neurons; Kainic Acid; Membrane Potential, Mitochondrial; Mesencephalon; Mice; Neuromuscular Depolarizing Agents; Neurotoxins; Quinoxalines; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate

The ionotropic glutamate receptorantagonists include two types: MK-801, antagonist of N-methyl-D-asparticacid (NMDA) receptor, and NBQX, antagonist of non-NMDA receptor.The above-mentioned ionotropic antagonists can block the glutamate and its corresponding receptor binding to produce analgesic effect. The objective of this research was to study two antagonists in analgesic effect on rat behavior,as well as to investigate the down-regulation and up-regulation of cyclooxygenase-2 (COX-2) and Janus-activated kinase (Jak3) in collagen-induced arthritis (CIA) rat serum and tissue fluid after the application of these antagonists, that is, the effect on molecular biology.. This study used the ionotropic glutamate receptors as the target and established CIA rat model. Vivo studies were used to observe changes in behavior and molecular biology of the CIA rat.Behavioral assessment includedmechanical allodynia and joint swelling in the CIA rat,where themechanical allodynia was measured using the paw-withdrawal threshold (PWT) with VonFrey filaments according to the "Up-Down" method,and the drainage volume was used to assess joint swelling. Then the blood samples taken from the heart of the rat and the tissue homogenate were collected to detect the down-regulation and up-regulation of COX-2 and Jak3 in the serum and tissue fluid after the antagonists wereused.. Using MK-801, NBQX alone or using the combination of these two antagonists,these three methods all could alleviate pain(P<0.01).The analgesic effect lasted more than 24 h.Both antagonists reached the peak of analgesia at the end of 4 hours post-injection. NBQX had stronger analgesic effect than MK-801 (P<0.05).Whether alone or combined use of these two antagonists,could not change the CIA rats' swelling of the joint (P>0.05). MK-801 could decrease the expression of COX-2 (P<0.01).At the same time, NBQX did not have this effect (P>0.05). Using MK-801, NBQX alone or combination of these two antagonists could not affect the increased expression of Jak3 caused by the CIA (P>0.05).. MK-801 and NBQX could both alleviate pain, NBQX was much better than MK-801. Neither MK-801 nor NBQX had the effect on the swelling of the joint. NMDA receptor and COX-2 inflammatory pathways had certain interactions. For Jak3, it could not be found to have cross-function with ionotropic glutamate signaling pathways by this experiment.

Topics: Analgesics; Animals; Arthritis, Experimental; Cyclooxygenase 2; Dizocilpine Maleate; Edema; Excitatory Amino Acid Antagonists; Gene Expression Regulation; Hyperalgesia; Janus Kinase 3; Male; Pain; Quinoxalines; Rats; Receptors, N-Methyl-D-Aspartate

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

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

Excitotoxicity plays a critical role in neurodegenerative disease. Cytosolic calcium overload and mitochondrial dysfunction are among the major mediators of high level glutamate-induced neuron death. Here, we show that the transient opening of mitochondrial permeability transition pore (tMPT) bridges cytosolic calcium signaling and mitochondrial dysfunction and mediates glutamate-induced neuron death. Incubation of the differentiated motor neuron-like NSC34D cells with glutamate (1mM) acutely induces cytosolic calcium transient (30% increase). Glutamate also stimulates tMPT opening, as reflected by a 2-fold increase in the frequency of superoxide flash, a bursting superoxide production event in individual mitochondria coupled to tMPT opening. The glutamate-induced tMPT opening is attenuated by suppressing cytosolic calcium influx and abolished by inhibiting mitochondrial calcium uniporter (MCU) with Ru360 (100 μM) or MCU shRNA. Further, increased cytosolic calcium is sufficient to induce tMPT in a mitochondrial calcium dependent manner. Finally, chronic glutamate incubation (24h) persistently elevates the probability of tMPT opening, promotes oxidative stress and induces neuron death. Attenuating tMPT activity or inhibiting MCU protects NSC34D cells from glutamate-induced cell death. These results indicate that high level glutamate-induced neuron toxicity is mediated by tMPT, which connects increased cytosolic calcium signal to mitochondrial dysfunction.

Topics: Animals; Calcium Signaling; Cell Line, Transformed; Cyclosporine; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Glutamic Acid; Hybrid Cells; Membrane Potential, Mitochondrial; Mice; Mitochondria; Mitochondrial Membrane Transport Proteins; Mitochondrial Permeability Transition Pore; Motor Neurons; Oxidative Stress; Quinoxalines; Receptors, AMPA; Ruthenium Compounds; Time Factors; Transfection

We investigated the link between direct activation of inhibitory neurons, local neuronal activity, and hemodynamics. Direct optogenetic cortical stimulation in the sensorimotor cortex of transgenic mice expressing Channelrhodopsin-2 in GABAergic neurons (VGAT-ChR2) greatly attenuated spontaneous cortical spikes, but was sufficient to increase blood flow as measured with laser speckle contrast imaging. To determine whether the observed optogenetically evoked gamma aminobutyric acid (GABA)-neuron hemodynamic responses were dependent on ionotropic glutamatergic or GABAergic synaptic mechanisms, we paired optogenetic stimulation with application of antagonists to the cortex. Incubation of glutamatergic antagonists directly on the cortex (NBQX and MK-801) blocked cortical sensory evoked responses (as measured with electroencephalography and intrinsic optical signal imaging), but did not significantly attenuate optogenetically evoked hemodynamic responses. Significant light-evoked hemodynamic responses were still present after the addition of picrotoxin (GABA-A receptor antagonist) in the presence of the glutamatergic synaptic blockade. This activation of cortical inhibitory interneurons can mediate large changes in blood flow in a manner that is by and large not dependent on ionotropic glutamatergic or GABAergic synaptic transmission. This supports the hypothesis that activation of inhibitory neurons can increase local cerebral blood flow in a manner that is not entirely dependent on levels of net ongoing neuronal activity.

Topics: Animals; Cerebrovascular Circulation; Channelrhodopsins; Dizocilpine Maleate; Electroencephalography; Excitatory Amino Acid Antagonists; Female; GABA Antagonists; GABAergic Neurons; gamma-Aminobutyric Acid; Interneurons; Lasers; Male; Mice; Optogenetics; Photic Stimulation; Quinoxalines

We investigated the role of peripheral NMDA receptors (NMDARs) in antidromic nerve stimulation-induced tactile hypersensitivity outside the skin area innervated by stimulated nerve. Tetanic electrical stimulation (ES) of the decentralized L5 spinal nerve, which induced enlargement of plasma extravasation, resulted in tactile hypersensitivity in the L4 plantar dermatome of the hind-paw. When intraplantar (i.pl.) injection was administered into the L4 dermatome before ES, NMDAR and group-I metabotropic Glu receptor (mGluR) antagonists and group-II mGluR agonist but not AMPA/kainate receptor antagonist prevented ES-induced hypersensitivity. I.pl. injection of PKA or PKC inhibitors also prevented ES-induced hypersensitivity. When the same injections were administered after establishment of ES-induced hypersensitivity, hypersensitivity was partially reduced by NMDAR antagonist only. In naïve animals, i.pl. Glu injection into the L4 dermatome induced tactile hypersensitivity, which was blocked by NMDAR antagonist and PKA and PKC inhibitors. These results suggest that the peripheral release of Glu, induced by antidromic nerve stimulation, leads to the expansion of tactile hypersensitive skin probably via nociceptor sensitization spread due to the diffusion of Glu into the skin near the release site. In addition, intracellular PKA- and PKC-dependent mechanisms mediated mainly by NMDAR activation are involved in Glu-induced nociceptor sensitization and subsequent hypersensitivity.

Topics: Animals; Cyclic AMP-Dependent Protein Kinases; Dizocilpine Maleate; Electric Stimulation; Glutamic Acid; Hyperalgesia; Isoquinolines; Male; Protein Kinase C; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Sulfonamides; Tibial Nerve

When exposed to a hypoxic environment the body's first response is a reflex increase in ventilation, termed the hypoxic ventilatory response (HVR). With chronic sustained hypoxia (CSH), such as during acclimatization to high altitude, an additional time-dependent increase in ventilation occurs, which increases the HVR. This secondary increase persists after exposure to CSH and involves plasticity within the circuits in the central nervous system that control breathing. Currently these mechanisms of HVR plasticity are unknown and we hypothesized that they involve glutamatergic synapses in the nucleus tractus solitarius (NTS), where afferent endings from arterial chemoreceptors terminate. To test this, we treated rats held in normoxia (CON) or 10% O2 (CSH) for 7 days and measured ventilation in conscious, unrestrained animals before and after microinjecting glutamate receptor agonists and antagonists into the NTS. In normoxia, AMPA increased ventilation 25% and 50% in CON and CSH, respectively, while NMDA doubled ventilation in both groups (P < 0.05). Specific AMPA and NMDA receptor antagonists (NBQX and MK801, respectively) abolished these effects. MK801 significantly decreased the HVR in CON rats, and completely blocked the acute HVR in CSH rats but had no effect on ventilation in normoxia. NBQX decreased ventilation whenever it was increased relative to normoxic controls; i.e. acute hypoxia in CON and CSH, and normoxia in CSH. These results support our hypothesis that glutamate receptors in the NTS contribute to plasticity in the HVR with CSH. The mechanism underlying this synaptic plasticity is probably glutamate receptor modification, as in CSH rats the expression of phosphorylated NR1 and GluR1 proteins in the NTS increased 35% and 70%, respectively, relative to that in CON rats.

Topics: Acclimatization; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Chemoreceptor Cells; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Hypoxia; Male; N-Methylaspartate; Pulmonary Ventilation; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Glutamate; Reflex; Solitary Nucleus

The hippocampus is an important brain region that is involved in neurological disorders such as Alzheimer disease, schizophrenia, and epilepsy. Ionotropic glutamate receptors-namely,N-methyl-D-aspartate (NMDA) receptors (NMDARs), α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptors (AMPARs), and kainic acid (KA) receptors (KARs)-are well known to be involved in these diseases by mediating long-term potentiation, excitotoxicity, or both. To predict the therapeutic efficacy and neuronal toxicity of drug candidates acting on these receptors, physiologically relevant systems for assaying brain region-specific human neural cells are necessary. Here, we characterized the functional differentiation of human fetal hippocampus-derived neural stem/progenitor cells-namely, HIP-009 cells. Calcium rise assay demonstrated that, after a 4-week differentiation, the cells responded to NMDA (EC50= 7.5 ± 0.4 µM; n= 4), AMPA (EC50= 2.5 ± 0.1 µM; n= 3), or KA (EC50= 33.5 ± 1.1 µM; n= 3) in a concentration-dependent manner. An AMPA-evoked calcium rise was observed in the absence of the desensitization inhibitor cyclothiazide. In addition, the calcium rise induced by these agonists was inhibited by antagonists for each receptor-namely, MK-801 for NMDA stimulation (IC50= 0.6 ± 0.1 µM; n= 4) and NBQX for AMPA and KA stimulation (IC50= 0.7 ± 0.1 and 0.7 ± 0.03 µM, respectively; n= 3). The gene expression profile of differentiated HIP-009 cells was distinct from that of undifferentiated cells and closely resembled that of the human adult hippocampus. Our results show that HIP-009 cells are a unique tool for obtaining human hippocampal neural cells and are applicable to systems for assay of ionotropic glutamate receptors as a physiologically relevant in vitro model.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Calcium; Cell Differentiation; Cell Line; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; Electrophysiology; Gene Expression Regulation; Hippocampus; Humans; Kainic Acid; N-Methylaspartate; Neural Stem Cells; Quinoxalines; Receptors, Ionotropic Glutamate

Zinc chloride (0.02 mM, 3h) did not have any influence on the survival of cerebellar granule neurons (CGNs) incubated in balanced salt solution (BSS). However, in the absence of glucose ZnCl(2) caused severe neuronal damage, decreasing cell survival to 12±2%. Either the blockade of ionotropic glutamate NMDA-receptors with MK-801 or APV or supplementation the medium with ruthenium red (mitochondrial Ca(2+) uniporter blocker) almost entirely protected CGNs from the toxic effect of ZnCl(2) during glucose deprivation (GD). However, NBQX (AMPA/kainate glutamate receptor blocker) did not show protective effect. Measurements of intracellular calcium ions concentration using fluorescent probe (Fluo-4 AM) and zinc ions (FluoZin-3AM) demonstrated that 1.5h-exposure to GD induced intensive increase of Fluo-4 fluorescence and small increase of FluoZin-3 fluorescence in neurons. The supplementation of medium with ZnCl(2) caused equal increase of FluoZin-3 fluorescence at both GD and normoglycemia, whereas the potentiation of Fluo-4 fluorescence by zinc was observed only under GD and could be prevented by MK-801. However, neither MK-801 nor NBQX could influence [Zn(2+)](i) increase caused by zinc addition under GD, while ruthenium red did cause significant increase of [Zn(2+)](i). This data implies that zinc ions during GD induce an additional overload of CGNs with calcium ions that get transported through activated NMDA-channel. Zinc and calcium ions accumulate in mitochondria and amplify individual destructive action on these organelles leading to neuronal death.

Topics: Animals; Calcium; Cell Death; Cells, Cultured; Cerebellum; Chlorides; Coloring Agents; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glucose; Neurons; Neuroprotective Agents; Quinoxalines; Rats; Rats, Wistar; Ruthenium Red; Zinc Compounds

It is known that enrichment of glutamatergic transmission in the nucleus tractus solitarii (NTS) plays an important role in central cardiovascular regulation. Our previous study demonstrated that nicotine decreased blood pressure and heart rate in the NTS probably acting via the nicotinic acetylcholine receptors (nAChRs)-Ca²⁺-calmodulin-eNOS-NO signaling pathway. The possible relationship between glutamate and nicotine in the NTS for cardiovascular regulation is poorly understood. This study investigated the involvement of glutamate receptors in the cardiovascular effects of nicotine in the NTS. Nicotine (a non-selective nAChRs agonist), MK801 (a non-competitive NMDA receptor antagonist), APV (a competitive NMDA receptor antagonist), or NBQX (a selective AMPA receptor antagonist) was microinjected into the NTS of anesthetized Wistar-Kyoto rats. Microinjection of nicotine (1.5 pmol) into the NTS produced decreases in blood pressure and heart rate. The hypotensive and bradycardic effects of nicotine were abolished by prior administration of MK801 (1 nmol) and APV (10 nmol), but was completely restored after 60 min of recovery. In contrast, prior administration of NBQX (10 pmol) into the NTS did not alter the cardiovascular effects of nicotine. The nitrate (served as total NO) production in response to nicotine microinjection into the NTS was suppressed by prior administration of APV. These results suggest that the hypotensive and bradycardic effects of nicotine in the NTS might be mediated through NMDA receptors, and that the nAChRs-NMDA receptor-NO pathway could be involved.

Topics: Animals; Blood Pressure; Bradycardia; Dizocilpine Maleate; Drug Interactions; Excitatory Amino Acid Antagonists; Glutamic Acid; Heart Rate; Hypotension; Male; Nicotine; Nicotinic Agonists; Quinoxalines; Rats; Rats, Inbred WKY; Receptors, N-Methyl-D-Aspartate; Signal Transduction; Solitary Nucleus; Valine

Nitric oxide ((•)NO) is a labile endogenous free radical produced upon glutamatergic neuronal activity in hippocampus by neuronal nitric oxide synthase (nNOS), where it acts as a modulator of both synaptic plasticity and cell death associated with neurodegeneration. The low CNS levels and fast time dynamics of this molecule require the use of rapid analytical methods that can more accurately describe its signaling in vivo. This is critical for understanding how the kinetics of (•)NO-dependent signaling pathways is translated into physiological or pathological functions. In these studies, we used (•)NO selective microelectrodes coupled with rapid electrochemical recording techniques to characterize for the first time the concentration dynamics of (•)NO endogenously produced in hippocampus in vivo following activation of ionotropic glutamate receptors. Both L-glutamate (1-100 mM) and N-methyl-D-aspartate (NMDA; 0.01-5 mM) produced transient, dose-dependent increases in extracellular (•)NO concentration. The production of (•)NO in the hippocampus by glutamate was decreased by the nNOS inhibitor 7-NI. Intraperitoneal administration of the NMDA receptor blocker, MK-801, and the inhibitor of α-amino-3-hydroxy-5-methyl-4-isoazolepropionic acid (AMPA) receptor, NBQX, applied locally greatly attenuated glutamate-evoked overflow of (•)NO. Thus, (•)NO overflow elicited by activation of glutamate receptors appeared to result from an integrated activation of ionotropic glutamate receptors, both of the NMDA and AMPA receptors subtypes. Additionally, distinct concentration dynamics was observed in the trisynaptic loop with stronger and longer lasting effects of glutamate activation on (•)NO overflow seen in the CA1 region as compared with the dentate gyrus. Overall, the results provide a quantitative and temporal basis for a better understanding of (•)NO activity in the rat hippocampus.

Topics: Animals; CA1 Region, Hippocampal; Dentate Gyrus; Dizocilpine Maleate; Electrochemical Techniques; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Glutamic Acid; Male; Microelectrodes; N-Methylaspartate; Neurons; Nitric Oxide; Nitric Oxide Synthase Type I; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate

During development there is an activity-dependent switch in synaptic N-Methyl-D-aspartate (NMDA) receptor subunit composition from predominantly GluN2B to GluN2A, though the precise role of this switch remains unknown. By deleting GluN2 subunits in single neurons during synaptogenesis, we find that both GluN2B and GluN2A suppress AMPA receptor expression, albeit by distinct means. Similar to GluN1, GluN2B deletion increases the number of functional synapses, while GluN2A deletion increases the strength of unitary connections without affecting the number of functional synapses. We propose a model of excitatory synapse maturation in which baseline activation of GluN2B-containing receptors prevents premature synapse maturation until correlated activity allows induction of functional synapses. This activity also triggers the switch to GluN2A, which dampens further potentiation. Furthermore, we analyze the subunit composition of synaptic NMDA receptors in CA1 pyramidal cells, provide electrophysiological evidence for a large population of synaptic triheteromeric receptors, and estimate the subunit-dependent open probability.

Topics: Animals; Dendrites; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Hippocampus; Mice; Patch-Clamp Techniques; Piperidines; Protein Subunits; Pyramidal Cells; Quinoxalines; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Synapses

We previously have identified phenotypic differences in susceptibility to hippocampal seizure-induced cell death among two inbred strains of mice. We have also reported that the age-related increased susceptibility to the neurotoxic effects of seizure-induced injury is regulated in a strain-dependent manner. In the present study, we wanted to begin to determine the pharmacological mechanism that contributes to variability in the response to the neurotoxic effects of kainate. Thus, we compared the effects of the NMDA receptor antagonist, MK-801 and of the AMPA receptor antagonist NBQX on hippocampal damage in the kainate model of seizure-induced excitotoxic cell death in young, middle-aged, and aged C57BL/6 and FVB/N mice, when given 90 min following kainate-induced status epilepticus. Following kainate injections, mice were scored for seizure activity and brains from mice in each age and antagonist group were processed for light microscopic histopathologic evaluation 7 days following kainate administration to evaluate the severity of seizure-induced injury. Administration of MK-801 significantly reduced the extent of hippocampal damage in young, mature and aged FVB/N mice, while application of NBQX was only effective at attenuating cell death in young and aged mice throughout all hippocampal subfields. Our results suggest that both NMDA and non-NMDA receptors are involved in kainate-induced cell death in the mouse and suggest that aging may differentially affect the ability of neuroprotectants to protect against hippocampal damage. Differences in the effectiveness of these two antagonists could result from differential regulation of glutamatergic neurotransmitter systems or ion channel specificity.

Topics: Age Factors; Analysis of Variance; Animals; Brain; Cell Count; Cell Death; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Fluorescent Antibody Technique; Kainic Acid; Mice; Neurons; Neuroprotective Agents; Quinoxalines; Seizures; Severity of Illness Index; Species Specificity

Munc18 is considered as an intracellular protein that plays an important role in exocytosis of neurotransmitters. Previous studies have demonstrated the presence of autoantibodies against Munc18 in a subgroup of Rasmussen's encephalitis patients. However, the machinery of Munc18 autoimmunity is still elusive. The present study was aimed to investigate Munc18 release from primary cultured neurons, Munc18 distribution on the outer plasma membrane of neurons, and the neurotoxicity of Munc18 antibody.. The cerebral cortical neurons from embryonic day 17 Sprague-Dawley rats were prepared and cultured in neurobasal medium. The proteins in culture medium were precipitated with 10 % trichloroacetic acid, and analyzed by immunoblotting. The proteins on neuronal surface were biotinylated with EZ-Link-sulfo-NHS-LC-Biotin, and collected with avidin-conjugated agarose beads followed by immunoblotting analysis. For cell surface immunofluorescent staining, the living neurons were labeled with anti-Munc18 antibody at 4 degrees C. Neuronal injury was assessed by lactate dehydrogenase(LDH) release.. Munc18 was detected in culture medium by immunoblotting analysis. After treatment with 50 micromol/L glutamate for 1 h, Munc18 content in medium was increased. Meanwhile, beta-actin and syntaxin1 were not detected in culture medium, and LDH release was not significantly increased. Moreover, glutamate enhanced Munc18 distribution on outer plasma membrane. Living neuron staining also demonstrated the localization of Munc18 on neuronal surface after glutamate treatment, especially at contacting regions between neurons. Glutamate-induced increase of surface Munc18 distribution was suppressed by NMDA receptor antagonist MK801, but not by AMPA receptor antagonist NBQX. Moreover, compared with c-Fos antibody, Munc18 antibody could induce neuronal injury, when culture medium contained the components of serum.. A portion of Munc18 can be released from neurons or distributed on neuronal surface, which can be enhanced by glutamate treatment via activation of NMDA receptors. Besides, Munc18 antibody-induced neuronal injury depends on the serum components.

Topics: Animals; Biotinylation; Cell Death; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Interactions; Embryo, Mammalian; Excitatory Amino Acid Antagonists; Gene Expression Regulation; Glutamic Acid; L-Lactate Dehydrogenase; Munc18 Proteins; Neurons; Protein Transport; Quinoxalines; Rats; Rats, Sprague-Dawley; Time Factors

The rodent hippocampus exhibits population activities called sharp waves (SPWs) during slow wave sleep and wake immobility. SPWs are important for hippocampal-cortical communication and memory consolidation, and abnormal sharp wave-ripple complexes are closely related to epileptic seizures. Although the SPWs are known to arise from the CA3 circuit, the local mechanisms underlying their generation are not fully understood. We hypothesize that endogenous adenosine is a local regulator of hippocampal SPWs. We tested this hypothesis in thick mouse hippocampal slices that encompass a relatively large hippocampal circuit and have a high propensity of generating spontaneous in vitro SPWs. We found that application of adenosine A1 receptor antagonists induced in vitro SPWs and that such induction was sensitive to blockade by NMDA receptor antagonists. By contrast, an increase in endogenous adenosine via pharmacological inhibition of adenosine transporters or adenosine degrading enzymes suppressed spontaneous in vitro SPWs. We thus suggest that the initiation and incidence of sharp wave-like population events are under tight control by the activity of endogenously stimulated A1 receptors.

Topics: Adenine; Adenosine; Adenosine A1 Receptor Antagonists; Animals; Dizocilpine Maleate; Electric Stimulation; Excitatory Postsynaptic Potentials; Hippocampus; In Vitro Techniques; Inhibitory Postsynaptic Potentials; Membrane Potentials; Mice; Mice, Inbred C57BL; Microelectrodes; Nucleoside Transport Proteins; Patch-Clamp Techniques; Pyramidal Cells; Quinoxalines; Receptor, Adenosine A1; Receptors, N-Methyl-D-Aspartate; Theophylline; Xanthines

The major aim of this study was to elucidate the relationship between nitric oxide (NO) and generalized epilepsy. Mice lacking the neuronal nitric oxide synthase (nNOS) gene (nNOS(-/-)) were used in this study to determine the relationship between nNOS alpha and NO in pentylentetrazole (PTZ)-induced convulsions. nNOS(-/-) mice exhibited severe convulsions following injection with a subconvulsive dose of PTZ (40 mg/kg i.p.) and convulsive doses were lethal in all of the mice (60 mg/kg i.p.) following tonic convulsions. The results were confirmed by using selective nNOS inhibitors in wild-type (nNOS(+/+)) mice. The higher doses of the nNOS inhibitors 1-[2-(trifluoromethyl)phenyl] imidazole (TRIM) and 3-bromo-7-nitroindazole (3Br7NI) inhibited clonic-tonic convulsions induced by a convulsive dose of PTZ (60 mg/kg) in nNOS(+/+) mice. In contrast, either TRIM or 3Br7NI at lower doses enhanced convulsions following injection with a subconvulsive dose of PTZ (40 mg/kg) in nNOS(+/+) mice similar to nNOS(-/-) mice treated with PTZ. Such a proconvulsant effect was observed in nNOS(+/+) mice pretreated with nNOS inhibitors but not other NOS inhibitors. These results indicate that NO may be regarded as an anticonvulsant or a proconvulsant substance in relation to convulsions induced by PTZ in mice. Pretreatment with N-methyl-d-aspartate (NMDA) receptor antagonists (5S,10R)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]-cyclohepten-5,10-imine maleate (MK-801), (E)-(+/-)-2-amino-4-methyl-5-phospho no-3-pentenoic acid ethyl ester, CGP39551) and DL-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor antagonist (2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide, NBQX) inhibited a subconvulsive dose of PTZ-induced convulsions in nNOS(-/-) mice, demonstrating that convulsions induced by PTZ are modulated by endogenous NO production and ionotropic glutamate receptor-mediated stimulation. These results suggest a negative or positive modulation of neuronal interactions by basal or enhanced NO production, respectively.

Topics: Animals; Convulsants; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Gene Expression Regulation; Imidazoles; Indazoles; Mice; Mice, Inbred C57BL; Mice, Knockout; N-Methylaspartate; Neuroprotective Agents; Nitric Oxide; Nitric Oxide Synthase Type I; Pentylenetetrazole; Quinoxalines; Seizures

In the present study, we investigated the effects of mebudipine and dibudipine, two new Ca(2+) channel blockers, on primary murine cortical neurons exposed to oxygen-glucose deprivation/reperfusion. The experiments were performed on cells after 11-16 days of culture. To initiate oxygen-glucose deprivation /reperfusion, the culture medium was replaced by glucose-free medium, and the cells were transferred to a humidified incubation chamber in a mixture of 95% N(2) and 5% CO(2) at 37 degrees C for 30 min. The cultures were pretreated with mebudipine and dibudipine 3 h prior to oxygen-glucose deprivation/reperfusion, in order to explore their effects on neurons under oxygen-glucose deprivation conditions. Cell viability and nitric oxide (NO) production were assessed by MTT assay and the modified Griess method, respectively. Exposure of murine cortical neuronal cells to 30 min oxygen-glucose deprivation significantly decreased cell viability and increased NO production. Pretreatment of the cultures with mebudipine and dibudipine significantly increased cell viability and decreased NO generation in a dose-dependent manner. However, the drugs had no protective effect in cells subjected to oxygen-glucose deprivation for 60 min. Pretreatment of cultures with MK-801 (10 microM), a non-competitive NMDA antagonist, decreased neuronal death after 30-min oxygen-glucose deprivation, while application of NBQX (30 microM), a selective AMPA-kainate receptor antagonist, partially attenuated the cell injury. oxygen-glucose deprivation -induced cytotoxicity and NO production were also inhibited by N-nitro-L-arginine methyl ester (L-NAME), a nitric oxide synthase inhibitor and MK-801. We conclude that mebudipine and dibudipine could protect cortical neurons against oxygen-glucose deprivation /reperfusion-induced cell injury in a dose-dependent manner, and that this could be mediated partially by decreased NO production.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Cell Death; Cell Survival; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Dose-Response Relationship, Drug; Embryo, Mammalian; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Formazans; Glucose; Hypoxia; Mice; N-Methylaspartate; Neurons; Neuroprotective Agents; NG-Nitroarginine Methyl Ester; Nifedipine; Nitric Oxide; Nitric Oxide Synthase; Nitrites; Quinoxalines; Receptors, Kainic Acid; Reperfusion Injury; Tetrazolium Salts; Time Factors

Glutamate plays an essential role in neuronal cell death in many neurological disorders. In this study, we examined both glucose metabolism and cerebral blood flow in the same rat following infusion of glutamate or ibotenic acid using the dual-tracer technique. The effects of MK-801, an NMDA receptor antagonist, and NBQX, an AMPA-kainate receptor antagonist, on the changes in the glucose metabolism and cerebral blood flow induced by glutamate were also examined.. The rats were microinjected with glutamate (1 micromol/microl, 2 microl) or ibotenic acid (10 microg/microl, 1 microl) into the right striatum, and dual-tracer autoradiograms of [(18)F]FDG and [(14)C]IMP were obtained. MK-801 and NBQX were injected intravenously about 45 and 30 min, respectively, after the infusion of glutamate.. De-coupling of blood flow and metabolism was noted in the glutamate-infused hemisphere (as assessed by no alteration of [(18)F]FDG uptake and significant decrease of [(14)C]IMP uptake). Pretreatments with MK-801, NBQX, or combined use of MK-801 and NBQX did not affect the de-coupling of the blood flow and metabolism induced by glutamate. A histochemical study revealed that about 20% neuronal cell death had occurred in the striatum at 105 min after the infusion of glutamate. In addition, a significant increase of the [(18)F]FDG uptake and decrease of [(14)C]IMP uptake were also seen in the rat brain infused with ibotenic acid.. These results indicate that glutamate and ibotenic acid caused a significant de-coupling of blood flow and glucose metabolism in the intact rat brain during the early phase of neurodegeneration. It is necessary to evaluate the relation between metabotropic glutamate receptors and de-coupling of blood flow and metabolism.

Topics: Amphetamines; Animals; Autoradiography; Brain; Carbon Radioisotopes; Cerebrovascular Circulation; Dizocilpine Maleate; Fluorodeoxyglucose F18; Glucose; Glutamic Acid; Male; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate

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

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

Further understanding of pathophysiology of postoperative acute pain is necessary for its better management. The methodology of current threshold (CT) determination by using sine-wave stimuli at 3 frequencies has been used to selectively and quantitatively analyze the function of the subsets of fibers (i.e., frequency of 5, 250, and 2000Hz recruits C-, Adelta-, and Abeta-fibers, respectively). This study investigated how surgical incision would affect the CTs, and then assessed the efficacy of intrathecal pharmacotherapy. The CT required to evoke a paw withdrawal response was assessed over time at stimulus frequencies of 5Hz (CT5), 250Hz (CT250), and 2000Hz (CT2000) in rats that had undergone surgical incision of the plantar skin and muscle. The CTs at all frequencies significantly decreased immediately after the incision. The decreased thresholds gradually recovered during the first week post-surgery. CT5 and CT250 (but not CT2000) remained significantly low even on day 7 post-surgery. Morphine at 5microg/10microL i.t. significantly reversed CT5 and CT250. NBQX (alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid [AMPA]/kainate receptor antagonist) at 1.9 or 3.8microg/10microL i.t. significantly increased the thresholds over the pre-surgery threshold levels at all frequencies. MK-801 (N-methyl d-aspartate [NMDA] receptor antagonist) up to 13.5microg/10microL i.t. did not significantly affect CTs at any frequencies. In conclusion, a broad spectrum of sensory fibers (Abeta, Adelta, and C) is sensitized at the spinal and/or peripheral level in the postoperative acute pain state. Spinal AMPA/kainate receptors but not NMDA receptors play a significant role in this sensitization.

Topics: Animals; Dizocilpine Maleate; Foot; Infusions, Parenteral; Male; Models, Animal; Morphine; Narcotics; Nerve Fibers, Myelinated; Nerve Fibers, Unmyelinated; Pain Threshold; Pain, Postoperative; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Subarachnoid Space

The influence of estradiol (E2) treatment on temporomandibular joint (TMJ) nociceptive processing in the caudal trigeminal sensory brain stem complex was assessed in ovariectomized female rats by quantitative Fos-immunoreactivity (Fos-LI). After 2 days of daily injections of high (HE2) or low (LE2) dose E2 rats were anesthetized and the small fiber excitant, mustard oil (MO, 0-20%), was injected into the TMJ and after 2 h brains were processed for Fos-LI. TMJ-evoked Fos-LI in laminae I-II at the trigeminal subnucleus caudalis/upper cervical cord (Vc/C1-2) junction and the dorsal paratrigeminal region (dPa5) was significantly greater in HE2 than LE2 rats, while Fos-LI produced at the ventral trigeminal interpolaris/caudalis transition region (Vi/Vc(vl)) was similar. E2 treatment also modified the influence of N-methyl-D-aspartate (NMDA) and AMPA receptor antagonists on TMJ-evoked Fos-LI. The NMDA antagonist, MK-801, dose-dependently reduced the Fos-LI response at the Vc/C1-2 junction in HE2 rats, while only high dose MK-801 was effective in LE2 rats. MK801 reduced equally the Fos-LI response at the Vi/Vc transition in both groups, while only minor effects were seen at the dPa5 region. The AMPA receptor antagonist, NBQX, reduced Fos-LI at the Vc/C(1-2) and Vi/Vc(vl) regions in HE2 rats, while only high dose NBQX was effective in LE2 rats. NBQX did not reduce Fos-LI at the dPa5 region in either group. These results suggest that estrogen status plays a significant role in TMJ nociceptive processing at the Vc/C1-2 junction mediated, in part, through ionotropic glutamate receptor-dependent mechanisms.

Topics: Animals; Dizocilpine Maleate; Dose-Response Relationship, Drug; Estradiol; Estrogens; Excitatory Amino Acid Antagonists; Female; Gene Expression Regulation; Medulla Oblongata; Mustard Plant; Ovariectomy; Plant Oils; Proto-Oncogene Proteins c-fos; Quinoxalines; Rats; Rats, Sprague-Dawley; Stimulation, Chemical; Temporomandibular Joint

Cocaine strengthens behaviors associated with its administration. The stress response by individuals that are defeated in a brief aggressive confrontation can also promote enduring behavioral consequences similar to those of stimulants.. The study intends to find whether intermittent episodes of defeat promote cocaine's reinforcing effects by triggering N-methyl-D: -aspartic acid (NMDA)-receptor-mediated plasticity in the ventral tegmental area (VTA).. Long-Evans rats were investigated after four social defeats in three experiments. Two experiments examined systemic or intra-VTA antagonism of NMDA receptors during stress on the later expression of behavioral sensitization and cocaine self-administration during fixed and progressive ratio (PR) schedules of reinforcement (0.3 mg/kg/infusion), including a novel 24-h variable-dose continuous access binge (0.2, 0.4, and 0.8 mg/kg/infusion, delivered in an irregular sequence). Third, the expression of receptor proteins NR1 (NMDA) and GluR1 [alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)] were examined in VTA and nucleus accumbens.. Intermittent defeats augment locomotor responses to cocaine and increase cocaine taking. Rates of responding during binges are increased after defeat stress. These effects are prevented when NMDA or AMPA receptor antagonists are administered before defeats. VTA infusions of the NMDA antagonist AP-5 (5 nmol/side) before stress prevents locomotor sensitization to cocaine and intensified responding for cocaine during a PR schedule or binge. Episodic defeats increase GluR1 AMPA subunit protein expression in the VTA.. Social stress stimulates NMDA receptors in the VTA, and this neural action of defeat may be essential for prompting a later increase in cocaine intake during binges.

Topics: 2-Amino-5-phosphonovalerate; Animals; Blotting, Western; Cocaine-Related Disorders; Conditioning, Operant; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Male; Quinoxalines; Rats; Rats, Long-Evans; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Reinforcement Schedule; Self Administration; Social Environment; Stress, Psychological; Ventral Tegmental Area

Glutamate, the major excitatory neurotransmitter, can cause the death of neurons by a mechanism known as excitotoxicity. This is a calcium-dependent process and activation of the NMDA receptor subtype contributes mainly to neuronal damage, due to its high permeability to calcium. Activation of calpain, a calcium-dependent cysteine protease, has been implicated in necrotic excitotoxic neuronal death. We have investigated the contribution of NMDA and non-NMDA ionotropic receptors to calpain activation and neuronal death induced by the acute administration of glutamate into the rat striatum. Calpain activity was assessed by the cleavage of the cytoskeletal protein, alpha-spectrin. Caspase-3 activity was also studied because glutamate can also lead to apoptosis. Results show no caspase-3 activity, but a strong calpain activation involving both NMDA and non-NMDA receptors. Although neuronal damage is mediated mainly by the NMDA receptor subtype, it can not be attributed solely to calpain activity.

Topics: Animals; Calpain; Caspase 3; Corpus Striatum; Cysteine Proteinase Inhibitors; Dipeptides; Dizocilpine Maleate; Enzyme Activation; Excitatory Amino Acid Antagonists; Male; Neurons; Neuroprotective Agents; Quinoxalines; Rats; Rats, Wistar; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Spectrin

Clinical evidence demonstrates altered glutamatergic neurotransmission in patients suffering from obsessive-compulsive disorder (OCD). We examined the effects of glutamate-related drugs on marble-burying behavior, which is an animal model of OCD. The uncompetitive N-methyl-d-aspartate (NMDA) antagonists memantine (10 mg/kg, i.p.) and amantadine (30 mg/kg, i.p.) significantly inhibited marble-burying behavior without affecting locomotor activity in mice. Similarly, the uncompetitive NMDA receptor antagonist 5R,10S-(+)-5-methyl-10,11-dihydro-5H-dibenzo(a,d)cyclohepten-5,10-imine hydrogen maleate (MK-801, 0.3 mg/kg, i.p.) inhibited marble-burying behavior. However, MK-801 at the same dose markedly increased locomotor activity. By contrast, the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptor antagonist 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo-benzo[f]quinoxaline-7-sulfonamide (NBQX) and the glutamate release inhibitor riluzole showed no effect on marble-burying behavior and significant suppression of locomotor activity. MK-801 (0.3 mg/kg, i.p.) and memantine (10 mg/kg, i.p.) significantly disrupted prepulse inhibition as an operational measure of sensorimotor gating. By contrast, amantadine (30 mg/kg, i.p.) did not affect prepulse inhibition. These findings suggest that amantadine could be a useful drug for the treatment of OCD.

Topics: Acoustic Stimulation; Amantadine; Animals; Behavior, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Male; Memantine; Mice; Mice, Inbred ICR; Motor Activity; Obsessive-Compulsive Disorder; Quinoxalines; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Reflex, Startle; Riluzole

The naturally occurring sex difference in dendritic spine number on hypothalamic neurons offers a unique opportunity to investigate mechanisms establishing synaptic patterning during perinatal sensitive periods. A major advantage of the rat as a model of sexual differentiation is that treatment of neonatal females with estradiol will permanently induce the male phenotype. During the development of other systems, exuberant innervation is followed by activity-dependent pruning necessary for elimination of spurious synapses. In contrast, we demonstrate that estradiol-induced organization in the hypothalamus involves the induction of new synapses on dendritic spines. Activation of estrogen receptors by estradiol triggers a nongenomic activation of PI3 kinase that results in enhanced glutamate release from presynaptic neurons. Subsequent activation of ionotropic glutamate receptors activates MAP kinases, thereby inducing dendritic spine formation. These results reveal a transneuronal mechanism by which estradiol acts during a sensitive period to establish a profound and lasting sex difference in hypothalamic synaptic patterning.

Topics: Animals; Animals, Newborn; Dendritic Spines; Dizocilpine Maleate; Drug Interactions; Estradiol; Excitatory Amino Acid Antagonists; Female; Hypothalamus; In Vitro Techniques; Male; N-Methylaspartate; Neurons; Patch-Clamp Techniques; Phosphatidylinositol 3-Kinases; Pregnancy; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Sex Characteristics; Silver Staining

Imbalances of the glutamatergic system are implicated in the pathophysiology of various basal ganglia disorders, but few is known about their role in dystonia, a common neurological syndrome in which involuntary muscle co-contractions lead to twisting movements and abnormal postures. Previous systemic administrations of glutamate receptor antagonists in dtsz hamsters, an animal model of primary paroxysmal dystonia, exerted antidystonic effects and electrophysiological experiments pointed to an enhanced corticostriatal glutamatergic activity. In order to examine the pathophysiological relevance of these findings, we performed striatal microinjections of the alpha-amino-2,3-dihydro-5-methyl-3-oxo-4-isoxazolepropanoic acid (AMPA) receptor antagonist 2,3-dioxo-6-nitro-7-sulfamoylbenzo(f)quinoxaline (NBQX) and the N-methyl-D-aspartate (NMDA) receptor antagonists D(-)-2-amino-5-phosphopentanoic acid (AP-5), (R)-(+)-3-amino-1-hydroxypyrrolidin-2-one (HA-966) and dizocilpine (MK-801). The striatal application of NBQX reduced the severity and increased the latency to onset of dystonia significantly only at a dosage of 0.08 microg per hemisphere, lower (0.03 microg) and higher dosages (0.16 microg and 0.32 microg) failed to exert comparable effects on the severity. None of the striatal injected NMDA receptor antagonists influenced the severity of the dystonic attacks in the mutant hamster. The combined application of NBQX (0.08 microg) with AP-5 (1.0 microg) failed to exert synergistic antidystonic effects, but the beneficial effect on the severity of dystonia of the single application of NBQX was reproduced. Therefore, corticostriatal glutamatergic overactivity mediated by AMPA receptors, but not by NMDA receptors, is possibly important for the manifestation of dystonic attacks in the dtsz hamster mutant.

Topics: Animals; Basal Ganglia; Caudate Nucleus; Cricetinae; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Dystonia; Excitatory Amino Acid Antagonists; Microinjections; Movement; Muscle Contraction; Mutation; Posture; Putamen; Pyrrolidinones; Quinoxalines; Reaction Time; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Severity of Illness Index; Valine

The role of AMPA receptors (AMPARs) in generation and propagation of respiratory rhythm is well documented both in vivo and in vitro, whereas the functional significance of NMDA receptors (NMDARs) in preBötzinger complex (preBötC) neurons has not been explored. Here we examined the interactions between AMPARs and NMDARs during spontaneous respiratory rhythm generation in slices from neonatal rats in vitro. We tested the hypothesis that activation of NMDARs can drive respiratory rhythm in the absence of other excitatory drives. Blockade of NMDARs with dizocilpine hydrogen maleate (MK-801, 20 microM) had a negligible effect on respiratory rhythm and pattern under standard conditions in vitro, whereas blockade of AMPARs with NBQX (0.5 microM) completely abolished respiratory activity. Removal of extracellular Mg2+ to relieve the voltage-dependent block of NMDARs maintained respiratory rhythm without a significant effect on period, even in the presence of high NBQX concentrations (

Topics: Action Potentials; Animals; Animals, Newborn; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Glutamic Acid; In Vitro Techniques; Magnesium; Neurons; Patch-Clamp Techniques; Periodicity; Potassium; Quinoxalines; Rats; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Respiratory Center; Respiratory Mechanics; Synaptic Transmission

Most in vitro models are only used to assess short-term effects of test compounds. However, as demonstrated here, hippocampal slice cultures can be used for long-term studies. The test compound used was the metabotropic glutamate receptor antagonist, L(+)-2-amino-3-phosphonopropionic acid (L-AP3), which is known to be toxic in vivo after subchronic, but not acute, administration. Degenerative effects were monitored by measuring the cellular uptake of propidium iodide (PI; continuously present in the medium) and lactate dehydrogenase (LDH) leakage, and by using a panel of histological stains. Hippocampal slices, derived from 2-3 day old rats and grown for 3 weeks, were subsequently exposed for the next 3 weeks to 0, 10 or 100microM L-AP3, with PI (2microM) in the culture medium. Exposure to 100microM L-AP3 induced severe toxicity after 4-6 days, shown by massive PI uptake, LDH leakage, changes in MAP2 and GFAP immunostaining, and in Nissl and Timm staining. In contrast, 10microM L-AP3 did not induce detectable neuronal degeneration. Treatment with the NMDA receptor antagonist, MK-801, or the AMPA/KA receptor antagonist NBQX, together with 100microM L-AP3, reduced neurodegeneration down to close to control values. It is concluded that continuous incubation of hippocampal slice cultures with PI is technically feasible for use in studies of inducible neuronal degeneration over time.

Topics: Alanine; Animal Testing Alternatives; Animals; Coloring Agents; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hippocampus; L-Lactate Dehydrogenase; Propidium; Quinoxalines; Rats; Staining and Labeling; Tissue Culture Techniques

When excitotoxic mechanisms are blocked, severe or prolonged hypoxia and hypoxia-ischemia can still kill neurons, by a mechanism which is poorly understood. We studied this "non-excitotoxic hypoxic death" in primary cultures of rat dentate gyrus neurons. Many neurons subjected to hypoxia in the presence of blockers of ionotropic glutamate receptors developed the electron microscopic features of necrosis. They showed early mitochondrial swelling, loss of mitochondrial membrane potential and cytoplasmic release of cytochrome c, followed by activation of caspase-9, and by caspase-9-dependent activation of caspase-3. Caspase inhibitors were neuroprotective. These results suggest that "non-excitotoxic hypoxic neuronal death" requires the activation in many neurons of a cell death program originating in mitochondria and leading to necrosis.

Topics: Adenosine Triphosphate; Analysis of Variance; Animals; Animals, Newborn; Caspases; Cell Survival; Cells, Cultured; Dentate Gyrus; Dizocilpine Maleate; Dose-Response Relationship, Drug; Enzyme Activation; Hypoxia; L-Lactate Dehydrogenase; Microscopy, Electron, Transmission; Microscopy, Immunoelectron; Mitochondria; Necrosis; Neurons; Neuroprotective Agents; Quinoxalines; Rats; Sodium Cyanide

Allodynia or hyperalgesia induced by peripheral nerve injury may be involved in changes in the sensitivity of neurotransmitters at the spinal cord level. In order to clarify the functional role of neurotransmitters in peripheral nerve injury, we used rats with nerve injury induced by chronic constriction of the sciatic nerve (CCI rat model) and estimated the effects of the intrathecal injection of drugs known to affect glutamate and tachykinin receptors. In sham-operated rats, the NMDA receptor agonist NMDA and AMPA-kinate receptor agonist RS-(5)-bromowillardin reduced withdrawal latency. The non-competitive NMDA receptor antagonist MK-801, competitive NMDA receptor antagonist AP-5 and AMPA-kinate receptor antagonist NBQX increased withdrawal latency. Substance P (SP) increased the withdrawal latency but only transitorily. The NK1 receptor antagonist RP67580 increased withdrawal latency, but the NK2 receptor antagonist SR48968 did not show an effect. In CCI rats, RS-(5)-bromowillardin reduced withdrawal latency, but NMDA did not show an effect. NBQX increased withdrawal latency, while MK-801 and AP-5 showed little or no effect. SP reduced withdrawal latency, and both RP67580 and SR48968 increased it. These results indicate that the alteration in sensitivity of ionotropic glutamate receptors and tachykinin receptors in the spinal cord contribute to development and maintenance of nerve injury-evoked neuropathic pain.

Topics: Alanine; Analgesics; Animals; Behavior, Animal; Benzamides; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Indoles; Isoindoles; Male; N-Methylaspartate; Pain; Pain Measurement; Piperidines; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, Kainic Acid; Receptors, Tachykinin; Sciatic Nerve; Spinal Cord; Substance P; Valine

Excitatory amino acids play an important role in generation of epileptic seizures. To study the participation of different types of their receptors in cortical epileptic afterdischarges, a noncompetitive NMDA receptor antagonist dizocilpine and a competitive AMPA receptor antagonist NBQX were used. Adult rats with implanted epidural stimulation and registration electrodes were pretreated either with NBQX (30 or 60 mg/kg i.p.) or with dizocilpine (0.1 or 0.5 mg/kg i.p.) and low-frequency stimulation of sensorimotor cortical area was repeatedly applied with stepwise increased current intensities. Lower dose of NBQX unexpectedly decreased thresholds for elicitation of spike-and-wave afterdischarges (ADs), clonic seizures accompanying this type of ADs and for transition into the second, limbic type of ADs. Lower dose of dizocilpine increased these three thresholds. Higher doses of either drug did not significantly change threshold intensities. Duration of ADs was also influenced by the two antagonists in opposite directions: higher dose of NBQX resulted in prolongation of ADs mainly due to an increased duration of the spike-and-wave part of ADs whereas dizocilpine shortened ADs in a dose-dependent manner affecting both types of ADs. In addition, NBQX did not influence interhemispheric responses meanwhile dizocilpine moderately suppressed these evoked potentials. According to our results, NMDA receptors are important for generation of cortical epileptic afterdischarges meanwhile the role of AMPA receptors is not clear and has to be analyzed.

Topics: Animals; Cerebral Cortex; Dizocilpine Maleate; Dose-Response Relationship, Drug; Dose-Response Relationship, Radiation; Electric Stimulation; Electroencephalography; Epilepsy; Evoked Potentials; Excitatory Amino Acid Antagonists; Quinoxalines; Rats; Rats, Wistar

Abstract We have investigated the role of Ca2+ accumulation and neuronal injury in cerebellar granule neurons after glutamate receptor overactivation. After the removal of the free cytosolic Ca2+ we identified an extensive second Ca2+ fraction (SCF) that is retained within the neurons after glutamate receptor overactivation. The SCF reaches a plateau within 10 min with the magnitude of this SCF accumulation reflecting the extent of the neuronal injury that occurs within the neurons. The existence of this SCF is sensitive to both NMDA receptor antagonists and mitochondrial inhibitors but is unaffected by agents that deplete endoplasmic reticulum Ca2+, indicating that this Ca2+ fraction may be located within the mitochondria. Through the isolation of mitochondria from cerebellar granule neurons treated with glutamate we have shown that the majority of the SCF is mitochondrial in location. On the removal of the glutamate stimulus the SCF recovers at a slower rate than the free Ca2+ concentration within the neuron. This is intriguing, as it implies a capacity to remember previous excitatory events. Most significantly we have shown that a short pre-application of subthreshold glutamate or kainate blocks both SCF Ca2+ accumulation and extensive neuronal injury in response to high concentrations of glutamate. These findings may be relevant to the observations of pre-conditioning in the brain and heart.

Topics: Age Factors; Alamethicin; Analysis of Variance; Animals; Animals, Newborn; Brain Diseases; Calcium; Calcium Isotopes; Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; Cell Count; Cell Death; Cell Fractionation; Cells, Cultured; Cerebellum; Diagnostic Imaging; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Female; Fura-2; Glutamic Acid; Glycine; Hydro-Lyases; Indoles; Intracellular Space; Ionophores; Male; Mitochondria; Neurons; Propidium; Quinoxalines; Rats; Rats, Wistar; Time Factors

Recent experiments from this laboratory demonstrated synergistic effects of AMPA/kainate receptor blockade and D(2/3) dopamine (DA) receptor stimulation on brain stimulation reward and locomotor activity.. Using place conditioning, this study explored further the interaction between DA and glutamate (Glu) using the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801, the AMPA/kainate receptor antagonist NBQX, and the D(2/3) DA receptor agonist 7-OH-DPAT.. Effects of these compounds, alone and combined, were measured in male Sprague--Dawley rats using an unbiased two-compartment place conditioning procedure.. 7-OH-DPAT (0.03--5.0 mg kg(-1), s.c.) administered immediately prior to conditioning was ineffective; when administered 15 min prior to conditioning, only the highest dose (5.0 mg kg(-1), s.c.) induced conditioned place preference (CPP). Acquisition of 7-OH-DPAT-induced CPP was blocked by MK-801 (0.06 or 0.13 mg kg(-1), i.p.) or NBQX (0.5 microg) microinjected into the nucleus accumbens (NAS) shell subregion. Intra-NAS shell administration of 7-OH-DPAT (5.0 microg) or NBQX (0.5 microg), alone or combined, failed to induce place conditioning, and this lack of effect was not due to state dependency. Administration of MK-801 or 7-OH-DPAT (5.0 mg kg(-1)) during the conditioning phase acutely increased horizontal activity, but neither compound, alone or combined, induced conditioned locomotor effects.. Acquisition of place conditioning induced by systemic administration of 7-OH-DPAT is blocked by systemic NMDA receptor antagonism by MK-801 or by the AMPA/kainate receptor antagonist NBQX microinjected into the NAS shell subregion.

Topics: Animals; Choice Behavior; Conditioning, Psychological; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Male; Nucleus Accumbens; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, Dopamine D2; Receptors, Dopamine D3; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Tetrahydronaphthalenes

Apolipoprotein E is a genetic risk factor for Alzheimer's disease, and the apoE protein is associated with beta-amyloid deposits in Alzheimer's disease brain. We examined signaling pathways stimulated by apoE in primary neurons in culture. ApoE and an apoE-derived peptide activated several intracellular kinases, including prominently extracellular signal-regulated kinase 1/2 (ERK1/2). ERK1/2 activation by apoE was blocked by an inhibitor of the low-density lipoprotein receptor family, the specific NMDA glutamate receptor antagonist MK 801 and other calcium channel blockers. Activation of apoE receptors also induced tyrosine phosphorylation of Dab1, an adaptor protein of apoE receptors, but experiments in Dab1 knockout neurons demonstrated that Dab1 was not necessary for ERK activation. In contrast, apoE treatment of primary neurons decreased activation of c-Jun N-terminal kinase, a kinase that interacts with another apoE receptor adaptor protein, c-Jun N-terminal kinase-interacting protein. This change also depended on interactions with the low-density lipoprotein receptor family but was independent of calcium channels. c-Jun N-terminal kinase deactivation by apoE was blocked by gamma-secretase inhibitors and pertussis toxin. These results demonstrate that apoE affects several signaling cascades in neurons: increased disabled phosphorylation, activation of the ERK1/2 pathway (dependent on calcium influx via the NMDA receptor) and inhibition of the c-Jun N-terminal kinase 1/2 pathway (dependent on gamma-secretase and G proteins).

Topics: Analysis of Variance; Androstadienes; Animals; Apolipoproteins E; Blotting, Western; Calcium Channel Blockers; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Drug Interactions; Embryo, Mammalian; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Flavonoids; JNK Mitogen-Activated Protein Kinases; MAP Kinase Kinase 4; Mice; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinase Kinases; Models, Neurological; Nerve Tissue Proteins; Neurons; Phosphatidylinositol 3-Kinases; Phosphorylation; Protein Isoforms; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Quinoxalines; Signal Transduction; src-Family Kinases; Time Factors; Wortmannin

Surgery commonly causes pain and neural plasticity that are unique compared to other persistent pain problems. To more precisely study central sensitization and plasticity, we examined the role of ionotropic EAA receptors in dorsal horn neuron sensitization early after incision. Sensitization, in the form of increased background activity, increased mechanosensitivity or pinch receptive field expansion, was induced by plantar incision 1 h later in 30 neurons. (+)-5-Methyl-10,11-dihydro-5H-dibenzo(a,d)cyclohepten-5,10-imine (MK-801) or 1 mM 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo[f]quinoxaline-7-sulfonamide (NBQX) was administered through a microdialysis fiber to block NMDA and nonNMDA EAA receptors, respectively. Dorsal horn neuron sensitization was reexamined 1 h later. Spinal administration of NBQX blocked AMPA-induced excitation but did not affect excitation by NMDA. NBQX decreased background activity in the neurons that developed sustained increased activity after incision. The median decrease caused by NBQX was from 2.3 to 0.0 imp/s. Spinal administration of 5 mM MK-801 blocked NMDA-induced excitation but did not affect excitation by AMPA. The median change (from 2.6 to 1.1 imp/s) in background activity increased by incision was not significantly affected by MK-801. The responses to mechanical stimuli were enhanced after incision in wide dynamic range (WDR) neurons. NBQX eliminated these responses but MK-801 had no effect. The pinch receptive field (RF) expansion into uninjured areas of the paw and hindquarters occurred after incision. Only 1 of 13 neurons exhibited RF expansion after spinal NBQX administration; 9 of 12 neurons had RF expansion remaining after MK-801. Thus, nonNMDA receptors are critical and NMDA-independent factors influence the increased responsiveness of dorsal horn neurons that occur early after incision.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Male; N-Methylaspartate; Neuronal Plasticity; Pain, Postoperative; Physical Stimulation; Posterior Horn Cells; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

The functional properties of neurotransmission in the medial preoptic nucleus (MPN) were studied in a brain slice preparation from young male rats. The aims were to evaluate the thin slice preparation for studying evoked synaptic responses in MPN neurons, to characterize the fast responses triggered by activation of presynaptic nerve fibers in the MPN, and to identify the involved receptor types. Presynaptic stimulation within the MPN evoked postsynaptic voltage and current responses that were blocked by 200 microM Cd2+ or by 2.0 microM tetrodotoxin and were attributed to action potential-evoked transmitter release. The relation to stimulus strength and comparison with spontaneous synaptic currents suggested that in many cases only one presynaptic nerve fiber was excited by the stimulus. Furthermore, the transmission was probabilistic in nature, with frequent failures. Thus, response probability, most likely reflecting transmitter release probability, could be evaluated in the thin slice preparation. Evoked excitatory postsynaptic currents recorded under voltage-clamp conditions were, due to kinetics, I-V relation, and pharmacological properties, attributed to AMPA/kainate receptors and NMDA receptors, whereas inhibitory currents were attributed to GABAA receptors. No responses that could be attributed to glycine or other types of primary transmitters were detected. Although serotonin (5-HT) did not appear to function as a primary transmitter, glutamate- as well as GABA-mediated transmission was suppressed by 500 microM 5-HT, with a clear reduction in response probability observed. 5-HT also reduced the frequency, but not the amplitude, of spontaneous postsynaptic currents and was therefore ascribed a presynaptic site of action.

Topics: Animals; Dizocilpine Maleate; In Vitro Techniques; Preoptic Area; Quinoxalines; Rats; Rats, Sprague-Dawley; Synaptic Transmission

Antagonists for spinal N-methyl-D-aspartate (NMDA) and amino-hydroxy-methtyl-isoxazolepropionate (AMPA) receptors are effective in attenuating acute nociception or injury-induced hyperalgesia. The antinociception of spinal gabapentin is developed in injury-induced hyperalgesia without affecting acute nociception. The authors evaluated the effects of intrathecal gabapentin, NMDA antagonist (MK801) and AMPA antagonist (NBQX) in the formalin test which shows injury-induced hyperalgesia as well as acute pain. We further assessed the interactions between gabapentin and either MK801 or NBQX. Male Sprague-Dawley rats were implanted with intrathecal catheters. To evoke pain, 50 microL of 5% formalin solution was injected into the hindpaw. The interaction was investigated by a fixed dose analysis or an isobolographic analysis. MK801 and NBQX suppressed flinching responses during phase 1 of the formalin test, while gabapentin had little effect on phase 1. All three agents decreased the phase 2 flinching response. A fixed dose analysis in phase 1 showed that gabapentin potentiated the antinociceptive effect of MK801 and NBQX. Isobolographic analysis in phase 2 revealed a synergistic interaction after coadministration of gabapentin-MK801 or gabapentin-NBQX. Correspondingly, spinal gabapentin with NMDA or AMPA antagonist may be useful in managing acute pain and injury-induced hyperalgesia.

Topics: Amines; Analgesics; Animals; Cyclohexanecarboxylic Acids; Dizocilpine Maleate; Drug Interactions; Excitatory Amino Acid Antagonists; Gabapentin; gamma-Aminobutyric Acid; Hyperalgesia; Injections, Spinal; Male; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate

Postoperative pain significantly impacts patient recovery. However, postoperative pain management remains suboptimal, perhaps because treatment strategies are based mainly on studies using inflammatory pain models. We used a recently developed mouse model of incisional pain to investigate peripheral and spinal mechanisms contributing to heat hyperalgesia after incision. Behavioral experiments involving TRPV1 KO mice demonstrate that, as previously observed in inflammatory models, TRPV1 is necessary for heat (but not mechanical) hyperalgesia after incision. However, in WT mice, neither the proportion of TRPV1 immunoreactive neurons in the DRG nor the intensity of TRPV1 staining in the sciatic nerve was different from that in controls up to 4 days after incision. This result was corroborated by immunoblot analysis of sciatic nerve in rats subjected to an incision, and is distinct from that following inflammation of the rat hind paw, a situation in which TRPV1 expression levels in sciatic nerve increases. In the absence of heat exposure, spinal c-Fos staining was similar between incised TRPV1 KO and WT mice. However, differences in c-Fos staining between heat exposed TRPV1 KO and WT mice after incision suggest that the incision-mediated enhancement of heat-evoked signaling to the spinal cord involves a TRPV1-dependent mechanism. Finally, heat hyperalgesia after incision was reversed by antagonism of spinal non-NMDA receptors, unlike inflammatory hyperalgesia, which is mediated via NMDA receptors . Thus, TRPV1 is important for the generation of thermal hyperalgesia after incision. Our observations suggest that all experimental pain models may not be equally appropriate to guide the development of postoperative pain therapies.

Topics: Animals; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Hot Temperature; Hyperalgesia; Ion Channels; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Neurogenic Inflammation; Nociceptors; Pain, Postoperative; Physical Stimulation; Proto-Oncogene Proteins c-fos; Quinoxalines; Skin; TRPV Cation Channels

The cellular mechanisms coupling mechanical loading with bone remodeling remain unclear. In the CNS, the excitatory amino acid glutamate (Glu) serves as a potent neurotransmitter exerting its effects via various membrane Glu receptors (GluR). Nerves containing Glu exist close to bone cells expressing functional GluRs. Demonstration of a mechanically sensitive glutamate/aspartate transporter protein and the ability of glutamate to stimulate bone resorption in vitro suggest a role for glutamate linking mechanical load and bone remodeling. We used immunohistochemical techniques to identify the expression of N-methyl-d-aspartate acid (NMDA) and non-NMDA (AMPA or kainate) ionotropic GluR subunits on bone cells in vivo. In bone sections from young adult rats, osteoclasts expressed numerous GluR subunits including AMPA (GluR2/3 and GluR4), kainic acid (GluR567) and NMDA (NMDAR2A, NMDAR2B and NMDAR2C) receptor subtypes. Bone lining cells demonstrated immunoexpression for NMDAR2A, NMDAR2B, NMDAR2C, GluR567, GluR23, GluR2 and GluR4 subunits. Immunoexpression was not evident on osteocytes, chondrocytes or vascular channels. To investigate the effects of mechanical loading on GluR expression, we used a Materials Testing System (MTS) to apply 10 N sinusoidal axial compressive loads percutaneously to the right limbs (radius/ulna, tibia/fibula) of rats. Each limb underwent 300-load cycles/day (cycle rate, 1 Hz) for 4 consecutive days. Contralateral, non-loaded limbs served as controls. Mechanically loaded limbs revealed a load-induced loss of immunoexpression for GluR2/3, GluR4, GluR567 and NMDAR2A on osteoclasts and NMDAR2A, NMDAR2B, GluR2/3 and GluR4 on bone lining cells. Both neonatal rabbit and rat osteoclasts were cultured on bone slices to investigate the effect of the NMDA receptor antagonist, MK801, and the AMPA/kainic acid receptor antagonist, NBQX, on osteoclast resorptive activity in vitro. The inhibition of resorptive function seen suggested that both NMDAR and kainic acid receptor function are required for normal osteoclast function. While the exact role of ionotropic GluRs in skeletal tissue remains unclear, the modulation of GluR subunit expression by mechanical loading lends further support for participation of Glu as a mechanical loading effector. These ionotropic receptors appear to be functionally relevant to normal osteoclast resorptive activity.

Topics: Acid Phosphatase; Animals; Animals, Newborn; Biomechanical Phenomena; Bone and Bones; Bone Resorption; Calcification, Physiologic; Dizocilpine Maleate; Down-Regulation; Excitatory Amino Acid Antagonists; Female; Forelimb; Immunohistochemistry; Isoenzymes; Leg Bones; Osteoclasts; Osteocytes; Pliability; Protein Subunits; Quinoxalines; Rats; Rats, Long-Evans; Rats, Wistar; Receptors, AMPA; Receptors, Glutamate; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Tartrate-Resistant Acid Phosphatase; Weight-Bearing

GABA(A) receptor activation during brain development is a critical source of excitation. This is due to the positive equilibrium potential for chloride relative to resting membrane potential, resulting in membrane depolarization sufficient to open voltage sensitive calcium channels. The gonadal steroid estradiol has pronounced trophic effects on the developing hippocampus, promoting cell survival and synaptogenesis. In the current study, we investigated the effect of estradiol on GABA(A) receptor-mediated calcium transients in cultured neonatal hippocampal neurons, from Sprague-Dawley rats, using the calcium sensitive dye, Fura-2-AM. Treatment of hippocampal neurons with physiological levels of estradiol significantly increased the peak amplitude of calcium transients, increased the number of cells responding to the GABA(A) agonist muscimol with membrane depolarization, and delayed the rate of clearance of free intracellular calcium. These effects were significantly attenuated by pretreatment with the oestrogen receptor antagonist ICI-182,780. This suggests that estradiol, via its action on the oestrogen receptor, prolongs the developmental duration of depolarizing GABA. Estradiol likely maintains GABA-mediated excitation by promoting increased protein levels of the active/phosphorylated form of the chloride cotransporter Na+K+2CL- and L-type voltage sensitive calcium channels containing the alpha1C subunit. We propose that a component of the trophic effects of estradiol on hippocampal development results from enhanced calcium influx subsequent to GABA(A) receptor activation.

Topics: Animals; Bicuculline; Blotting, Western; Calcium; Calcium Channel Blockers; Calcium Channels, L-Type; Cells, Cultured; Diagnostic Imaging; Diltiazem; Dizocilpine Maleate; Drug Administration Schedule; Drug Interactions; Embryo, Mammalian; Estradiol; Excitatory Amino Acid Antagonists; Female; Fura-2; GABA Agonists; GABA Antagonists; Glutamic Acid; Hippocampus; K Cl- Cotransporters; Muscimol; Neurons; Nimodipine; Potassium Chloride; Pregnancy; Quinoxalines; Rats; Receptors, GABA-A; Sodium-Potassium-Chloride Symporters; Solute Carrier Family 12, Member 2; Symporters; Time Factors

Adenosine and excitatory amino acids have been known to be involved in modulating nociceptive transmission at the spinal level. The authors assessed the characteristics of the interaction of the adenosine-excitatory amino acid antagonist combinations in the spinal cord of rats on the formalin-induced nociception. Intrathecal NMDA antagonist ((5R, 10S)-(+)-5-methyl-10,11-dihydro-(5)H-dibenzo[a[,]d]cyclohepten-5,10-imine hydrogen maleate, MK801, 30 microg) and AMPA antagonist (2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[F]quinoxaline-7-sulfonamide, NBQX, 3 microg) decreased the total number of flinches during both phases in the formalin test. Intrathecal adenosine (300 microg) had little effect on the phase 1 flinching response, but decreased the phase 2 response. The fixed dose analysis and the isobolographic analysis revealed that adenosine interacts additively with MK801 and NBQX in the spinal cord.

Topics: Adenosine; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Disease Models, Animal; Dizocilpine Maleate; Drug Synergism; Excitatory Amino Acid Antagonists; Formaldehyde; Injections, Spinal; Male; N-Methylaspartate; Pain; Quinoxalines; Rats; Rats, Sprague-Dawley

Organotypic hippocampal slice cultures represent a feasible model for studies of cerebral ischemia and the role of ionotropic glutamate receptors in oxygen-glucose deprivation-induced neurodegeneration. New results and a review of existing data are presented in the first part of this paper. The role of glutamate transporters, with special reference to recent results on inhibition of glutamate transporters under normal and energy-failure (ischemia-like) conditions is reviewed in the last part of the paper. The experimental work is based on hippocampal slice cultures derived from 7 day old rats and grown for about 3 weeks. In such cultures we investigated the subfield neuronal susceptibility to oxygen-glucose deprivation, the type of induced cell death and the involvement of ionotropic glutamate receptors. Hippocampal slice cultures were also used in our studies on glutamate transporters reviewed in the last part of this paper. Neurodegeneration was monitored and/or shown by cellular uptake of propidium iodide, loss of immunocytochemical staining for microtubule-associated protein 2 and staining with Fluoro-Jade B. To distinguish between necrotic vs. apoptotic neuronal cell death we used immunocytochemical staining for active caspase-3 (apoptosis indicator) and Hoechst 33342 staining of nuclear chromatin. Our experimental studies on oxygen-glucose deprivation confirmed that CA1 pyramidal cells were the most susceptible to this ischemia-like condition. Judged by propidium iodide uptake, a selective CA1 lesion, with only minor affection on CA3, occurred in cultures exposed to oxygen-glucose deprivation for 30 min. Nuclear chromatin staining by Hoechst 33342 and staining for active caspase-3 showed that oxygen-glucose deprivation induced necrotic cell death only. Addition of 10 microM of the N-methyl-D-aspartate glutamate receptor antagonist MK-801, and 20 microM of the non-N-methyl-D-aspartate glutamate receptor antagonist 2,3-dihyroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline to the culture medium confirmed that both N-methyl-D-aspartate and non-N-methyl-D-aspartate ionotropic glutamate receptors were involved in the oxygen-glucose deprivation-induced cell death. Glutamate is normally quickly removed, from the extracellular space by sodium-dependent glutamate transporters. Effects of blocking the transporters by addition of the DL-threo-beta-benzyloxyaspartate are reviewed in the last part of the paper. Under normal conditions addition of DL-threo-beta-benzylo

Topics: Amino Acid Transport System X-AG; Analysis of Variance; Animals; Animals, Newborn; Aspartic Acid; Cell Death; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Interactions; Excitatory Amino Acid Antagonists; Excitatory Amino Acid Transporter 1; Excitatory Amino Acid Transporter 2; Glial Fibrillary Acidic Protein; Glucose; Hippocampus; Histocytochemistry; Hypoxia; Immunohistochemistry; In Vitro Techniques; Microtubule-Associated Proteins; Necrosis; Neurofilament Proteins; Neurons; Neuroprotective Agents; Propidium; Quinoxalines; Rats; Receptors, Glutamate; Time Factors

To study the effects of intrathecal (it) agonists and antagonists of N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) receptors and NMDAR1 antisense oligodeoxynucleotides (AS ODN) on the antinociception of propofol.. Hot-plate test (HPPT) and acetic acid-induced writhing test were used to measure the nociceptive thresholds in mice. The effects of intrathecal NMDA, AMPA, MK-801, NBQX, or NMDAR1 AS ODN on the antinociception of propofol were observed.. Propofol (25, 50 mg/kg, ip) displayed an appreciable antinociceptive effect in hot-plate test and acetic acid-induced writhing test. NMDA (12.5, 25 ng, it) or AMPA (1.25, 2.5 ng, it) exhibited no effects on the behavior but decreased HPPT significantly compared with basal HPPT and aCSF group (P<0.05, P<0.01). No effects on behavior and HPPT were obtained in NMDA (6.25 ng, it) or AMPA (0.625 ng, it) groups. NMDA (6.25, 12.5, and 25 ng, it) dose-dependently decreased the HPPT in propofol-treated group. AMPA (1.25, 2.5 ng, it) also decreased HPPT significantly. MK-801 (0.25, 0.5 microg, it) or NBQX (0.25, 0.5 microg, it) groups had no behavioral changes, two antagonists 0.5 microg but not 0.25 microg increased HPPT in conscious or propofol-treated mice. AS ODN (5, 10, and 20 microg, it) groups exhibited dose-dependent increased in HPPT in propofol-treated groups compared with aCSF group (P<0.05, P<0.01).. Both agonists NMDA and AMPA reversed the antinociception of propofol. MK-801, NBQX, and NMDAR1 AS ODN potentiated the antinociceptive effects of propofol. Propofol produced antinociception through an interaction with spinal NMDA and AMPA receptors in mice.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Analgesics; Animals; Dizocilpine Maleate; Female; Injections, Spinal; Male; Mice; N-Methylaspartate; Nociceptors; Pain Threshold; Propofol; Quinoxalines; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate

Approximately 10% of total zinc in the brain exists in synaptic vesicles of glutamatergic neurons; however, the function of vesicular zinc is poorly understood. The presynaptic action of zinc against excitatory and inhibitory neurotransmission was studied in rat hippocampus using in vivo microdialysis. When the hippocampal CA3 region was perfused with 10-300 microM ZnCl(2), the level of glutamate in the perfusate was decreased, whereas the level of gamma-aminobutyric acid (GABA) was increased. Chelation of endogenous zinc with CaEDTA increased the glutamate level in the perfusate but decreased the GABA level, suggesting that zinc released into the synaptic cleft acts differentially on glutamatergic and GABAergic neurons in the CA3 region. The increase of GABA level by zinc was antagonized by 2,3-dioxo-6-nitro-1,2.3,4-tetrahydrobenzo(f)quinoxaline-7-sulphonamide (NBQX), an antagonist of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)/kainate receptors, but not affected by MK801, an antagonist of N-methyl-D-aspartate (NMDA) receptors, and verapamil, a blocker of voltage-dependent calcium channels. The present study suggests that zinc enhances GABA release via potentiation of AMPA/kainate receptors in the CA3 region, followed by a decrease in presynaptic glutamate release in the same region. Zinc seems to be an inhibitory neuromodulator of glutamate release.

Topics: Animals; Calcium Channel Blockers; Chelating Agents; Chlorides; Dizocilpine Maleate; Down-Regulation; Excitatory Amino Acid Antagonists; gamma-Aminobutyric Acid; Glutamic Acid; Hippocampus; Male; Neural Inhibition; Neural Pathways; Presynaptic Terminals; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Synapses; Synaptic Transmission; Up-Regulation; Zinc; Zinc Compounds

To investigate the role of glutamate receptors in overactive bladder (OAB) caused by cerebral infarction (CI) we examined the effects of 2 different types of receptors antagonists on OAB induced by left middle cerebral artery (MCA) occlusion.. Female rats were intravenously injected with dizocilpine, an NMDA (N-methyl-D-aspartate) receptor antagonist, or NBQX (2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo(f)quinoxaline-7-sulfonamide), an AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor antagonist, before or after MCA occlusion. Awake rats were cystometrically examined for 8 hours. Detrusor strips were evaluated for force development in response to dizocilpine and NBQX.. In CI rats without pretreatment bladder capacity (BC) was significantly decreased after MCA occlusion and remained consistently below half that of pre-occlusion capacity. Dizocilpine (0.5 mg/kg intravenously) administered before MCA occlusion blocked the decrease in BC in awake rats 5 to 8 hours after MCA occlusion. In CI rats pretreated with NBQX (10 or 30 mg/kg intravenously) BC was not different from that in rats without pretreatment. Increasing doses of dizocilpine (0.01 to 10 mg/kg) or NBQX (0.1 to 30 mg/kg) increased rat BC 2 hours after MCA occlusion. NBQX did not change the BC of sham operated rats. No differences in the contractile response to dizocilpine or NBQX of detrusor strips from sham operated and CI rats were observed.. These results indicate that NMDA receptor has an essential role in the development of OAB after CI. AMPA receptor antagonist cannot block the development of OAB. However, AMPA receptor antagonist temporally inhibits OAB after it is established by CI.

Topics: Animals; Cerebral Infarction; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Female; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, Glutamate; Urinary Bladder Diseases

The neurotoxic action of the abuse drugs methamphetamine (METH) and 3,4-methylenedioxymethamphetamine (MDMA) on cerebellar granule neurones (CGNs) culture was examined. Treatment for 48 h with METH or MDMA (1-5 mM) induced a higher decrease in viability than 24 h treatment. z.VAD.fmk (100 microM) but not MK-801 nor NBQX recovered control viability values. In both cases, cell death was characterised as apoptotic rather than necrotic by morphology cell observation. Apoptosis measured by flow cytometry indicated an increase in the hypodiploid population after 48 h treatment with METH and MDMA. Apoptosis was reverted by the presence of z.VAD.fmk (100 microM) but not by 10 microM MK-801 or NBQX. Similar results were obtained by analysing nuclear chromatine condensation. These results ruled out excitotoxic participation in amphetamine derivative-induced neurotoxicity in CGNs. Participation of radical oxygen species (ROS) was evaluated using alpha-tocopherol (1-15 microM) and cytometric studies. The co-treatment with 4 mM METH or MDMA for 48 h partially reverted neurotoxic action and apoptotic features, indicating ROS implication in CGNs death by amphetamine derivatives. Alteration of mitochondrial function induced cytochrome C (Cyt C) release after 48-h treatment with METH and MDMA (4 mM). There was also indication of caspase-3-like activation, measured by immunoanalysis and biochemically. Finally, neurodegenerative action caused by amphetamine derivatives may be prevented by using caspase inhibitors.

Topics: alpha-Tocopherol; Amino Acid Chloromethyl Ketones; Animals; Apoptosis; Caspase 3; Caspases; Cerebellum; Cytochromes c; Dizocilpine Maleate; Drug Interactions; Enzyme Activation; Flow Cytometry; Methamphetamine; Microscopy, Fluorescence; Microscopy, Phase-Contrast; Mitochondria; N-Methyl-3,4-methylenedioxyamphetamine; Neurons; Neuroprotective Agents; Quinoxalines; Rats; Reactive Oxygen Species

The phosphorylation of calcium/calmodulin-dependent protein kinase (CaMK) II, induced by an increase in the intracellular Ca2+ concentration, is involved in the alteration of brain functions such as memory formation. In the present study, we examined the influence of various immobilization stress paradigms on the phosphorylation of CaMKII (phospho-CaMKII) and CaMKII levels in the rat hippocampus. Immunoblot and immunohistochemical analyses were performed to examine the levels of CaMKII and phospho-CaMKII. Real-time quantitative polymerase chain reaction (PCR) was performed to analyse the mRNA levels of N-methyl-D-aspartic acid (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subtypes. Acute (single) and repeated (4 d), but not chronic (14 d), stress exposure of 45 min or longer duration significantly increased phospho-CaMKII levels without affecting the levels of CaMKII. Pre-treatment with NBQX, a selective AMPA receptor antagonist, significantly prevented this stress-induced increase. In contrast, two NMDA receptor antagonists, LY235959 and MK-801, showed no inhibitory effect on phospho-CaMKII levels during acute stress. Neither acute nor chronic stress changed mRNA levels of NMDA and AMPA receptors. These results demonstrate that immobilization stress promotes the phosphorylation of CaMKII. The increase in the intracellular Ca2+ concentration by the activation of AMPA receptors may play a role in the stress-induced phospho-CaMKII in the rat hippocampus.

Topics: Animals; Blotting, Western; Calcium Channel Blockers; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Calcium-Calmodulin-Dependent Protein Kinases; Dizocilpine Maleate; DNA Primers; Excitatory Amino Acid Antagonists; Hippocampus; Immunohistochemistry; Isoenzymes; Isoquinolines; Male; Nimodipine; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Restraint, Physical; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Stress, Psychological; Up-Regulation

Numerous data indicate that nonsynaptic release of glutamate occurs both in normal and pathophysiological conditions. When reaching receptors in the postsynaptic density (PSD), glutamate (Glu) could affect the synaptic transmission. We have tested this possibility in the hippocampal CA1 synapses of rats, either by applying exogenous Glu to the CA1 neurons or by disruption of Glu transporter activity. L-Glu (400 microM) was directly applied to the hippocampal slices acutely isolated from the rats. It produced a strong inhibition of both ortho- and antidromically elicited action potentials fired by CA1 neurons while the excitatory postsynaptic current (EPSC) measured in these neurons remained totally unaffected. The optical isomer D-Glu which is not transported by the systems of Glu uptake inhibited not only orthodromic and antidromic spikes, but also EPSC. Non-specific glutamate transporter inhibitor DL-threo-beta-hydroxyaspartic acid (THA, 400 microM) mimicked the effects of exogenous Glu and produced strong inhibition of both orthodromic and antidromic spikes, without any influence on the amplitude of EPSCs. Dihydrokainate (DHK, 300 microM), selective inhibitor of GLT-1 subtype of glutamate transporter, exerted a significant inhibitory action on the orthodromically evoked spikes and also on the EPSC. Our results indicate that extrasynaptic and PSD membranes of CA1 neurons form separate compartments differing in the mechanisms and efficiency of external Glu processing: the protection of PSD markedly prevails.

Topics: 4-Aminopyridine; Amino Acid Transport System X-AG; Animals; Animals, Newborn; Aspartic Acid; Dizocilpine Maleate; Drug Interactions; Evoked Potentials; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Glutamic Acid; Hippocampus; In Vitro Techniques; Kainic Acid; Models, Neurological; N-Methylaspartate; Neural Inhibition; Neurons; Potassium Channel Blockers; Quinoxalines; Rats; Rats, Wistar; Synapses

1. Ingestion of a poisonous mushroom Clitocybe acromelalga is known to cause severe tactile pain (allodynia) in the extremities for a month and acromelic acid (ACRO), a kainate analogue isolated from the mushroom, produces selective damage of interneurons of the rat lower spinal cord when injected either systemically or intrathecally. Since ACRO has two isomers, ACRO-A and ACRO-B, here we examined their acute and late effects on induction of allodynia. 2. Intrathecal administration of ACRO-A and ACRO-B provoked marked allodynia by the first stimulus 5 min after injection, which lasted over the 50-min experimental period. Dose-dependency of the acute effect of ACRO-A on induction of allodynia showed a bell-shaped pattern from 50 ag x kg(-1) to 0.5 pg x kg(-1) and the maximum effect was observed at 50 fg x kg(-1). On the other hand, ACRO-B induced allodynia in a dose-dependent manner from 50 pg x kg(-1) to 50 ng x kg(-1). 3. N-methyl-d-aspartate (NMDA) receptor antagonists and Joro spider toxin, a Ca(2+)-permeable AMPA receptor antagonist, inhibited the allodynia induced by ACRO-A, but not by ACRO-B. However, other AMPA/kainate antagonists did not affect the allodynia induced by ACRO. 4. Whereas no neuronal damage was observed in the spinal cord in ACRO-A-treated mice, induction of allodynia by ACRO-A (50 fg x kg(-1)) and ACRO-B (50 ng x kg(-1)) was selectively lost 1 week after i.t. injection of a sublethal dose of ACRO-A (50 ng x kg(-1)) or ACRO-B (250 ng x kg(-1)). Higher doses of ACRO-A, however, could evoke allodynia dose-dependently from 50 pg x kg(-1) to 500 ng x kg(-1) in the ACRO-A-treated mice. The allodynia induced by ACRO-A (500 ng x kg(-1)) was not inhibited by Joro spider toxin or NMDA receptor antagonists. These properties of the late allodynia induced by ACRO-A were quite similar to those of the acute allodynia induced by ACRO-B. 5. ACRO-A could increase [Ca(2+)](i) in the deeper laminae, rather than in the superficial laminae, of the spinal cord. This increase was not blocked by the AMPA-preferring antagonist GYKI52466 and Joro spider toxin. 6. Taken together, these results demonstrate the stereospecificity of ACRO for the induction of allodynia and suggest the presence of a receptor specific to ACRO.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Basidiomycota; Benzodiazepines; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; Glutamates; Heterocyclic Compounds; Indoles; Injections, Spinal; Japan; Kainic Acid; Lumbosacral Region; Male; Mice; Mice, Inbred Strains; Mushroom Poisoning; Oximes; Pain; Quinoxalines; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Spider Venoms; Spinal Cord; Stereoisomerism; Structure-Activity Relationship; Time Factors

Carbon monoxide (CO) poisoning caused by CO exposure at 3000 ppm for 40 min resulted in stimulation of hydroxyl radical (*OH) generation (estimated by measuring 2,3-dihydroxybenzoic acid (2,3-DHBA) production from salicylic acid) in the striatum of free-moving rats, as determined by means of brain microdialysis. Pretreatment with a voltage-dependent Na+ channel blocker, tetrodotoxin (TTX), lowered the basal level of 2,3-DHBA and strongly suppressed the increase in 2,3-DHBA induced by CO poisoning. CO poisoning significantly, though only slightly, increased extracellular glutamate in the striatum, and glutamate (Glu) receptor antagonists, such as MK-801 (dizocilpine) and NBQX, failed to suppress the CO-induced increase in 2,3-DHBA. These findings suggest that CO poisoning may induce Na+ influx via the voltage-dependent Na+ channels, resulting in stimulation of *OH generation in rat striatum. This effect may be independent of Glu receptor activation by increased extracellular Glu.

Topics: Animals; Carbon Monoxide Poisoning; Corpus Striatum; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Hydroxybenzoates; Hydroxyl Radical; Male; Microdialysis; Quinoxalines; Rats; Rats, Sprague-Dawley; Tetrodotoxin; Wakefulness

It is widely believed that long-term depression (LTD) and its counterpart, long-term potentiation (LTP), involve mechanisms that are crucial for learning and memory. However, LTD is difficult to induce in adult cortex for reasons that are not known. Here we show that LTD can be readily induced in adult cortex by the activation of NMDA receptors (NMDARs), after inhibition of glutamate uptake. Interestingly there is no need to activate synaptic NMDARs to induce this LTD, suggesting that LTD is triggered primarily by extrasynaptic NMDA receptors. We also find that de novo LTD requires the activation of NR2B-containing NMDAR, whereas LTP requires activation of NR2A-containing NMDARs. Surprisingly another form of LTD, depotentiation, requires activation of NR2A-containing NMDARs. Therefore, NMDARs with different synaptic locations and subunit compositions are involved in various forms of synaptic plasticity in adult cortex.

Topics: 2-Amino-5-phosphonovalerate; Animals; Aspartic Acid; Cerebral Cortex; Dicarboxylic Acids; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Glutamic Acid; Long-Term Potentiation; Long-Term Synaptic Depression; N-Methylaspartate; Neurons; Neurotransmitter Uptake Inhibitors; Phenols; Picrotoxin; Piperidines; Protein Subunits; Pyrrolidines; Quinoxalines; Rats; Receptors, Metabotropic Glutamate; Receptors, N-Methyl-D-Aspartate

Intracellular ATP supply and ion homeostasis determine neuronal survival and degeneration after ischemic stroke. The present study provides a systematic investigation in organotypic hippocampal slice cultures of the influence of experimental ischemia, induced by oxygen-glucose-deprivation (OGD). The pathways controlling intracellular Na(+) and Ca(2+) concentration ([Na(+)](i) and [Ca(2+)](i)) and their inhibition were correlated with delayed cell death or protection. OGD induced a marked decrease in the ATP level and a transient elevation of [Ca(2+)](i) and [Na(+)](i) in cell soma of pyramidal neurons. ATP level, [Na(+)](i) and [Ca(2+)](i) rapidly recovered after reintroduction of oxygen and glucose. Pharmacological analysis showed that the OGD-induced [Ca(2+)](i) elevation in neuronal cell soma resulted from activation of both N-methyl-d-aspartate (NMDA)-glutamate receptors and Na(+)/Ca(2+) exchangers, while the abnormal [Na(+)](i) elevation during OGD was due to Na(+) influx through voltage-dependent Na(+) channels. In hippocampal slices, cellular degeneration occurring 24 h after OGD, selectively affected the pyramidal cell population through apoptotic and non-apoptotic cell death. OGD-induced cell loss was mediated by activation of ionotropic glutamate receptors, voltage-dependent Na(+) channels, and both plasma membrane and mitochondrial Na(+)/Ca(2+) exchangers. Thus, we show that neuroprotection induced by blockade of NMDA receptors and plasma membrane Na(+)/Ca(2+) exchangers is mediated by reduction of Ca(2+) entry into neuronal soma, whereas neuroprotection induced by blockade of AMPA/kainate receptors and mitochondrial Na(+)/Ca(2+) exchangers might result from reduced Na(+) entry at dendrites level.

Topics: Adenosine Triphosphate; Animals; Animals, Newborn; Boron Compounds; Calcium; Calcium Channel Blockers; Cell Death; Clonazepam; Dantrolene; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Fura-2; Glucose; Hippocampus; Hypoxia; In Situ Nick-End Labeling; Indoles; Intracellular Space; Ion Exchange; Lidocaine; Mibefradil; Nimodipine; Organ Culture Techniques; Quinoxalines; Rats; Rats, Wistar; Sodium; Sodium Channel Blockers; Thiazepines; Thiourea; Time Factors

Traumatic brain injury (TBI) involves alterations in neuronal physiology, often complicated by secondary hypoxic or hypotensive events. Excitotoxicity is an important process induced in both TBI and hypoxic or ischemic insults to the brain. We investigated two hypotheses: (1) excitotoxicity is more prominent following combined mechanical and hypoxic injury than either alone; (2) both AMPA and NMDA receptor activation mediate combined mechanical and hypoxic injury. Media in primary mixed neuronal cultures were replaced with conditioned media containing MK801 (NMDA antagonist) and/or NBQX (AMPA/kainate antagonist). Cultures were then subjected to mechanical injury. Afterward, media were exchanged for hypoxic media containing the antagonist, and plates were placed in hypoxia chambers for 7 h. At 24 h following hypoxia, LDH release, trypan blue uptake, and morphologic changes were assessed. Blockade had no effect after mechanical injury. After hypoxia, MK801 and combined MK801/NBQX decreased LDH and trypan blue to control levels. NBQX alone after hypoxia had less impact. After combined mechanical injury and hypoxia, both MK801 and NBQX partially reduced LDH and trypan blue. Combining the antagonists led to reduction to control values for both endpoints. We conclude that excitotoxic processes are more prominent after combined than isolated injuries in neurons and that increased cell death is mediated by both NMDA and AMPA receptor activation following combined injuries.

Topics: Animals; Anticonvulsants; Cell Hypoxia; Cell Survival; Cells, Cultured; Dizocilpine Maleate; Female; Neurons; Pregnancy; Quinoxalines; Rats; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate

Hypoxia of the brain may alter further seizure susceptibility in a different way. In this study, we tried to answer the question how episode of convulsion induced by hypoxia (HS) changes further seizure susceptibility, and how N-methyl-D-aspartic acid (NMDA) and AMPA/KA receptor antagonists influence this process. Adult Albino Swiss mice exposed to hypoxia (5% O(2)) developed clonic/tonic convulsions after about 340 s. Mice which underwent 10 s but not 5 s seizures episode subsequently exhibited significantly increased seizure susceptibility to low doses (equal ED(16)) of bicuculline (BCC) and NMDA during a 3-week observation period. No morphological signs of brain tissue damage were seen in light microscope on the third day after a hypoxia-induced seizure (HS). Learning abilities assessed in passive avoidance test as well as spontaneous alternation were not disturbed after an HS episode. Pretreatment with AMPA/KA receptor antagonist NBQX effectively prolonged latency to HS and given immediately after seizure episode also attenuated subsequent convulsive susceptibility rise, however, NMDA receptor antagonist, MK-801, appeared to be ineffective. These results suggest that a seizure episode induced by hypoxia, depending on its duration, may play an epileptogenic role. The AMPA/KA receptor antagonist prolongs the latency to HS, and given after this episode, prevents the long-term epileptogenic effect.

Topics: Animals; Avoidance Learning; Bicuculline; Brain Chemistry; Dizocilpine Maleate; Epilepsy; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Female; GABA Antagonists; gamma-Aminobutyric Acid; Hypoxia; Mice; N-Methylaspartate; Quinoxalines; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Seizures

The role of AMPA and NMDA receptors in the shell of the nucleus accumbens in turning behaviour of rats was investigated. Unilateral injection of the AMPA receptor agonist, AMPA (0.25, 0.4, 0.5 and 1 microg), into the shell of the nucleus accumbens dose-dependently produced contraversive pivoting, namely tight head-to-tail turning marked by abnormal hindlimb backward stepping, while injection of AMPA (0.5 microg) into the core produced only a marginal effect. This shell-specific AMPA effect was dose-dependently inhibited by the AMPA receptor antagonist, NBQX (1 and 10 ng), which alone did not produce turning behaviour. The AMPA-induced pivoting was also dose-dependently inhibited by the non-competitive NMDA receptor antagonist, MK-801 (0.1 and 0.5 microg). Neither MK-801 (0.1, 0.5 and 5 microg) nor the NMDA receptor agonist, NMDA (0.5 and 1 microg), injected unilaterally into the shell, produced turning behaviour. Unilateral injection of a mixture of dopamine D(1) (SKF 38393, 5 microg) and D(2) (quinpirole, 10 microg) receptor agonists into the shell has been found to elicit contraversive pivoting. The dopamine D(1)/D(2) receptor antagonist, cis-(Z)-flupentixol (1 and 10 microg), injected into the shell, in doses known to block dopamine D(1)/D(2) receptor-mediated pivoting, also significantly inhibited AMPA (0.5 microg)-induced pivoting. Moreover, both NBQX (1 and 10 ng) and MK-801 (0.1 and 0.5 microg), injected into the shell, significantly inhibited dopamine D(1)/D(2) receptor-mediated pivoting. It is therefore concluded that unilateral stimulation of AMPA receptors in the shell of the nucleus accumbens can elicit contraversive pivoting, and that both AMPA and dopamine D(1)/D(2) receptors play a critical role in shell-specific pivoting in contrast to NMDA receptors that at best play only a modulatory role.

Topics: 2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine; Animals; Dizocilpine Maleate; Dopamine Agonists; Dopamine Antagonists; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Flupenthixol; Injections, Intraventricular; Male; Movement; Nucleus Accumbens; Quinoxalines; Quinpirole; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Dopamine; Receptors, Dopamine D1; Receptors, Dopamine D2; Receptors, N-Methyl-D-Aspartate

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

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

In this study we investigated the effects of deltamethrin on the expression of c-Fos and c-Jun in the cerebral cortex of rats. Immunohistochemical analysis demonstrated that the immunoreactivity for c-Fos was markedly increased in the cerebral cortex 5 h after deltamethrin treatment, and maintained at an increased level at 24 h, even though little immunoreactivity for c-Fos was seen in the same brain region of control rats. The immunostaining for c-Jun was also dramatically elevated in the same brain region, showing the same time course of c-Fos expression after deltamethrin treatment. Further, both MK-801, an N-methyl-D-aspartate (NMDA) receptor antagonist, and NBQX, an alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)/kainate (KA) receptor antagonist, attenuated deltamethrin-elicited prolonged expression of c-Fos and c-Jun. Since the persistent expression of c-Fos and c-Jun is unusual, and has been reported before in conditions involving neurodegeneration, our results are consistent with a model that deltamethrin induces neurodegeneration through a glutamate-dependent pathway.

Topics: Animals; Cerebral Cortex; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Immunohistochemistry; Insecticides; Male; Nitriles; Proto-Oncogene Proteins c-fos; Proto-Oncogene Proteins c-jun; Pyrethrins; Quinoxalines; Rats; Rats, Sprague-Dawley

Two main features make microelectrode arrays (MEAs) a valuable tool for electrophysiological measurements under the perspective of pharmacological applications, namely: (i) they are non-invasive and permit, under appropriate conditions, to monitor the electrophysiological activity of neurons for a long period of time (i.e. from several hours up to months); (ii) they allow a multi-site recording (up to tens of channels). Thus, they should allow a high-throughput screening while reducing the need for animal experiments. In this paper, by taking advantages of these features, we analyze the changes in activity pattern induced by the treatment with specific substances, applied on dissociated neurons coming from the chick-embryo spinal cord. Following pioneering works by Gross and co-workers (see e.g. Gross and Kowalski, 1991. Neural Networks, Concepts, Application and Implementation, vol. 4. Prentice Hall, NJ, pp. 47-110; Gross et al., 1992. Sensors Actuators, 6, 1-8.), in this paper analysis of the drugs' effects (e.g. NBQX, CTZ, MK801) to the collective electrophysiological behavior of the neuronal network in terms of burst activity, will be presented. Data are simultaneously recorded from eight electrodes and besides variations induced by the drugs also the correlation between different channels (i.e. different area in the neural network) with respect to the chemical stimuli will be introduced (Bove et al., 1997. IEEE Trans. Biomed. Eng., 44, 964-977.). Cultured spinal neurons from the chick embryo were chosen as a neurobiological system for their relative simplicity and for their reproducible spontaneous electrophysiological behavior. It is well known that neuronal networks in the developing spinal cord are spontaneously active and that the presence of a significant and reproducible bursting activity is essential for the proper formation of muscles and joints (Chub and O'Donovan, 1998. J. Neurosci., 1, 294-306.). This fact, beside a natural variability among different biological preparations, allows a comparison also among different experimental session giving reliable results and envisaging a definition of a bioelectronic 'neuronal sensory system'.

Topics: Algorithms; Animals; Benzothiadiazines; Biosensing Techniques; Cells, Cultured; Cells, Immobilized; Chick Embryo; Chickens; Dizocilpine Maleate; Microelectrodes; Nerve Net; Quinoxalines; Signal Processing, Computer-Assisted

Classical in vitro and in vivo models and electrophysiological techniques were used to investigate the role of AMPA- and NMDA-type glutamate receptors in various components of spinal segmental reflex potentials. In the rat hemisected spinal cord preparation, the AMPA antagonists NBQX and GYKI 52466 abolished the monosynaptic reflex (MSR) potential but caused only partial inhibition of the motoneuronal population EPSP. NMDA antagonists had no noticeable effect on the MSR in normal medium, but markedly depressed the late part of EPSP. However, an NMDA receptor antagonist sensitive monosynaptic response was recorded in magnesium-free medium at complete blockade of the AMPA receptors. In spinalized rats, the AMPA antagonists completely blocked all components of the dorsal root stimulation evoked potential. MK-801 (2mg/kg, i.v.) reduced monosynaptic responses in a frequency dependent way, with no effect at 0.03 Hz and 22% inhibition at 0.25 Hz. The reduction of the di- and polysynaptic reflex components was about 30% and did not depend on stimulation frequency. Long-latency reflex discharge responses, especially when evoked by train stimulation, were more sensitive to MK-801 than the polysynaptic reflex. These results suggest that glutamate activates MSR pathways through AMPA receptors. However, under certain conditions, NMDA receptors can modulate this transmission through plastic changes in the underlying neuronal circuits. AMPA and NMDA receptors play comparable roles in the mediation of longer latency reflex components.

Topics: Action Potentials; Animals; Animals, Newborn; Anti-Anxiety Agents; Benzodiazepines; Dizocilpine Maleate; Dose-Response Relationship, Drug; Electric Stimulation; Excitatory Amino Acid Antagonists; In Vitro Techniques; Magnesium; Models, Neurological; Piperazines; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Reflex, Monosynaptic; Spinal Cord; Synapses; Time Factors; Valine

This study investigated the role of ionotropic and metabotropic glutamate receptors in the deficits in brain reward function, as measured by elevations in intracranial self-stimulation (ICSS) reward thresholds, associated with nicotine withdrawal. The group II metabotropic glutamate (mGluII) receptor agonist LY314582 [a racemic mixture of LY354740 ([+]-2-aminobicyclo[3.1.0]hexane-2,6-dicarboxylic acid])] (2.5-7.5 mg/kg) precipitated withdrawal-like elevations in ICSS thresholds, a sensitive measure of reward function, in nicotine-dependent but not control rats. LY314582 did not affect response latencies, a measure of performance in the ICSS paradigm. Bilateral microinfusion of LY314582 (10-100 ng/side) into the ventral tegmental area likewise precipitated dose-dependent threshold elevations in nicotine-dependent rats. Furthermore, a single injection of the mGluII receptor antagonist LY341495 (2S-2-amino-2-[1S,2S-2-carboxycyclopropan-1-yl]-3-[xanth-9-yl]propionic acid) (1 mg/kg) attenuated the threshold elevations observed in rats undergoing spontaneous nicotine withdrawal. mGluII receptors are primarily located on glutamatergic terminals throughout the mesocorticolimbic system, where they act as inhibitory autoreceptors. To investigate whether mGluII receptors contributed to nicotine withdrawal by decreasing glutamatergic transmission, we next examined whether direct blockade of postsynaptic glutamate receptors precipitated withdrawal-like reward deficits in nicotine-dependent rats. The alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA)/kainate receptor antagonist 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(f)quinoxaline (NBQX; 0.01-1 mg/kg) precipitated withdrawal-like threshold elevations in nicotine-dependent but not control rats, whereas 6-methyl-2-[phenylethynyl]-pyridine (MPEP; 0.01-3 mg/kg) and dizocilpine (MK-801; 0.01-0.2 mg/kg), antagonists at metabotropic glutamate 5 and N-methyl-d-aspartate receptors, respectively, did not. Overall, these data demonstrate that mGluII receptors play an important role in the reward deficits associated with nicotine withdrawal. Furthermore, it is likely that mGluII receptors generate this reward deficit, at least in part, by decreasing glutamate transmission at AMPA/kainate receptors.

Topics: Animals; Bridged Bicyclo Compounds; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Male; Nicotine; Pyridines; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Glutamate; Receptors, Kainic Acid; Receptors, Metabotropic Glutamate; Receptors, N-Methyl-D-Aspartate; Reward; Substance Withdrawal Syndrome; Ventral Tegmental Area

Prolonged inhibition of glutamate reuptake by L-trans-pyrrolidine-2,4-dicarboxylate (PDC), a specific glutamate transporter blocker, reduced the number of GABA positive neurons in a primary cerebellar culture by 54%. The disappearance of immunostaining for GABA was gradual and was partially prevented by the N-methyl-D-aspartate (NMDA) receptor blocker, MK-801, and the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptor antagonist, NBQX. Combined blockade of NMDA and AMPA receptors restored the original proportion of GABAergic neurons observed in control cultures. Following the PDC exposure, expression of other GABAergic markers, such as glutamic acid decarboxylase (GAD) and vesicular GABA transporter (VGAT) was also dramatically decreased in an AMPA receptor-dependent manner. Loss of GABA or GAD immunostaining is commonly regarded as a sign of degeneration of GABAergic neurons. However, none of the GABAergic neurons were positive for propidium iodide uptake or showed abnormal nuclear morphology. Based on the above data we conclude that prolonged activation of ionotropic glutamate receptors by endogenously released glutamate was not toxic to cerebellar GABAergic neurons, but lead to the loss of their characteristic neurotransmitter phenotype.

Topics: Animals; Animals, Newborn; Carrier Proteins; Cell Death; Cells, Cultured; Cerebellum; Chromatography, High Pressure Liquid; Dicarboxylic Acids; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Interactions; Excitatory Amino Acid Antagonists; Fluorescent Dyes; GABA Plasma Membrane Transport Proteins; gamma-Aminobutyric Acid; Gene Expression; Glutamate Decarboxylase; Glutamic Acid; Indicators and Reagents; Membrane Proteins; Membrane Transport Proteins; Neurofilament Proteins; Neurons; Neurotransmitter Uptake Inhibitors; Organic Anion Transporters; Organic Chemicals; Propidium; Pyrrolidines; Quinoxalines; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Time Factors

Glucose is the main substrate that fulfills energy brain demands. However, in some circumstances, such as diabetes, starvation, during the suckling period and the ketogenic diet, brain uses the ketone bodies, acetoacetate and beta-hydroxybutyrate, as energy sources. Ketone body utilization in brain depends directly on its blood concentration, which is normally very low, but increases substantially during the conditions mentioned above. Glutamate neurotoxicity has been implicated in neurodegeneration associated with brain ischemia, hypoglycemia and cerebral trauma, conditions related to energy failure, and to elevation of glutamate extracellular levels in brain. In recent years substantial evidence favoring a close relation between glutamate neurotoxic potentiality and cellular energy levels, has been compiled. We have previously demonstrated that accumulation of extracellular glutamate after inhibition of its transporters, induces neuronal death in vivo during energy impairment induced by glycolysis inhibition. In the present study we have assessed the protective potentiality of the ketone body, acetoacetate, against glutamate-mediated neuronal damage in the hippocampus of rats chronically treated with the glycolysis inhibitor, iodoacetate, and in hippocampal cultured neurons exposed to a toxic concentration of iodoacetate. Results show that acetoacetate efficiently protects against glutamate neurotoxicity both in vivo and in vitro probably by a mechanism involving its role as an energy substrate.

Topics: Acetoacetates; Adenosine Triphosphate; Animals; Cell Survival; Cells, Cultured; Dicarboxylic Acids; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Administration Routes; Drug Administration Schedule; Drug Interactions; Embryo, Mammalian; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Female; Glutamic Acid; Glycolysis; Hippocampus; Iodoacetates; Male; Neurons; Neuroprotective Agents; Neurotransmitter Uptake Inhibitors; Pregnancy; Pyrrolidines; Pyruvic Acid; Quinoxalines; Rats; Rats, Wistar; Time Factors

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

In the rat, postsynaptic 5-hydroxytryptamine2A receptors medial prefrontal cortex control the activity of the serotonergic system through changes in the activity of pyramidal neurons projecting to the dorsal raphe nucleus. Here we extend these observations to mouse brain. The prefrontal cortex expresses abundant 5- hydroxytryptamine2A receptors, as assessed by immunohistochemistry, Western blots and in situ hybridization procedures. The application of the 5-hydroxytryptamine2A/2C agonist DOI (100 microm) by reverse dialysis in the medial prefrontal cortex doubled the local release of 5-hydroxytryptamine. This effect was reversed by coperfusion of tetrodotoxin, and by the selective 5-hydroxytryptamine2A receptor antagonist M100907, but not by the 5-hydroxytryptamine2C antagonist SB-242084. The effect of DOI was also reversed by prazosin (alpha1-adrenoceptor antagonist), BAY x 3702 (5-hydroxytryptamine1A receptor agonist), NBQX (alpha-amino-3-hydroxy-5-methyl-4-isoxazole-4-propionate/kainic acid antagonist) and 1S,3S-ACPD (mGluR II/III agonist), but not by dizocilpine (N-methyl-d-aspartate antagonist). alpha-Amino-3-hydroxy-5-methyl-4-isoxazole-4-propionate mimicked the 5-hydroxytryptamine elevation produced by DOI, an effect also reversed by BAY x 3702. Likewise, the coperfusion of classical (chlorpromazine, haloperidol) and atypical antipsychotic drugs (clozapine, olanzapine) fully reversed the 5-hydroxytryptamine elevation induced by DOI. These observations suggest that DOI increases 5-hydroxytryptamine release in the mouse medial prefrontal cortex through the activation of local 5-hydroxytryptamine2A receptors by an impulse-dependent mechanism that involves/requires the activation of local alpha-amino-3-hydroxy-5-methyl-4-isoxazole-4-propionate receptors. This effect is reversed by ligands of receptors present in the medial prefrontal cortex, possibly in pyramidal neurons, which are involved in the action of antipsychotic drugs. In particular, the reversal by classical antipsychotics may involve blockade of alpha1-adrenoceptors, whereas that of atypical antipsychotics may involve 5-hydroxytryptamine2A receptors and alpha1-adrenoceptors.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Aminopyridines; Anesthetics, Local; Animals; Antipsychotic Agents; Benzopyrans; Blotting, Western; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Interactions; Excitatory Amino Acid Antagonists; Fluorobenzenes; Immunohistochemistry; In Situ Hybridization; Indoles; Indophenol; Male; Mice; Mice, Inbred C57BL; Microdialysis; Piperidines; Prazosin; Prefrontal Cortex; Quinoxalines; Receptor, Serotonin, 5-HT2A; Receptors, Serotonin; RNA, Messenger; Serotonin; Serotonin Antagonists; Serotonin Receptor Agonists; Tetrodotoxin; Thiazoles; Time Factors

We investigated the effect of agmatine on cell viability of rat cerebellar granule neurons in a high-K+ (27.5 mM) medium. Exposure of cultured rat cerebellar granule neurons to agmatine (200-800 microM) resulted in a significant decrease in cell viability. Agmatine-induced neuronal death began to occur 6-12 h after addition, and gradually progressed. The agmatine neurotoxicity was attenuated by N-methyl-D-aspartate (NMDA) receptor antagonists and by enzymatic degradation of L-glutamate with glutamic pyruvic transaminase. Furthermore, a significant increase in extracellular L-glutamate concentration was detected before cell death occurred. In addition, agmatine-induced glutamate release and cell death were both blocked by pretreatment with botulinum toxin C, which is known to specifically inhibit the exocytosis. The agmatine neurotoxicity was not observed when extracellular K+ concentration was lower (10 mM). These results suggest that agmatine induces glutamate release through the exocytosis and thereby causes NMDA receptor-mediated neuronal death in conditions in which extracellular K+ concentrations are elevated.

Topics: Agmatine; Animals; Botulinum Toxins; Cell Death; Cell Survival; Cells, Cultured; Cerebellum; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; L-Lactate Dehydrogenase; Neurons; Potassium; Quinoxalines; Rats; Tetrazolium Salts; Thiazoles

To determine whether both the N-methyl-D-aspartate (NMDA) and non-NMDA receptors in the knee joint contribute to the induction and/or maintenance of arthritic pain, we examined the effects of intra-articular injection of NMDA receptor antagonist dizocilpine (MK-801) and non-NMDA receptor antagonist 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo[f]quinoxaline (NBQX) on the decrease in weight load induced by carrageenan injection into the knee joint cavity in rats. Injection of MK-801 (0.75 and 1.5 mM) and NBQX (0.25, 0.625 and 2.5 mM) immediately prior to carrageenan injection (2%, 40 microl) significantly prevented the pain-related behavior. However, injection of MK-801 (0.75 and 1.5 mM) and NBQX (0.625 and 2.5 mM) 5 h after carrageenan injection had no effect on pain-related behavior. These results suggest that both the NMDA and non-NMDA receptors in the knee joint are involved in the induction, but not maintenance, of arthritic pain.

Topics: Animals; Arthritis, Experimental; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Joints; Male; Pain; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Time Factors

The purpose of this study was to examine the effects of inhibiting ionotropic glutamate receptor subtypes on measures of oxidative stress events at acute times following traumatic spinal cord injury (SCI). Rats received a moderate contusion injury and 15 min later were treated with one of two doses of 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo-benzol[f]quinoxaline-7-sulfonamide disodium (NBQX), MK-801, or the appropriate vehicle. At 4 h following injury, spinal cords were removed and a crude synaptosomal preparation obtained to examine mitochondrial function using the MTT assay, as well as measures of reactive oxygen species (ROS), lipid peroxidation, and glutamate and glucose uptake. We report here that intraspinal treatment with either 15 or 30 nmol of NBQX improves mitochondrial function and reduces the levels of ROS and lipid peroxidation products. In contrast, MK-801, given intravenously at doses of 1.0 or 5.0 mg/kg, was without effect on these same measures. Neither drug treatment had an effect on glutamate or glucose uptake, both of which are reduced at acute times following SCI. Previous studies have documented that drugs acting on non-N-methyl-D-aspartate (NMDA) receptors exhibit greater efficacy compared to NMDA receptor antagonists on recovery of function and tissue sparing following traumatic spinal cord injury. The results of this study provide a potential mechanism by which blockade of the non-NMDA ionotropic receptors exhibit positive effects following traumatic SCI.

Topics: Animals; Dizocilpine Maleate; Female; Glucose; Glutamic Acid; Injections, Spinal; Lipid Peroxidation; Mitochondria; Neuroprotective Agents; Quinoxalines; Rats; Rats, Long-Evans; Reactive Oxygen Species; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Spinal Cord Injuries; Synaptosomes; Thiobarbituric Acid Reactive Substances; Thoracic Vertebrae

Upon systemic administration of apomorphine, a potent dopamine agonist, pigeons show a bout of pecking behaviour. When the drug is repeatedly administered a sensitization takes place that is associated with pronounced discrimination learning. Here we show that intra-cerebral injections of apomorphine in the periphery of the nucleus accumbens of pigeons also elicit pecking. We additionally show that injections of 5-amino-phosphonohepatnoic acid, a NMDA-glutamate receptor blocker, into the Acc impairs the performance of a learned visual discrimination incorporating pecking as a choice response. We conclude that, as it is the case in mammals, the control mechanisms of learned sensory-motor behaviour in birds involves dopaminergic and glutamatergic synaptic transmission within the nucleus accumbens area.

Topics: Animals; Apomorphine; Behavior, Animal; Columbidae; Discrimination Learning; Dizocilpine Maleate; Dopamine; Dopamine Agonists; Excitatory Amino Acid Antagonists; Glutamic Acid; Injections; Nucleus Accumbens; Quinoxalines; Receptors, N-Methyl-D-Aspartate; Stereotaxic Techniques; Stimulation, Chemical

To understand the pathogenesis of diffuse axonal injury, we investigated the temporal and spatial profiles of neuronal degeneration in impact-acceleration injury in rats using Fluoro-Jade (FJ) staining. Impact-acceleration injury was produced in Wistar rats by the method described by Marmarou et al. with some modifications. Animals were sacrificed 1, 2, 7, 14, or 28 days after injury. Paraffin-embedded coronal sections were stained with HE or FJ, or analyzed immunohistochemically for GFAP or amyloid precursor protein (APP). FJ-positive degenerative neurons were found primarily in the dorsal brainstem and thalamus from 1 to 2 days following injury and these were associated with GFAP expression. However, FJ-positive cells were rarely found after 7 days. In all rats, significant expression of APP was observed primarily in the cingulum, cerebral peduncle and pontomedullary junction. FJ also stained these injured axons. Intrathecal administration of both NMDA and AMPA/kinate glutamate receptor antagonists MK-801 and NBQX, respectively, reduced the neuronal injury. NBQX showed more significant effects on axonal injury than MK-801. These observations indicate that not only axonal damage, but also primary neuronal damage occurs in this impact-acceleration injury model. It is also suggested that NBQX can act both directly on neuronal cells and white matter and that NMDA could have a significant protective effect against not only neuronal, but also axonal injury.

Topics: Amyloid beta-Protein Precursor; Animals; Brain Injuries; Brain Stem; Dizocilpine Maleate; Glial Fibrillary Acidic Protein; Male; Neurons; Quinoxalines; Rats; Rats, Wistar; Thalamus

This study characterizes the receptor subtypes and investigates some of the mechanisms by which glutamate, injected intraplantarly (i.pl.) into the mouse paw, produces nociception and paw oedema. I.pl. injection of glutamate induced a rapid-onset, dose-related pain response associated with oedema formation, with mean ED(50) values of 2.6 (1.6-4.3) and 0.5 (0.4-0.7) micromol/kg, respectively. Pretreatment with Chicago sky blue 6B (100 microg/kg), an inhibitor of glutamate uptake, caused a significant (about sixfold) reduction of the mean ED(50) value for glutamate-induced nociception, but not paw oedema. NMDA receptor antagonist MK 801, given by systemic (i.p.), intracerebroventricular (i.c.v.), i.pl. or intrathecal (i.t.) routes, produced graded inhibition of glutamate-induced nociception. Non-NMDA receptor antagonists NBQX or GAMS, metabotropic antagonist E4CPG, and also the antagonist that acts at the NMDA receptor-associated glycine binding site felbamate, significantly inhibited the nociception induced by glutamate. L(omega)-N-nitro-arginine (given i.p., i.t., i.pl. or i.c.v.) prevented the nociception and paw oedema caused by glutamate, an effect that was reversed by L-arginine but not by D-arginine. S-nitroso-N-acetyl-D,L-penicillamine (SNAP), given i.pl., greatly potentiated glutamate-induced nociception and oedema formation. Finally, the i.pl. injection of glutamate was accompanied by a graded increase in the nitrite levels of the hindpaw exudate. It is concluded that the nociception caused by i.pl. injection of glutamate probably involves the activation of NMDA and non-NMDA receptors by a mechanism which largely depends on the activation of L-arginine-nitric oxide pathway. Glutamate-induced paw oedema seems to be primarily mediated by non-NMDA ionotropic glutamate receptors and release of nitric oxide.

Topics: Animals; Azo Compounds; Coloring Agents; Dizocilpine Maleate; Dose-Response Relationship, Drug; Edema; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Extremities; Glutamic Acid; Glutamine; Male; Mice; Nitric Oxide; Nitric Oxide Donors; Nitrites; Nitroarginine; Nociceptors; Penicillamine; Quinoxalines; Trypan Blue

Recent reports have suggested that proper maturation of synapses in the hippocampus requires activation of NMDA receptors. We previously demonstrated that neonatal ethanol exposure results in a lasting reduction in synaptic strength in the hippocampus. To determine if this reduction was due to ethanol's effects on NMDA receptors, we investigated long-term changes in synaptic properties resulting from administration of NMDA receptor antagonists to neonatal animals. Rats were injected daily from PND 4-9 with either the noncompetitive NMDA receptor antagonist MK-801, the competitive NMDA receptor antagonist CPP, or the AMPA receptor antagonist NBQX. Control rats were either injected daily with physiological saline during the same period or left to develop normally. Hippocampal slices were prepared from nembutal-anesthetized animals between PND 35 and PND 40. The maximum pEPSP and PS values were not significantly different between controls and NMDA antagonist-treated animals. However, slices from animals injected with NMDA receptor antagonists required higher stimulus currents to attain comparable pEPSPs. The ratio of the slope of the pEPSP to the amplitude of the presynaptic volley was also reduced, as were pEPSP responses to specific stimulus currents. None of these effects were observed in slices prepared from animals treated with the AMPA receptor antagonist NBQX. Glutamate receptor antagonism did not produce lasting changes in long-term potentiation or paired-pulse facilitation. These results indicate activation of NMDA receptors during development is necessary for proper development of synapses.

Topics: Animals; Behavior, Animal; Cell Differentiation; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Female; Hippocampus; Long-Term Potentiation; Neuronal Plasticity; Neurons; Organ Culture Techniques; Piperazines; Pregnancy; Quinoxalines; Rats; Receptors, N-Methyl-D-Aspartate; Synapses; Synaptic Transmission

NMDA glutamate receptor antagonists are used in clinical anesthesia, and are being developed as therapeutic agents for preventing neurodegeneration in stroke, epilepsy, and brain trauma. However, the ability of these agents to produce neurotoxicity in adult rats and psychosis in adult humans compromises their clinical usefulness. In addition, an NMDA receptor hypofunction (NRHypo) state might play a role in neurodegenerative and psychotic disorders, like Alzheimer's disease and schizophrenia. Thus, understanding the mechanism underlying NRHypo-induced neurotoxicity and psychosis could have significant clinically relevant benefits. NRHypo neurotoxicity can be prevented by several classes of agents (e.g. antimuscarinics, non-NMDA glutamate antagonists, and alpha(2) adrenergic agonists) suggesting that the mechanism of neurotoxicity is complex. In the present study a series of experiments was undertaken to more definitively define the receptors and complex neural circuitry underlying NRHypo neurotoxicity. Injection of either the muscarinic antagonist scopolamine or the non-NMDA antagonist NBQX directly into the cortex prevented NRHypo neurotoxicity. Clonidine, an alpha(2) adrenergic agonist, protected against the neurotoxicity when injected into the basal forebrain. The combined injection of muscarinic and non-NMDA Glu agonists reproduced the neurotoxic reaction. Based on these and other results, we conclude that the mechanism is indirect, and involves a complex network disturbance, whereby blockade of NMDA receptors on inhibitory neurons in multiple subcortical brain regions, disinhibits glutamatergic and cholinergic projections to the cerebral cortex. Simultaneous excitotoxic stimulation of muscarinic (m(3)) and glutamate (AMPA/kainate) receptors on cerebrocortical neurons appears to be the proximal mechanism by which the neurotoxic and psychotomimetic effects of NRHypo are mediated.

Topics: Adrenergic alpha-Agonists; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Carbachol; Carbazoles; Cerebral Cortex; Clonidine; Dizocilpine Maleate; Drug Interactions; Excitatory Amino Acid Antagonists; Female; Kainic Acid; Models, Neurological; Muscarinic Antagonists; Nerve Tissue Proteins; Neurons; Neuroprotective Agents; Phenazocine; Prosencephalon; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Glutamate; Receptors, Muscarinic; Receptors, N-Methyl-D-Aspartate; Receptors, sigma; Scopolamine

Glutamate release in ischaemia triggers neuronal death. The major glial glutamate transporter, GLT-1, might protect against glutamate-evoked death by removing extracellular glutamate, or contribute to death by reversing and releasing glutamate. Previous studies of the role of GLT-1 in ischaemia have often used the GLT-1 blocker dihydrokainate at concentrations that affect transporters other than GLT-1 and which affect kainate, N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors. In hippocampal slices from postnatal day 14 mice lacking GLT-1, the current response of area CA1 pyramidal cells to superfused AMPA and NMDA (which are not taken up) was unaffected, whereas the response to 100 microm glutamate was more than doubled relative to that in wild-type littermates, a finding consistent with a decrease in glutamate uptake. In response to a few minutes of simulated ischaemia, pyramidal cells in wild-type mice showed a large and sudden inward glutamate-evoked current [the anoxic depolarization (AD) current], which declined to a less inward plateau. In mice lacking GLT-1, the time to the occurrence of the AD current, its amplitude, the size of the subsequent plateau current and the block of the plateau current by glutamate receptor blockers were all indistinguishable from those in wild-type mice. We conclude that GLT-1 does not contribute significantly to glutamate release or glutamate removal from the extracellular space in early simulated ischaemia. These data are consistent with glutamate release being by reversal of neuronal transporters, and with uptake into glia being compromised by the ischaemia-evoked fall in the level of ATP needed to convert glutamate into glutamine.

Topics: 2-Amino-5-phosphonovalerate; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Excitatory Amino Acid Transporter 2; Glutamic Acid; Hippocampus; Hypoxia-Ischemia, Brain; Kainic Acid; Membrane Potentials; Mice; Mice, Inbred C57BL; Mice, Knockout; N-Methylaspartate; Organ Culture Techniques; Patch-Clamp Techniques; Pyramidal Cells; Quinoxalines; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate

Polychlorinated biphenyls (PCBs) are widespread persistent environmental contaminants that display a complex spectrum of toxicological properties, including neurotoxicity. The present study investigates the effects of the PCB mixtures Aroclor 1242 (A1242) and Aroclor 1254 (A1254), and the PCB congeners 126 (3,3',4,4',5,-PeCB) and 153 (2,2',4,4',5,5'-HxCB) on formation of reactive oxygen species (ROS) and cell death in cultured rat cerebellar granule cells. The increase of ROS and induction of cell death were assayed using the fluorescent probe 2,7-dichlorofluorescin diacetate (DCFH-DA) and the trypan blue exclusion assay, respectively. A1242 and A1254 and PCB 153 induced a concentration-dependent increase in cell death and ROS formation. A1254 was selected for mechanistic studies. When the cerebellar granule cells were exposed to 15 microM A1254 for 12 h, 95% of the cells died. Both PCB-mediated cell death and the increase of the ROS formation were inhibited by MK-801, demonstrating the importance of the N-methyl-D-aspartate receptor. Inhibitors of nitric oxide synthase and phospholipase A2 led to a significant reduction of the DCF fluorescence and cell death. The mitochondrial permeability transition pore blocker cyclosporin A and the antioxidant vitamin E also increased survival and reduced ROS formation. The results show a connection between cell death and free radical formation.

Topics: Animals; Cell Death; Cells, Cultured; Cerebellum; Chlorodiphenyl (54% Chlorine); Cyclosporine; Cytoplasmic Granules; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Fatty Acids, Unsaturated; Male; Neuroprotective Agents; NG-Nitroarginine Methyl Ester; Phosphodiesterase Inhibitors; Quinoxalines; Rats; Rats, Wistar; Reactive Oxygen Species; Receptors, N-Methyl-D-Aspartate; Signal Transduction

The vulnerability of cultured cortical neurons to oxidative injury is an inverse function of the extracellular Mg2+ concentration. In order to test the hypothesis that depolarization-enhanced release of reduced glutathione (GSH) contributes to this phenomenon, we assessed the effect of Mg2+ deprivation on cellular and medium glutathione levels. Incubation of mixed neuronal and glial cultures in Mg2+-free medium resulted in a decline in cellular total glutathione (GSx) within 8 h, without change in oxidized glutathione (GSSG); no effect was seen in pure glial cultures. This decrease in cellular GSx was associated with a progressive increase in GSx but not GSSG in the culture medium. Cellular GSH loss was not attenuated by concomitant treatment with antioxidants (ascorbate, Trolox, or deferoxamine), but was prevented by the NMDA receptor antagonist MK-801. Mg2+ deprivation for over 24 h produced neuronal but not glial death, with release of about 40% of neuronal lactate dehydrogenase by 48-60 h. Most of this cytotoxicity was prevented by treatment with either antioxidants or MK-801. These results suggest that Mg2+ deprivation causes release of neuronal reduced glutathione via a mechanism involving excessive NMDA receptor activation. If prolonged, cellular GSH depletion ensues, leading to oxidative neuronal death.

Topics: Animals; Cell Death; Cerebral Cortex; Coculture Techniques; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Free Radicals; Glutathione; Lipid Peroxidation; Magnesium; Mice; Neuroglia; Neurons; Neurotoxins; Oxidative Stress; Quinoxalines; Receptors, N-Methyl-D-Aspartate

Experiments were carried out in 5-day-old male and female rats in order to assess the respective roles of distinct gamma-aminobutyric acid (GABA) receptor subtypes in the control of growth hormone (GH) secretion in the newborn rat and to identify the potential sexual dimorphism of GABA actions on GH release. The interplay of GABA and excitatory acids (EAAs) to control GH secretion was also analysed. Effects of specific GABA(A), GABA(B) and GABA(C) agonists upon GH secretion as well as of co-administration of GABA and agonists and antagonists of EAA receptors were monitored in 5-day-old male rats. The data confirm that GABA stimulates GH secretion in 5-day-old rats, but indicate that the effect is sexually dimorphic, being greater in females. Stimulation of GH secretion by GABA is mainly mediated by GABA(A) receptors, while GABA(C) appears to be ineffective. Stimulatory effects of GH by GABA and muscimol, a GABA(A) receptor agonist, are blocked by pretreatment with (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclophepten-5-10-imine hydrogen maleate (MK-801), an antagonist of N-methyl-D-aspartate (NMDA) receptors, but not by 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo-benzo[f]-quinoxaline-7-sulfonamide (NBQX), an antagonist of (+/-)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors. Simultaneous administration of GABA and NMDA was additive on GH release while basal GH secretion was not affected by either bicuculline or the respective blockers of GABA(A) and GABA(B) receptors. In conclusion, our results suggest that cross-talk between inhibitory (GABA) and EAAs is involved in the control of GH secretion in neonates.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Animals, Newborn; Baclofen; Bicuculline; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Female; GABA Agonists; GABA Antagonists; gamma-Aminobutyric Acid; Glutamic Acid; Growth Hormone; Growth Hormone-Releasing Hormone; Male; Muscimol; N-Methylaspartate; Neurosecretory Systems; Quinoxalines; Rats; Rats, Wistar; Receptors, GABA; Receptors, N-Methyl-D-Aspartate; Sex Characteristics

The potential toxic effects of the metabotropic glutamate receptor agonist (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (ACPD) and its interactions with the N-methyl-D-aspartate (NMDA) receptor were studied in hippocampal brain slice cultures, using densitometric measurements of the cellular uptake of propidium iodide (PI) to quantify neuronal degeneration. Cultures exposed to ACPD, showed a concentration (2-5 mM) and time (1-4 days) dependent increase in PI uptake in CA1, CA3 and dentate subfields after 24 h and 48 h of exposure, with CA1 pyramidal cells being most sensitive. The neurodegeneration induced by 2 mM ACPD was completely abolished by addition of 10 microM of the NMDA receptor antagonist (5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine (MK-801), while 20 microM of the 2-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA)/kainic acid receptor antagonist 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide (NBQX) had no effect. Co-exposing cultures to a subtoxic dose of 300 microM ACPD together with 10 microM NMDA, which at this dose is known to induce a fairly selective degeneration of CA1 pyramidal cells, significantly increased the PI uptake in both CA1 and CA3, compared to cultures exposed to 10 microM NMDA only. Adding the 300 microM ACPD as pretreatment for 30 min followed by a 30 min wash in normal medium before the ACPD/NMDA co-exposure, eliminated the potentiation of NMDA toxicity. The potentiation was also blocked by addition of 10 or 100 microM 2-methyl-6-(phenylethynyl)pyridine (MPEP) (mGluR5 antagonist) during the co-exposure, while a corresponding addition of 10 or 100 microM 7-(hydroxyimino)cyclopropa[b]chromen-1a-carboxylate ethyl ester (CPCCOEt) (mGluR1 antagonist) had no effect. We conclude that, stimulation of metabotropic glutamate receptors with ACPD at concentrations of 2 mM or higher induces a distinct subfield-related and time and concentration dependent pattern of hippocampal degeneration, and that ACPD at subtoxic concentrations modulates NMDA-induced excitotoxicity through the mGluR5 receptor in a time dependent way.

Topics: Animals; Coloring Agents; Cycloleucine; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Hippocampus; Immunohistochemistry; In Vitro Techniques; Microtubule-Associated Proteins; N-Methylaspartate; Nerve Tissue Proteins; Neurotoxins; Propidium; Quinoxalines; Rats; Rats, Wistar; Receptors, Metabotropic Glutamate; Receptors, N-Methyl-D-Aspartate; Staining and Labeling

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

The effects of three glutamate receptor antagonists, (5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]-cyclohepten-5,10-imine hydrogen maleate (MK-801) for the N-methyl-D-aspartate receptor, 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo-benzo[f] quinoxaline-7-sulfonamide (NBQX) for the alpha-amino-3-hydroxy-5methyl-4-isoxazole propionate /kinate receptor and (S)-alpha-methyl-4-carboxyphenylglycine (MCPG) for the metabotropic receptor, on c-fos and c-jun mRNA expression were investigated in cultured cortical glial cells following traumatic scratch injury. Expression of the two genes along the edges of wounds detected by in situ hybridization was not affected by MK-801 and NBQX. However, 100 and 500 microM of MCPG remarkably reduced the hybridization signals for both c-fos and c-jun mRNAs. The present results suggest that group I metabotropic glutamate receptors might have some association with immediate early gene induction after in vitro traumatic injury in glial cells.

Topics: Animals; Benzoates; Brain Injuries; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Fetus; Gene Expression Regulation; Genes, Immediate-Early; Gliosis; Glycine; Nerve Regeneration; Neuroglia; Proto-Oncogene Proteins c-fos; Proto-Oncogene Proteins c-jun; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Metabotropic Glutamate; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Transcriptional Activation

The effect of L-glutamate (Glu) on human lymphocyte function was studied by measuring anti-CD(3) monoclonal antibody (mAb) or phytohaemagglutinin (PHA)-induced intracellular Ca(2+) ([Ca(2+)](i)) rise (Fura-2 method), and cell proliferation (MTT assay). Glu (0.001 - 100 microM) did not modify basal lymphocyte [Ca(2+)](i), but significantly potentiated the effects of anti-CD(3) mAb or PHA. Maximal [Ca(2+)](i) rises over resting cells were: 165+/-8 and 247+/-10 nM at 3.0x10(-2) mg ml(-1) anti-CD(3) mAb; 201+/-4 and 266+/-9 nM at 5.0x10(-2) mg ml(-1) PHA, in the absence or presence of 1 microM Glu, respectively. The Glu effect showed a bell-shape concentration-dependent relationship, with a maximum (+90+/-3% for anti-CD(3) mAb and +57+/-2% for PHA over Glu-untreated cells) at 1 microM. Non-NMDA receptor agonists (1 microM) showed a greater efficacy (+76+/-2% for (S)-AMPA; +78+/-4% for KA), if compared to NMDA (+46+/-2%), or Glu itself. Ionotropic Glu receptor antagonists completely inhibited the effects of the corresponding specific receptor agonists (1 microM). The IC(50) values calculated were: 0.9 microM for D-AP5; 0.6 microM for (+)-MK801; 0.3 microM for NBQX. Both NBQX and KYNA were able to abolish Glu effect. The IC(50s) calculated were: 3.4 microM for NBQX; 0.4 microM for KYNA. Glu (0.1 - 1 mM) did not change the resting cell proliferation, whereas Glu (1 mM) significant inhibited (-27+/-4%) PHA (1.0x10(-2) mg ml(-1))-induced lymphocyte proliferation at 72 h. In conclusion, human lymphocytes express ionotropic Glu receptors functionally operating as modulators of cell activation.

Topics: 2-Amino-5-phosphonovalerate; Adult; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Antibodies, Monoclonal; Calcium; CD3 Complex; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Synergism; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Glutamic Acid; Humans; Kainic Acid; Lymphocytes; N-Methylaspartate; Phytohemagglutinins; Quinoxalines; Receptors, Glutamate

This study investigated the influence of temperature or glutamate antagonism on the immediate outcome of perinatal asphyxia. Perinatal asphyxia was produced by water immersion of fetus-containing uterus horns removed by cesarean section from ready to deliver rats. The uterus horns were kept in a water bath for different time periods, before the pups were delivered and stimulated to breathe. After delivery, the pups were assessed for behavior and for systemic glutamate, aspartate, lactate and pyruvate levels measured with in vivo microdialysis, or ex vivo for energy-rich phosphates, including adenosine triphosphate (ATP), in brain, heart and kidney. In a series of experiments, asphyxia was initiated in a water bath at 37 degrees C, before the pup-containing uterus horns were moved for different time intervals to a 15 degrees C bath. In another series of experiments, the mothers were treated with N-methyl-D-aspartate (NMDA) antagonist, dizocilpine (MK-801), or alpha-amino-3-hydroxy-methylisoxazole-4-propionic acid (AMPA) antagonist,2,3-dihydroxy-6-nitro-7-sulfamoyl benzo(f) quinoxalin NBQX) 1 h before hysterectomy and asphyxia at 37 degrees C. The rate of survival rapidly decreased following exposure to more than 16 min of asphyxia, and no survival could be observed after 22 min of asphyxia. An LD50 was estimated to occur at approximately 19 min of asphyxia. The outcome was paralleled by a decrease in ATP in kidney, followed by a decrease in heart and brain. A maximal decrease in ATP was observed after 20 min of asphyxia in all tissues. Systemic microdialysis revealed that glutamate, aspartate and pyruvate levels were increased with a peak after 5 min of asphyxia. In contrast, lactate levels increased along with the length of the insult. Survival was increased when the pup-containing uterus horns were moved from a 37 degrees C to a 15 degrees C bath, at 15 min of asphyxia (the LD50 was thus increased to 30 min). If the shift occurred at 10 or 5 min of asphyxia, the LD50 increased to 80 or 110 min, respectively. The effect of glutamate antagonism was minor compared to hypothermia; the best effect (an increase in the LD50 to approximately 22 min) was observed after combining AMPA and NMDA antagonists.

Topics: Adenosine Triphosphate; Animals; Animals, Newborn; Aspartic Acid; Asphyxia Neonatorum; Behavior, Animal; Brain; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Heart; Humans; Hypothermia, Induced; Hypoxia-Ischemia, Brain; Infant, Newborn; Kidney; Lactic Acid; Maternal Behavior; Microdialysis; Pyruvic Acid; Quinoxalines; Rats; Receptors, Glutamate; Survival Rate; Treatment Outcome

The excitotoxic profiles of (RS)-2-amino-3-(3-hydroxy-5-tert-butylisoxazol-4-yl)propionic acid (ATPA), (RS)-2-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA), kainic acid (KA) and N-methyl-D-aspartate (NMDA) were evaluated using cellular uptake of propidium iodide (PI) as a measure for induced, concentration-dependent neuronal damage in hippocampal slice cultures. ATPA is in low concentrations a new selective agonist of the glutamate receptor subunit GluR5 confined to KA receptors and also in high concentrations an AMPA receptor agonist. The following rank order of estimated EC(50) values was found after 2 days of exposure: AMPA (3.7 mM)>NMDA (11 mM)=KA (13 mM)>ATPA (33 mM). Exposed to 30 microM ATPA, 3 microM AMPA and 10 microM NMDA, CA1 was the most susceptible subfield followed by fascia dentata and CA3. Using 8 microM KA, CA3 was the most susceptible subfield, followed by fascia dentata and CA1. In 100 microM concentrations, all four agonists induced the same, maximal PI uptake in all hippocampal subfields, corresponding to total neuronal degeneration. Using glutamate receptor antagonists, like GYKI 52466, NBQX and MK-801, inhibition data revealed that AMPA excitotoxicity was mediated primarily via AMPA receptors. Similar results were found for a high concentration of ATPA (30 microM). In low GluR5 selective concentrations (0.3-3 microM), ATPA did not induce an increase in PI uptake or a reduction in glutamic acid decarboxylase (GAD) activity of hippocampal interneurons. For KA, the excitotoxicity appeared to be mediated via both KA and AMPA receptors. NMDA receptors were not involved in AMPA-, ATPA- and KA-induced excitotoxicity, nor did NMDA-induced excitotoxicity require activation of AMPA and KA receptors. We conclude that hippocampal slice cultures constitute a feasible test system for evaluation of excitotoxic effects and mechanisms of new (ATPA) and classic (AMPA, KA and NMDA) glutamate receptor agonists. Comparison of concentration-response curves with calculation of EC(50) values for glutamate receptor agonists are possible, as well as comparison of inhibition data for glutamate receptor antagonists. The observation that the slice cultures respond with more in vivo-like patterns of excitotoxicity than primary neuronal cultures, suggests that slice cultures are the best model of choice for a number of glutamate agonist and antagonist studies.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Anti-Anxiety Agents; Benzodiazepines; Dizocilpine Maleate; Dose-Response Relationship, Drug; Hippocampus; In Vitro Techniques; Isoxazoles; Kainic Acid; Microtubule-Associated Proteins; N-Methylaspartate; Neuroprotective Agents; Neurotoxins; Nissl Bodies; Nuclear Proteins; Propidium; Propionates; Quinoxalines; Rats; Rats, Wistar

Previous studies have shown that identified neurons of the nucleus of the solitary tract (NST) are excited by the cytokine tumor necrosis factor-alpha (TNF-alpha). Vagal afferent connections with the NST are predominantly glutaminergic. Therefore, we hypothesized that TNF-alpha effects on NST neurons may be via modulation of glutamate neurotransmission. The present study used activation of the immediate early gene product c-Fos as a marker for neuronal activation in the NST. c-Fos expression was evaluated after microinjections of TNF-alpha in the presence or absence of either the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor antagonist 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo-benzo[f]quinoxaline-7-sulfonamide disodium (NBQX) or the N-methyl-D- aspartate (NMDA) antagonist MK-801. To assess the specificity of the interaction between TNF-alpha and glutamate, c-Fos expression was also evaluated after injection of oxytocin (OT) (which has a direct excitatory effect in this area of the brain stem) in the presence and absence of NBQX or MK-801. c-Fos labeling was significantly increased in the NST after TNF-alpha exposure. Coinjection of either NBQX or MK-801 with TNF-alpha prevented significant c-Fos induction in the NST. Microinjections of OT also induced significant NST c-Fos elevation, but this expression was unaffected by coinjection of either antagonist with OT. These data lead us to conclude that TNF-alpha activation of NST neurons depends on glutamate and such an interaction is not generalized to all agonists that act on the NST.

Topics: Animals; Dizocilpine Maleate; Drug Interactions; Excitatory Amino Acid Antagonists; Male; Microinjections; Neurons; Oxytocin; Proto-Oncogene Proteins c-fos; Quinoxalines; Rats; Rats, Long-Evans; Receptors, AMPA; Solitary Nucleus; Tumor Necrosis Factor-alpha

We previously reported that intra-4th-ventricular (i.4th.v.) administration of a non-NMDA receptor antagonist, NBQX, abolished vagally induced retching. This study was undertaken to ascertain whether or not the neuronal response in the solitary tract nucleus (NTS) to vagal stimulation and the vago-vagal gastric reflexes induced by non-emetic stimulation are also abolished by NBQX with a similar latency as in the case of retching. Ketamine and thiopental- or chloralose-anesthetized dogs were decerebrated, and the dorsal surface of the medulla was exposed. This study consisted of two series of experiments. In the first series, extracellular neuronal responses in the NTS to pulse-train vagal stimulation were recorded. Effects of NBQX on the neural response and vagally induced fictive retching were observed. In the second series, effects of glutamate receptor antagonists on gastric corpus responses to esophageal or gastric antral distension were observed. Retching was abolished 5-15 min after an i.4th.v. application of NBQX. and the neuronal responses disappeared within 14 min after application in nine of 10 NTS neurons. On the other hand, corpus contractility was inhibited by esophageal distension, and inhibited and/or enhanced by antral distension. While the inhibitory responses disappeared within 17 min after NBQX, the enhanced response remained even after NBQX and vagotomy, but was abolished by i.v. administration of hexamethonium. These results suggest that adaptive relaxation in the corpus, as well as retching, may be mediated by glutaminergic vagal afferents and non-NMDA receptors in the NTS.

Topics: Action Potentials; Animals; Catheterization; Dizocilpine Maleate; Dogs; Esophagus; Excitatory Amino Acid Antagonists; Ganglionic Blockers; Glutamic Acid; Hexamethonium; Neurons, Afferent; Quinoxalines; Reflex; Solitary Nucleus; Stomach; Vagotomy; Vagus Nerve; Vomiting

Excitotoxicity is implicated in the pathogenesis of several neurologic diseases, such as chronic neurodegenerative diseases and stroke. Recently, it was reported that excitotoxicity has a relationship to apoptotic neuronal death, and that the mitochondrial toxin, 3-nitropropionic acid (3-NP), could induce apoptosis in the striatum. Although striatal lesions produced by 3-NP could develop through an excitotoxic mechanism, the exact relationship between apoptosis induction and excitotoxicity after 3-NP treatment is still not clear. The authors investigated the role of excitotoxicity and oxidative stress on apoptosis induction within the striatum after intraperitoneal injection of 3-NP. The authors demonstrated that removal of the corticostriatal glutamate pathway reduced superoxide production and apoptosis induction in the denervated striatum of decorticated mice after 3-NP treatment. Also, the N-methyl-D-aspartate (NMDA) receptor antagonist, MK-801, prevented apoptosis in the striatum after 3-NP treatment for 5 days, whereas the non-NMDA receptor antagonist, 2,3-dihydroxy-6-nitro-7-sulphamoyl-benzo(F)quinoxaline, was ineffective. The authors also evaluated the initial type of neuronal death by 3-NP treatment for different durations from 1 to 5 days. In early striatal damage, apoptotic neuronal death initially occurred after 3-NP treatment. Our data show that excitotoxicity related to oxidative stress initially induces apoptotic neuronal death in mouse striatum after treatment with 3-NP.

Topics: Animals; Apoptosis; Caspases; Corpus Striatum; Decerebrate State; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; In Situ Nick-End Labeling; Mice; Mice, Inbred C57BL; Mitochondria; Nervous System; Neurotoxins; Nitro Compounds; Oxidative Stress; Propionates; Quinoxalines; Receptors, N-Methyl-D-Aspartate; Superoxides

The neuroprotective efficacy of antioxidant molecules against iodoacetate (IAA) neurotoxicity in rat cerebellar granule cell (CGC) cultures was investigated. Transient exposure to IAA caused a concentration-dependent decrease in cell viability (ED50 = 9.8 microM). Dizocilpine maleate (MK-801), and 1,2,3,4-tetrahydro-6-nitro-2,3-dioxobenzo[f]quinoxaline-7-sulfonamide (NBQX), failed to prevent IAA toxicity. Certain antioxidant molecules were shown to be neuroprotective against IAA when combined with MK-801 but were ineffective when administered alone. (S)-(-)-Trolox, butylated hydroxytoluene (BHT), and U-83836E exhibited EC50 values of 78, 5.9, and 0.25 microM, respectively, in the presence of 10 microM MK-801. IAA also induced an increase in intracellular oxidative stress, which was quenched by the antioxidants (in the presence of MK-801) in cultures loaded with the oxidant sensitive dye 2'7'-dichlorodihydrofluorescein diacetate (DCFH-DA).

Topics: Animals; Antioxidants; Butylated Hydroxytoluene; Cells, Cultured; Cerebellar Cortex; Chromans; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Synergism; Fluoresceins; Fluorescent Dyes; Iodoacetates; Neuroprotective Agents; Neurotoxins; Oxidative Stress; Piperazines; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

Bilateral intracerebroventricular infusion of dl-homocysteic acid (DL-HCA) (600 nmol on each side) to immature 12-day-old rats induced generalized clonic-tonic seizures, recurring frequently for at least 90 min, with a high rate of survival. Electrographic recordings from sensorimotor cortex, hippocampus, and striatum demonstrated isolated spikes in the hippocampus and/or striatum as the first sign of dl-HCA action. Generalization of epileptic activity occurred during generalized clonic-tonic seizures, but electroclinical correlation was very low; dissociation between EEG pattern and motor phenomena was common. Seizures were accompanied by large decreases of cortical glucose and glycogen and by approximately 7- to 10-fold accumulation of lactate. ATP and phosphocreatine (PCr) levels remained unchanged even during longlasting (3 h) convulsions. Metabolite levels became normalized during the recovery period (24 h). The examination of the effect of selected antagonists of NMDA [AP7 (18.5 and 37 mg/kg, respectively), MK-801 (0.5 mg/kg)] and non-NMDA [NBQX (10, 15 and 30 mg/kg, respectively)] receptors revealed that seizures could be attenuated or prevented (depending on the dose employed) by antagonists of both NMDA and non-NMDA receptors, as evaluated not only according to the suppression of behavioral manifestations of seizures, but also in terms of the protection of metabolite changes accompanying seizures. All antagonists employed, when given alone in the same doses as those used for seizure protection, did not influence metabolite levels, with the exception of increased glucose concentrations. Furthermore, the pronounced anticonvulsant effect could be achieved by the combined treatment with low subthreshold doses of NMDA (AP7) and non-NMDA (NBQX) receptor antagonists, which may be of potential significance for a new approach to the treatment of epilepsy.

Topics: 2-Amino-5-phosphonovalerate; Age Factors; Animals; Cerebral Cortex; Disease Models, Animal; Dizocilpine Maleate; Electroencephalography; Energy Metabolism; Epilepsy; Excitatory Amino Acid Antagonists; Glucose; Homocysteine; Injections, Intraventricular; Male; Neuroprotective Agents; Quinoxalines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Seizures

Despite its importance in the cerebellum, the functions of the orphan glutamate receptor delta2 are unknown. We examined a mutant delta2 receptor channel in lurcher mice that was constitutively active in the absence of ligand. Because this mutation was within a highly conserved motif (YTANLAAF), we tested its effect on several glutamate receptors. Mutant delta2 receptors showed distinct channel properties, including double rectification of the current-voltage relationship, sensitivity to a polyamine antagonist and moderate Ca 2+ permeability, whereas other constitutively active mutant glutamate channels resembled wild-type channels in these respects. Moreover, the kinetics of ligand-activated currents were strikingly altered. We conclude that the delta2 receptor has a functional ion channel pore similar to that of glutamate receptors. The motif may have a role in the channel gating of glutamate receptors.

Topics: Amino Acid Sequence; Amino Acid Substitution; Animals; Anti-Anxiety Agents; Antihypertensive Agents; Benzodiazepines; Benzothiadiazines; Cell Line; Conserved Sequence; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Glutamic Acid; Humans; Ion Channel Gating; Kainic Acid; Kidney; Mice; Mice, Neurologic Mutants; Molecular Sequence Data; Mutagenesis; Neuromuscular Depolarizing Agents; Patch-Clamp Techniques; Purkinje Cells; Quinoxalines; Receptors, Glutamate; Transfection

NMDA or AMPA/kainate receptor antagonists have been shown to provide neuroprotection following in vitro spinal cord injury, but the mechanisms by which these agents improve behavioral recovery and protect axonal function remains unclear. We hypothesized that treatment of spinal cord injury with these drugs would attenuate glutamate excitatory transmission by blocking the effects of glutamate receptors at the injury site or would improve spinal cord blood flow. To test these hypotheses, we observed the effects of locally administered MK-801 (30 nmol) or NBQX (5 or 15 nmol) into the injured spinal cord on axonal conduction and post-traumatic ischemia of the cord. The outcome measures were multimodality evoked potentials and blood flow in an acute compression injury model in rats. We found that locally administered MK-801 or NBQX 15 min after spinal cord injury attenuated the amplitude, delayed the latency of sensory evoked potentials and increased the sensory conduction time across the injury site, but did not improve blood flow during the 4-h period of observation. These results demonstrate that the NMDA and non-NMDA receptor antagonists produced a blockade of glutamate excitatory transmission in the afferent pathways at the injury site. It is suggested that the neuroprotection provided by these agents following spinal cord injury is mediated through blockade of glutamate ionotropic receptors in the injured spinal cord, but is not related to improvement of SCBF.

Topics: Animals; Cerebellum; Dizocilpine Maleate; Evoked Potentials; Evoked Potentials, Motor; Evoked Potentials, Somatosensory; Excitatory Amino Acid Antagonists; Male; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Regional Blood Flow; Spinal Cord; Spinal Cord Injuries

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

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

Although neurons often fire in bursts, most of what is known about glutamate signaling and postsynaptic receptor activation is based on experiments using single stimuli. Here we examine the activation of ionotropic glutamate receptors by bursts at the parallel fiber to stellate cell synapse. We show that brief stimulus trains generate prolonged AMPA receptor (AMPAR)- and NMDA receptor (NMDAR)-mediated EPSCs recorded in whole-cell voltage clamp. These EPSCs contrast with the rapid AMPAR-mediated EPSC evoked by a single stimulus. The prolonged AMPAR-mediated EPSC is promoted by high-frequency and high-intensity trains and can persist for hundreds of milliseconds. This EPSC is also increased by l-trans-2,4-PDC, an inhibitor of glutamate transporters, suggesting that these transporters usually limit the synaptic response to trains. These prolonged EPSCs reflect both receptor properties and a long-lasting glutamate signal. In addition, several experiments demonstrate that glutamate spillover can contribute to receptor activation. First, imaging stimulus-evoked changes in presynaptic calcium establishes that distinct parallel fiber bands can be activated. Second, activation of parallel fibers that do not directly synapse onto a given stellate cell can evoke indirect AMPAR- and NMDAR-mediated EPSCs in that cell. Third, experiments using the use-dependent NMDAR blocker MK-801 show that these indirect EPSCs reflect glutamate spillover in response to trains. Together, these findings indicate that stimulus trains can generate a sustained and widespread glutamate signal that can in turn evoke large and prolonged EPSCs mediated by ionotropic glutamate receptors. These synaptic properties may have important functional consequences for stellate cell firing.

Topics: 2-Amino-5-phosphonovalerate; Animals; Cerebellum; Dicarboxylic Acids; Dizocilpine Maleate; Electric Stimulation; Evoked Potentials; Excitatory Amino Acid Antagonists; Glutamic Acid; In Vitro Techniques; Nerve Fibers; Neurons; Neurotransmitter Uptake Inhibitors; Pyrrolidines; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Synapses; Synaptic Transmission; Time Factors

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

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

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

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

1. The main purpose of the present study was to investigate the effects of the neuroprotective agent riluzole on the electrically evoked release of [(3)H]-glutamate ([(3)H]-Glu) in mouse neocortical slices. The reported selectivity of riluzole for excitatory amino acids was tested in release experiments with further neurotransmitters. Also distinct species, mouse, rat and man were compared. 2. [(3)H]-Glu was formed endogenously during incubation of slices with [(3)H]-glutamine ([(3)H]-Gln). Released [(3)H]-Glu and tissue [(3)H]-Glu was separated by anion exchange chromatography. Electrically evoked [(3)H]-Glu release was strongly diminished by tetrodotoxin (TTX) and Ca(2+)-withdrawal. 3. Riluzole (100 microM) depressed the release of [(3)H]-Glu up to 77% (IC(50)=19.5 microM). Riluzole was also able to inhibit strongly the electrically evoked release of [(3)H]-acetylcholine ([(3)H]-ACh) (at 100 microM by 92%, IC(50)=3.3 microM, and [(3)H]-dopamine ([(3)H]-DA) (at 32 microM by 72%, IC(50)=6.8 microM). However, the release of [(3)H]-serotonin ([(3)H]-5-HT) was less diminished (at 100 microM by 53%, IC(50)=39.8 microM). Riluzole up to 100 microM did not affect [(3)H]-noradrenaline ([(3)H]-NA) release. 4. Between species, i.e. in mouse, rat and human neocortex, no significant differences between the effects of riluzole could be observed. 5. The NMDA-receptor blocker MK-801 (1 microM) and the AMPA/Kainate-receptor blocker NBQX (1 microM) did neither affect the electrically evoked [(3)H]-ACh release nor its inhibition by riluzole, indicating that effects of riluzole on transmitter release were neither due to modulation of ionotropic Glu receptors, nor due to indirect inhibition of Glu release through these receptors. 6. Taken together, riluzole inhibits the release of distinct neurotransmitters differently, but is not selective for the excitatory amino acid Glu.

Topics: Acetylcholine; Animals; Calcium; Dicarboxylic Acids; Dizocilpine Maleate; Dopamine; Dose-Response Relationship, Drug; Electric Stimulation; Excitatory Amino Acid Antagonists; Glutamic Acid; Humans; In Vitro Techniques; Mice; Neostriatum; Neuroprotective Agents; Neurotransmitter Agents; Norepinephrine; Pyrrolidines; Quinoxalines; Rats; Rats, Wistar; Riluzole; Serotonin; Tetrodotoxin; Tritium

Drugs that block spinal excitatory amino acid receptor activation may prevent pain after surgery. The authors previously studied the effect of excitatory amino acid receptor antagonists after incision. In the present study, we examined the role of N-methyl-d-aspartate (NMDA), non-NMDA, and metabotropic glutamate receptors (mGluRs) on the development of pain behavior after plantar incision.. Rats with lumbar intrathecal catheters were anesthetized with halothane. Fifteen minutes before an incision was made, drug [40 nmol MK-801; 20 nmol NBQX; or 200 nmol (+)-MCPG] or vehicle was injected intrathecally followed by an infusion of the same drug for 75 min. Withdrawal thresholds to calibrated von Frey filaments applied adjacent to the wound and response frequencies to a blunt mechanical stimulus applied directly to the wound were measured before incision and 1, 2, 4, and 6 h after incision and then once daily for 6 days.. Preincision treatments with antagonists against the NMDA (MK-801) and group I and II metabotropic receptors [(+)-MCPG] did not inhibit the development of mechanical hyperalgesia caused by incision. Preincision treatment with the non-NMDA receptor antagonist NBQX increased withdrawal thresholds at 1 and 2 h and on postoperative day 1 compared with the vehicle group; response frequencies were reduced 1 and 2 h after incision and on postoperative day 2 (P < 0.05). In an additional group, postincision treatment with NBQX was similar to preincision treatment.. Spinal NMDA and mGluR antagonists may not be useful for preventing postsurgical pain. Spinal non-NMDA receptor antagonists reduced pain behaviors, but a preventive effect using preincision treatment was not apparent.

Topics: Analgesia; Anesthesia, Inhalation; Anesthetics, Inhalation; Animals; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Foot; Halothane; Injections, Spinal; Male; Motor Activity; Pain, Postoperative; Quinoxalines; Rats; Rats, Sprague-Dawley

The role of glutamate receptors in the inferior colliculus (IC) in audiogenic and audiogenic-like seizures was investigated in adult rats with transient neonatal hypothyroidism by 0.02% propylthiouracil (PTU) treatment through mother's milk (PTU rats) and in naive rats treated intracisternally with N-methyl-d-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazole-proprionic acid (AMPA), or cyclothiazide, an inhibitor of rapid AMPA receptor desensitization. All rats showed audiogenic or audiogenic-like seizures characterized by running fit (RF) and generalized tonic-clonic seizures (GTCS). While systemically administered MK-801 inhibited GTCS, intracisternally administered NBQX inhibited RF and GTCS in both audiogenic and audiogenic-like seizures. Auditory stimulation shortened the latency to GTCS induced by AMPA, but not NMDA, at a subclinical dose and further elongated the shortened duration of RF, but not GTCS, induced by MK-801 pretreatment. Furthermore, Northern blot analysis was used to evaluate the expression of the immediate-early gene c-fos in the IC following induction of audiogenic or audiogenic-like seizures. The significant induction of c-fos mRNA by audiogenic seizures in PTU rats or by AMPA- or cyclothiazide-induced seizures in naive rats was prominent in the IC. MK-801 suppressed c-fos mRNA expression in the IC induced by audiogenic seizures in PTU rats or by AMPA-induced seizures in naive rats. NBQX suppressed the expression of c-fos mRNA in the IC induced by AMPA-induced seizures but did not suppress c-fos mRNA in PTU rats or rats with cyclothiazide-induced seizures. Auditory stimuli failed to affect c-fos mRNA induction by AMPA. The present study suggests that audiogenic-like seizures can be reproduced by glutamate receptor agonists in which AMPA receptors are primarily linked to the initiation of audiogenic seizures (RF) while NMDA receptors presumably located within the IC are involved in the propagation of GTCS in audiogenic seizures.

Topics: Acoustic Stimulation; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Animals, Suckling; Benzothiadiazines; Dizocilpine Maleate; Drug Administration Routes; Excitatory Amino Acid Antagonists; Female; Hypothyroidism; Inferior Colliculi; Injections, Intraventricular; Male; Maternal Exposure; N-Methylaspartate; Propylthiouracil; Proto-Oncogene Proteins c-fos; Quinoxalines; Rats; Rats, Sprague-Dawley; Reaction Time; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Seizures

The present study describes the effect of (S)-2,3-dihydro-[3, 4]cyclopentano-1,2,4-benzothiadiazine-1,1-dioxide (S18986-1), a positive allosteric modulator of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors with cognitive-enhancing effects, on (S)-AMPA-induced [3H]noradrenaline release in rat hippocampal and frontal cortex slices. (S)-AMPA significantly increased [3H]noradrenaline release in rat hippocampus and frontal cortex slices, whereas S18986-1 (3-1000 microM) alone, was inactive. However, S18986-1 between 30 and 1000 microM potently enhanced (+200%) (S)-AMPA-mediated [3H]noradrenaline release in both hippocampal and frontal cortex slices. The capacity of S18986-1 to potentiate [3H]noradrenaline release was specific for AMPA receptors as S18986-1 failed to potentiate either kainate and N-methyl-D-aspartate (NMDA)-mediated release of [3H]noradrenaline in rat hippocampal slices. Moreover, 1, 2,3,4-tetrahydro-6-nitro-2,3-dioxo-benzo[f]quinoxaline-7-sulfonamide (NBQX) and 1-(4-aminophenyl)-3-methylcarbamoyl-4-methyl-3, 4-dihydro-7,8-methylenedioxy-5H-2,3-benzodiazepine (GYKI-53655) but not (5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5, 10-imine ((+)-MK-801), inhibited (S)-AMPA and S18986-induced stimulation of (S)-AMPA-mediated [3H]noradrenaline release. In addition, S18986-1-induced stimulation of (S)-AMPA-evoked [3H]noradrenaline release was markedly attenuated in the presence of tetrodotoxin (1 microM) and in Ca(2+)-free buffer. S18986-1 enhanced (S)-AMPA-mediated [3H]noradrenaline release to a greater extent than its corresponding (R)-enantiomer S19024-1 and racemic mixture S17951-1. However, positive allosteric modulators of AMPA receptors such as aniracetam failed to potentiate AMPA-mediated noradrenaline release in hippocampal slices, whereas cyclothiazide potently enhanced (S)-AMPA-mediated [3H]noradrenaline release. These results suggest that the capacity of S18986-1 to enhance AMPA receptor-mediated release of noradrenaline in rat hippocampus and frontal cortex, could contribute to the cognition enhancing mechanisms of S18986-1.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Benzodiazepines; Benzothiadiazines; Calcium; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Synergism; Excitatory Amino Acid Antagonists; Frontal Lobe; Hippocampus; In Vitro Techniques; Male; Norepinephrine; Pyrrolidinones; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Stereoisomerism; Tetrodotoxin; Tritium

The secretion of PRL is controlled by different hypothalamic signals. Depending on the experimental model, PRL secretion increases or decreases after activation of N-methyl-d -aspartic acid and kainate receptors. Recently we have described that activation of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors inhibits PRL secretion in prepubertal male rats. The aim of present study was to examine (1) the physiological relevance of this finding, (2) the possible age-related changes observed after activation or blockade of AMPA receptors, (3) the specificity of the AMPA effect, (4) the hypothalamic and/or pituitary localization of AMPA action, and (5) the mechanism(s) of action of AMPA agonists. In a first set of experiments, neonatal males (5 and 10 days old) and prepubertal (23 days old) male rats were injected with AMPA (1, 2.5 or 5 mg/kg) or the antagonist of AMPA receptors 1,2,3, 4-tetrahydro-6-nitro-2,3-dioxo-! benzo (f) quinoxaline-7-sulfonamide (NBQX; 0.25 or 0.50 mg/kg). Serum PRL concentrations decreased significantly 15 and 30 min after i.p. administration of AMPA in prepubertal male rats, while the inhibitory effect of AMPA was not observed in 5- and 10-day-old males. The effect of AMPA was abolished by NBQX but not by MK-801 (a selective antagonist of NMDA receptors). NBQX alone (0.25 or 0.50 mg/kg) had no effect on PRL release. In vitro, AMPA slightly stimulated PRL secretion by hemipituitaries from prepubertal males, suggesting that the hypothalamus is likely the site of action for the reported inhibitory action of AMPA on PRL release. In this sense, the blockade of AMPA effects in animals pretreated with domperidone (a dopaminergic antagonist) or alpha-methyl-p-tyrosine (an inhibitor of dopamine synthesis) suggests that an increase in the release of hypothalamic dopamine is probably the mechanism i! nvolved in the effect of AMPA. In a second set of experiments, the effects of AMPA (2.5 mg/kg i.p.) and NBQX (0.5 mg/kg i.p. and 20 or 40 nmol i.c.v.) were tested in freely moving adult male rats sampled during periods of 2, 3 or 6 h. In contrast with data obtained in prepubertal rats, neither AMPA nor NBQX affected PRL secretion. In conclusion, these data indicate that activation of AMPA receptors inhibits PRL secretion in prepubertal male rats. This effect probably involves the release of dopamine from the hypothalamus and disappears in adulthood.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Analysis of Variance; Animals; Animals, Newborn; Depression, Chemical; Dizocilpine Maleate; Male; Pituitary Gland; Prolactin; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Sexual Maturation

We previously created a transgenic mouse model of comorbid Tourette's syndrome and obsessive-compulsive disorder (TS+OCD), by expressing a neuropotentiating cholera toxin (CT) transgene in a subset of dopamine D1 receptor-expressing (D1+) neurons thought to induce cortical and amygdalar glutamate output. To test glutamate's role in the TS+OCD-like disorder of these transgenic mice (D1CT-7 line), the effects of glutamate receptor-binding drugs on their behavior were examined. MK-801, a non-competitive NMDA receptor antagonist that indirectly stimulates cortical-limbic glutamate output, aggravated a transgene-dependent abnormal behavior (repetitive climbing and leaping) in the D1CT-7 mice at doses insufficient to induce stereotypies, and more readily induced stereotypies and limbic seizure behaviors at high doses. NBQX, a seizure-inhibiting AMPA receptor antagonist, reduced only the MK-801-dependent stereotypic and limbic seizure behavior of D1CT-7 mice, but not their transgene-dependent behaviors. These data imply that TS+OCD-like behavior is mediated by cortical-limbic glutamate, but that AMPA glutamate receptors are not an essential part of this behavioral circuit. Our findings lead to the prediction that the symptoms of human Tourette's syndrome and obsessive-compulsive disorder are elicited by excessive forebrain glutamate output.

Topics: Animals; Cerebral Cortex; Comorbidity; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Humans; Limbic System; Male; Mice; Mice, Inbred BALB C; Mice, Transgenic; Motor Activity; Obsessive-Compulsive Disorder; Quinoxalines; Seizures; Tourette Syndrome

Glutamate promotes neuronal survival during brain development and destroys neurons after injuries in the mature brain. Glutamate antagonists are in human clinical trials aiming to demonstrate limitation of neuronal injury after head trauma, which consists of both rapid and slowly progressing neurodegeneration. Furthermore, glutamate antagonists are considered for neuroprotection in chronic neurodegenerative disorders with slowly progressing cell death only. Therefore, humans suffering from Huntington's disease, characterized by slowly progressing neurodegeneration of the basal ganglia, are subjected to trials with glutamate antagonists. Here we demonstrate that progressive neurodegeneration in the basal ganglia induced by the mitochondrial toxin 3-nitropropionate or in the hippocampus by traumatic brain injury is enhanced by N-methyl-d-aspartate antagonists but ameliorated by alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate antagonists. These observations reveal that N-methyl-d-aspartate antagonists may increase neurodestruction in mature brain undergoing slowly progressing neurodegeneration, whereas blockade of the action of glutamate at alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate receptors may be neuroprotective.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Brain Injuries; Cell Death; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Memantine; N-Methylaspartate; Neurons; Neuroprotective Agents; Neurotoxins; Nitro Compounds; Piperazines; Propionates; Quinoxalines; Rats; Rats, Wistar; Wounds and Injuries

Increased amounts of bilirubin, the end product of heme degradation, are known to be detrimental to the central nervous system, especially in preterm newborns. In an attempt to delineate the cellular mechanisms by which unconjugated bilirubin exerts its toxic effects on neuronal cells in the developing brain, bilirubin (0.25-5 microM) was added to the extracellular medium of 6-day-old primary cultured neurons from the embryonic rat forebrain, and cell alterations were studied over the ensuing 96 h. Bilirubin decreased cell viability dose dependently with an ED(50) around 1 microM. At the dose of 0.5 microM, it triggered delayed cell death that affected 24% of the neurons. Nuclear incorporation of the fluorescent dye DAPI (4,6-diamidino-2-phenylindole) depicted the presence of apoptosis (16%). Apoptosis features were confirmed by DNA fragmentation reflected by a progressive loss of [(3)H]thymidine and sequential changes in macromolecular synthesis, as shown by the time course of [(3)H]leucine incorporation, as well as by the beneficial effects of cycloheximide and caspase inhibitors. In parallel, treatments with glutamate receptor antagonists showed that MK-801, but not NBQX, protected neurons against bilirubin neurotoxicity, suggesting a role for NMDA receptors in bilirubin effects. Coupled with previous work about glutamate toxicity in the same culture model, these data support the hypothesis that low levels of free bilirubin may promote programmed neuronal death corresponding to an apoptotic process which involves caspase activation and requires the participation of NMDA receptors, along with bilirubin-induced inhibition of protein kinase C activity.

Topics: Animals; Apoptosis; Bilirubin; Caspases; Cells, Cultured; Cycloheximide; Cysteine Proteinase Inhibitors; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Fluorescent Dyes; Indoles; Male; Neurons; Oligopeptides; Pregnancy; Prosencephalon; Protein Kinase C; Protein Synthesis Inhibitors; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Thymidine; Tritium

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

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

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

In addition to its intracellular antioxidant role, reduced glutathione (GSH) is released by CNS cells and may mediate or modulate excitatory neurotransmission. Although extracellular GSH levels rise in the ischemic cortex, its effect on the viability of energy-compromised neurons has not been defined. In this study, we tested the hypothesis that exogenous GSH would increase the vulnerability of cultured cortical neurons to azide-induced chemical hypoxia combined with glucose deprivation. Thirty minutes azide exposure in a glucose-free buffer was tolerated by most neurons, with release of less than 10% of neuronal LDH over the subsequent 21-25 h. Concomitant treatment with 10-100 microM GSH increased cell death in a concentration-dependent fashion, to 71.6+/-5.1% of neurons at 100 microM; GSH alone was nontoxic. Injury was blocked by the selective N-methyl-d-aspartate (NMDA) antagonist MK-801 but not by the AMPA/kainate antagonist NBQX. The sulfhydryl reducing agent mercaptoethanol (10-100 microM) mimicked the action of GSH; however, the zinc chelator ethylenediaminetetraacetic acid (EDTA) was ineffective. Two GSH analogues that lack a sulfhydryl group, S-hexylglutathione (SHG) and oxidized glutathione (GSSG), were inactive per se but attenuated the effect of both GSH and mercaptoethanol. These results suggest that micromolar concentrations of GSH enhance neuronal loss due to energy depletion by altering the extracellular redox state, resulting in increased NMDA receptor activation.

Topics: Animals; Cells, Cultured; Dizocilpine Maleate; Edetic Acid; Excitatory Amino Acid Antagonists; Glucose; Glutathione; Glutathione Disulfide; Hypoxia, Brain; Mercaptoethanol; Mice; Neurons; Oxidation-Reduction; Quinoxalines; Synaptic Transmission

There is mounting evidence that inflammation and associated excitotoxicity may play important roles in various neurodegenerative disorders, such as bacterial infections, Alzheimer's disease, AIDS dementia, and multiple sclerosis. The immunogen E. coli lipopolysaccharide (LPS, endotoxin) has been widely used to stimulate immune/inflammatory responses both systemically and in the CNS. Here, we show that exposure of parietal cortical slices from adult rats to LPS triggered very rapid (<2.5 min) and sustained releases of the neurotransmitters glutamate and noradrenaline, and of the neuromodulator adenosine. The responses to LPS declined rapidly following removal of the LPS and exhibited no tachyphylaxis to repeated exposures to LPS. The detoxified form of LPS had no effect. LPS-evoked release of [3H]noradrenaline, but not of glutamate or adenosine, appears to be partly due to the released glutamate acting at ionotropic receptors on the noradrenergic axons present in the cortical slices. LPS appears to release glutamate, which then acts at non-NMDA receptors to remove the voltage-sensitive Mg2+ block of NMDA receptors, thus permitting NMDA receptors to be activated and noradrenaline release to proceed. It seems possible that rapid, inappropriate excitation may occur in the immediate vicinity of gram-negative bacterial infections in the brain. If similar inappropriate excitations are also triggered by those immunogens specifically associated with Alzheimer's disease (beta-amyloid), AIDS dementia (gp120 and gp41), or multiple sclerosis (myelin basic protein), they might explain some of the acute, transient neurological and psychiatric symptoms associated with these disorders.

Topics: Adenosine; Age Factors; Animals; Cerebral Cortex; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Lipopolysaccharides; Male; Neuroprotective Agents; Norepinephrine; Organ Culture Techniques; Parietal Lobe; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Sympathomimetics; Tritium

The neurotoxic actions of kainate and domoate were studied in cultured murine neocortical neurons at various days in culture and found to be developmentally regulated involving three components of neurotoxicity: (1) toxicity via indirect activation of N-methyl-D-aspartate (NMDA) receptors, (2) toxicity mediated by alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptors, and (3) toxicity that can be mediated by kainate receptors when desensitization of the receptors is blocked. The indirect action at NMDA receptors was discovered because (5R, 10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-im ine (MK-801), an NMDA receptor antagonist, was able to block part of the toxicity. The activation of NMDA receptors is most likely a secondary effect resulting from glutamate release upon kainate or domoate stimulation. 1-(4-Aminophenyl)-3-methylcarbamyl-4-methyl-3,4-dihydro-7,8-ethyle nedioxy-5H-2,3-benzodiazepine (GYKI 53655), a selective AMPA receptor antagonist, abolished the remaining toxicity. These results indicated that kainate- and domoate-mediated toxicity involves both the NMDA and the AMPA receptors. Pretreatment of the cultures with concanavalin A to prevent desensitization of kainate receptors led to an increased neurotoxicity upon stimulation with kainate or domoate. In neurons cultured for 12 days in vitro a small but significant neurotoxic effect was observed when stimulated with agonist in the presence of MK-801 and GYKI 53655. This indicates that the toxicity is produced by kainate receptors in mature cultures. Examining the subunit expression of the kainate receptor subunits GluR6/7 and KA2 did, however, not reveal any major change during development of the cultures.

Topics: Animals; Benzodiazepines; Blotting, Western; Cell Survival; Cells, Cultured; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Kainic Acid; Mice; Neocortex; Neuromuscular Depolarizing Agents; Neurotoxins; Quinoxalines; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate

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

Glutamatergic transmission within the nucleus tractus solitarii (nTS) is critical to full expression of hypoxia-induced cardiorespiratory responses in the rat. To further examine the role of N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) glutamate receptors in these responses, double-labeling studies of immunoreactivity for c-Fos protein and either NMDA (NR1) or AMPA (mGluR2/3) receptor expression were conducted in normoxic rats and in hypoxic rats receiving vehicle, MK801, or NBQX. Hypoxia markedly increased c-Fos immunoreactivity within nTS neurons which in the vast majority co-labeled for NMDA. Furthermore, MK801 markedly attenuated such responses. In contrast, co-localization of AMPA and c-Fos occurred in only a small proportion of neurons, and NBQX failed to modify hypoxia-induced c-Fos enhancements. These data suggest a predominant role for NMDA but not for AMPA glutamate receptors in nTS mediated components of the hypoxic response.

Topics: Air; Amino Acid Sequence; Animals; Cell Hypoxia; Consciousness; Dizocilpine Maleate; Gene Expression Regulation; Male; Molecular Sequence Data; Neurons; Proto-Oncogene Proteins c-fos; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Solitary Nucleus

Postmortem alterations in the neuronal cytoskeleton resemble some aspects of the cytoskeletal disruption associated with neurodegenerative disorders, and are also similar to those observed following ischemia and produced by excitotoxins in vivo and in vitro. This suggests the involvement of excitotoxic mechanisms during the postmortem interval. The purpose of this study was to determine if extracellular levels of glutamate are elevated postmortem. Extracellular levels of GABA and taurine were also monitored using in vivo microdialysis. These three amino acids were analyzed using high-performance liquid chromatography. When postmortem rat brain temperature cooled rapidly to near room temperature, dialysate concentrations of glutamate were not increased in the hippocampal CA1 region during a 2-h postmortem interval, although increased extracellular levels of GABA and taurine were observed. In contrast, maintenance of brain temperature at 37 degrees C resulted in a 12-to-40 fold elevation in extracellular glutamate levels 20-120 min postmortem. In addition, the elevation in dialysate taurine concentration was greater than that observed in rats in which postmortem brain temperature was not maintained. Excitatory amino acid antagonists, NBQX (2, 3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline) and MK-801 (dizocilpine, (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cylohepten-5, 10-imine hydrogen maleate blocked the additional elevation in taurine associated with maintaining brain at 37 degrees C, but had less robust effects against glutamate and GABA release. The results indicate that extracellular concentrations of glutamate, taurine and GABA increase in postmortem rat brain when physiologic temperatures are maintained, but that these increases are blunted when brain temperature decreases. After death, the human brain cools much more slowly than does the rat brain. Therefore, extracellular glutamate levels are likely to increase in the postmortem human brain and may contribute to excitotoxic neuronal damage occurring in the interval between death and autopsy.

Topics: Animals; Autopsy; Body Temperature Regulation; Brain; Chromatography, High Pressure Liquid; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; gamma-Aminobutyric Acid; Glutamic Acid; Hippocampus; Humans; Male; Microdialysis; Postmortem Changes; Quinoxalines; Rats; Rats, Sprague-Dawley; Taurine

Two patterns of transforming growth factor-beta1 (TGF-beta1) expression were identified in brains of normotensive rats following permanent occlusion of the middle cerebral artery (MCAO). First, a relative increase of TGF-beta1 mRNA by 37% was found at 12 h after MCAO in the ipsilateral cingulate cortex as compared to the homotopic contralateral region. The cingulate cortex is located distant from the ischemic territory. Treatment with the glutamate receptor antagonists MK-801 and NBQX did not reduce this expression (34% and 26% increase, respectively). Therefore, peri-infarct depolarization waves were probably not responsible for induction. Secondly, an increase of TGF-beta1 mRNA by 116% was found at 7 days after MCAO within infarcted tissue. This expression was not reduced by the glutamate receptor antagonists MK-801 (increase 140%) and NBQX (increase 137%), either. TGF-beta1 mRNA expression in the cingulate cortex at 12 h after MCAO is possibly mediated by neurons and astroglia and may support cell survival. Expression in the infarcted tissue at 7 days after MCAO is most likely related to the invasion of monocytes and may be involved in the downregulation of inflammatory events, in neoangiogenesis, and in formation of a glial scar around the infarct.

Topics: Animals; Arterial Occlusive Diseases; Brain; Cerebral Arteries; Chronic Disease; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Male; Quinoxalines; Rats; Rats, Inbred F344; RNA, Messenger; Transforming Growth Factor beta

Altered glutamatergic neurotransmission appears to be central to the pathophysiology of Parkinson's disease; consequently, considerable effort has been made to elucidate neuroprotective mechanisms against such toxicity. In the present study, the possible neuroprotective effect of glutamate receptor antagonists against MPP+ neurotoxicity on dopaminergic terminals of rat striatum was investigated. Different doses of glutamate receptor antagonists were coinfused with 1.5 microg of MPP+ into the striatum; kynurenic acid, a nonselective antagonist of glutamate receptors (30 and 60 nmol), partially protected dopaminergic terminal degeneration in terms of rescue of dopamine levels and tyrosine hydroxylase immunohistochemistry. Dizocilpine, a channel blocker of the NMDA receptor (1, 4, and 8 nmol), and 7-chlorokynurenic acid, a selective antagonist at the glycine site of the NMDA receptor (1 and 10 nmol), failed to protect dopaminergic terminals from MPP+ toxicity. However, 6-cyano-7-nitroquinoxaline-2,3-dione (0.5 and 1 nmol) and 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(f)quinoxaline (1 nmol), two AMPA-kainate receptor antagonists, protected against MPP toxicity. Our findings suggest that the toxic effects of MPP+ on dopaminergic terminals are not mediated through a direct interaction with the NMDA subtype of glutamate receptor, but with the AMPA-kainate subtype.

Topics: 1-Methyl-4-phenylpyridinium; 3,4-Dihydroxyphenylacetic Acid; 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Corpus Striatum; Dizocilpine Maleate; Dopamine; Dopamine Agents; Excitatory Amino Acid Antagonists; Female; Homovanillic Acid; Kynurenic Acid; Microinjections; Neuroprotective Agents; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Tyrosine 3-Monooxygenase

The novel serotonin receptor subtypes, 5-HT6 and 5-HT7, are located in limbic regions and have nanomolar affinities for atypical antipsychotics. These factors have led some to speculate about the involvement of 5-HT6 and 5-HT7 receptors in schizophrenia. However, relatively little is known about these receptor subtypes, including the regulation of their expression in limbic regions. In particular, the regulation of extracellular serotonin levels in the striatum and hippocampal formation by glutamate receptors led us to examine the effects of systemic ionotropic glutamate receptor modulator treatment on 5-HT6 and 5-HT7 receptor expression in these regions. MK-801 treatment induced a dose-dependent decrease in striatal 5-HT6 receptor mRNA levels; similarly, both aniracetam and NBQX treatments also led to decreases in striatal 5-HT6 receptor mRNA levels. Hippocampal 5-HT6 and 5-HT7 receptor expression were not dramatically affected by any of the treatments. To our knowledge, this is the first demonstration of the regulation of striatal 5-HT6 receptor mRNA expression, and provides neurochemical anatomical evidence for the interaction of serotonergic and glutamatergic systems. Furthermore, although these two neurotransmitter systems are separately implicated in schizophrenia, the glutamatergic regulation of the expression of a receptor subtype associated with schizophrenia suggests that alterations in serotonin receptor expression in schizophrenia may result, in part, from altered glutamatergic activity.

Topics: Animals; Corpus Striatum; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hippocampus; Male; Nootropic Agents; Pyrrolidinones; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Glutamate; Receptors, Serotonin; RNA, Messenger; Schizophrenia

Administration of beta-amyloid fragment 25-35 (Abeta25-35) to cultured rat cerebellar granule cells (CGC) or cortical neurons caused cell death that was characterized by morphological and nuclear changes consistent with apoptosis. Inhibition of NMDA receptors produced a mild exacerbation of Abeta25-35 toxicity in cortical neurons; a similar effect was induced by AMPA/kainate receptor inhibition in CGC. Selective activation of group I metabotropic glutamate receptors (mGluR) by dihyroxyphenylglycine (DHPG) had no effect on Abeta25-35-induced apoptosis in either cell type, and was unaffected by blockade of ionotropic glutamate receptors. In contrast, selective inhibition of group I mGluR by (RS)-1-aminoindan-1,5-dicarboxylic acid (AIDA) exacerbated Abeta toxicity in cortical neurons, whereas this treatment was without effect on CGC. However, AIDA significantly increased Abeta-induced apoptosis in CGC in the presence of either NMDA or AMPA/kainate receptor inhibition; blockade of both ionotropic glutamate receptor classes further increased the exacerbation of apoptosis following treatment with AIDA. These findings suggest that Abeta25-35-induced neuronal injury leads to activation of group I mGluR, which attenuates the resulting apoptosis.

Topics: Amyloid beta-Peptides; Animals; Apoptosis; Cells, Cultured; Cerebellum; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Neocortex; Neurons; Peptide Fragments; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Metabotropic Glutamate

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

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

After the transection of the Schaffer collateral pathway in hippocampal slice cultures, reactive sprouting is induced in the CA3 area, and eventually synaptic transmission between areas CA1 and CA3 is restored. Using this model, we have studied the role of ionotropic glutamate receptors in the initiation of axonal sprouting and the regeneration of functional synapses. We show that neither reactive sprouting nor functional recovery of synaptic transmission occur in the presence of the non-N-methyl-D-aspartate (NMDA) receptor antagonist 6-nitro-7-sulfamoylbenzoquinoxaline-2,3-dione (CNQX). In contrast, the NMDA receptor antagonists methyl-10, 11-dihydro-5-H-dibenzocyclohepten-5,10-imine (MK-801) or 3-(RS)-2-carboxypiperazine-4-yl)-propyl-1-phosphonic acid (CPP) did not interfere with these processes. Moreover, we observed that the application of NMDA receptor antagonists induced massive axonal sprouting and an increase in the frequency of miniature excitatory postsynaptic currents in unlesioned cultures. Our results thus indicate that NMDA and non-NMDA receptors exert a differential effect on reactive sprouting and the recovery of synaptic transmission after injury in the hippocampus. Activation of non-NMDA receptors appears necessary for these processes to occur, whereas activation of NMDA receptors suppresses growth-associated protein -43 expression and axonal outgrowth.

Topics: Animals; Axons; Culture Techniques; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; GAP-43 Protein; Glial Fibrillary Acidic Protein; Hippocampus; Nerve Regeneration; Piperazines; Quinoxalines; Rats; Rats, Wistar

It is generally believed that nuclear condensation and fragmentation as well as DNA fragmentation reflect the events related to the neuronal apoptosis. Our report demonstrates that severe oxygen-glucose deprivation (OGD) induced condensation and fragmentation of nuclear chromatin of neurones in primary cultures of cerebellar granule cells without intemucleosomal DNA fragmentation. DNA fragmentation detected by TUNEL assay was seen only after mild OGD or after addition of colchicine but not after severe OGD. Thus, at least in primary cerebellar granule cell cultures, the chromatin condensation and fragmentation cannot be considered as a hallmark of apoptosis but rather reflect the neuronal death despite of its form.

Topics: Animals; Cell Death; Cerebellum; Colchicine; Dizocilpine Maleate; DNA Fragmentation; Excitatory Amino Acid Antagonists; Glucose; In Situ Nick-End Labeling; Microscopy, Confocal; Neurons; Oxygen; Quinoxalines; Rats; Rats, Wistar

A double-tracer autoradiographic method for simultaneous measurement of regional glucose utilization (rCMRglc) and regional protein synthesis (PS) in consecutive brain sections is described and applied to study the metabolism of the ischemic penumbra 2 h after occlusion of the middle cerebral artery (MCAO) in rats. In halothane anesthesia, the left middle cerebral artery was permanently occluded. Two hours after MCAO an i.v. bolus injection of 14C-deoxyglucose and 3H-leucine was given and circulated for 45 min. Two sets of brain sections were processed for quantitative autoradiography. Neighboring brain sections exposed an X-ray film (3H-insensitive), and a 3H-sensitive for determination of rCMRglc and PS, respectively. Sections for PS determination were washed in trichloroacetic acid (TCA) prior to film exposure in order to remove 14C-deoxyglucose and unincorporated 3H-leucine. Regional rates of PS and glucose utilization were measured by densitometric image analysis. Normal rates of metabolism were defined as mean +/- 2 SD of values in the non-ischemic cortex. The volumes of ischemic cortex displaying normal rates of PS and glucose utilization, respectively, were measured. The cortical volume with normal PS was significantly less than that of normal rCMRglc: 142 (127-147) mm3 vs. 203 (184-206) mm3. Treatment with the glutamate antagonists MK-801 (1 mg kg-1) and NBQX (30 mg kg-1 x 2) did not significantly change this, although MK-801 tended to reduce the size of the metabolic penumbra calculated as the difference between ischemic cortex with reduced PS and ischemic cortex with reduced rCMRglc.

Topics: Animals; Autoradiography; Blood Pressure; Brain; Carbon Radioisotopes; Deoxyglucose; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glucose; Ischemic Attack, Transient; Leucine; Male; Nerve Tissue Proteins; Quinoxalines; Radioisotope Dilution Technique; Rats; Rats, Wistar; Reference Values; Tritium

Metabotropic glutamate (mGlu) receptors have been implicated in a number of physiological and pathological responses to glutamate, but the exact role of group I mGlu receptors in causing postischaemic injury is not yet clear. In this study, we examined whether the recently-characterized and relatively selective mGlu1 receptor antagonists 1-aminoindan-1,5-dicarboxylic acid (AIDA) and (S)-(+)-2-(3'-carboxybicyclo[1.1.1]pentyl)-glycine (CBPG) could reduce neuronal death in vitro, following oxygen-glucose deprivation (OGD) in murine cortical cell and rat organotypic hippocampal cultures, and in vivo, after global ischaemia in gerbils. When present in the incubation medium during the OGD insult and the subsequent 24 h recovery period, AIDA and CBPG significantly reduced neuronal death in vitro. The extent of protection was similar to that observed with the nonselective mGlu receptor antagonist (+)-alpha-methyl-4-carboxyphenylglycine [(+)MCPG] and with typical ionotropic glutamate (iGlu) receptor antagonists. Neuroprotection was also observed when AIDA or CBPG were added only after the OGD insult was terminated. Neuronal injury was not attenuated by the inactive isomer (-)MCPG, but was significantly enhanced by the nonselective mGlu receptor agonist (1S,3R)-1-aminocyclopentane-1, 3-dicarboxylic acid [(1S,3R)-ACPD] and the group I mGlu receptor agonist 3,5-dihydroxyphenylglycine (3,5-DHPG). The antagonists (+)MCPG, AIDA and CBPG were also neuroprotective in vivo, because i. c.v. administration reduced CA1 pyramidal cell degeneration examined 7 days following transient carotid occlusion in gerbils. Our results point to a role of mGlu1 receptors in the pathological mechanisms responsible for postischaemic neuronal death and propose a new target for neuroprotection.

Topics: Animals; Animals, Newborn; Astrocytes; Benzoates; Bridged Bicyclo Compounds; Cell Death; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Gerbillinae; Glycine; Indans; Ischemic Attack, Transient; Mice; Neuroprotective Agents; Neurotoxins; Organ Culture Techniques; Pyramidal Cells; Quinoxalines; Receptors, Metabotropic Glutamate; Resorcinols

Quantitative autoradiography was used to characterise the binding of selective radiolabelled antagonists for the N-methyl-D-aspartate (NMDA) receptor and the alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) receptor in the dorsal, intermediate and ventral subregions of the grey matter of the upper thoracic spinal cord in male and female lean and obese-diabetic (ob/ob) mice. The density of binding sites for both receptor subtypes was greater in diabetic mice, in all three subregions of the grey matter, than the corresponding subregions in the lean mice. The affinity of the binding site for the NMDA antagonist was significantly higher in obese mice than lean mice, consistent with the presence of two subpopulations of NMDA receptors with different ligand binding affinities in obese mice. The increase in expression of the glutamate receptor subtypes, and altered ligand affinity for the NMDA receptor subtype in the obese mice may be causally involved in the peripheral neuropathies which can accompany diabetes mellitus.

Topics: Animals; Autoradiography; Dizocilpine Maleate; Female; Kinetics; Male; Mice; Mice, Obese; Neurons; Obesity; Quinoxalines; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Sex Characteristics; Spinal Cord; Thinness; Tritium

Changes of evoked potentials under the influence of NBQX (a non-NMDA receptor antagonist), MK-801 (a NMDA receptor antagonist) and GDEE (a nonselective antagonist of glutamate receptor) were studied. GDEE augmented potentiation and was marked progression of potentiation with increasing number of stimuli. There was no potentiation of responses in relation to the number of stimulus in a series in experiments with both MK-801 and NBQX. Interpretation of results with NBQX and MK-801 is difficult.

Topics: Animals; Dizocilpine Maleate; Evoked Potentials; Excitatory Amino Acid Antagonists; Glutamates; Male; Quinoxalines; Rats; Receptors, AMPA

The influence of N-methyl-D-aspartate (NMDA) or alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptor antagonists in combination with halothane anaesthesia on the respiratory system was investigated. Under 1.5% halothane anaesthesia, respiratory parameters including respiratory rate, minute volume, tidal volume, inspiratory and expiratory duration were measured before and after drug administration in rats. The AMPA receptor antagonists, 6-(1H-imidazol-1-yl)-7-nitro-2,3-(1H,4H)-quinoxalinedione hydrochloride, YM90K (5 and 10 mg/kg) and 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX, 15 mg/kg), which were administered intravenously for 30 min, significantly reduced the respiratory rate (P < 0.01) and minute volume (P < 0.01) and increased the tidal volume (P < 0.05) compared with values obtained before drug administration. None of these drugs affected respiratory parameters in the absence of anaesthesia. A NMDA receptor antagonist, MK-801 (0.5 mg/kg), which was administered intravenously for 30 min, also significantly reduced respiratory rate (P < 0.01), minute volume (P < 0.01) and tidal volume (P < 0.01) and prolonged inspiratory duration (P < 0.05). These results suggest that both AMPA and NMDA receptor antagonists cause respiratory depression under halothane anaesthesia in rats, although the mechanisms may be different for the two types of antagonists.

Topics: Anesthetics, Inhalation; Animals; Blood Gas Analysis; Dizocilpine Maleate; Drug Synergism; Excitatory Amino Acid Antagonists; Halothane; Hemodynamics; In Vitro Techniques; Male; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Respiratory Mechanics

Pesticides acting at GABAA receptors may induce convulsions in man and animals, but the mechanisms responsible for their convulsant activity are not fully explained. The following excitatory amino acid antagonists were studied for their protective action in mice intoxicated with chlorinated hydrocarbon insecticide lindane (gamma-hexachlorocyclohexane): the competitive NMDA antagonist: 3-(2-carboxypiperazine-4-yl)propenyl-1- phosphonic acid (D-CPPene, 20 mg/kg), the non-competitive NMDA antagonist: dizocilpine (MK-801, 0.4 mg/kg), the glycine site antagonist of NMDA receptor: 2-phenyl-1,3-propane-diol dicarbamate (felbamate, 400 mg/kg) and the competitive AMPA antagonist: 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX, 100 mg/kg). Systemic administration of an antagonist prior to lindane resulted in a strong anticonvulsant effect. D-CPPene, MK-801 and NBQX produced a marked increase of CD50 values of lindane for clonic convulsions. All the antagonists protected animals against tonic convulsions. Toxicity of lindane was potently reduced, as assessed 2, 24 and 120 hr after administration of the pesticide. Our results demonstrate that excitatory amino acid antagonists reduce convulsant properties and toxicity of lindane, suggesting that excitatory amino acid neurotransmission may be involved in its central action.

Topics: Animals; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Felbamate; Hexachlorocyclohexane; Insecticides; Lethal Dose 50; Male; Mice; Phenylcarbamates; Propylene Glycols; Quinoxalines; Seizures

The present study tested the hypothesis that the increase in extracellular striatal dopamine during hypoxia is least partly associated with activation of N-methyl-D-aspartate (NMDA) and/or non-NMDA excitatory amino acid receptors. Studies were performed in anesthetized and mechanically ventilated 2-3 days old piglets. Hypoxic insult was induced by decreasing the oxygen fraction in inspired gas (FiO2) from 22 to 7% for 1 h, followed by 1 h reoxygenation at 22%. Cortical oxygen pressure was measured optically by oxygen dependent quenching of phosphorescence, and extracellular striatal dopamine was measured using in vivo microdialysis. The microdialysis probes were perfused with Ringer solution +/- 50 microM (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801) or 50 microM 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX). One hour of hypoxia decreased the cortical oxygen pressure from 46 +/- 3 Torr to 10 +/- 1.8 Torr. In striatum perfused with Ringer, statistically significant increase in extracellular dopamine, to 1050 +/- 310% of control, was observed after 20 min of hypoxia. By 40 min of hypoxia the extracellular level of dopamine increased to 4730 +/- 900% of control; by the end of the hypoxic period the values increased to 18,451 +/- 1670% of control. The presence of MK-801 in the perfusate significantly decreased the levels of extracellular dopamine during hypoxia. At 20, 40 and 60 min of hypoxia extracellular level of dopamine increased to 278 +/- 94% of control, 1530 +/- 339% of control and 14,709 +/- 1095 of control, respectively. The presence of NBQX caused a statistically significant decrease, by about 30%, in the extracellular dopamine compared to control, only at the end of the hypoxic period. It can be concluded that in striatum of newborn piglets, the excitatory NMDA receptors but not the non-NMDA receptors may be modulating the changes in extracellular levels of dopamine. The NMDA receptor antagonist, MK-801, may exert part of its reported neuroprotective effect to hypoxic stress in striatum by decreasing the levels of extracellular dopamine.

Topics: Animals; Animals, Newborn; Corpus Striatum; Dizocilpine Maleate; Dopamine; Excitatory Amino Acid Antagonists; Extracellular Space; Hypoxia; Oxygen; Oxygen Consumption; Pressure; Quinoxalines; Receptors, Amino Acid; Swine

Hyperbaric O2 exposure causes seizures by an unknown mechanism. Cerebral blood flow (CBF) may affect seizure latency, although no studies have demonstrated a direct relationship. Awake rats (male, Sprague-Dawley, 350-450 g), instrumented for measuring electroencephalographic activity (EEG) and CBF (laser-Doppler flowmetry), were exposed to 100% O2 at 4 or 5 atm (gauge pressure) until EEG seizures. Compression with O2 caused vasoconstriction to about 70% of control flow that was maintained for various times. CBF then suddenly, but transiently, increased at a time that was reliably related to seizure latency (r=0.8, p<0.01). Additional animals were treated with agents that have diverse pharmacology and their effects on CBF and latency were measured. Glutamate receptor antagonists MK-801 (1 or 4 mg/kg) and ketamine (20-100 mg/kg) significantly increased CBF by 60-80% and decreased seizure latency from about 17+/-8 min (+/-S.D.) in controls to 5+/-1 and 6+/-2 min, respectively. In opposite, a nitric oxide synthase (NOS) inhibitor, N-nitro-L-arginine (NNA)(25 mg/kg) decreased CBF by about 25% and increased time to seizure to 60+/-16 min. If these effects occur in humans, non-invasive measurement of CBF could potentially improve the safety and reliability of hyperbaric O2 usage in clinical and diving applications. It also appears that the effect of drugs on seizure latency can be explained, at least in part, by their effect on CBF.

Topics: Analysis of Variance; Animals; Cerebrovascular Circulation; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Hyperbaric Oxygenation; Ketamine; Male; Nitric Oxide Synthase; Nitroarginine; Quinoxalines; Rats; Rats, Sprague-Dawley; Reaction Time; Seizures

We investigated whether the neuropeptide galanin affects the nitric oxide synthase/cyclic GMP pathway in rat hippocampus by measuring in vivo the extracellular cyclic GMP levels during microdialysis. Galanin (2.5 and 3.5 nmol; i.c.v.) dose-dependently raised the extracellular levels of cyclic GMP in the ventral but not the dorsal hippocampus. The effect of 3.5 nmol galanin was blocked by local application of tetrodotoxin and inhibited by the high-affinity galanin antagonist M40 (galanin-[1-12]-Pro3-[Ala-Leu]2-Ala amide). The non-competitive N-methyl-D-aspartate receptor antagonist dizocilpine maleate (30 microM infused into the ventral hippocampus or 0.2 mg/kg, i.p.) and the competitive one, 3-([R]-carboxypiperazin-4-yl)-propyl-phosphonic acid (50 microM infused), but not local perfusion of the AMPA antagonist 6-nitro-7-sulphamoylbenzo(f)quinoxaline-2,3-dione (15 microM) abolished the galanin-evoked cyclic GMP response in the hippocampus. Inhibitors of nitric oxide synthase, L-Arg(NO2)-OMe.HCl and 7-nitroindazole monosodium salt, applied locally, blocked the galanin-induced increase in hippocampal extracellular cyclic GMP. This increase was also prevented by local application of 1H-(1,2,4)oxadiazolo(4,3a) quinoxalin-1-one, a selective inhibitor of soluble guanylyl cyclase. The galanin receptors mediating the rise in cyclic GMP reside outside the hippocampus, as galanin (0.35-3 nmol) locally applied had no effect. The results provide in vivo evidence that galanin stimulates the N-methyl-D-aspartate receptor/nitric oxide synthase/cyclic GMP pathway in the ventral hippocampus, which may be of importance in memory processes.

Topics: Animals; Cyclic GMP; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Galanin; Hippocampus; Locomotion; Male; Microdialysis; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Oxadiazoles; Peptide Fragments; Piperazines; Quinoxalines; Rats; Rats, Inbred Strains; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Tetrodotoxin

A new AMPA receptor antagonist, Ro 48-8587, was characterized pharmacologically in vitro. It is highly potent and selective for AMPA receptors as shown by its effects on [3H]AMPA, [3H] kainate, and [3H] MK-801 binding to rat brain membranes and on AMPA- or NMDA-induced depolarization in rat cortical wedges. [3H]Ro 48-8587 bound with a high affinity (KD = 3 nM) to a single population of binding sites with a Bmax of 1 pmol/mg of protein in rat whole brain membranes. [3H]Ro 48-8587 binding to rat whole brain membranes was inhibited by several compounds with the following rank order of potency: Ro 48-8587 > 6-nitro-7-sulphamoylbenzo[f] quinoxaline-2,3-dione (NBQX) > YM 90K > 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) > quisqualate > AMPA > glutamate > kainate > NMDA. The distribution and abundance of specific binding sites (approximately 95% of total) in sections of rat CNS, revealed by quantitative receptor radioautography and image analysis, indicated a very discrete localization. Highest binding values were observed in cortical layers (binding in layers 1 and 2 > binding in layers 3-6), hippocampal formation, striatum, dorsal septum, reticular thalamic nucleus, cerebellar molecular layer, and spinal cord dorsal horn. At 1 nM, the values for specific binding were highest in the cortical layers 1 and 2 and lowest in the brainstem (approximately 2.6 and 0.4 pmol/mg of protein, respectively). Ro 48-8587 is a potent and selective AMPA receptor antagonist with improved binding characteristics (higher affinity, selectivity, and specific binding) compared with those previously reported.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Anti-Anxiety Agents; Benzodiazepines; Brain Chemistry; Dizocilpine Maleate; Electrophysiology; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Imidazoles; In Situ Hybridization; Kainic Acid; Kinetics; Male; N-Methylaspartate; Quinazolines; Quinoxalines; Radioligand Assay; Rats; Rats, Inbred Strains; Receptors, AMPA; RNA, Messenger; Tritium

We determined the conditions (immaturity, species, anesthesia, receptor blockade selectivity) under which glutamate receptor blockade produces respiratory depression in mammals. In unrestrained 0- to 2-day-old neonate and adult mice and cats, ventilation was measured by the barometric method, before and after separate or sequential administration of a non-NMDA receptor antagonist, NBQX (2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(F)quinoxaline, 2-200 mg kg(-1) in mice, 10-40 mg kg(-1) in cats), and a NMDA receptor antagonist, dizocilpine (3 mg kg(-1) in mice, 0.15-1.0 mg kg(-1) in cats). NBQX or dizocilpine alone did not decrease ventilation in awake adults, but NBQX strongly depressed ventilation in neonate awake mice and in adult anesthetized animals. Given together, dizocilpine and NBQX always profoundly depressed ventilation by producing a lethal apnea in neonate mice, and an apneustic pattern of breathing in adults of both species and in neonate cats. We conclude that blockade of either NMDA or non-NMDA receptors is innocuous in awake adults. The factors which may potentiate respiratory depression are (1) anesthesia, (2) immaturity, and (3) combined blockade of both receptors types. The mechanism of depression is species-dependent and age-dependent.

Topics: Age Factors; Animals; Animals, Newborn; Apnea; Cats; Dizocilpine Maleate; Drug Synergism; Excitatory Amino Acid Antagonists; Injections, Intraperitoneal; Injections, Subcutaneous; Mice; Plethysmography; Pulmonary Ventilation; Quinoxalines; Receptors, Glutamate; Species Specificity

The isolated spinal cord of the neonatal mouse spontaneously generates two different motor patterns of continuous rhythmic bursting: one in lumbar ventral roots in earliest postnatal preparations (P0-2) and another at the sacral level at later postnatal times (P3-5). Lumbar rhythmic motor discharges clearly alternate on contralateral roots and are in a frequency range (approximately 1 Hz) usually described for locomotor-like activity, while sacral motor sequences include mixed synchrony and irregular bilateral alternation that differ from typical locomotor-like activity. A significant decrease in the frequency and increase in the duration of spontaneous rhythmic bursts occur between lumbar and sacral motor patterns. In quiescent preparations from both postnatal periods, perfusion with Mg(2+)-free medium systematically induces a rhythmic activity at both lumbar and sacral level. Temporal characteristics of motor patterns under Mg(2+)-free medium are similar to spontaneous rhythms. Activating NMDA receptor channels by diminishing their Mg2+ block appears to be an efficient way of decreasing the threshold for genesis of the spinal rhythm in mouse. Bath application of NMDA and non-NMDA receptor antagonists blocks Mg(2+)-free-induced rhythmic activities in an irreversible or reversible manner, respectively. The effects of Mg(2+)-free medium and of glutamate antagonists provide evidence for the excitatory amino acid (EAA) dependence of both rhythmic motor patterns. Finally, the possibility that the recording of two different motor patterns may reflect a rostrocaudal developmental process is discussed.

Topics: Animals; Animals, Newborn; Dizocilpine Maleate; Electrophysiology; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Female; Locomotion; Magnesium; Membrane Potentials; Mice; Motor Neurons; N-Methylaspartate; Periodicity; Pregnancy; Quinoxalines; Spinal Cord

At many central excitatory synapses, AMPA receptors relay the electrical signal, whereas activation of NMDA receptors is conditional and serves a modulatory function. We show here quite a different role for NMDA receptors at dendrodendritic synapses between mitral and granule cells in the rat olfactory bulb. In whole-cell patch-clamp recordings in bulb slices, stimulation of mitral cells elicited slowly decaying, GABAA receptor-mediated reciprocal IPSCs that reflected prolonged GABA release from granule cells. Although granule cells had a normal complement of AMPA and NMDA receptors, the IPSC was completely blocked by the NMDA receptor antagonist D,L-AP-5, suggesting that NMDA receptor activation is an absolute requirement for dendrodendritic inhibition. The AMPA receptor antagonist 1,2,3,4-tetrahydro-6-nitro-2, 3-dioxobenzo[f]quinoxaline-7-sulfonamide (NBQX) had no effect on IPSCs in the absence of extracellular magnesium but modestly reduced IPSCs in 1 mM magnesium, indicating that the primary effect of the AMPA receptor-mediated depolarization was to facilitate the unblocking of NMDA receptors. Granule cell voltage recordings indicated that effective spike stimulation in granule cells depended on the slow NMDA receptor kinetics. Granule cells also showed a pronounced delay between synaptic stimulation and action potential generation, suggesting that their intrinsic membrane properties underlie the ineffectiveness of brief AMPA receptor-mediated EPSPs. NMDA receptors also seem to have a central role in dendrodendritic inhibition in vivo, because intraperitoneal dizocilpine maleate (MK-801) injection in young adult rats resulted in disinhibition of mitral cells as measured by the generation of c-fos mRNA. The unique dependence of dendrodendritic inhibition on slow EPSPs generated by NMDA receptors suggests that olfactory information processing depends on long-lasting reciprocal and lateral inhibition.

Topics: 2-Amino-5-phosphonovalerate; Animals; Dendrites; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; In Vitro Techniques; Magnesium; Male; Neural Inhibition; Olfactory Bulb; Patch-Clamp Techniques; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

Functional interaction between ionotropic and metabotropic glutamate receptors (iGluR and mGluR respectively) was studied in cerebellar granule cell cultures using quisqualate (QA), the most potent agonist of phosphoinositide hydrolysis coupled mGluR, and N-methyl-D-aspartate (NMDA) or kainate (KA) that activate different classes of iGluR. Two h exposure to NMDA or KA resulted in a marked reduction (about 75%) of QA-evoked PI hydrolysis. The efficacy of the two agonists was about the same, but the potencies were different (IC50 for NMDA about 35 microM and for KA about 70 microM). NMDA-induced depression of QA-stimulated PI hydrolysis was relatively long lasting but reversible. Recovery required protein synthesis. In nominally Ca2+-free medium both NMDA and KA failed to attenuate QA-stimulated PI hydrolysis. The effect of NMDA was prevented by the NMDA receptor antagonist MK801, but not by the wide spectrum protein kinase inhibitor staurosporin nor by the nitric oxide synthase inhibitor N omega-nitro-L-arginine. Cycloheximide and concanavalin A were also ineffective. The effect of KA was prevented by the selective non-NMDA receptor antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX). Voltage sensitive Ca2+ channel antagonists together with MK801 did not counteract the inhibition by KA of the QA response. Both NMDA and KA attenuated PI hydrolysis evoked by the muscarinic receptor agonist carbachol (about 30%), indicating that the activation of iGluRs exerts a relatively general inhibitory effect on the function of different PLC-coupled metabotropic receptors. Consistent with this observation is that treatments either with KA and NMDA induced an inhibition (about 30%) of NaF-stimulated PI hydrolysis which occurs through the direct activation of G proteins. Our observations show that ionotropic glutamate receptor stimulation induces a long lasting suppression of QA-evoked PI breakdown through a Ca2+ dependent mechanism which seems to involve receptor coupled transduction systems downstream from mGluR. Such a Ca2+-dependent cross-talk involving ionotropic and metabotropic receptors may play a role in certain events of synaptic plasticity.

Topics: Animals; Calcium; Cells, Cultured; Cerebellum; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; GTP-Binding Proteins; Hydrolysis; Kainic Acid; N-Methylaspartate; Phosphatidylinositols; Quinoxalines; Quisqualic Acid; Rats; Rats, Wistar; Receptors, Glutamate; Receptors, Metabotropic Glutamate; Receptors, N-Methyl-D-Aspartate; Sodium Fluoride

Expression of brain-derived neurotrophic factor (BDNF) may play a role in the mechanism of neuronal cell death after cerebral ischemia. We investigated the changes in levels of mRNAs encoding BDNF and its promoters in the rat brain after transient forebrain ischemia. Transient forebrain ischemia was induced by occlusion of bilateral common carotid arteries and systemic hypotension for 8 min. The alterations in BDNF gene expression in the hippocampus and in the cerebral cortex were examined by in situ hybridization using a mouse BDNF cDNA probe and cDNA probes including exon-specific promoters. BDNF transcripts were rapidly enhanced after the ischemic insult, both in the hippocampus and the cerebral cortex. NBQX suppressed the enhanced gene expression of BDNF markedly in the dentate gyrus (DG). In contrast, MK-801 had little effect on BDNF expression. In the piriform cortex, MK-801 or NBQX reduced the expression only moderately. After the ischemic insult, promoter specific BDNF 5'-exon I and exon III were increased remarkably in the DG. The increase in exon I in DG was suppressed partially by MK-801 and NBQX, while the increase in exon III in CA3 was suppressed by MK-801 but that in DG was not suppressed by either antagonist. In the piriform cortex, exon III was increased remarkably and this increase was not influenced by either agonist. These results suggest that the gene expression of BDNF was enhanced by transient ischemia both in the hippocampus and the cerebral cortex and that the cerebral ischemia stimulated at least two different promoter- and neuron type-specific pathways regulating expression of the BDNF gene mediated by glutamate receptors of non-NMDA type and NMDA type.

Topics: Animals; Brain-Derived Neurotrophic Factor; Cerebral Cortex; Dizocilpine Maleate; DNA Probes; Excitatory Amino Acid Antagonists; Exons; Gene Expression; Hippocampus; In Situ Hybridization; Ischemic Attack, Transient; Male; Mice; Promoter Regions, Genetic; Quinoxalines; Rats; Rats, Wistar; RNA, Messenger; Tissue Distribution

The developmental expression of calcium (Ca2+)-permeable alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and kainate receptors in cultured neocortical neurons was evaluated by using cobalt uptake, a histochemical method that identifies cells expressing Ca2+-permeable, non-N-methyl-D-aspartate (non-NMDA) receptors. At a concentration of 500 microM, AMPA was found to stimulate cobalt uptake only late in development, resulting in staining of 2.7%+/-0.3% of the neurons maintained in culture for 12 days in vitro (DIV). When AMPA receptor desensitization was blocked with 50 microM cyclothiazide, the developmental profile of cobalt uptake mediated by 25 microM AMPA changed dramatically. The cobalt staining now appeared in young cultures (5 DIV), and the percentage of stained cells increased from 3.4%+/-0.2% at 5 DIV to 21.7%+/-1.6% at 12 DIV. The effect of 200 microM kainate was similar to that seen with 25 microM AMPA plus 50 microM cyclothiazide, resulting in 17.7%+/-0.3% stained neurons at 12 DIV. The cobalt uptake was specific to AMPA and kainate receptors because NMDA receptors and voltage-gated calcium channels were found not to mediate any cobalt staining. In addition, 10 microM 6-nitro-7-sulphamoylbenzo-[f]-quinoxaline-2,3-dione (NBQX) was able to prevent all staining at 5 and 8 DIV and most of the staining at 12 DIV, indicating that the non-NMDA ionotropic glutamate receptors are involved in cobalt uptake into the neurons. The AMPA receptor-selective antagonist GYKI 53655 was used to differentiate between cobalt influx through AMPA- or kainate-preferring receptors. After pretreatment with concanavalin A (con A), an inhibitor of kainate receptor desensitization, cobalt uptake was assessed after stimulation by 200 microM kainate in the presence of 25 microM GYKI 53655. No cobalt staining was observed under these conditions, indicating that most if not all of the cobalt influx induced by kainate was mediated through AMPA receptor channels.

Topics: Animals; Calcium; Cells, Cultured; Cobalt; Concanavalin A; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Mice; Neocortex; Neurons; Permeability; Quinoxalines; Receptors, AMPA; Receptors, Kainic Acid; Staining and Labeling

Evidence is accumulating that glutamate, a major neurotransmitter, exerts potent neurotoxic activity during ischemia. In our laboratory, a delayed-onset paraplegia model using rabbits has been developed and described. The severity of the ischemic event in this model, i.e., extracellular glutamate overload, is believed to influence the etiology of this borderline lesion. We hypothesized that glutamate receptor antagonists (MK-801, NBQX) would attenuate the delayed neuronal dysfunction that follows spinal cord ischemia. Infrarenal aortic segments from 18 New Zealand white rabbits were isolated for 5 minutes and infused at a rate of 2 ml/min. Group I (n = 6) received normothermic L-glutamate (20 mM). Group II (n = 6) received 3 mg of MK-801 and normothermic L-glutamate (20 mM). Group III (n = 6) received 3 mg of NBQX and normothermic L-glutamate (20 mM). Neurologic function was assessed at 6, 24, and 48 hours after surgery according to the modified Tarlov scale. After 48 hours, the rabbits were euthanized and spinal cords were harvested for histologic examination. The neurologic function of three rabbits in group I showed acure paraplegia and the other three showed delayed-onset paraplegia, whereas all group II animals had nearly intact neurologic function and all group III animals showed mild neurologic disturbance. Histologic examination of spinal cords from rabbits in group I showed evidence of moderate spinal cord injury with necrosis of central gray matter and adjacent white matter and axonal swelling, whereas spinal cords from group II showed small and localized spinal cord injuries and those from group III revealed no evidence of cord injury. These results indicate that MK-801 and NBQX exert different neuroprotective effects related to different mechanisms of glutamate neurotoxicity mediated by the NMDA receptor and non-NMDA receptor, which initiate a deleterious cascade of biochemical events that ultimately results in delayed-onset paraplegia.

Topics: Animals; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Glutamates; Ischemia; Neuroprotective Agents; Quinoxalines; Rabbits; Spinal Cord

N-methyl-D-aspartate (NMDA) stimulated release of [3H]noradrenaline (NA) from prelabelled rat spinal cord slices. The release was partially insensitive to tetrodotoxin (TTX) and was inhibited by the NMDA antagonist MK-801. Alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) also evoked release of [3H]NA, which was enhanced by blocking AMPA receptor desensitization with cyclothiazide. AMPA-evoked release was inhibited by the non-NMDA antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(f)-quinoxaline (NBQX) but was not affected by TTX. NMDA and AMPA showed synergistic effects, indicating co-existence of NMDA and AMPA receptors on noradrenergic terminals. Kainate evoked [3H]NA release only at high concentrations and the release was not potentiated by blocking kainate receptor desensitization with concanavalin A. Thus, the results indicate that there are stimulatory presynaptic NMDA and AMPA receptors on noradrenergic axon terminals in the spinal cord and that they interact synergistically to evoke release of [3H]NA.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Benzothiadiazines; Concanavalin A; Dizocilpine Maleate; In Vitro Techniques; Kainic Acid; Male; N-Methylaspartate; Norepinephrine; Presynaptic Terminals; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Spinal Cord; Tetrodotoxin

The molecular basis for developing symptomatic epilepsy (epileptogenesis) remains ill defined. We show here in a well characterized hippocampal culture model of epilepsy that the induction of epileptogenesis is Ca2+-dependent. The concentration of intracellular free Ca2+ ([Ca2+]i) was monitored during the induction of epileptogenesis by prolonged electrographic seizure activity induced through low-Mg2+ treatment by confocal laser-scanning fluorescent microscopy to directly correlate changes in [Ca2+]i with alterations in membrane excitability measured by intracellular recording using whole-cell current-clamp techniques. The induction of long-lasting spontaneous recurrent epileptiform discharges, but not the Mg2+-induced spike discharges, was prevented in low-Ca2+ solutions and was dependent on activation of the N-methyl-D-aspartate (NMDA) receptor. The results provide direct evidence that prolonged activation of the NMDA-Ca2+ transduction pathway causes a long-lasting plasticity change in hippocampal neurons causing increased excitability leading to the occurrence of spontaneous, recurrent epileptiform discharges.

Topics: 2-Amino-5-phosphonovalerate; Animals; Animals, Newborn; Benzoates; Calcium; Cells, Cultured; Dizocilpine Maleate; Egtazic Acid; Epilepsy; Excitatory Amino Acid Antagonists; Glycine; Hippocampus; Magnesium; Membrane Potentials; Microscopy, Confocal; Neurons; Nifedipine; Patch-Clamp Techniques; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Signal Transduction

The diazoxide derivative 7-chloro-3-methyl-3,4-dihydro-2H-1,2,4-benzothiadiazine-S,S-dioxide (IDRA21) enhances memory and learning in rodents, most likely by potentiating AMPAergic synaptic activity. We examined IDRA21's effect upon AMPAergic synaptic currents and whole-cell glutamate currents in cultured rat hippocampal neurons to determine whether IDRA21 was a partial modulator of AMPA receptor desensitization and deactivation. Comparable to cyclothiazide, IDRA21 prolonged AMPAergic autaptic currents (5.6 times control, EC50 150 microM) and slowed the rate of AMPA deactivation (3 times control) following 1-ms applications of 1 mM glutamate to excised, outside-out membrane patches. IDRA21 also augmented autaptic GABA currents by 27 +/- 8.1%, although it had two opposing effects, reducing the peak amplitude versus prolonging autaptic GABA currents. IDRA21 (200 microM) inhibited whole-cell GABA currents elicited by exogenously applied 1 mM GABA by 41 +/- 11%. At sufficient concentrations, IDRA21 reduced AMPA receptor desensitization and slowed the rate of deactivation, most consistent with full agonist activity with lower potency compared to cyclothiazide. IDRA21 slightly augments GABAergic synaptic currents.

Topics: Animals; Animals, Newborn; Antihypertensive Agents; Benzothiadiazines; Cells, Cultured; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Glutamic Acid; Hippocampus; Neural Inhibition; Neurons; Neurotoxins; Patch-Clamp Techniques; Quinoxalines; Rats; Receptors, AMPA; Receptors, GABA; Synapses; Synaptic Transmission

The aim of this study was to assess whether a drug which combines an antagonistic action at both NMDA and non-NMDA receptors offers advantages for treatment of epileptic seizures compared to drugs which antagonize only one of these ionotropic glutamate receptors. The novel glutamate receptor antagonist LU 73068 (4,5-dihydro-1-methyl-4-oxo-7-trifluoromethylimidazo[1,2a]quinoxal ine-2-carbonic acid) binds with high affinity to both the glycine site of the NMDA receptor (Ki 185 nM) and to the AMPA receptor (Ki 158 nM). Furthermore, binding experiments with recombinant kainate receptor subunits showed that LU 73068 binds to several of these subunits, particularly to rGluR7 (Ki 104 nM) and rGluR5 (Ki 271 nM). In comparison, the prototype non-NMDA receptor antagonist NBQX (2,3-dihydroxy-6-nitro-7-sulphamoyl-benzo[f]quinoxaline) binds with high affinity to AMPA receptors only. Both NBQX and LU 73068 were about equieffective after i.p. injection in mice to block lethal convulsions induced by AMPA or NMDA. In the rat amygdala kindling model of temporal lobe epilepsy, LU 73068 dose-dependently increased the focal seizure threshold (afterdischarge threshold, ADT). When rats were stimulated with a current 20% above the individual control ADT, LU 73068 completely blocked seizures with an ED50 of 4.9 mg kg(-1). Up to 20 mg kg(-1), only moderate adverse effects, e.g. slight ataxia, were observed. NBQX, 10 mg kg(-1), and the glycine/NMDA site antagonist L-701,324 (7-chloro-4-hydroxy-3-(3-phenoxy)phenyl-quinoline-2(1H)one), 2.5 or 5 mg kg(-1), exerted no anticonvulsant effects in kindled rats when administered alone, but combined treatment with both drugs resulted in a significant ADT increase. The data indicate that combination of glycine/NMDA and non-NMDA receptor antagonism in a single drug is an effective means of developing a potent and effective anticonvulsant agent.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Anticonvulsants; Disease Models, Animal; Dizocilpine Maleate; Drug Synergism; Epilepsy; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Imidazoles; Kindling, Neurologic; Male; Mice; Mice, Inbred Strains; N-Methylaspartate; Quinolones; Quinoxalines; Receptors, AMPA; Receptors, Glycine; Receptors, N-Methyl-D-Aspartate; Tritium

Rats neonatally treated with 0.02% propylthiouracil (PTU) through mother's milk showed a high incidence of audiogenic seizures after maturation. These audiogenic seizures were differently modified by MK-801 and NBQX; while intraperitoneal MK-801 equally inhibited running fit (RF) and generalized tonic-clonic seizure (GTCS), NBQX administered into cisterna ambiens significantly inhibited RF but not GTCS. The possible involvement of glutamate receptors in the inferior colliculus was further investigated using naive Sprague-Dawley rats injected with NMDA, AMPA or cyclothiazide, known as an inhibitor of desensitization of AMPA action. All drugs tested successfully induced RF followed by GTCS, resembling audiogenic seizures in PTU-treated rats. However, sound stimulation could augment AMPA-induced, but not NMDA-induced GTCS. Systemic administration with MK-801 potently blocked GTCS induced by AMPA/cyclothiazide, but the same drug failed to block RF after intracisternal injection with AMPA/cyclothiazide. Furthermore, intracisternal administration with NBQX significantly inhibited only RF induced by AMPA/cyclothiazide. The present study suggests that: 1) glutamate receptors in the brainstem, possible in the inferior colliculus, play a crucial role in audiogenic seizures, namely the initiation of RF and propagation into GTCS; and 2) the initiation mechanism is regulated by both NMDA and AMPA receptors, whereas propagation is mainly controlled by NMDA receptors.

Topics: Acoustic Stimulation; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Anticonvulsants; Benzothiadiazines; Brain Stem; Dizocilpine Maleate; Epilepsy, Tonic-Clonic; Excitatory Amino Acid Agonists; Inferior Colliculi; N-Methylaspartate; Neuroprotective Agents; Propylthiouracil; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Seizures

The postsynaptic neuronal dendrite is selectively vulnerable to hypoxic-ischemic brain injury and glutamate receptor overactivation. We explored the glutamate receptor pharmacology and ionic basis of rapid, reversible alterations in dendritic shape which occur in cultured neurons exposed to glutamate. Dendrite morphology was assessed with the fluorescent membrane tracer, DiI, or immunofluorescence labeling of the somatodendritic protein, MAP2. Cortical cultures derived from 15-day-old mouse embryos underwent segmental dendritic beading when exposed to NMDA, AMPA, or kainate, but not to metabotropic glutamate receptor agonists. Varicosity formation in response to NMDA or kainate application was substantially attenuated in reduced sodium buffer (substituted with N-methyl-D-glucamine). Furthermore, veratridine-induced sodium entry mimicked excitotoxic alterations in dendrites and additionally caused varicosity formation in axons. Solutions deficient in chloride (substituted with Na methylsulfate) and antagonists of chloride-permeable GABA/glycine receptors reduced NMDA- or kainate-induced varicosity formation. An increase in dendrite volume was observed as varicosities formed, and varicosity formation was attenuated in sucrose-supplemented hypertonic media. Despite marked structural changes affecting virtually all neurons, dendrite shape returned to normal within 2 h of terminating glutamate receptor agonist application. Neurons exposed to kainate recovered more rapidly than those exposed to NMDA, and neurons exposed to NMDA in calcium-free buffer recovered more rapidly than cells treated with NMDA in normal buffer. While sodium, chloride, and water entry contribute to excitotoxic dendritic injury acutely, calcium entry through NMDA receptors results in lasting structural changes in damaged dendrites.

Topics: Animals; Brain Injuries; Bridged Bicyclo Compounds; Calcium; Cells, Cultured; Chlorides; Coculture Techniques; Cytosol; Dendrites; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Kainic Acid; Mice; Microscopy, Fluorescence; N-Methylaspartate; Quinoxalines; Receptors, Glutamate; Sodium; Veratridine

The effects of the N-methyl-D-aspartate (NMDA) and non-NMDA receptor antagonists MK-801 and NBQX, respectively, on salivary secretion and retching induced by vagal stimulation were studied in decerebrate dogs. Vagal stimulation induced an increase in salivary secretion and fictive retching. Intra-4th ventricular application of vehicle or MK-801 did not change either response, while NBQX completely abolished both responses. These results suggest that non-NMDA receptors mediate both responses in the solitary nucleus.

Topics: Anesthesia; Animals; Decerebrate State; Dizocilpine Maleate; Dogs; Electric Stimulation; Excitatory Amino Acid Antagonists; Lingual Nerve; Phrenic Nerve; Quinoxalines; Reaction Time; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Salivation; Solitary Nucleus; Synaptic Transmission; Vagus Nerve; Vomiting

Effects of the glutamate receptor agonists, N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), on the activator protein-1 (AP-1) DNA binding activity were studied in primary cultures of rat cerebellar granule cells. Application of NMDA as well as of AMPA produced a concentration-dependent enhancement of AP-1 binding. Further examination revealed that only a brief exposure (10 min) to NMDA or AMPA was required for the initiation of a significant, four- to sixfold enhancement of AP-1 DNA binding activity. Blockade of the desensitization of AMPA receptors by cyclothiazide further reduced the exposure time needed to activate the AP-1 complex. The time needed to achieve a maximal increase of AP-1 binding activity varied depending on the glutamate receptor agonist used. NMDA gave maximal AP-1 stimulation after 60 min exposure, whereas stimulation with AMPA alone reached a maximum after 240 min exposure. When AMPA was applied together with cyclothiazide the maximal enhancement of AP-1 binding was reached much faster, within 120 min. Supershift analysis with specific antibodies against the members of Fos and Jun protein families (c-Fos, Fos B, c-Jun, Jun B, Jun D) revealed that the NMDA-induced AP-1 complex was composed predominantly of Jun D and c-Fos. The composition of the AP-1 complex activated by AMPA alone was similar to that produced by NMDA, but with an additional contribution of Fos B. In contrast, application of AMPA plus cyclothiazide induced an AP-1 transcription with contribution of Jun D, c-Fos, Fos B, c-Jun and Jun B proteins. These findings indicate that glutamate is able to enhance AP-1 DNA binding activity in cerebellar granule cells through both NMDA and AMPA glutamate receptors.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Benzothiadiazines; Cells, Cultured; Cerebellum; Dizocilpine Maleate; DNA-Binding Proteins; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Glycine; Kinetics; N-Methylaspartate; Neurons; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Glutamate; Transcription Factor AP-1

Repetitive spreading depression (SD) waves, involving depolarization of neurons and astrocytes and up-regulation of glucose consumption, is thought to lower the threshold of neuronal death during and immediately after ischemia. Using rat models for SD and focal ischemia we investigated the expression of cyclooxygenase-1 (COX-1), the constitutive form, and cyclooxygenase-2 (COX-2), the inducible form of a key enzyme in prostaglandin biosynthesis and the target enzymes for nonsteroidal anti-inflammatory drugs. Whereas COX-1 mRNA levels were undetectable and uninducible, COX-2 mRNA and protein levels were rapidly increased in the cortex, especially in layers 2 and 3 after SD and transient focal ischemia. The cortical induction was reduced by MK-801, an N-methyl-D-aspartic acid-receptor antagonist, and by dexamethasone and quinacrine, phospholipase A2 (PLA2) inhibiting compounds. MK-801 acted by blocking SD whereas treatment with PLA2 inhibitors preserved the wave propagation. NBQX, an alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid/kainate-receptor antagonist, did not affect the SD-induced COX-2 expression, whereas COX-inhibitors indomethacin and diclofenac, as well as a NO synthase-inhibitor, NG-nitro-L-arginine methyl ester, tended to enhance the COX-2 mRNA expression. In addition, ischemia induced COX-2 expression in the hippocampal and perifocal striatal neurons and in endothelial cells. Thus, COX-2 is transiently induced after SD and focal ischemia by activation of N-methyl-D-aspartic acid-receptors and PLA2, most prominently in cortical neurons that are at a high risk to die after focal brain ischemia.

Topics: Animals; Cerebral Cortex; Corpus Striatum; Cortical Spreading Depression; Cyclooxygenase 1; Cyclooxygenase 2; Dexamethasone; Dizocilpine Maleate; Endothelium, Vascular; Enzyme Induction; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Functional Laterality; Hippocampus; Immunohistochemistry; In Situ Hybridization; Ischemic Attack, Transient; Isoenzymes; Male; Membrane Proteins; Neurons; Phospholipases A; Phospholipases A2; Prostaglandin-Endoperoxide Synthases; Quinacrine; Quinoxalines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Transcription, Genetic

Glycine is an inhibitory neurotransmitter in the spinal cord and also acts as a permissive cofactor required for activation of the N-methyl-D-aspartate (NMDA) receptor. We have found that high concentrations of glycine (10 mM) cause marked hyperexcitability and neurotoxicity in organotypic hippocampal slice cultures. The hyperexcitability, measured using intracellular recording in CA1 pyramidal neurons was completely blocked by the NMDA receptor antagonist MK-801 (10 microM), but not by the AMPA receptor antagonist DNQX (100 microM). The neurotoxicity caused by glycine occurred in all regions of hippocampal cultures but was most marked in area CA1. There was significant CA1 neuronal damage in cultures exposed to 10 mM glycine for 30 min or longer (P < 0.01) or those exposed to 4 mM glycine for 24 h compared to control cultures (P < 0.01). The NMDA antagonists MK-801 (10 microM) and APV (100 microM) significantly reduced glycine-induced neuronal damage in all hippocampal subfields (P < 0.01). The AMPA antagonists CNQX, DNQX, and NBQX (100 microM) had no effect on glycine-induced neuronal damage. High concentrations of glycine therefore appear to enhance the excitability of hippocampal slices in an NMDA receptor-dependent manner. The neurotoxic actions of glycine are also blocked by NMDA receptor antagonists.

Topics: 2-Amino-5-phosphonovalerate; 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Animals, Newborn; Benzoxazines; Cell Death; Dizocilpine Maleate; Electrophysiology; Excitatory Amino Acid Antagonists; Glycine; Hippocampus; Neurons; Neurotoxins; Organ Culture Techniques; Oxazines; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

Seizures were induced in immature 18-day-old rats by i.p. administration of homocysteine (11 mmol/kg) and the effects of selected antagonists of NMDA receptors [MK-801 (0.5 mg/kg), AP7 (0.33 mmol/kg), CGP 40116 (10 mg/kg)] and non-NMDA receptors [GDEE (4 mmol/kg), NBQX (two doses, 30 mg/kg each)] were studied. The effect of MgSO4 (two doses, 2 mmol/kg each) was also tested. The anticonvulsant effect was evaluated not only from the behavioral manifestations of seizures, but also in terms of some indicators of brain energy metabolism. Rat pups were sacrificed during generalized clonic-tonic seizures, corresponding to 16-45 min after homocysteine administration. Comparable time intervals were used for sacrificing the pups which had received the protective drugs. In contrast to neonatal rats, in which only NMDA antagonists could prevent homocysteine-induced seizures, both NMDA and non-NMDA receptor antagonists exerted an anticonvulsant effect in 18-day-old rats. In addition, the pronounced anticonvulsant effect could be achieved by the combined treatment with low subthreshold doses of NMDA (MK-801) and non-NMDA (NBQX) receptor antagonists. The protection was evident not only in suppressing behavioral symptoms of seizures, but also in preventing most of the metabolic changes accompanying seizures, mainly glycogen degradation. More than a sevenfold accumulation of lactate occurring during seizures was markedly reduced by all the tested drugs, but was not completely eliminated. All antagonists, when given alone in the same doses as those used for seizure protection, remained without any effect on lactate levels. Comparison of the present data with previous findings concerning neonatal rats suggests that there may be a developmental change in anticonvulsant efficacy of non-NMDA receptor antagonists against homocysteine-induced seizures in rats.

Topics: 2-Amino-5-phosphonovalerate; Age Factors; Animals; Anticonvulsants; Behavior, Animal; Brain Chemistry; Dizocilpine Maleate; Dose-Response Relationship, Drug; Energy Metabolism; Excitatory Amino Acid Antagonists; Glutamates; Homocysteine; Male; Neuroprotective Agents; Quinoxalines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Seizures

The participation of NMDA and non-NMDA receptors in domoic acid-induced neurotoxicity was investigated in cultured rat cerebellar granule cells (CGCs). Neurons were exposed to 300 microM L-glutamate or 10 microM domoate for 2 h in physiologic buffer at 22 degrees C followed by a 22-h incubation in 37 degrees C conditioned growth media. Excitotoxic injury was monitored as a function of time by measurement of lactate dehydrogenase (LDH) activity in both the exposure buffer and the conditioned media. Glutamate and domoate evoked, respectively, 50 and 65% of the total 24-h increment in LDH efflux after 2 h. Hyperosmolar conditions prevented this early response but did not significantly alter the extent of neuronal injury observed at 24 h. The competitive NMDA receptor antagonist D(-)-2-amino-5-phosphonopentanoic acid and the non-NMDA receptor antagonist 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(f)quinoxaline (NBQX) reduced glutamate-induced LDH efflux totals by 73 and 27%, respectively, whereas, together, these glutamate receptor antagonists completely prevented neuronal injury. Domoate toxicity was reduced 65-77% when CGCs were treated with competitive and noncompetitive NMDA receptor antagonists. Unlike the effect on glutamate toxicity, NBQX completely prevented domoate-mediated injury. HPLC analysis of the exposure buffer revealed that domoate stimulates the release of excitatory amino acids (EAAs) and adenosine from neurons. Domoate-stimulated EAA release occurred almost exclusively through mechanisms related to cell swelling and reversal of the glutamate transporter. Thus, whereas glutamate-induced injury is mediated primarily through NMDA receptors, the full extent of neurodegeneration is produced by the coactivation of both NMDA and non-NMDA receptors. Domoate-induced neuronal injury is also mediated primarily through NMDA receptors, which are activated secondarily as a consequence of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA)/kainate receptor-mediated stimulation of EAA efflux.

Topics: Animals; Cells, Cultured; Cerebellum; Culture Media, Conditioned; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Excitatory Amino Acids; Glutamic Acid; Kainic Acid; Kinetics; L-Lactate Dehydrogenase; Neurons; Neurotoxins; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Tetrodotoxin

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

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

L-Glutamate, N-methyl-D-aspartate, DL-alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) and kainate increased the release of neuropeptide Y-like immunoreactivity from primary cultures of rat hippocampal neurons incubated in Mg2+(1.2 mM)-containing medium. The neuropeptide Y-like immunoreactivity released by 100 microM glutamate was mainly accounted for by neuropeptide Y (1-36), but consisted in part (about 20%) of peptide YY. The effect of 100 microM glutamate on neuropeptide Y-like immunoreactivity release was largely (about 70%) prevented by the N-methyl-D-aspartate receptor antagonist dizocilpine maleate (10 microM), while the remainder (about 30%) was sensitive to the AMPA/ kainate receptor antagonist 6-nitro-7-sulphamoylbenzo(f)quinoxaline-2-3-dione (10 microM). The AMPA(100 microM)-evoked release of neuropeptide Y-like immunoreactivity was strongly antagonized by 6-nitro-7-sulphamoylbenzo(f)quinoxaline-2-3-dione and by 1-aminophenyl-4-methyl-7,8-methylenedioxy-5H-2,3-benzodiazepine, but it was in part (15-20%) sensitive to dizocilpine. The releases of neuropeptide Y-like immunoreactivity elicited by glutamate, N-methyl-D-aspartate, AMPA and kainate were all strongly Ca(2+)-dependent. Tetrodotoxin (1 microM) abrogated the N-methyl-D-aspartate-evoked release and partly inhibited the release caused by glutamate, but did not modify significantly AMPA- or kainate-evoked release. Removal of Mg2+ from the medium caused increase of neuropeptide Y-like immunoreactivity release, an effect prevented by dizocilpine maleate or 7-Cl-kynurenate. Cyclothiazide (10 microM), a drug known to prevent AMPA receptor desensitization, enhanced the neuropeptide Y-like immunoreactivity release elicited by 100 microM AMPA, but not that caused by 100 microM kainate. However, when used at a lower concentration (50 microM), kainate elicited a response that was potentiated significantly by cyclothiazide. It is concluded that glutamate can stimulate Ca(2+)-dependent release of neuropeptide Y from hippocampal neurons mainly through N-methyl-D-aspartate receptors and, less so, by activating cyclothiazide-sensitive receptors of the AMPA-preferring type.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Anti-Anxiety Agents; Antibody Specificity; Antihypertensive Agents; Benzodiazepines; Benzothiadiazines; Calcium; Cells, Cultured; Chromatography, High Pressure Liquid; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Glutamic Acid; Hippocampus; Kainic Acid; Magnesium; Neurons; Neuropeptide Y; Quinoxalines; Radioimmunoassay; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Tetrodotoxin

Neurotoxicity mediated by glutamate is thought to play a role in neurodegenerative disorders, and alterations in cytoskeletal proteins are possibly involved in the mechanisms of neuronal death occurring in Alzheimer's disease. In the present work we studied the neurotoxic effects of the intrahippocampal injections of the glutamate transport inhibitor dihydrokainate as compared to those of kainate, as well as the concomitant changes in the microtubule-associated protein MAP2. Neuronal alterations were assessed at 3, 12, 24, and 48 h by Nissl staining and immunocytochemistry of MAP2. At 3 h, both compounds induced neuronal damage that was correlated with loss of dendritic MAP2 immunoreactivity. Neuronal damage was more evident at 12 h and 24 h after drug injection, and at these times an accumulation of MAP2 in the somata of pyramidal neurons was observed. The effects of dihydrokainate were restricted to the CA1 region and totally prevented by the N-methyl-D-aspartate receptor antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801), but not by the non-NMDA receptor antagonist 2,3-dihydro-6-nitro-7-sulphamoyl-benzo(f)-quinoxaline (NBQX). In contrast, kainate-induced alterations included CA1, CA3, and CA4 subfields, and the changes in CA1 were prevented by NBQX, while MK-801 was ineffective. These results suggest that early MAP2 disruption may be a marker of the excitotoxicity due to activation of different glutamate receptors located in discrete hippocampal regions.

Topics: Animals; Biological Transport; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Hippocampus; Immunohistochemistry; Kainic Acid; Male; Microtubule-Associated Proteins; Neurons; Quinoxalines; Rats; Rats, Wistar

1. Nitric oxide (NO) synthase activity was studied in slices of human temporal cortex samples obtained in neurosurgery by measuring the conversion of L-[3H]-arginine to L-[3H]-citrulline. 2. Elevation of extracellular K+ to 20, 35 or 60 mM concentration-dependently augmented L-[3H]-citrulline production. The response to 35 mM KCl was abolished by N(G)-nitro-L-arginine (100 microM) demonstrating NO synthase specific conversion of L-arginine to L-citrulline. Increasing extracellular MgCl2 concentration up to 10 mM also prevented the K+ (35 mM)-induced NO synthase activation, suggesting the absolute requirement of external calcium ions for enzyme activity. 3. However, the effect of high K+ (35 mM) on citrulline synthesis was insensitive to the antagonists of ionotropic and metabotropic glutamate receptors dizocilpine (MK-801), 6-nitro-7-sulphamoylbenzo(f)-quinoxaline-2-3-dione (NBQX) or L-2-amino-3-phosphonopropionic acid (L-AP3) as well as to the nicotinic receptor antagonist, mecamylamine. 4. The 35 mM K+ response was insensitive to omega-conotoxin GVIA (1 microM) and nifedipine (100 microM), but could be prevented in part by omega-agatoxin IVA (0.1 and 1 microM). The inhibition caused by 0.1 microM omega-agatoxin IVA (approximately 30%) was enhanced by adding omega-conotoxin GVIA (1 microM) or nifedipine (100 microM). Further inhibition (up to above 70%) could be observed when the three Ca2+ channel blockers were added together. Similarly, synthetic FTX 3.3 arginine polyamine (sFTX) prevented (50% at 100 microM) the K+-evoked NO synthase activation. This effect of sFTX was further enhanced (up to 70%) by adding 1 microM omega-conotoxin GVIA plus 100 microM nifedipine. No further inhibition could be observed upon addition of MK-801 or/and NBQX. 5. It was concluded that elevation of extracellular [K+] causes NO synthase activation by external Ca2+ entering cells mainly through channels of the P/Q-type. Other Ca2+ channels (L- and N-type) appear to contribute when P/Q-channels are blocked.

Topics: Alanine; Arginine; Calcium Channel Blockers; Calcium Channels; Citrulline; Dizocilpine Maleate; Dose-Response Relationship, Drug; Enzyme Activation; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Humans; In Vitro Techniques; Mecamylamine; Nicotinic Antagonists; Nitric Oxide Synthase; Nitroarginine; Potassium; Potassium Chloride; Quinoxalines; Temporal Lobe

We have studied in a well-characterized in vitro neuronal system, cultures of cerebellar granule cells, the toxicity of polyamines endogenously present in the brain: spermine, spermidine, and putrescine. Twenty-four-hour exposure of mature (8 days in vitro) cultures to 1-500 microM spermine resulted in a dose-dependent death of granule cells, with the half-maximal effect being reached below 50 microM concentration. Putrescine was moderately toxic but only at 500 microM concentration. Spermidine was tested at 50 and 100 microM concentration and its toxicity was evaluated to be about 50% that of spermine. Neuronal death caused by spermine occurred, at least in part, by apoptosis. Spermine toxicity was completely prevented by competitive (CGP 39551) and noncompetitive (MK-801) antagonists of the NMDA receptor, but was unaffected by a non-NMDA antagonist (NBQX) or by antagonists of the polyamine site present on the NMDA receptor complex, such as ifenprodil. A partial protection from spermine toxicity was obtained through the simultaneous presence of free radical scavengers or through inhibition of the free radical-generating enzyme nitric oxide synthase, known to be partially effective against direct glutamate toxicity. The link between spermine toxicity and glutamate was further strengthened by the fact that, under culture conditions in which glutamate toxicity was ineffective or much reduced, spermine toxicity was absent or very much decreased. Exposure to spermine was accompanied by a progressive accumulation of glutamate in the medium of granule cell cultures. This was attributed to glutamate leaking out from dying or dead cells and was substantially prevented by the simultaneous presence of MK-801 or CGP 39551. The present results demonstrate that polyamines are toxic to granule cells in culture and that this toxicity is mediated through the NMDA receptor by interaction of exogenously added polyamines with endogenous glutamate released by neurons in the medium. The involvement of brain polyamines, in particular spermine and spermidine, in excitotoxic neuronal death is strongly supported by our present results.

Topics: 2-Amino-5-phosphonovalerate; Animals; Apoptosis; Aspartic Acid; Butylated Hydroxytoluene; Cells, Cultured; Cerebellar Cortex; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Female; Free Radical Scavengers; Glutamic Acid; L-Lactate Dehydrogenase; Male; Nerve Tissue Proteins; Neurons; Neuroprotective Agents; Nitric Oxide Synthase; Nitroarginine; Piperidines; Putrescine; Quinoxalines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Spermidine; Spermine; Vitamin E

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

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

Ion-selective microelectrodes were used to study acute effects of N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy- 5-methyl-4-isoxazole (AMPA) receptor blockade on posttraumatic calcium disturbances. An autoradiographic technique with 45 Ca2+ was used to study calcium disturbances at 8, 24, and 72 h. Compression contusion trauma of the cerebral cortex was produced by a 21-g weight dropped from a height of 35 cm onto a piston that compressed the brain 2 mm. Pre- and posttrauma interstitial [Ca2+] ([Ca2+]e) concentrations were measured in the perimeter, i.e., the shear stress zone (SSZ) and in the central region (CR) of the trauma site. For the [Ca2+]e studies the animals were divided into controls and groups pretreated with dizocilipine maleate (MK-801) or with 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo[F]quinoxaline (NBQX). In all groups, [Ca2+]e decreased from pretrauma values of approximately 1 mM to posttraumatic values of 0.1 mM in both the CR and the SSZ. This was followed by a slow restitution toward pretraumatic levels during the 2-h observation period. There was no significant difference in recovery pattern between controls and pretreated animals. Accumulation of 45Ca2+ and serum proteins was seen in the entire SSZ, while neuronal necrosis was confined to a narrow band within the SSZ. The CR was unaffected apart from occasional eosinophilic neurons and showed no accumulation of 45Ca2+. Posttraumatic treatment with MK-801 or NBQX had no obvious effect on neuronal injury in the SSZ. We conclude that (a) acute [Ca2+]e disturbances in compression contusion brain trauma are not affected by blockade of NMDA or AMPA receptors, (b) 45Ca2+ accumulation in the SSZ reflects mainly protein accumulation due to blood-brain barrier breakdown rather than cell death, and (c) acute cellular Ca2+ over-load per se does not seem to be a major determinant of cell death after cerebral trauma in our model.

Topics: Animals; Autoradiography; Brain Injuries; Calcium; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Ion Channels; Male; Necrosis; Neurons; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate

3-Nitropropionic acid (3-NPA) is a metabolic poison that produces lesions of striatal intrinsic neurones such as gamma-aminobutyric acid (GABA) neurones. This study was carried out to determine whether 3-NPA would impair the ability of striatal GABAergic neurones to withstand hypoglycaemic stress. 3-NPA (500 microM) did not affect [3H]GABA release from striatal slices under normal (11 mM) glucose concentrations. When the glucose concentration was lowered to 0.3 mM, however, 3-NPA greatly potentiated the leakage of [3H]GABA from the slices. Blockage of N-methyl-D-aspartate (NMDA) or alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors with 1 microM 5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine (MK-801) or 10 microM 2,3-dihydroxy-6-nitro-7-sulpha-moylbenzo[F]quinoxaline (NBQX), respectively, or a combination of both, had no effect. However, blockade of voltage-dependent sodium channels with tetrodotoxin totally antagonized the [3H]GABA overflow induced by the combination of 3-NPA and hypoglycaemis. Riluzole (10 to 100 microM), a neuroprotective agent that stabilizes the inactivated state of the voltage-dependent sodium channel, also dose-dependently antagonized the increase in [3H]GABA release induced by the combination of the two stresses.

Topics: Animals; Corpus Striatum; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; gamma-Aminobutyric Acid; Glucose; Hypoglycemia; Isotope Labeling; Male; Neurons; Neuroprotective Agents; Neurotoxins; Nitro Compounds; Propionates; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Regression Analysis; Sodium Channels; Tritium

Recent evidence implicates the endogenous excitatory neurotransmitters, glutamate (Glu) and aspartate, in the pathophysiology of traumatic injury in the adult CNS, but it is not known whether similar excitotoxic mechanisms mediate traumatic injury in the immature CNS. Therefore, we developed a model of brain contusion injury in infant rats and used this model to study the nature and evolution of the acute cytopathologic changes and to evaluate the ability of Glu receptor antagonists to protect the immature brain against such changes. Seven-day-old rat pups were subjected to contusion injury and were killed 0, 0.5, 1, 2, 4, and 6 h later for histologic evaluation of the brain. Physical tearing of the dura and minor disruption of underlying brain tissue was noted at 0 h. At 30 min a discrete zone of neuronal necrosis began to appear at the border of the trauma site; this zone progressively expanded over a period of 4 h. The cytopathologic changes closely resembled the type of changes Glu is known to cause; these changes consisted of swollen dendrites, degenerating neurons with pyknotic nuclei and markedly swollen cytoplasm, and dark cells with vacuolated cytoplasm. The noncompetitive N-methyl-D-aspartate (NMDA) antagonist, dizocilpine maleate, when administered 30 min before or 1 h after trauma, significantly attenuated the lesion. The competitive NMDA antagonist, 3-((-2)-carboxypiperazine-4-yl)-propyl-1-phosphonate, was also neuroprotective. The alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate/kainate receptor antagonist 2,3-dihydro-6-nitro-7-sulfamoyl-benzo(f)quinoxaline did not significantly suppress the lesion when given as three treatments (30 mg/kg each) 30 min before plus 15 and 75 min after the insult. These findings suggest that traumatic injury in the infant rat brain is mediated by endogenous excitotoxins (Glu and aspartate) acting at NMDA receptors and can be substantially mitigated by timely treatment with NMDA receptor antagonists.

Topics: Animals; Brain Injuries; Dendrites; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Piperazines; Quinoxalines; Rats; Rats, Sprague-Dawley; Time Factors; Wounds and Injuries

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

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

The spinal cord dorsal horn contains neural mechanisms which can greatly facilitate pain. It is well established that excitatory amino acids, aspartate and glutamate, are involved in the spinal transmission of nociceptive information and in the development of hyperalgesia. In the present study, intrathecal (i.t.) administration of alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA), a structural analog of L-glutamate, produced a dose-dependent behavioural syndrome characterized by caudally directed biting in mice. We demonstrated that peripheral pre-administration of the AMPA receptor antagonists 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(F)quinoxaline (NBQX, 10-100 mg/kg s.c.) and 1-(4-aminophenyl)-3-methylcarbamoyl-4-methyl-3,4-dihydro-7, 8-methylene-dioxy-5H-2,3-benzo-diazepine-HCl (GYKI 53655, 3-10 mg/kg s.c.), and also of the NMDA receptor antagonist 5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5, 10-imine maleate (MK 801, 0.3-1 mg/kg s.c.) reversed this effect. These findings suggest that the hyperalgesia induced by the i.t. injection of AMPA in mice involves the activation of both NMDA and non-NMDA excitatory amino acid receptor sites.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Behavior, Animal; Benzodiazepines; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Hyperalgesia; Injections, Spinal; Male; Mice; Mice, Inbred Strains; Nociceptors; Quinoxalines; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate

To evaluate the contribution of apoptotic mechanisms to excitotoxin-induced neurodegeneration as well as to characterize the glutamate receptor subtypes involved, biochemical and morphological effects of intrastriatally administered NMDA receptor agonist N-methyl-D-aspartate (NMDA) or quinolinic acid (QA) were studied. Receptor autoradiography showed that NMDA (75-300 nmol) caused a loss of 18-68% of striatal D1 dopamine (DA) and 10-43% of NMDA receptors 7 days after drug administration. Treatment with QA (60-240 nmol) also led to a loss of 60-73% of D1 DA and 37-44% of NMDA receptors in the ipsilateral striatum. Agarose gel electrophoresis revealed that both NMDA and QA induced internucleosomal DNA fragmentation in the striatum 12 to 48 h after drug administration. NMDA- and QA-induced internucleosomal DNA fragmentation was attenuated by the protein synthesis inhibitor cycloheximide in a dose-dependent manner. Terminal transferase-mediated deoxyuridine triphosphate (d-UTP)-digoxigenin nick end labeling (TUNEL)-positive nuclei were found in the ipsilateral striatum in response to NMDA or QA treatment. In addition, many fragmented nuclei were observed in the NMDA or QA-treated striatum and propidium iodide staining showed profound nuclear condensation in the NMDA or QA-treated striatum. NMDA- and QA-induced internucleosomal DNA fragmentation and TUNEL-positive nuclei as well as nuclear condensation were abolished by the NMDA receptor antagonist MK-801, but not by the AMPA/KA receptor antagonist NBQX. MK-801, but not NBQX, also prevented NMDA or QA-induced striatal cell death. These results suggest that apoptotic mechanisms are involved in excitotoxin-induced striatal cell death. The initiation of an apoptotic cascade by NMDA or QA appears to be mediated by stimulation of NMDA but not AMPA/KA receptors.

Topics: Animals; Apoptosis; Autoradiography; Deoxyuracil Nucleotides; Dizocilpine Maleate; DNA Fragmentation; DNA Nucleotidylexotransferase; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Genetic Techniques; Male; Nucleic Acid Hybridization; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Stimulation, Chemical

Because of the critical role of excitatory amino acids (EAAs) in epileptogenesis and seizure-induced brain damage, EAA antagonists are now being considered as a possible therapy for seizures. However, during development EAAs play a pivotal role in learning, memory, and brain plasticity. To evaluate the long-term effects of a short course of EAA antagonists on the developing brain, a non-NMDA antagonist, NBQX, or a NMDA antagonist, MK-801, were administered over 7 days by osmotic pumps stereotaxically implanted into the lateral ventricles of normal 10 day old rats. Alternatively, 10 and 20 day old rats received a 7 day course of intraperitoneal (i.p.) NBQX. One month later, the NBQX-, MK-801-treated rats, and controls underwent a series of behavioral studies: handling test, open field, and Morris water maze. Flurothyl inhalation was used to test seizure susceptibility in all groups. Although all of the rats treated with NBQX via osmotic pumps has spontaneous seizure, rats surviving infusion of EAAs had no deficits in learning, memory, or behavior and did not differ from controls in seizure susceptibility with flurothyl. In the developing animal, a short-term course of EAA antagonists leads to no long-term adverse effects on behavior or seizure susceptibility.

Topics: Animals; Animals, Newborn; Brain; Dizocilpine Maleate; Evaluation Studies as Topic; Excitatory Amino Acid Antagonists; Exploratory Behavior; Infusion Pumps, Implantable; Injections, Intraperitoneal; Male; Maze Learning; Neuroprotective Agents; Pilot Projects; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Seizures

The effects of NBQX (1.56-7.5 mg/kg, i.p.), a competitive antagonist at the AMPA type of glutamate receptor, were studied in two operant behavioural paradigms, differential reinforcement of low response rates (DRL), and delayed matching to position (DMTP), which have been shown to be sensitive to the antagonists of the NMDA type of glutamate receptor. Additionally, the non-competitive AMPA antagonist, GYKI 52466 (7.5-15 mg/kg, i.p.), was studied in the DRL procedure. As a positive control, the non-competitive NMDA antagonist, MK 801 (0.0125-0.1 mg/kg, i.p.) was studied in both procedures. During performance of the DRL schedule, MK 801 increased response rates in a dose dependent manner, and decreased the number of reinforcers obtained. The increase in response rates could be attributed to both a shift in the median inter-response time (IRT) to shorter intervals, and to a marked, dose dependent increase in the occurrence of bursts of responses (responses occurring within 3 s of a previous response). In contrast, NBQX and GYKI 52466 both decreased response rates in a dose dependent fashion, and did not shift the distribution of the IRTs, or increase the occurrence of burst responding. In the DMTP procedure, accuracy of matching decreased with increasing delay (up to 30 s, between presentation of sample and opportunity to respond). NBQX disrupted responding at a dose of 7.5 mg/kg, but lower doses were ineffective in influencing accuracy of performance of the discrimination. In contrast, MK 801 (0.1 and 0.2 mg/kg) reduced accuracy of matching at all delays, while tending to increase the speed of responding. These data demonstrate differences in the effects of AMPA and NMDA antagonists on performance of well trained operant behaviour.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Anti-Anxiety Agents; Benzodiazepines; Conditioning, Operant; Discrimination, Psychological; Dizocilpine Maleate; Dose-Response Relationship, Drug; Male; N-Methylaspartate; Quinoxalines; Rats; Rats, Wistar; Reinforcement, Psychology

A chronic allodynia-like response to mechanical stimulation was observed in rats after severe spinal cord ischemia. This allodynia-like response was not relieved by most conventional analgesics used for treating chronic neuropathic pain. The present experiments evaluated the effects of systemically administered excitatory amino acid receptor antagonists, including the non-competitive N-methyl-D-aspartate (NMDA) receptor/channel blockers MK-801 and dextromethorphan, the competitive NMDA receptor antagonist CGS 19755 and a competitive antagonist of the alpha-amino-3-hydroxyl-5-methyl-4-isoxazolepropionic acid (AMPA) receptor NBQX, on the chronic allodynia-like response in spinally injured rats. Systemic MK-801, dextromethorphan and CGS 19755 dose-dependently relieved the mechanical allodynia-like response. Systemic MK-801 and CGS 19755, but not dextromethorphan, also induced severe motor impairment at analgesic doses. All three NMDA antagonists increased spontaneous motor activity. Systemic NBQX reduced muscle tone and caused sedation. The mechanical allodynia was only relieved by NBQX at a sedative dose. It is concluded that systemic NMDA, but not AMPA, receptor antagonists may have an analgesic effect upon the chronic allodynia-like response. However, the analgesic effect of all NMDA antagonists was associated with side effects. Dextromethorphan, which is clinically tolerated and produced less side effects, may be useful for treating chronic pain associated with central nervous system injury.

Topics: Animals; Behavior, Animal; Chronic Disease; Dextromethorphan; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Female; Hyperalgesia; Ischemia; Motor Activity; Pipecolic Acids; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Regional Blood Flow; Spinal Cord; Spinal Cord Injuries

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

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

The purpose of the present study was to examine the relationship between elevation in intracellular Ca2+ concentration ([Ca2+]i) and development of neuronal damage after cytotoxic hypoxia in vitro. Chick telencephalic neurons were exposed to NaCN 1 mM for up to 2 h. [Ca2+]i was assessed by means of fura-2 based microfluorometry and viability was measured by means of trypan blue exclusion on the same relocated cells for a period 24 h after initiation of hypoxia. Exposure to sodium cyanide resulted in an up to 10 fold increase in [Ca2+]i and led to subsequent neuronal damage. According to [Ca2+]i and viability neurons in four different stages could be revealed. The percentage of neurons showing elevated [Ca2+]i paralleled exactly the percentage of neuronal damage. The elevation in [Ca2+]i clearly preceded neuronal damage suggesting a time window for pharmacological intervention. The NMDA antagonists dizocilpine, memantine and amantadine were capable of reducing the percentage of neurons showing elevated [Ca2+]i and attenuated neuronal damage. Dizocilpine proved to be the most potent and amantadine to be the weakest antagonist whereas the alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionate (AMPA) antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo-(F)-quinoxaline (NBQX) was ineffective. Under our experimental conditions, measurement of [Ca2+]i was able to predict the extent of neuronal damage as well as the neuroprotective potency of drugs.

Topics: Amantadine; Analysis of Variance; Animals; Biomarkers; Calcium; Cell Hypoxia; Cell Survival; Cells, Cultured; Chickens; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Kinetics; Memantine; N-Methylaspartate; Neurons; Neuroprotective Agents; Quinoxalines; Sodium Cyanide; Telencephalon; Time Factors

Cerebral pentose phosphate pathway (PPP) plays a role in the biosynthesis of macromolecules, antioxidant defense and neurotransmitter metabolism. Studies on this potentially important pathway have been hampered by the inability to easily quantitate its activity, particularly in vivo. In this study we review the use of [1,6-13C2,6,6-2H2]glucose for measuring the relative activities of the PPP and glycolysis in a single incubation in cultured neurons and in vivo, when combined with microdialysis techniques. PPP activity in primary cerebrocortical cultures and in the caudate putamem of the rat in vivo was quantitated from data obtained by GC/MS analysis of released labeled lactate following metabolic degradation of [1,6-13C2,6,6-2H2]glucose. Exposure of cultures to H2O2 resulted in stimulation of PPP activity in a concentration-dependent fashion and subsequent cell death. Chelation of iron during H2O2 exposure exerted a protective effect thus implicating the participation of the Fenton reaction in mediating damage caused by the oxidative insult. Partial inhibition of glutathione peroxidase, but not catalase, was extremely toxic to the cultures reflecting the pivotal role of GPx in H2O2 detoxification. These results demonstrate the ability to dynamically monitor PPP activity and its response to oxidative challenges and should assist in facilitating our understanding of antioxidant pathways in the CNS.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Carbon Isotopes; Caudate Nucleus; Cells, Cultured; Cerebral Cortex; Deferoxamine; Deuterium; Dizocilpine Maleate; Fetus; Gas Chromatography-Mass Spectrometry; Glucose; Hydrogen Peroxide; Iron; Kainic Acid; Kinetics; N-Methylaspartate; Neuroglia; Neurons; Pentose Phosphate Pathway; Putamen; Quinoxalines; Rats

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

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

The effects of the AMPA-receptor antagonists NBQX and GYKI 52466 were compared with those of the NMDA-receptor channel blocker dizocilpine in two drug discrimination tests. In the first, rats were trained to discriminate morphine (2 mg/kg) from saline and in the second, to discriminate ketamine (7 mg/kg) from saline, using a two-lever food reinforced method. NBQX (1-6 mg/kg) did not substitute for either morphine or ketamine, even at a dose which reduced response rates (6 mg/kg). Likewise, the noncompetitive antagonist GYKI 52466 (5 and 10 mg/kg) produced only saline lever responding in the ketamine trained rats. When tested in combination with the training drug, NBQX (4.5 mg/kg) did not alter the morphine generalisation gradient, and similarly, neither NBQX (3 mg/kg) nor GYKI 52466 (5 and 10 mg/kg) interacted with the ketamine cue. In contrast, dizocilpine (0.05 mg/kg) significantly disrupted discrimination of morphine and produced clear drug lever responding (0.0125-0.1 mg/kg) in ketamine trained rats. These results suggest that AMPA-receptor antagonists and non-competitive NMDA-antagonists have different stimulus properties.

Topics: Analysis of Variance; Animals; Anti-Anxiety Agents; Behavior, Animal; Benzodiazepines; Conditioning, Operant; Discrimination Learning; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Ketamine; Male; Morphine; Narcotic Antagonists; Quinoxalines; Rats; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Reinforcement Schedule

A disturbed cellular calcium homeostasis is suggested to play a pivotal role in neuronal damage. Energy deficiency causes depolarization of the neuronal membrane and Ca2+ enters the cells through different ion channels, the voltage-sensitive L-type Ca2+ channels and the NMDA-operated channels being the main gates. In the present study we used primary cultures of rat hippocampal neurons to demonstrate that the dihydropyridine calcium antagonist nimodipine, the non-competitive NMDA antagonists dizocilpine and memantine, as well as the AMPA antagonist NBQX (2,3-dihydroxy-6-nitro -7-sulfamoyl-benzo(F)quinoxaline), attenuate the glutamate-induced neuronal damage dose-dependently. Nimodipine applied simultaneously with NMDA-antagonists and NBQX, respectively, resulted in somewhat greater neuroprotection of glutamate-treated neurons compared with the effects of these agents applied singly. The type of interaction is best described by an independent action in combination, which means that the relative effects of nimodipine were not enhanced. Therefore, it can be considered as a lack of potentiation.

Topics: Animals; Cells, Cultured; Dizocilpine Maleate; Drug Interactions; Excitatory Amino Acid Antagonists; Glutamic Acid; Hippocampus; Memantine; Neurons; Neurotoxins; Nimodipine; Quinoxalines; Rats; Rats, Inbred F344

Although the mechanism of neuronal death in neurodegenerative diseases remains unknown, it has been hypothesized that relatively minor metabolic defects may predispose neurons to N-methyl-D-aspartate (NMDA) receptor-mediated excitotoxic damage in these disorders. To further investigate this possibility, we have characterized the excitotoxic potential of the reversible succinate dehydrogenase (SDH) inhibitor malonate. After its intrastriatal stereotaxic injection into male Sprague-Dawley rats, malonate produced a dose-dependent lesion when assessed 3 days after surgery using cytochrome oxidase histochemistry. This lesion was attenuated by coadministration of excess succinate, indicating that it was caused by specific inhibition of SDH. The lesion was also prevented by administration of the noncompetitive NMDA antagonist MK-801. MK-801 did not induce hypothermia, and hypothermia itself was not neuroprotective, suggesting that the neuroprotective effect of MK-801 was due to blockade of the NMDA receptor ion channel and not to any nonspecific effect. The competitive NMDA antagonist LY274614 and the glycine site antagonist 7-chlorokynurenate also profoundly attenuated malonate neurotoxicity, further indicating an NMDA receptor-mediated event. Finally, the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) antagonist NBQX (2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(f)-quinoxaline) was ineffective at preventing malonate toxicity at a dose that effectively reduced S-AMPA toxicity, indicating that non-NMDA receptors are involved minimally, if at all, in the production of the malonate lesion. We conclude that inhibition of SDH by malonate results in NMDA receptor-mediated excitotoxic neuronal death. If this mechanism of "secondary" or "weak" excitotoxicity plays a role in neurodegenerative disease, NMDA antagonists and other "antiexcitotoxic" strategies may have therapeutic potential for these diseases.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Body Temperature; Brain Diseases; Cell Death; Corpus Striatum; Dizocilpine Maleate; Dose-Response Relationship, Drug; Glutamic Acid; Glycine; Isoquinolines; Male; Malonates; Neurons; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Succinate Dehydrogenase

High doses of 3,4-methylenedioxymethamphetamine (MDMA) have been shown to cause long-lasting depletions of central serotonin (5-HT) which are indicative of neuronal toxicity. The noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist dizocilpine (DZ) attenuates depletions of 5-HT induced by MDMA. Because DZ has been shown to induce hypothermia in rat models of ischemia, the purpose of this study was to assess whether DZ and two other glutamate antagonists, CGS 19755 (CGS) and 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX), protect against MDMA-induced 5-HT depletions by induction of hypothermia. Male Sprague-Dawley rats were injected with either saline (SAL), DZ (2.5 mg/kg), CGS (25.0 or 50.0 mg/kg x 2 injections) or NBQX (30.0 mg/kg x 2 injections or 55.0 mg/kg x 3 injections) followed by either MDMA (40.0 mg/kg) or SAL. Core body temperature (TEMP) was monitored for 4 h or longer using radiotelemetry. Base-line TEMP was between 37.0 and 37.6 degrees C. Administration of DZ with MDMA decreased TEMP to 34.0 +/- 0.39 degrees C within 2 h of the MDMA injection, and also protected against serotonergic toxicity. Neither SAL/MDMA nor DZ/SAL had an effect on TEMP over the same period. When rats were treated with DZ/MDMA and TEMP was maintained between 38.4 degrees C and 40.4 degrees C for 4 h, protection against 5-HT depletion was abolished. Coadministration of the competitive NMDA antagonist CGS with MDMA-resulted in a decrease in TEMP to 34.5 +/- 0.27 degrees C, and provided partial protection against 5-HT depletions.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Body Temperature; Brain Chemistry; Dizocilpine Maleate; Hydroxyindoleacetic Acid; Male; N-Methyl-3,4-methylenedioxyamphetamine; Pipecolic Acids; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Serotonin

We report that a subtoxic dose of the succinate dehydrogenase (SDH) inhibitor malonate greatly enhances the neurotoxicity of three different excitatory amino acid agonists: N-methyl-D-aspartate (NMDA), S-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (S-AMPA), and L-glutamate. In male Sprague-Dawley rats, intrastriatal stereotaxic injection of malonate alone (0.6 mumol), NMDA alone (15 nmol), S-AMPA alone (1 nmol), or glutamate alone (0.6 mumol) produced negligible toxicity as assessed by measurement of lesion volume. Coinjection of subtoxic malonate with NMDA produced a large lesion (15.2 +/- 1.4 mm3), as did coinjection of malonate with S-AMPA (11.0 +/- 1.0 mm3) or glutamate (12.8 +/- 0.7 mm3). Administration of the noncompetitive NMDA antagonist MK-801 (5 mg/kg i.p.) completely blocked the toxicity of malonate plus NMDA (0.5 +/- 0.3 mm3). This dose of MK-801 had little effect on the lesion produced by malonate plus S-AMPA (9.0 +/- 0.7 mm3), but it attenuated the toxicity of malonate plus glutamate by approximately 40% (7.5 +/- 0.9 mm3). Coinjection of the AMPA antagonist 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(f)-quinoxaline (NBQX; 2 nmol) had no effect on malonate plus NMDA or malonate plus glutamate toxicity (12.3 +/- 1.8 and 14.0 +/- 0.9 mm3, respectively) but greatly attenuated malonate plus S-AMPA toxicity (1.5 +/- 0.9 mm3). Combination of the two antagonists conferred no additional neuroprotection in any paradigm. These results indicate that metabolic inhibition exacerbates both NMDA receptor- and non-NMDA receptor-mediated excitotoxicity.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Brain; Brain Diseases; Dizocilpine Maleate; Drug Synergism; Glutamic Acid; Male; Malonates; N-Methylaspartate; Quinoxalines; Rats; Rats, Sprague-Dawley; Succinate Dehydrogenase

Anoxic depolarization (AD) and failure of ion homeostasis play an important role in ischemia-induced neuronal injury. In the present study, different drugs with known ion-channel-modulating properties were examined for their ability to interfere with cardiac-arrest-elicited AD and with the changes in the extracellular ion activity in rat brain. Our results indicate that only drugs primarily blocking membrane Na+ permeability (NBQX, R56865, and flunarizine) delayed the occurrence of AD, while compounds affecting cellular Ca2+ load (MK-801 and nimodipine) did not influence the latency time. The ischemia-induced [Na+]e reduction was attenuated by R56865. Blockade of the ATP-sensitive K+ channels with glibenclamide reduced the [K+]e increase upon ischemia, indicating an involvement of the KATP channels in ischemia-induced K+ efflux. The KATP channel opener cromakalim did not affect the AD or the [K+]e concentration. The ischemia-induced rapid decline of extracellular calcium was attenuated by receptor-operated Ca2+ channel blockers MK-801 and NBQX, but not by the voltage-operated Ca2+ channel blocker nimodipine, R56865, and flunarizine.

Topics: Adenosine Triphosphate; Animals; Benzothiazoles; Calcium; Dizocilpine Maleate; Flunarizine; Glyburide; Heart Arrest; Hypoxia; Ion Channels; Male; Nimodipine; Piperidines; Potassium; Potassium Channels; Quinoxalines; Rats; Rats, Wistar; Sodium; Thiazoles

Clinically, and in experimental models, perinatal hypoxic encephalopathy is commonly associated with seizures. We previously described a rat model in which hypoxia induces seizures and permanently increases in seizure susceptibility in immature rats [postnatal day (P) 10-12] but not in older rats. In the present study, we compared the effect of pretreatment with the excitatory amino acid antagonists MK-801 and NBQX versus lorazepam in our rat model of perinatal hypoxia. Animals exposed to hypoxia at P10 without treatment have frequent seizures during hypoxia and subsequently exhibit increased seizure susceptibility to flurothyl. Treatment with 6-nitro-7-sulfamoylbenzo(f)quinoxaline-2,3-dione (NBQX 20 mg/kg) effectively suppressed hypoxia-induced seizures in immature rats and also protected against permanent changes in flurothyl threshold in adulthood, whereas treatment with MK-801 (1 mg/kg) or lorazepam (LZP 1 mg/kg) did not prevent these hypoxia-related epileptogenic effects. These results suggest that activation of alpha-amino-3-hydroxy-5-methyl-4-isoxazol propionic acid (AMPA) receptors may partly mediate the age-dependent epileptogenic effect of hypoxia in the perinatal period.

Topics: Age Factors; Animals; Dizocilpine Maleate; Epilepsy; Excitatory Amino Acid Antagonists; Flurothyl; Hypoxia, Brain; Lorazepam; Male; Quinoxalines; Rats; Receptors, AMPA

The effect of pretreatment with an AMPA receptor antagonist, NBQX, on MK-801-induced alterations in glucose use was examined using [14C]-2-deoxyglucose autoradiography. NBQX (7 mg/kg) had minimal effect on glucose utilisation in all anatomical regions examined. The intravenous administration of MK-801 (0.2 mg/kg) induced increases in glucose use in the limbic system and cingulate cortex. MK-801 reduced glucose utilisation in the sensory motor and auditory cortices. Pretreatment with NBQX attenuated the MK-801-induced hypermetabolism in the posterior cingulate cortex. The decreases in glucose utilisation induced by MK-801 were not exacerbated by the pretreatment with NBQX. The interaction between NBQX and MK-801 suggests a possible method of attenuating some of the adverse effects of the non-competitive NMDA receptor antagonists in the posterior cingulate cortex.

Topics: Animals; Autoradiography; Behavior, Animal; Deoxyglucose; Dizocilpine Maleate; Gyrus Cinguli; Limbic System; Male; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Stereotyped Behavior; Tissue Distribution

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

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

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 behavioural and convulsant effects of imipenem (Imi), a carbapenem derivative, were studied after intraperitoneal (i.p.) or intracerebroventricular (i.c.v.) administration in DBA/2 mice, a strain genetically susceptible to sound-induced seizures. The anticonvulsant effects of some excitatory amino acid antagonists and muscimol (Msc), a GABAA agonist, against seizures induced by i.p. or i.c.v. administration of Imi were also evaluated. The present study demonstrated that the order of anticonvulsant activity in our epileptic model, after i.p. administration, was (+)-5-methyl-10,11-dihydro-5H-dibenzo(a,d)-cyclohepten-5,10-imine maleate (MK-801) > (+/-)(E)-2-amino-4-methyl-5-phosphono-3-pentenoate ethyl ester (CGP 39551) > 3-((+/-)-2-carboxypiperazin-4-yl)propenyl-1-phosphonic acid (CPPene) > 3-((+/-)-2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CCP) > 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(F)-quinoxaline (NBQX). Ifenprodil, a compound acting on the polyamine site of NMDA receptor complex was unable to protect against seizures induced by Imi, suggesting that the poliamine site did not exert a principal role in the genesis of seizures induced by Imi. In addition, the order of anticonvulsant potency in our epileptic model, after i.c.v. administration, was CPPene > MK-801 > Msc > (-)-2-amino-7-phosphonic acid (AP7) > gamma-D-glutamylaminomethylsulphonate (gamma-D-GAMS) > NBQX > kynurenic acid (KYNA) > 6-cyano-7-nitro-quinoxaline-2,3-dione (CNQX). The relationship between the different site of action and the anticonvulsant activity of these derivatives was discussed. Although the main mechanism of Imi induced seizures cannot be easily determined, potential interactions with the receptors of the excitatory amino acid neurotransmitters exists. In fact, antagonists of excitatory amino acids are able to increase the threshold for the seizures or to prevent the seizures induced by Imi. In addition, Imi acts on the central nervous system by inhibition of GABA neurotransmission and Msc, a selective GABAA agonist, was able to protect against seizures induced by Imi.

Topics: 2-Amino-5-phosphonovalerate; 6-Cyano-7-nitroquinoxaline-2,3-dione; Amino Acids; Animals; Anticonvulsants; Dizocilpine Maleate; Excitatory Amino Acids; Glutamine; Imipenem; Kynurenic Acid; Mice; Mice, Inbred DBA; Muscimol; Piperazines; Piperidines; Quinoxalines; Seizures

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

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

In order to study the possible contribution of the substantia nigra (SN) in the positive interaction between dopamine D1 receptor agonists and glutamate antagonists in unilaterally 6-hydroxydopamine (6-OHDA) lesioned rats, the effect of the D1 agonist, SKF 38393, was studied in combination with intranigral infusions of glutamate antagonists of the NMDA (MK 801, CPP) or AMPA (NBQX) type of receptor. Local infusion into the SN of the 6-OHDA lesioned side of MK 801, CPP or NBQX at doses inducing no or minimal behavioral effects significantly increased the turning behavior and the expression of c-fos induced, in the lesioned caudate-putamen (CPu), by a parenteral administration of SKF 38393. The same result was obtained after intra-SN infusion of the GABA agonist, muscimol. High doses of MK 801, CPP or muscimol infused into the SN produced intense contralateral turning per se and induced a sparse c-fos expression in the lesioned CPu which was antagonized by parenteral administration of MK 801. The results indicate that a depression of SN pars reticulata efferent neurons potentiates D1-mediated responses and suggest that this area may play a role in the positive interaction between glutamate antagonists and D1 receptor agonists.

Topics: 2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine; Animals; Dizocilpine Maleate; Immunohistochemistry; Injections; Male; Neostriatum; Oxidopamine; Piperazines; Proto-Oncogene Proteins c-fos; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, Dopamine D1; Receptors, N-Methyl-D-Aspartate; Stereotyped Behavior; Substantia Nigra; Sympathectomy, Chemical

Combinations of dopaminergic agonists with glutamate receptor antagonists have been suggested to be a possible alternative treatment of Parkinson's disease. To gain further insights into this possibility, the antagonist of the competitive AMPA-type glutamate receptor NBQX and the ion-channel blocker of the NMDA glutamate receptor (+)-MK-801 in combination with the dopamine D1 receptor agonists: SKF 38393, SKF 82958 and dihydrexidine; the dopamine D2 receptor agonist bromocriptine and the dopamine-precursor L-DOPA were tested in rats pretreated with reserpine and alpha-methyl-p-tyrosine. MK-801 on its own induced locomotor behaviour and potentiated the antiakinetic effects of dihydrexidine and L-DOPA but not of the other dopamine agonists tested. NBQX neither on its own nor coadministered with the dopamine agonists tested had an antiakinetic effect. These results indicate that agents, blocking the ion-channel of the NMDA receptor, might be useful adjuvants to some but not all dopaminomimetics in therapy of Parkinson's disease. The same does not seem to be true for the AMPA-antagonist NBQX.

Topics: Animals; Biogenic Monoamines; Bromocriptine; Dizocilpine Maleate; Dopamine Agonists; Drug Combinations; Excitatory Amino Acid Antagonists; Male; Motor Activity; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Dopamine D1

Systemic administration of relatively high doses of amphetamine or cocaine induces expression of c-fos in the rat striatum and nucleus accumbens. The doses of drugs used in such experiments are substantially higher than those needed to produce reward or sensitization. Therefore, it was determined if low doses of amphetamine capable of eliciting reward and sensitization increase levels of c-Fos protein in the nucleus accumbens. Amphetamine, 1 mg/kg, stimulated locomotor activity and increased the number of nucleus accumbens cells immunohistochemically positive for c-Fos protein to approximately 800 cells per section from a control of approximately 100 cells per section. Since glutamate antagonists modify various responses to amphetamine, it was then determined whether activation of glutamate receptors is involved in the induction of c-Fos protein by low doses of amphetamine. The NMDA receptor antagonist MK-801 by itself stimulated locomotor activity but did not alter baseline levels of c-Fos protein. When given before amphetamine, both locomotor activity and extent of c-fos induction were greater than from amphetamine alone. The AMPA receptor antagonist NBQX by itself had no effect on locomotor activity but increased slightly the number of cells positive for c-Fos protein in the nucleus accumbens. When given before amphetamine, locomotor activity was completely attenuated, and the extent of c-fos induction was greater than from amphetamine alone. We conclude that low doses of amphetamine do increase abundance of c-Fos protein in the nucleus accumbens. This action does not correlate with locomotor activity and is independent of activation of glutamate receptors.

Topics: Amphetamines; Animals; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Immunohistochemistry; Male; Nucleus Accumbens; Oligonucleotides, Antisense; Proto-Oncogene Proteins c-fos; Quinoxalines; Rats; Rats, Sprague-Dawley

Intrahippocampal injection of the endogenous excitotoxin quinolinic acid (QUIN) induces seizures together with local, delayed neurodegeneration in specific cell layers. In situ hybridization histochemistry was used to study the spatio-temporal pattern of expression of neurotrophins (NTFs) after this treatment. As in other excitatory paradigms, nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) mRNA levels increased dramatically and transiently in dentate gyrus after the administration of 120 nmol of QUIN to the left hippocampus. BDNF, but not NGF, mRNA also increased in the hippocampal pyramidal cell layer, mainly in the CA1 field. Neurotrophin-3 (NT3) mRNA levels decreased in dentate gyrus, practically disappeared around 12 h after the insult and returned to basal levels four days later. A very different pattern of expression of NTFs was found locally: (a) upregulation of NGF and BDNF mRNAs expression was prevented in a spherical region of 1-2 mm diameter around the injection site, (b) a delayed increase in NT3 mRNA levels, beginning at 12 h and lasting for at least 4 days after the administration of QUIN, was found in the same region, in cell layers showing neurodegeneration. Pretreatment with the non-competitive NMDA antagonist MK-801 (2 mg/kg, 30 min before the insult), partially blocked the increase in both BDNF and NGF mRNAs, as well as the decrease in NT3, in the contralateral hippocampus. However, this treatment did not prevent the QUIN-induced local downregulation of NGF and BDNF. Treatment with the AMPA/kainate antagonist NBQX (30 mg/kg, 15 and 5 min before, and 10 min after the insult) did not influence the effect of QUIN upon NGF or BDNF mRNA levels, although it partially prevented the hippocampal contralateral decrease in NT3 mRNA. In conclusion, the present study strongly supports previous work concerning different regulation of BDNF/NGF respect to NT3 in seizure inducing paradigms. Moreover, the different and to some extent opposite regulation of NTFs in the hippocampal region contiguous to the injection site, respect to the remaining hippocampus, suggests a differential regulation of NTFs in QUIN-induced neurodegenerative and seizural processes. Finally, our pharmacological data, (i) show that the upregulation of NGF and BDNF mRNAs, indirectly induced by QUIN, is not mediated by AMPA receptors, and (ii) suggest other effects for QUIN, apart from the stimulation of NMDA receptors.

Topics: Animals; Brain; Brain-Derived Neurotrophic Factor; Dizocilpine Maleate; Gene Expression Regulation; Hippocampus; In Situ Hybridization; Male; Microinjections; Nerve Growth Factors; Nerve Tissue Proteins; Neurotrophin 3; Quinolinic Acid; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Reference Values; RNA, Messenger

This study examined the interaction between various glutamate antagonists and selective D1 (SKF 38393) and D2 (RU 24213) dopamine agonists in the production of locomotion in the reserpine-treated mouse. Firstly, in normal mice, the NMDA channel blocker MK 801 (0.1-1.6 mg/kg) caused a biphasic stimulation/depression of locomotor activity, whereas the competitive NMDA antagonists CGP 40116 (0.25-8 mg/kg) and CPP (0.2-20 mg/kg), and the NMDA glycine site antagonist HA 966 (0.4-10 mg/kg) inhibited locomotion monophasically. These compounds caused varying degrees of muscle weakness and impairment of posture and gait, whilst the AMPA receptor blocker NBQX (0.2-25 mg/kg) had no significant effect on unconditioned mouse motor behaviour. None of the antagonists reversed reserpine-induced akinesia by themselves, but they all potentiated the locomotor movements induced by 30 mg/kg SKF 38393. Movements remained fluent with low doses of CPP, HA 966 and NBQX, but became ataxic with MK 801 and CGP 40116, with sedation prevailing at high doses of all the antagonists, as in normal mice. CPP and NBQX also combined synergistically with SKF 38393 to promote tonic convulsions. By contrast, RU 24213-induced locomotion was dose-dependently depressed by MK 801, CGP 40116 and HA 966, but was unaffected by CPP or NBQX. These differential effects of NMDA and AMPA antagonists on D1 and D2 motor responding in the monoamine-depleted mouse are discussed in terms of possible mechanisms and sites of action within the brain, and the implications for their putative use as adjuvants to L-dopa in antiparkinson therapy.

Topics: 2-Amino-5-phosphonovalerate; 2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine; Animals; Dizocilpine Maleate; Dopamine Agonists; Male; Mice; Mice, Inbred Strains; Motor Activity; Phenethylamines; Piperazines; Pyrrolidinones; Quinoxalines; Receptors, AMPA; Receptors, Dopamine D1; Receptors, Dopamine D2; Receptors, N-Methyl-D-Aspartate; Reserpine

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

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

Homocysteine induces seizures in adult, as well as in immature, experimental animals, but the mechanism of its action is still unknown. The aim of the present study was to examine whether homocysteine in immature animals may act via excitatory amino acids receptors. Seizures were induced in 7-day-old rats by ip administration of homocysteine (16.5 mmol/kg) and the effects of selected antagonists at NMDA and non-NMDA receptor sites were investigated. The anticonvulsant effect was evaluated not only in terms of behavioral changes, but also in terms of some indicators of brain energy metabolism. Rat pups were sacrificed during generalized clonic-tonic seizures, corresponding approximately to 15-30 min after homocysteine administration. Comparable time intervals were used for sacrificing pups in the groups with protective drugs. Non-NMDA antagonists, L-glutamic acid diethylester (GDEE) (4 mmol/kg, ip) and 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo (F) quinoxaline (NBQX) (two doses, 30 mg/kg each, ip), failed to protect neonatal rats against homocysteine-induced seizures. Although NBQX prevented the tonic phase, the severity of clonic movements was even more pronounced. Metabolic changes accompanying the seizures (decreases of glucose and glycogen and a rise of lactate) were also not influenced by GDEE or NBQX pretreatment. On the contrary, NMDA antagonists, both competitive (AP7, 0.33 mmol/kg, ip) and noncompetitive (MK-801, 0.5 mg/kg, ip), had a clear-cut anticonvulsant effect. They not only suppressed the behavioral signs of seizures, but also prevented most of the metabolic changes accompanying seizures.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 2-Amino-5-phosphonovalerate; Amino Acids; Animals; Animals, Newborn; Anticonvulsants; Dizocilpine Maleate; Glutamates; Homocysteine; Male; Neuroprotective Agents; Quinoxalines; Rats; Rats, Wistar; Receptors, Amino Acid; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Seizures

The severity and rapidity of acute, glutamate-induced energy failure were compared in live cerebral cortical slices. In each experiment 80 live cerebral cortical slices (350 microns thick) were obtained from neonatal Sprague-Dawley rats, suspended and perfused in a nuclear magnetic resonance (NMR) tube, and studied at 4.7 T with interleaved 31P/1H NMR spectroscopy. NMR spectra, obtained continually, were determined as 5-min averages. Slices were perfused for 60 min with artificial cerebrospinal fluid (ACSF) containing either glutamate alone or glutamate mixed with one of three glutamate-receptor antagonists: kynurenate, dizocilpine (MK-801), and 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(F)quinoxaline (NBQX). Dose-dependent decreases in high-energy phosphates were studied during glutamate exposure (0.5 to 10 mM), with and without antagonist protection. Energy recovery after glutamate exposures was measured during a 60-min washout with glutamate-free, antagonist-free ACSF. Reversible and irreversible energy failures were characterized by changes in intracellular pH, and by changes in relative concentrations of ATP, phosphocreatine (PCr), and inorganic phosphate. No changes were observed in intracellular levels of N-acetylaspartate and lactate. Some special studies were also done using R-(-)-2-amino-5-phosphonovaleric acid (100 microM) and tetrodotoxin (1 mM) to examine glutamate receptor specificity in this tissue model. Dizocilpine (150 microM) best ameliorated the energy failure caused by 2.0 mM glutamate. With dizocilpine the maximum ATP decrease was only 6 +/- 5%, instead of 35 +/- 7%.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenosine Triphosphate; Animals; Animals, Newborn; Cerebral Cortex; Dizocilpine Maleate; Energy Metabolism; Glutamates; Glutamic Acid; In Vitro Techniques; Intracellular Membranes; Kynurenic Acid; Neurotoxins; Phosphocreatine; Quinoxalines; Rats; Rats, Sprague-Dawley

The AMPA antagonists NBQX (2.5, 5, 10 mg/kg) and GYKI 52466 (4.8, 8 mg/kg) were investigated in haloperidol (0.5 mg/kg)-induced catalepsy in the rat. The effects of AMPA antagonists administered either alone or in combination with the noncompetitive NMDA antagonist dizocilpine (0.02 mg/kg), with the dopamine D-2 agonist quinpirole (1 mg/kg) or with L-DOPA (50, 100 mg/kg plus benserazide) were tested. NBQX or GYKI 52466 did not exert anticataleptic effects, neither alone nor in combination with dizocilpine, quinpirole or L-DOPA. Thus, in the rat inhibition of AMPA receptors with NBQX or GYKI 52466 does not have effects predictive for an antiparkinsonian potential.

Topics: Animals; Anti-Anxiety Agents; Antipsychotic Agents; Benzodiazepines; Catalepsy; Dizocilpine Maleate; Drug Interactions; Ergolines; Haloperidol; Levodopa; Male; Pilot Projects; Quinoxalines; Quinpirole; Rats; Rats, Sprague-Dawley; Receptors, AMPA

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

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

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

WHILE exposure times of several hours or more are needed for kainate to induce widespread degeneration of most cortical or basal forebrain neurons, basal forebrain cholinergic neurons, as identified by choline acetyltransferase immunocytochemistry, were substantially damaged by brief (45 min) kainate exposures. This rapidly triggered damage to basal forebrain cholinergic neurons is Ca2+ dependent. Also, basal forebrain cholinergic neurons have unusually large elevations in intracellular Ca2+ concentration in response to kainate, and generally exhibit kainate-activated Co2+ uptake, suggesting that they possess Ca(2+)-permeable AMPA/kainate receptor-gated channels. The heightened vulnerability of basal forebrain cholinergic neurons to kainate toxicity may reflect rapid Ca2+ entry through Ca(2+)-permeable AMPA/kainate channels.

Topics: Animals; Calcium; Cell Death; Cells, Cultured; Choline O-Acetyltransferase; Cobalt; Dizocilpine Maleate; Fetus; Immunohistochemistry; Kainic Acid; Kinetics; Mice; Neurons; Neurotoxins; Prosencephalon; Quinoxalines; Receptors, AMPA; Time Factors

MUSCARINIC, NMDA and metabotropic glutamate receptor antagonists were tested for anticonvulsant effects in a continuous hippocampal stimulation model in rats in order to identify the receptors involved in the initiation of electrically induced status epilepticus (SE). Whereas the muscarinic receptor antagonists scopolamine and atropine and the metabotropic receptor antagonist L(+)-2-amino-3-phosphonopropionic acid (AP3) did not affect SE initiation, the N-methyl-D-aspartate (NMDA) antagonist dizocilpine (MK-801) (1 mg kg-1 i.p.) and the non-NMDA ionotropic receptor antagonist 2,3-dihydroxy-6-nitro-7-sulphamoylbenzo(F)-quinoxaline (NBQX) (two doses of 50 micrograms i.c.v.) prevented the induction of SE. It has been shown in a previous study that non-NMDA ionotropic receptors are involved in SE maintenance and it is now suggested that activation of NMDA receptors may principally initiate electrically induced SE, although non-NMDA ionotropic receptors may also be involved.

Topics: Animals; Atropine; Cerebral Ventricles; Dizocilpine Maleate; Electric Stimulation; Hippocampus; Injections, Intraperitoneal; Injections, Intraventricular; Male; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Scopolamine; Status Epilepticus

Brain-derived neurotrophic factor belongs to the neurotrophin family of trophic factors. Recently we have described four promoters in the BDNF gene. We have analyzed the hippocampal expression pattern of BDNF mRNA's containing different 5' exons during postnatal development. This analysis has revealed distinct regulation of promoters I and II compared to promoters III and IV, which suggests the presence of common regulatory elements for these clusters of promoters. Induction of different BDNF promoters after treatment with kainic acid combined with glutamate antagonists MK801 and NBQX discloses the differential participation of different glutamate receptor subtypes in regulation of the BDNF gene in the hippocampus.

Topics: Animals; Brain-Derived Neurotrophic Factor; Dizocilpine Maleate; Exons; Gene Expression Regulation; Hippocampus; Kainic Acid; Male; Nerve Growth Factors; Nerve Tissue Proteins; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, Glutamate; Ribonucleases; RNA, Messenger

We investigated the anticonvulsant and adverse effects of various dose combinations of the AMPA receptor antagonist NBQX (2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(F)quinoxaline) and the low-affinity, rapidly channel blocking NMDA receptor antagonist memantine in the kindling model of epilepsy. While memantine was ineffective when given alone, co-administration with NBQX markedly potentiated the increase in focal seizure threshold induced by NBQX alone. This synergistic (i.e. over-additive) interaction was seen at doses of both drugs which did not induce behavioural adverse effects. The data substantiate that combinations of AMPA and NMDA receptor antagonists provide a new strategy for the treatment of epileptic seizures.

Topics: Animals; Anticonvulsants; Dizocilpine Maleate; Drug Synergism; Female; Kindling, Neurologic; Memantine; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA

Brain edema caused by glutamate excitotoxicity was studied in well oxygenated neonatal cerebrocortical brain slices (350 mu thick). Slices exposed to 60 minutes of 2 mM glutamate, with or without glutamate antagonists (dizocilpine, kynurenate, or NBQX), were allowed to recover for 60 minutes. The protocol was identical to that in noninvasive multinuclear NMR spectroscopy studies (31P/1H/19F) of live slices. Percent water and swelling were determined invasively in isolated slices by wet and dry weight measurements before and after glutamate exposure. Edema was detectable within minutes in all experiments with glutamate exposures, but not in untreated control slices. Dizocilpine, kynurenate, and NBQX differently affected swelling, which correlated with PCr and ATP loss in separate NMR studies. Synaptic glutamate receptor activation appears to initiate events causing both edema and energy failure. Multiple glutamate receptor types seem to be involved. No glutamate antagonist provided greater protection against both edema and energy loss than dizocilpine. Dizocilpine might also block voltage-dependent Na+ channels, and provide protection via mechanisms other than NMDA-receptor dependent channel antagonism.

Topics: Animals; Brain Edema; Cerebral Cortex; Culture Techniques; Dizocilpine Maleate; Energy Metabolism; Glutamic Acid; Kynurenic Acid; Magnetic Resonance Spectroscopy; Organ Size; Quinoxalines; Rats; Receptors, AMPA; Receptors, Glutamate; Sodium Channels

In the adult female rat, the densities of dendritic spines and synapses on hippocampal CA1 pyramidal cells are dependent upon the ovarian steroid estradiol; moreover, spine and synapse density fluctuate naturally as ovarian steroid levels vary across the estrous cycle. To determine whether the effects of estradiol on dendritic spine density require activation of specific neurotransmitter systems, we have treated animals concurrently with estradiol and one of four selective neurotransmitter receptor antagonists: MK 801, a noncompetitive NMDA receptor antagonist; CGP 43487, a competitive NMDA receptor antagonist; NBQX, an AMPA receptor antagonist; or scopolamine, a muscarinic receptor antagonist. Our results indicate that the effects of estradiol can be blocked by treatment with either of the NMDA receptor antagonists, but treatment with an AMPA or muscarinic receptor antagonist has no effect on spine density. Thus, we have concluded that estradiol exerts its effect on hippocampal dendritic spine density via a mechanism requiring activation specifically of NMDA receptors.

Topics: 2-Amino-5-phosphonovalerate; Animals; Dendrites; Dizocilpine Maleate; Estradiol; Female; Hippocampus; Ovariectomy; Quinoxalines; Rats; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Scopolamine

We studied the effects of the non-NMDA antagonist NBQX and of the NMDA antagonist dizocilpine (MK-801), administered separately or together, on the respiratory function of conscious cats. NBQX (20 mg kg-1) did not affect minute ventilation nor the timing of inspiratory and expiratory phases, but the addition of a small dose of dizocilpine (0.15 mg kg-1) induced inspiratory pauses and respiratory arrest in the inspiratory phase (apneusis). Similarly, larger doses of either NBQX or dizocilpine did not induce apneusis but the addition of a small dose of the other compound provoked an apneusis. Thus, a blockade of either non-NMDA or NMDA receptors is well tolerated, but the combined blockade of both receptor types severely disrupts the respiratory function in the cat.

Topics: Animals; Cats; Dizocilpine Maleate; Quinoxalines; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Respiration Disorders

Excitatory amino acid receptor antagonists have been proposed as novel therapeutic agents to be used with levopoda in the treatment of Parkinson's disease. We examined the neural substrates for the interaction between levodopa and antagonists of either the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid or N-methyl-D-aspartate type of excitatory amino acid receptor using 2-deoxyglucose autoradiography. Thus, we compared the effects of the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (10 mg/kg, i.v.) and the N-methyl-D-aspartate antagonist MK-801 (0.1 mg/kg, i.v.) on cerebral metabolic responses to levodopa (25 mg/kg, i.v., with 12.5 mg/kg benserazide) in rats with a unilateral nigrostriatal pathway lesion. Levodopa increased glucose utilization ipsilateral to the lesion in substantia nigra pars reticula (up to 104%), entopeduncular nucleus (up 90%) and subthalamic nucleus (up 30%), indicating that levodopa alters striatal output through the striatonigral, striatoentopeduncular and striatopallidal pathways. Levodopa also decreased metabolic rate in lateral habenula (down 39%), a target of projections from entopeduncular nucleus, implying a reduction in basal ganglia output. 2,3-Dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline and MK-801 by themselves did not affect glucose utilization in any of these regions. Pretreatment with 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline reduced the effect of levodopa in substantia nigra pars reticulata but not in entopeduncular nucleus or subthalamic nucleus, while MK-801 attenuated the effect of levodopa in all three of these structures; neither 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline nor MK-801 altered the effect of levodopa in lateral habenula. When given at the same doses to a separate group of lesioned animals, neither 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline nor MK-801 affected rotational behavior elicited by levodopa. These findings indicate that alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid and N-methyl-D-aspartate receptor antagonists differentially modify dopamine receptor-mediated striatal output. alpha-Amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor blockade may preferentially attenuate the effect of dopamine receptor activation on the striatonigral pathway, while N-methyl-D-aspartate blockade appears to reduce the actions of dopamine on the striatonigral, s

Topics: Animals; Autoradiography; Brain; Carbon Radioisotopes; Deoxyglucose; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Levodopa; Male; Motor Activity; Organ Specificity; Oxidopamine; Quinoxalines; Rats; Rats, Sprague-Dawley

1. Activation of N-methyl-D-aspartate (NMDA) glutamate receptors in the brainstem network of respiratory neurones is required to terminate inspiration in the absence of lung afferents, but it is not required in the inspiratory motor act of lung inflation. In the present study we examined the involvement of non-NMDA ionotropic glutamate receptors in these two mechanisms in the adult mammal. 2. Adult cats were either decerebrated or anaesthetized with sodium pentobarbitone, paralysed and ventilated. Inspiratory motor output was recorded from the phrenic nerve and central respiratory activity from neurones in the bulbar ventral respiratory group. 3. In decerebrate vagotomized cats, ionophoretic application of 2,3-dihydroxy-6-nitro-7-sulphamoylbenzo(F)quinoxaline (NBQX) onto single respiratory neurones decreased their spontaneous discharge rate and abolished the excitatory effect of exogenously applied (RS) alpha-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid (AMPA) but not NMDA. 4. In these animals, intravenous infusion (12 mg kg-1) of the non-NMDA receptor blockers GYKI 52466 (1-(4-aminophenyl)-4-methyl-7,8-methylene-dioxy-5-H-2,3-benzodi aze pine) or NBQX: (1) decreased (in 10/15 cats) or abolished (in 5/15 cats) the inspiratory-related discharge of the phrenic nerve; (2) did not prolong the inspiratory phase; (3) reduced or abolished the spontaneous discharge of respiratory neurones; and (4) profoundly decreased the excitatory effects of AMPA but not NMDA ionophoresed onto these neurones. When both the phrenic nerve and the recorded respiratory neurone were silenced, neuronal excitation by ionophoretic application of NMDA first revealed a subthreshold respiratory modulation without lengthening of the inspiratory phase, then respiratory modulation became undetectable. 5. Additional blockade of NMDA receptors by a small dose (0.15 mg kg-1) of dizocilpine (MK-801), abolished the phrenic nerve activity which persisted after NBQX (apnoea), but the discharge or the subthreshold modulation of the bulbar respiratory neurones showed a lengthening of the inspiratory phase (apneusis). 6. Elevation of FA,CO2 increased or re-established phrenic nerve discharges after blockade of non-NMDA receptors or of both NMDA and non-NMDA receptors. 7. Small doses of NBQX or GYKI 52466 induced apnoea in five of five cats anaesthetized with sodium pentobarbitone. 8. In decerebrate animals with intact vagi, GYKI 52466 and NBQX depressed the Hering-Breuer expiratory-lengthening

Topics: Action Potentials; Anesthesia; Animals; Anti-Anxiety Agents; Benzodiazepines; Blood Pressure; Carbon Dioxide; Cats; Dizocilpine Maleate; N-Methylaspartate; Neurons; Phrenic Nerve; Pulmonary Alveoli; Quinoxalines; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Reflex; Respiration; Synaptic Transmission; Time Factors

The purpose of our study was to examine whether a significant interaction occurs between NMDA and non-NMDA receptor antagonists on respiratory function. For this purpose chloralose-anesthetized cats were used and respiratory minute volume (VE), tidal volume (Vt) respiratory rate (f), inspiratory and expiratory durations, and end tidal CO2 (FeCO2) were monitored. In some animals, phrenic nerve activity was also continuously recorded. In five spontaneously breathing animals, the NMDA receptor antagonist MK-801 was administered in a dose of 0.1 mg/kg i.v., and produced decreases in VE, Vt, f and increases in inspiratory duration and FeCO2. Using these five animals exhibiting respiratory effects from prior MK-801 dosing, we then administered the non-NMDA receptor antagonist NBQX (2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline) i.v. in a dose of 3 mg/kg. This dose is too low to produce a neuroprotective effect in animal models of brain ischemia. In each of the five animals NBQX administration produced an immediate impairment of respiration, culminating in apneusis within 55 s after i.v. injection. In terms of phrenic nerve discharge, inspiratory duration was increased approximately 4-fold by MK-801, and with the addition of NBQX, continuous discharge of the phrenic nerve occurred. Finally, NBQX given i.v. to animals not pretreated with MK-801 had only a slight depressant effect on respiratory activity. These results obtained with co-administration of low doses of two drugs that block NMDA and non-NMDA receptors raise the spector that combined use of these agents to ameliorate disorders in neurological function may be extremely deleterious to respiratory function.

Topics: Animals; Blood Pressure; Cats; Dizocilpine Maleate; Female; Heart Rate; Injections, Intravenous; Male; Phrenic Nerve; Quinoxalines; Receptors, AMPA; Respiration; Respiratory Function Tests

Brain protection during open heart surgery in the neonate and infant remains inadequate. Effects of the excitatory neurotransmitter antagonists MK-801 and NBQX on recovery of brain cellular energy state and metabolic rates were evaluated in 34 4-week-old piglets (10 MK-801, 10 NBQX, 14 controls) undergoing cardiopulmonary bypass and hypothermic circulatory arrest at 15 degrees C nasopharyngeal temperature for 1 h, as is used clinically for repair of congenital heart defects. MK-801 (dizocilpine) (0.75 mg/kg) or NBQX [2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline] (25 mg/kg) was given intravenously before cardiopulmonary bypass. Equivalent doses were placed in the cardiopulmonary bypass prime plus continuous infusions after reperfusion (0.15 mg kg-1h-1 and 5 mg kg-1h-1). Changes in high-energy phosphate concentrations and pH were analyzed by magnetic resonance spectroscopy in 17 animals until 225 min after reperfusion. Cerebral blood flow determined by radioactive microspheres as well as cerebral oxygen and glucose consumption were studied in 17 other animals. Cerebral blood flow and oxygen consumption were depressed relative to control by both MK-801 and NBQX at baseline. Recovery of phosphocreatine (p = 0.010), ATP (p = 0.030), and intracellular pH (p = 0.004) was accelerated by MK-801 and retarded by NBQX over the 45 min of rewarming reperfusion and the first hour of normothermic reperfusion. The final recovery of ATP at 3 h and 45 min reperfusion was significantly reduced by NBQX (46 +/- 26% baseline, mean +/- SD) versus control (81 +/- 19%) and MK-801 (75 +/- 8%) (p = 0.030). Cerebral oxygen consumption recovered to 105 +/- 30% baseline in group MK-801 and 94 +/- 31% in control but only to 61 +/- 22% in group NBQX (p = 0.070). Cerebral blood flow stayed significantly lower in group NBQX relative to control. Thus, MK-801 accelerates recovery of cerebral high-energy phosphates and metabolic rate after cardiopulmonary bypass and hypothermic circulatory arrest in the immature animal. At the dosage used NBQX exerts an adverse effect.

Topics: Animals; Brain; Cerebrovascular Circulation; Dizocilpine Maleate; Energy Metabolism; Glucose; Heart Arrest, Induced; Hydrogen-Ion Concentration; Intracellular Membranes; Lactates; Lactic Acid; Oxygen; Phosphates; Quinoxalines; Swine; Swine, Miniature

Deposit of beta-amyloid protein (A beta) in Alzheimer's disease brain may contribute to the associated neurodegeneration. We have studied the neurotoxicity of A beta in primary cultures of murine cortical neurons, with the aim of identifying pharmacologic ways of attenuating the injury. Exposure of cultures to A beta (25-35 fragment; 3-25 microM) generally triggers slow, concentration-dependent neurodegeneration (over 24-72 h). With submaximal A beta-(25-35) exposure (10 microM), substantial (> 40% within 48 h) degeneration often occurs and is markedly attenuated by the presence of the Ca2+ channel blockers nimodipine (1-20 microM) and Co2+ (100 microM) during the A beta exposure. However, A beta neurotoxicity is not affected by the presence of glutamate receptor antagonists. We suggest that Ca2+ influx through voltage-gated Ca2+ channels may contribute to A beta-induced neuronal injury and that nimodipine and Co2+, by attenuating such influx, are able to attenuate A beta neurotoxicity.

Topics: Amyloid beta-Peptides; Animals; Calcium Channel Blockers; Cells, Cultured; Cerebral Cortex; Dizocilpine Maleate; Fetus; Mice; Neurons; Neurotoxins; Nimodipine; Quinoxalines

The cytoprotective effects of MK-801 and NBQX, selective N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptor antagonists, respectively, were compared both singularly and in combination in models of transient severe forebrain and transient focal cerebral ischemia. After 10 minutes of four-vessel occlusion ischemia, the sodium salt of NBQX (30 mg/kg IP) given at the time of reperfusion and, subsequently, 15 and 30 minutes later produced a dramatic reduction in CA1 hippocampal necrosis at 7 days. This effect was not obtained with the intraperitoneal administration of either MK-801 (1 mg/kg x 3) or the combination of both NBQX and MK-801 given at the same time intervals. This effect of intraperitoneal NBQX alone was reproduced in a two-vessel occlusion/hypotension model using this same drug administration. Delayed treatment with both NBQX and GYKI 52466, but neither MK-801 nor the combination of NBQX and MK-801 given after a delay, produced a significant reduction in the mean volume of neocortical infarction after transient focal ischemia. We conclude that the AMPA receptor may play a more important role than the NMDA receptor in both selective ischemic necrosis of hippocampal neurons and in neocortical infarction. AMPA antagonists should be subjected to clinical stroke trials.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Anti-Anxiety Agents; Benzodiazepines; Cerebral Infarction; Cerebrovascular Disorders; Dizocilpine Maleate; Drug Administration Schedule; Injections, Intraperitoneal; Male; N-Methylaspartate; Quinoxalines; Rats; Rats, Wistar; Reperfusion

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

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

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

Intracerebroventricular (i.c.v.) infusion in mice of the selective metabotropic excitatory amino acid receptor agonist 1S,3R-1- aminocyclopentane-1,3-dicarboxylate (1S,3R-ACPD) (0.6-575 nmol/min) dose dependently induced face washing and scratching. In contrast, the subtype-specific ionotropic excitatory amino acid receptor agonists N-methyl-D-aspartate (NMDA), kainate and alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) (0.3-3.0 nmol/min) dose dependently induced clonic convulsions. I.c.v. infusion of the non-selective metabotropic receptor agonists ibotenate (6 nmol/min) or quisqualate (30 nmol/min) induced clonic convulsions. However, when ionotropic receptors were blocked with (+)-5-methyl-10,11-dihydro-5H-dibenzo-(a,d)cyclohepten-5,10-imine maleate (MK-801, dizoclipine) (3 nmol/min) or 2,3-dihydroxy-6-nitro-7- sulfamoyl-benzo(f)-quinoxaline (NBQX) (9 nmol/min), respectively, face washing and scratching behavior emerged. Neither MK-801 or NBQX (ED50 value > 100 nmol/min), nor the putative metabotropic receptor antagonist L-amino-3-phosphoro-propionic acid (L-AP3) (> 176 nmol/min); nor the dopamine receptor antagonists SCH 23390 (> 74 nmol/min), metoclopramide (> 89 nmol/min) and haloperidol (> 27 nmol/min) antagonized 1S,3R-ACPD-induced scratching (144 nmol/min). These results suggest that the behavioral consequences of i.c.v. infusion of 1S,3R-ACPD in mice reflect a selective activation of metabotropic receptors that differs from the behavioral changes observed with i.c.v. infusion of ionotropic receptor agonists.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Behavior, Animal; Cycloleucine; Dizocilpine Maleate; Dose-Response Relationship, Drug; Ibotenic Acid; Kainic Acid; Male; Mice; N-Methylaspartate; Quinoxalines; Quisqualic Acid; Receptors, Glutamate; Seizures

The aim of our study was to evaluate whether blockade of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors could reduce brain edema in two experimental models of edema following opening of the blood-brain barrier (BBB). The brain specific gravity was determined 2 h after opening the BBB by a 30-s infusion of protamine sulfate (10 mg in 200 microliters 0.9% NaCl) or arabinose (1.5 or 1.8 mol/L, 0.06 ml.s-1) into the right internal carotid artery. Cisternal CSF was withdrawn for albumin determination before the carotid infusion and before killing 2 h later. After infusion of protamine sulfate or arabinose, CSF albumin increased in all groups. The brain specific gravity was significantly lower in the right than in the left (control) frontal, parietal, and occipital cortex and striatum. NBQX (2,3-dihydroxy-6-nitro-7-sulfamoylbenxo(F)quinoxaline), an AMPA receptor antagonist, given intravenously 10 min after opening the BBB (5 mg/kg), significantly increased the specific gravity in the treated rats (p < 0.01 for the difference from control rats) without reducing CSF albumin or albumin extravasation in the brain as evaluated with Evans blue. We hypothesize that intracerebral (glial?) AMPA receptors may play a role in brain edema following opening of the BBB.

Topics: Animals; Blood-Brain Barrier; Brain Edema; Dizocilpine Maleate; Evans Blue; Excitatory Amino Acid Antagonists; Injections, Intravenous; Male; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Specific Gravity; Staining and Labeling

The selective alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(f)quinoxaline (NBQX) and the selective N-methyl-D-aspartate (NMDA) receptor antagonists MK 801 and ifenprodil were administered to Mongolian gerbils following a 5 min period of bilateral carotid artery occlusion. NBQX when given 4, 6 or 24 h after ischaemia gave a reduced loss of hippocampal CA1 neurones compared to control animals receiving vehicle only. Dizocilipine (MK 801) (1-10 mg/kg i.p.) and ifenprodil (a total of 45 mg/kg i.p.) gave no protection. The peak levels of NBQX obtained in the cerebrospinal fluid of gerbils receiving the neuroprotective dose (3 x 30 mg/kg i.p.) was 1 microM. In gerbil cortex slices, this concentration had no effect on NMDA-evoked depolarization, but had a moderate effect on kainate and gave a total blockade of AMPA depolarizations. It is concluded that antagonists of non-NMDA glutamate receptor subtypes, possibly AMPA, may be a useful therapeutic approach for cerebral ischaemia-related brain damage following global ischaemia.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Cerebral Cortex; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Gerbillinae; Hippocampus; Ibotenic Acid; Ischemic Attack, Transient; Kinetics; Male; Neurons; Piperidines; Quinoxalines; Receptors, AMPA; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate

The pharmacological specificity of the mGluR1 alpha subtype of the metabotropic glutamate receptor (mGluR) was examined in a cloned baby hamster kidney cell line (BHK-ts13) measuring [3H]glutamate binding and inositol phosphate (PI) hydrolysis. PI-hydrolysis was maximally stimulated by quisqualate (1112 +/- 105% of basal), glutamate (1061 +/- 70% of basal), ibotenate (1097 +/- 115% of basal) and beta-N-methylamino-L-alanine (BMAA) (1010 +/- 104% of basal). In contrast, the maximal stimulation of PI-hydrolysis by (1S,3R)-1-amino-cyclopentane-1,3-dicarboxylic acid (t-ACPD) was only 673 +/- 78% of the basal level. The relative order of potency was quisqualate > glutamate > ibotenate > t-ACPD > BMAA. Agonist-stimulated PI-hydrolysis was attenuated (25 +/- 4% inhibition) by L-2-amino-3-phosphonopropionic acid and partially blocked (44 +/- 7%) by pertussis toxin treatment. Saturation binding studies with [3H]glutamate on membranes prepared from BHK-ts13 cells expressing the mGluR1 alpha subtype showed that glutamate binds to a single affinity state of this receptor with a limited capacity (Kd = 296 nM, Bmax = 0.8 pmol/mg protein). In competition experiments, [3H]glutamate was displaced by quisqualate, glutamate, ibotenate, t-ACPD and BMAA with a rank order of potency similar to that found for stimulation of PI-hydrolysis.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Amino Acids, Diamino; Aminobutyrates; Animals; Binding, Competitive; Cell Line; Cell Membrane; Clone Cells; Cricetinae; Cyanobacteria Toxins; Dizocilpine Maleate; Glutamates; Glutamic Acid; Ibotenic Acid; Inositol Phosphates; Kidney; Kinetics; Neurotoxins; Pertussis Toxin; Quinoxalines; Quisqualic Acid; Receptors, Glutamate; Recombinant Proteins; Transfection; Virulence Factors, Bordetella

The neuroprotective effects of 2,3-dihydroxy-6-nitro-7- sulfamoylbenzo(f)quinoxaline (NBQX), GYKI 52466, and MK-801 were tested following severe forebrain ischemia. Wistar rats were subjected to 10 min of normothermic ischemia and reperfused for 7 days. Necrotic hippocampal CA1 neurons were counted and expressed as a percentage (mean +/- SD). In Experiment 1, saline-treated rats sustained 81 +/- 20% damage to dorsal CA1. Rats given NBQX 30 mg/kg i.p. x3 lost 21 +/- 27% (p < 0.01). Neither MK-801 1 mg i.p. x3 alone, nor in combination with the cytoprotective dose of NBQX protected CA1, with 83 +/- 18 and 54 +/- 34% damage, respectively (NS). Giving NBQX 90 mg/kg i.v. did not protect cells (94 +/- 5%) and resulted in nephrotoxicity. In Experiment 2, rats were given saline or three doses of NBQX 30 mg/kg i.p. immediately at reperfusion (RP) or after a 6-, 12-, or 24-h delay. Saline-treated rats suffered 79 +/- 16% injury. NBQX given immediately resulted in 17 +/- 17% injury, and even if treatment was delayed by either 6 or 12 h, there was marked protection with only 27 +/- 32 and 25 +/- 17% injury, respectively (all p < 0.01). Delaying the initiation of treatment to 24 h was not successful, resulting in 50 +/- 28% injury (NS). In Experiment 3, saline-treated rats lost 81 +/- 19% of CA1 cells, while those given GYKI 52466 10 mg/kg i.p. x5 starting immediately following RP lost 80 +/- 14%.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Anti-Anxiety Agents; Benzodiazepines; Brain Diseases; Dizocilpine Maleate; Hippocampus; Ischemic Attack, Transient; Male; Neurons; Prosencephalon; Quinoxalines; Rats; Rats, Wistar

Plant amino acids beta-N-oxalylamino-L-alanine (L-BOAA, present in Lathyrus sativus) and beta-N-methylamino-L-alanine (L-BMAA, present in Cycas circinalis) have been implicated in the pathogenesis of human neurological disorders lathyrism and amyotrophic lateral sclerosis-Parkinson's dementia complex of Guam (ALS-PD), respectively. In view of the conflicting reports that have emerged on the role of L-BMAA in ALS-PD, we reinvestigated the comparative toxicity of L-BMAA and L-BOAA. We report here the potent toxicity of L-BOAA as examined in an in vitro model consisting of sagittal slices of mouse brain. Incubation of sagittal slices of mouse brain with L-BOAA (1 pM) resulted in significant leakage of lactate dehydrogenase (LDH) and potassium from the slices into the medium. Under similar conditions, L-BMAA-induced LDH leakage from the slices into the medium was observed only at very high concentration of the toxin, namely 1 mM. N-Methyl-D-aspartate (NMDA) receptor antagonists ameliorated the toxic effects of L-BMAA, while non-NMDA receptor antagonists (quinoxalinediones) protected against the toxicity of L-BOAA. Incubation of slices with L-BOAA for 1 h resulted in extensive vacuolation and degeneration of neurons in the thalamus and brain stem, and to a lesser extent in the hippocampus and cerebellar nuclei. The large sized neurons appeared to be affected to a greater extent than the smaller ones. The neurons in other areas of the brain also revealed variable degree of degeneration with swelling of axons and dendrites.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Amino Acids, Diamino; Animals; beta-Alanine; Brain; Brain Chemistry; Cyanobacteria Toxins; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Histocytochemistry; In Vitro Techniques; L-Lactate Dehydrogenase; Mice; Neurotoxins; Paraffin Embedding; Plants; Potassium; Quinoxalines

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

Excitatory amino acid transmitters are involved in the initiation of seizures and their propagation. Most attention has been directed to synapses using N-methyl-D-aspartate (NMDA) receptors, although more recent evidence indicates potential roles for the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors as well. In the present experiments in amygdala-kindled rats, i.e. a model of partial epilepsy, competitive and uncompetitive NMDA antagonists exerted only weak anticonvulsant effects, whereas the AMPA antagonist 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(F)quinoxaline (NBQX) potently increased focal seizure threshold and inhibited seizure spread from the focus. These effects of NBQX were dramatically increased by pretreatment with low doses of NMDA antagonists, whereas adverse effects of NBQX were not potentiated. These data suggest that both non-NMDA and NMDA receptors are critically involved in the kindled state, and that combinations of AMPA and NMDA receptor antagonists provide a new strategy for treatment of epileptic seizures.

Topics: 2-Amino-5-phosphonovalerate; Amygdala; Animals; Anticonvulsants; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Synergism; Electric Stimulation; Epilepsies, Partial; Female; Kindling, Neurologic; Motor Activity; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Stereotyped Behavior

An assortment of glutamate antagonists with differing selectivities for NMDA and AMPA-type glutamate receptors, were tested for their effects in the mouse pilocarpine model of complex partial seizures. MK 801 (0.1-0.8 mg/kg) and high doses of HA 966 (50 mg/kg) were proconvulsant, whilst CGP 40116 (1-8 mg/kg) and low doses of HA 966 (0.4-10 mg/kg) inhibited pilocarpine-induced convulsions. CPP (5-20 mg/kg) and NBQX (1-50 mg/kg) were without effect. The dopamine D1 agonist SKF 38393 (10 mg/kg) facilitated the convulsant effects of low-dose pilocarpine (100 mg/kg). MK 801 (0.1-0.2 mg/kg) and HA 966 (50 mg/kg) interacted synergistically with SKF 38393 to promote the proconvulsant effects of D1 stimulation, whilst CPP (10-20 mg/kg) and HA 966 (10 mg/kg) had the opposite effect. CGP 40116 and NBQX were without effect. These results show that the convulsant qualities of MK 801 and SKF 38393, that have been detected in animal models of Parkinson's disease, can be reproduced in the pilocarpine model of epilepsy. Whilst the glutamate antagonists all interact synergistically with SKF 38393 to improve its antiparkinson activity, only MK 801 and high doses of HA 966 similarly potentiate the convulsions associated with D1 stimulation. An appropriate mixture of a glutamate antagonist and a D1 agonist could theoretically be used beneficially in the treatment of Parkinson's disease, without causing epilepsy as a side effect.

Topics: 2-Amino-5-phosphonovalerate; 2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine; Animals; Dizocilpine Maleate; Dopamine Agents; Excitatory Amino Acid Antagonists; Glutamic Acid; Male; Mice; Mice, Inbred Strains; Pilocarpine; Piperazines; Pyrrolidinones; Quinoxalines; Receptors, AMPA; Receptors, Dopamine D1; Receptors, N-Methyl-D-Aspartate; Seizures

A focal, unilateral thrombotic stroke was produced in the rat sensorimotor cortex. The time course of expression and localization of the immediate early inducible genes: c-fos, c-jun, zif268; nerve growth factor, brain-derived neurotrophic factor and the related tyrosine kinase high-affinity receptor (trkB) messenger RNAs were studied by in situ hybridization. The levels of messenger RNAs for c-fos, zif268, brain-derived neurotrophic factor (but not nerve growth factor) and trkB were consistently increased in cortex ipsilaterally to the lesion, while c-jun messenger RNA content was only slightly increased. The brain-derived neurotrophic factor messenger RNA was increased from 2 to 18 h following the stroke, mainly in cells having a normal morphological appearance. The trkB messenger RNA displayed temporal and spatial increases similar to brain-derived neurotrophic factor messenger RNA. The time course and pattern of expression of immediate early inducible gene and trophic factor messenger RNAs did not clearly support a causal relationship between these two families of factors. The observed messenger RNA increases were greatly attenuated by the non-competitive N-methyl-D-aspartate-sensitive glutamate receptor antagonist (+)-5-methyl-10,11-dihydroxy-5H-dibenzo(a,d)cyclohepten-5,10-imine , but substantially unaffected by the non-N-methyl-D-aspartate receptor antagonist 2,3-dihydroxy-6-nitrosulphanoylbenzoquinoxaline. The results suggest a major contribution of N-methyl-D-aspartate-sensitive glutamate receptor activation to the transcriptionally directed events subsequent to stroke. Future studies should clarify the contribution of these processes to either the progression of neuronal degeneration or the establishment of protective compensatory responses.

Topics: Animals; Brain; Brain-Derived Neurotrophic Factor; Cerebrovascular Disorders; Dizocilpine Maleate; DNA-Binding Proteins; Early Growth Response Protein 1; Gene Expression; Genes, fos; Genes, jun; Immediate-Early Proteins; In Situ Hybridization; Kinetics; Male; Membrane Proteins; Nerve Growth Factors; Nerve Tissue Proteins; Photochemistry; Protein-Tyrosine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-fos; Proto-Oncogene Proteins c-jun; Proto-Oncogenes; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptor, Ciliary Neurotrophic Factor; RNA, Messenger; Rose Bengal; Time Factors; Transcription Factors

Sound-induced seizures in genetically epilepsy-prone rats were used to compare the anticonvulsant effect of phenytoin and diazepam with compounds which decrease glutamatergic neurotransmission including excitatory amino acid antagonists acting at N-methyl-D-aspartate (NMDA) receptors: D(-)CPPene, CGP 37849 and MK 801 or at the glycine/NMDA site: ACPC (1-aminocyclopropane-dicarboxylic acid) (partial agonist) or non-NMDA receptors: NBQX (2,3-dihydroxy-6-nitro-7-sulfamoylbenzo[f]-quinoxaline.Li) and GYKI 52466 (1-(aminophenyl)-4-methyl-7,8-methylene-dioxy-5H-2,3-benzodiazepin e.HCl) or acting at sodium channels to decrease glutamate release: lamotrigine and BW 1003C87 (5(2,3,5-trichlorophenyl)-2,4-diaminopyrimidine ethane sulphonate). ED50 values against clonic seizures (in mumol/kg at the time of peak anticonvulsant effect) were: phenytoin 30.5 (2 h), diazepam 0.5 (0.5 h), MK 801 0.01 (4 h), D(-)CPPene 1.9 (4 h), CGP 37849 2 (1 h), GYKI 52466 24 (0.25 h), NBQX 40 (0.5 h), ACPC 1053 (0.5 h), BW 1003C87 2.2 (1 h), lamotrigine 4.8 (4 h). BW 1003C87, lamotrigine, MK 801, phenytoin, diazepam and CGP 37849 had the most favourable therapeutic indices (rotarod locomotor deficit ED50/anticonvulsant ED50).

Topics: 2-Amino-5-phosphonovalerate; Acoustic Stimulation; Amino Acids; Amino Acids, Cyclic; Analysis of Variance; Animals; Anticonvulsants; Behavior, Animal; Diazepam; Disease Models, Animal; Dizocilpine Maleate; Epilepsy; Female; Lamotrigine; Male; Motor Activity; Phenytoin; Pyrimidines; Quinoxalines; Rats; Rats, Sprague-Dawley; Time Factors; Triazines

We have studied the effect of two glutamate receptor antagonists on seizures and hippocampal neurone loss in the rat after systemic kainic acid administration. Intraperitoneal injection of the novel AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazolproprionic acid) receptor antagonist NBQX (6-nitro-7-sulphamoylbenzo(f)quinoxaline-2,3-dione) (30 mg/kg x 3 and 15 mg/kg x 3) administered 30 and 15 min. before and simultaneously with injection of kainic acid (5 mg/kg) intraperitoneally, dramatically enhanced the toxicity of kainic acid leading to death of all animals. When the NBQX dose was reduced to 8 mg/kg x 3, all animals survived and neurone damage in the hippocampus did not differ from control animals. When NBQX (30 mg/kg x 3) was administered 30- or 60 min after injection of kainic acid (8 mg/kg) intraperitoneally, no changes were observed concerning survival rates, seizure generation and neurone loss. Post-kainic acid treatment with the non-competitive NMDA receptor antagonist MK-801 (0.5 mg/kg and 1.0 mg/kg), 30 and 60 min. after intraperitoneally injection of kainic acid 8 mg/kg, abolished seizures in all animals and the neurone damage in the hippocampus was completely prevented. The results emphasize the importance of the NMDA-receptor activation for seizure generation and subsequent brain damage after intraperitoneally kainic acid. The paradoxical, unexpected effects of NBQX contrast to the protective effect of this compound after cerebral ischaemia and hypoglycaemia, conditions which are also characterized by glutamate-mediated damage. One possible explanation of the lowered seizure threshold to kainic acid after NBQX could be that NBQX is blocking AMPA receptors on interneurones more efficiently than on pyramidal cells.

Topics: Animals; Behavior, Animal; Brain; Dizocilpine Maleate; Drug Synergism; Kainic Acid; Male; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Seizures; Survival Rate

We have previously reported that transient spinal cord ischemia induced a behavioral hypersensitivity (allodynia) to innocuous cutaneous mechanical stimulation in rats. The spinal ischemia-induced allodynia was not relieved by morphine, but it was relieved by the gamma-aminobutyric acid (GABA)-B receptor agonist baclofen, indicating that the allodynia may be related to dysfunction of the spinal GABA-ergic inhibitory system. In the present study we report that systemic application of 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(f)quinoxaline (NBQX), an antagonist of the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor for excitatory amino acids, dose-dependently relieved allodynia after spinal cord ischemia. The analgesic effect of NBQX at a low dose (7.5 mg/kg) was not accompanied by motor deficits or sedation. On the other hand, the N-methyl-D-aspartate (NMDA) receptor antagonist dizocilpine (MK-801) only partially alleviated allodynia, even at doses that produced severe motor deficits. It is suggested that the abnormal, possibly painful, sensations elicited by innocuous mechanical stimulation observed after spinal cord ischemia may be mediated by excitatory amino acids, acting mainly on the AMPA receptor. Antagonists of excitatory amino acid receptors, especially at the AMPA site, may be effective in treating pain conditions where input from low threshold afferents triggers painful sensations.

Topics: Animals; Dizocilpine Maleate; Female; Pain; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Reperfusion Injury; Skin Physiological Phenomena; Spinal Cord; Vocalization, Animal

The role of N-methyl-D-aspartate (NMDA), non-NMDA glutamate, metabotropic and muscarinic receptors in the maintenance of status epilepticus (SE) was investigated. SE induced in rat brain by continuous electrical stimulation to the hippocampus was terminated by intracerebroventricular (i.c.v.) injection of the non-NMDA antagonists DNQX and NBQX, but not by the muscarinic antagonists scopolamine or atropine, or the metabotropic antagonist AP3. The NMDA antagonist, MK-801 suppressed motor seizure activity but did not terminate electrographic seizures when generalized SE was induced, suggesting that both non-NMDA and NMDA receptors maintain generalized convulsive SE. However, when limbic SE was induced, MK-801 also had an anticonvulsant effect suggesting differences in the mechanisms maintaining limbic SE and generalized SE.

Topics: Animals; Anticonvulsants; Atropine; Brain; Cerebral Ventricles; Dizocilpine Maleate; Electric Stimulation; Electroencephalography; Hippocampus; Injections, Intraventricular; Male; Quinoxalines; Rats; Rats, Wistar; Receptors, Glutamate; Receptors, Muscarinic; Scopolamine; Status Epilepticus

In the present immunocytochemical study, we investigated the mechanism of Fos protein induction and the regional distribution of the Fos protein in brains of spontaneously hypertensive rats subjected to 2 h of permanent middle cerebral artery occlusion (MCAO). Rats were administered either saline or a glutamate receptor antagonist; the non-competitive NMDA receptor antagonist MK-801 or the AMPA receptor antagonist NBQX which are known to be able to reduce infarct size in MCA occluded rats. The saline treated rats showed presence of Fos protein in nerve cell nuclei throughout the cortical and striatal infarct borderzone, but no staining in the infarct core or contralateral hemisphere. MK-801 almost totally abolished this expression of Fos protein whereas NBQX had no significant effect on Fos protein expression. It is suggested that the Fos protein induction is due to repeated spreading depressions mediated by NMDA receptors in the infarct borderzone, and that Fos protein due to its persistence in the tissue can be used as a histochemical marker of borderzone tissue at risk for eventually becoming recruited in the infarct.

Topics: Animals; Cerebral Cortex; Cerebral Infarction; Corpus Striatum; Cortical Spreading Depression; Dizocilpine Maleate; Energy Metabolism; Gene Expression Regulation; Male; Neurons; Proto-Oncogene Proteins c-fos; Quinoxalines; Rats; Rats, Inbred SHR; Receptors, N-Methyl-D-Aspartate

Excitatory amino acid antagonists have been proposed as novel therapeutic agents for Parkinson's disease due to their ability to reverse akinesia in animal models of this disorder. To further evaluate this therapeutic potential, we examined the effects of a N-methyl-D-aspartate (NMDA) and an alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor antagonist on catalepsy produced by dopamine D1 or D2 receptor antagonists in rats. Male Sprague-Dawley rats were injected with dizocilpine (MK-801 0.025, 0.05 or 0.1 mg/kg i.p.), 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(f)quinoxaline (NBQX 12.5 mg/kg i.p.) or saline prior to administration of either raclopride (2.5 mg/kg i.p.) or SCH 23390 (0.5 mg/kg i.p.). Catalepsy was evaluated with both grid and bar tests every 20 min for 2.7 h. MK-801 (0.1 mg/kg) reversed the catalepsy produced by either raclopride or SCH 23390 but did not stimulate locomotion when given alone at this dose. At 0.05 mg/kg, MK-801 markedly decreased SCH 23390-induced catalepsy, but did not affect the catalepsy produced by raclopride. In contrast, NBQX increased raclopride-induced catalepsy, but had no effect on catalepsy elicited by SCH 23390. These findings suggest that blockade of NMDA receptors, but not non-NMDA receptors, may reverse the catalepsy produced by dopamine receptor antagonists.

Topics: Animals; Benzazepines; Catalepsy; Dizocilpine Maleate; Dopamine Antagonists; Dopamine D2 Receptor Antagonists; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Male; Motor Activity; Quinoxalines; Raclopride; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, Dopamine D1; Receptors, N-Methyl-D-Aspartate; Salicylamides

Because hypoxia increases brain extracellular glutamate levels, we hypothesized that gasping and increased sympathetic activity during severe hypoxia result from glutamergic excitation. To test this hypothesis, we exposed anesthetized paralyzed vagotomized glomectomized cats to hypoxia before and after N-methyl-D-aspartate (NMDA) glutamergic blockade (MK-801, 1 mg/kg iv) or non-NMDA blockade (NBQX, 3 mg/kg iv) while monitoring phrenic neurogram (PN) and inspiratory-synchronous (ISSN) and tonic (TSN) activity in cervical sympathetic neurogram (SN). Before hypoxia, MK-801 caused apneusis and reduced PN and ISSN amplitude by 38 and 84%, respectively, but TSN activity was unaffected. During hypoxia, MK-801 had no effect on PN gasping or TSN activity but reduced ISSN amplitude during gasping. Before hypoxia, NBQX reduced PN and ISSN amplitude by 54 and 60%, respectively but did not affect inspiratory timing or TSN activity. Gasping activity in PN and ISSN and TSN activity during hypoxia were unaffected by NBQX. We conclude that 1) ionotropic glutamergic receptor activation is important for eupneic phrenic patterning but is not involved in genesis of gasping, 2) NMDA receptor activation is involved in integration of respiratory and sympathetic activity, and 3) changes in TSN activity are independent of ionotropic glutamergic receptor activation.

Topics: Animals; Cats; Dizocilpine Maleate; Electrophysiology; Excitatory Amino Acid Antagonists; Female; Hypercapnia; Hypoxia; Male; Phrenic Nerve; Quinoxalines; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Sympathetic Nervous System

We have studied the effect of intrahippocampal administration of quinolinic acid (QUIN) on the temporal expression of mRNAs encoding the immediate early genes (IEGs) c-fos and NGFI-A, by in situ hybridization histochemistry. After administration of QUIN to the left hippocampus, expression of mRNA of both IEGs was transiently stimulated. Maximal expression was found between 1 and 3 h. mRNA of both IEGs was simultaneously expressed in the ipsilateral and contralateral sides in the granule cell layer of the dentate gyrus, the pyramidal cell layer of the CA1 and CA3 fields as well as in the cortex. After pretreatment with the non-competitive NMDA antagonist MK-801 (2 mg/kg i.p. -30 min) the increased expression of both IEGs was partially prevented in the hippocampus and completely in the cortex. No inhibition was observed after treatment with the AMPA antagonist NBQX (30 mg/kg i.p. -15, -5 and +10 min). Additional delayed expression of both IEGs was observed in the ipsilateral hippocampus. This expression was related to cell damage. Twelve h after QUIN administration, c-fos and NGFI-A mRNAs were present in the dentate gyrus. After 4 days, only c-fos mRNA was observed in the dentate gyrus and CA1 field while no NGFI-A mRNA was detected. The present results show that the effect of QUIN is mediated by NMDA and not by AMPA receptors.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Behavior, Animal; Dizocilpine Maleate; Gene Expression Regulation; Genes, fos; Hippocampus; Ibotenic Acid; In Situ Hybridization; Injections, Intraperitoneal; Male; Oligonucleotide Probes; Quinolinic Acid; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

The effect of different glutamate-receptor antagonists on the induction of cortical spreading depression of Leao and of cortical anoxic membrane depolarization were investigated in the anaesthetized rat. Spreading depression (SD), elicited by mechanical stimulation of the cortical surface, was inhibited by the non-competitive N-methyl-D-aspartate (NMDA)-receptor blocker, (+-)-5-methyl-10,11-dihydro-5H-dibenzo(a,d)-cyclo-hepten-5,10-imi ne maleate (dizocilpine or MK-801), (0.30 mumol kg-1 (0.10 mg kg-1)), and the competitive NMDA-receptor antagonists; cis-4-phosphonomethyl-2-piperidine carboxylate (CGS 19755), (3.36 mumol kg-1 (0.75 mg kg-1)), D-(E)-2-amino-4-methyl-5-phosphono-3-pentenoic acid (CGP 40116), (1.20 mumol kg-1 (0.25 mg kg-1)) and its carboxylester CGP 43487, (6.30 mumol kg-1 (1.50 mg kg-1)). The alpha-amino-3-hydroxy-5-methyl-4-isoxazolepripriate (AMPA)-receptor blocker, 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F) quinoxaline (NBQX), administered as an intravenous dose of 29.76 and 89.29 mumol kg-1 (10 & 30 mg kg-1), which is sufficient to block seizures and protect against ischaemic brain damage, did not inhibit spreading depression. None of the drugs utilized inhibited the anoxic membrane depolarization. The data demonstrate that NMDA-receptor activation is essential for the initiation and propagation of spreading depression, while activation of AMPA-receptors is not obligatory. The observed initiation and propagation of SD, during AMPA-receptor blockade, suggest that activation of voltage-operated ion channels may contribute to release the magnesium block of the NMDA-receptor operated channel and to the initiation of SD.

Topics: 2-Amino-5-phosphonovalerate; Animals; Brain; Cell Membrane Permeability; Cortical Spreading Depression; Dizocilpine Maleate; Electrophysiology; Excitatory Amino Acid Antagonists; Ion Channels; Male; Pipecolic Acids; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate

We examined the effect of blockade of N-methyl-D-aspartate (NMDA) and non-NMDA subtype glutamate receptors on anoxic depolarization (AD) and cortical spreading depression (CSD). [K+]e and the direct current (DC) potential were measured with microelectrodes in the cerebral cortex of barbiturate-anesthetized rats. NMDA blockade was achieved by injection of (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate [MK-801; 3 and 10 mg/kg] or amino-7-phosphonoheptanoate (APH; 4.5 and 10 mg/kg). Non-NMDA receptor blockade was achieved by injection of 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(F)quinoxaline (NBQX; 10 and 20 mg/kg). MK-801 and APH blocked CSD, while NBQX did not. In control rats, the latency from circulatory arrest to AD was 2.1 +/- 0.1 min, while the amplitude of the DC shift was 21 +/- 1 mV, and [K+]e increased to 50 +/- 6 mM. All variables remained unchanged in animals treated with MK-801, APH, or NBQX. Finally, MK-801 (14 mg/kg) and NBQX (40 mg/kg) were given in combination to examine the effect of total glutamate receptor blockade on AD. This combination slightly accelerated the onset of AD, probably owing to circulatory failure. In conclusion, AD was unaffected by glutamate receptor blockade. In contrast, NMDA receptors play a crucial role for CSD.

Topics: 2-Amino-5-phosphonovalerate; Amino Acids; Animals; Cerebral Cortex; Cortical Spreading Depression; Dizocilpine Maleate; Electrophysiology; Ischemic Attack, Transient; Male; Quinoxalines; Rats; Rats, Inbred Strains; Receptors, Glutamate; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter

Glutamatergic transmission is an important factor in the development of neuronal death following transient cerebral ischemia. In this investigation the effects of N-methyl-D-aspartate (NMDA) and non-NMDA receptor antagonists on neuronal damage were studied in rats exposed to 10 min of transient cerebral ischemia induced by bilateral common carotid occlusion combined with hypotension. The animals were treated with a blocker of the ionotropic quisqualate or alpha-amino-3-hydroxy-5-methyl-4-isoxazole (AMPA) receptor, 2.3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX), given postischemia as an intraperitoneal bolus dose of 30 mg kg-1 followed by an intravenous infusion of 75 micrograms min-1 for 6 h, or with the noncompetitive NMDA receptor blocker dizocilpine (MK-801) given 1 mg kg-1 i.p. at recirculation and 3 h postischemia, or with the competitive NMDA receptor antagonist DL-(E)-2-amino-4-methyl-5-phosphono-3-pentenoic acid (CGP 40116), 5 mg kg-1, given intraperitoneally at recirculation. Treatment with NBQX provided a significant reduction of neuronal damage in the hippocampal CA1 area by 44-69%, with the largest relative decrease in the temporal part of the hippocampus. In neocortex a significant decrease in the number of necrotic neurons was also noted. No protection could be seen following postischemic treatment with dizocilpine or CGP 40116. Our data demonstrate that AMPA but not NMDA receptor antagonists decrease neuronal damage following transient severe cerebral ischemia in the rat and that the protection by NBQX may be dependent on the severity of the ischemic insult. We propose that the AMPA receptor-mediated neurotoxicity could be due to ischemia-induced changes in the control mechanisms of AMPA receptor-coupled processes or to changes of AMPA receptor characteristics.

Topics: 2-Amino-5-phosphonovalerate; Animals; Blood Glucose; Blood Pressure; Body Temperature; Cell Death; Dizocilpine Maleate; Hippocampus; Ischemic Attack, Transient; Male; Neurons; Quinoxalines; Rats; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter

The receptor mediating glutamate toxicity in vivo has not yet been identified. The effects of NMDA antagonist MK-801 and the AMPA antagonist NBQX were studied on glutamate toxicity in the arcuate nucleus of newborn and adult mice. Morphometric methods were used to determine the effects of antagonists on glutamate toxicity. In the developing arcuate, MK-801 abolished glutamate neurotoxicity whereas NBQX has no effect. MK-801, but not NBQX, afforded significant but not complete protection in mature arcuate neurons. The residual toxicity could not be blocked by co-administration of NBQX which paradoxically partially inhibited the protective effect of MK-801. The results show that throughout development, glutamate neurotoxicity in the arcuate nucleus is predominantly mediated by NMDA receptors.

Topics: Animals; Arcuate Nucleus of Hypothalamus; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Glutamates; Glutamic Acid; Male; Mice; Mice, Inbred Strains; Neurons; Quinoxalines

Increased extracellular concentrations of glutamate during episodes of cerebral ischemia may be due in part to a positive glutaminergic feedback loop. We evaluated the effect of selective AMPA or NMDA receptor antagonists on hippocampal extracellular concentrations of excitatory amino acids during ischemia and reperfusion. Thirteen New Zealand white rabbits were subjected to 10 min of global cerebral ischemia produced by neck tourniquet inflation (20 psi) combined with systemic hypotension during halothane (1-1.5%) anesthesia. Hippocampal extracellular concentrations of glutamate, aspartate, and glycine were monitored using in vivo microdialysis. NBQX (a selective AMPA receptor antagonist), MK801 (a noncompetitive NMDA receptor antagonist), or 5% dextrose was administered starting 1 h before ischemia. The NBQX group (n = 4) received 5 mg.kg-1 of NBQX intravenously (dissolved in 5% dextrose) over 5 min followed by an infusion of 5 mg.kg-1.h-1. The 5% dextrose group (n = 4) received an equivalent volume of 5% dextrose. The peak concentrations of glutamate, aspartate, and glycine in the early reperfusion period were 5-8-fold, 9-10-fold, and 4-5-fold higher than preischemic values, respectively. There were no significant differences, however, among the three groups in the concentrations of glutamate, aspartate, or glycine at any time during the study. These results do not support the existence of a positive feedback loop for glutamate mediated via AMPA or NMDA autoreceptors in the hippocampus during transient global ischemia or reperfusion.

Topics: Animals; Aspartic Acid; Dizocilpine Maleate; Glutamates; Glutamic Acid; Glycine; Hippocampus; Ischemic Attack, Transient; Quinoxalines; Rabbits; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter; Reperfusion

Quisqualic acid-mediated excitotoxicity has been attributed essentially to the activation of non-N-methyl-D-aspartate (non-NMDA) receptors. In the present study we demonstrate the possible involvement of both NMDA and non-NMDA receptors in quisqualic acid-induced toxicity in mouse brain slices, in vitro. Incubation of mouse brain sagittal slices with various concentrations of quisqualic acid resulted in significant increase in the leakage of lactate dehydrogenase and potassium from the slices into the medium. Prior incubation of mouse brain slices with NMDA (MK-801 or AP7) or non-NMDA receptor antagonists (GDEE or quinoxalinediones) protected against quisqualic acid-mediated toxicity. Slices prepared from animals pretreated in vivo with MK-801 (5 mg/kg b.wt.) were also resistant to the toxic effects of quisqualic acid, indicating the possible involvement of NMDA receptors in quisqualic acid toxicity.

Topics: 2-Amino-5-phosphonovalerate; 6-Cyano-7-nitroquinoxaline-2,3-dione; Amino Acids; Animals; Brain; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamates; Glutamic Acid; Ion Channel Gating; L-Lactate Dehydrogenase; Mice; Potassium; Quinoxalines; Quisqualic Acid; Receptors, AMPA; Receptors, Glutamate; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate

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

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

The developmental pharmacology of excitatory amino acid (EAA) receptors in the chick cochlear nucleus (nucleus magnocellularis, NM) was studied by means of bath application of drugs and recording of synaptically-evoked field potentials in brain slices taken from chicks aged embryonic day (E) 14 through hatching (E21). The abilities of various EAA agonists (N-methyl-D-aspartate [NMDA], kainic acid, and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid [AMPA]) to suppress postsynaptic responses by depolarization block and of EAA antagonists ((3-[RS]-2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid [CCP], dizocilpine [MK-801], 6-nitro-7-sulfamoyl-benzo(F)quinoxaline-2,3 dione [NBQX], 6-cyano-7-nitroquinoxaline-2,3-dione [CNQX] and 6,7-dinitroquinoxaline-2,3-dione [DNQX]) to suppress these responses directly were assessed quantitatively. The results support the existence of NMDA receptors in NM and suggest that the ability of these receptors to influence synaptically-evoked responses declines dramatically during the last week of embryonic life. The results similarly suggest that the non-NMDA receptors in NM undergo changes in density and/or function during a period of development when the cochlear nucleus is undergoing a variety of morphological and functional transformations.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Anticonvulsants; Chick Embryo; Cochlear Nerve; Dizocilpine Maleate; Dose-Response Relationship, Drug; Electric Stimulation; Embryonic and Fetal Development; Evoked Potentials; Ibotenic Acid; N-Methylaspartate; Piperazines; Quinolinic Acid; Quinolinic Acids; Quinoxalines; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter; Synapses; Synaptic Transmission

In situ hybridization was used to study expression of mRNAs for members of the nerve growth factor (NGF) family in the rat brain after 2 and 10 min of forebrain ischemia and 1 and 30 min of insulin-induced hypoglycemic coma. Two hours after the ischemic insults, the level of brain-derived neurotrophic factor (BDNF) mRNA was markedly increased in the granule cells of the dentate gyrus, and at 24 h it was still significantly elevated. NGF mRNA showed a pronounced increase 4 h after 2 min of ischemia but had returned to a control level at 24 h. Both 2 and 10 min of ischemia caused a clear reduction of the level of mRNA for neurotrophin 3 (NT-3) in the dentate granule cells and in regions CA2 and medial CA1 of the hippocampus 2 and 4 h after the insults. The increase of BDNF mRNA could be partially blocked by the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor antagonist NBQX but was not influenced by the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801. Both NBQX and MK-801 attenuated the decrease of NT-3 mRNA after ischemia. One and 30 min of hypoglycemic coma also induced marked increases in BDNF and NGF mRNA in dentate granule cells with maximal levels at 2 h. If the changes of mRNA expression lead to alterations in the relative availability of neurotrophic factors, this could influence functional outcome and neuronal necrosis following ischemic and hypoglycemic insults.

Topics: Animals; Brain; Brain-Derived Neurotrophic Factor; Dizocilpine Maleate; Gene Expression; Hippocampus; Insulin Coma; Ischemic Attack, Transient; Male; Nerve Growth Factors; Nerve Tissue Proteins; Neurotrophin 3; Nucleic Acid Hybridization; Proto-Oncogene Proteins c-fos; Quinoxalines; Rats; Rats, Inbred Strains; RNA, Messenger; Time Factors

In the central nervous system brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) are predominantly located in neurons. Here we demonstrate that the balance between the activity of the glutamatergic and gamma-aminobutyric acid (GABA)ergic systems controls the physiological levels of BDNF and NGF mRNAs in hippocampal neurons in vitro and in vivo. The blockade of the glutamate receptors and/or stimulation of the GABAergic system reduces BDNF and NGF mRNAs in hippocampus and NGF protein in hippocampus and septum. The reduction of NGF in the septum reflects the diminished availability of NGF in the projection field of NGF-dependent septal cholinergic neurons. These neurons do not synthesize NGF themselves but accumulate it by retrograde axonal transport. The refined and rapid regulation of BDNF and NGF synthesis by the glutamate and GABA transmitter systems suggests that BDNF and NGF might be involved in activity-dependent synaptic plasticity.

Topics: Animals; Brain-Derived Neurotrophic Factor; Diazepam; Dizocilpine Maleate; Embryo, Mammalian; Enzyme-Linked Immunosorbent Assay; Female; gamma-Aminobutyric Acid; Glutamates; Hippocampus; Kinetics; Male; Nerve Growth Factors; Nerve Tissue Proteins; Polymerase Chain Reaction; Quinoxalines; Rats; Rats, Inbred Strains; Receptors, GABA-A; Receptors, Glutamate; Receptors, Neurotransmitter; Reference Values; RNA, Messenger; Tetrodotoxin

In urethane-anesthetized rats, xylene applied locally to the skin of the hind paws was shown to induce reflex increases of blood pressure (33%) and heart rate (37%). The blood pressure elevation was dose dependently inhibited by the NMDA antagonist, MK-801 (0.3-1.0 mg/kg i.v.), and by the AMPA (D,L-alpha-amino-3-hydroxy-5-methyl-4-isoxalonepropionic acid) antagonist, NBQX (2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline 0.1-1.0 mg/kg per min). In contrast, only the latter compound was shown to block dose dependently the observed increase in heart rate. The results suggest that the two glutamate antagonists inhibit nociceptive impulse traffic at distinct anatomical sites and/or by different modes of actions.

Topics: Animals; Blood Pressure; Cardiovascular Physiological Phenomena; Cardiovascular System; Dizocilpine Maleate; Female; Heart Rate; Pain; Propranolol; Quinoxalines; Rats; Rats, Inbred Strains; Receptors, AMPA; Receptors, Neurotransmitter; Reflex; Skin; Skin Physiological Phenomena

Kindling, induced by repeated subconvulsive electrical or chemical stimulations leads to progressive and permanent amplification of seizure activity, culminating in generalized seizures. We report that kindling induced by electrical stimulation in the ventral hippocampus leads to a marked and transient increase in mRNA for NGF and BDNF in the dentate gyrus, the parietal cortex, and the piriform cortex. BDNF mRNA increased also in the pyramidal layer of hippocampus and in the amygdaloid complex. No change was seen in the level of HDNF/NT-3 mRNA. The increased expression of NGF and BDNF mRNAs was not influenced by pretreatment with the NMDA receptor antagonist MK801, but was partially blocked by the quisqualate, AMPA receptor antagonist NBQX. The presumed subsequent increase of the trophic factors themselves may be important for kindling-associated plasticity in specific neuronal systems in the hippocampus, which could promote hyperexcitability and contribute to the development of epileptic syndromes.

Topics: Animals; Brain; Brain-Derived Neurotrophic Factor; Dizocilpine Maleate; Epilepsy; Kindling, Neurologic; Male; Nerve Growth Factors; Nerve Tissue Proteins; Quinoxalines; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate; RNA, Messenger