6-cyano-7-nitroquinoxaline-2-3-dione and Nerve-Degeneration

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

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

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

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

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

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

Stabilizing the survival of oligodendrocytes and oligodendrocyte precursors within and near lesions in patients suffering from multiple sclerosis (MS) and other demyelinating diseases is an important therapeutic goal. Previous studies have identified a human-derived monoclonal IgM antibody designated rHIgM22 that induces remyelination in a mouse model of MS. We provide evidence that this antibody, directed against myelin, induces antiapoptotic signaling in premyelinating oligodendrocytes and reduces caspase-3 activation and caspase gene expression in mice undergoing antibody-induced remyelination. This effect was dependent on calcium entry via CNQX-sensitive channels and on lipid raft integrity, and was correlated with suppression of JNK signaling. We conclude that rHIgM22 may induce remyelination via rescue of oligodendrocytes, and suggest that such autoantibody-mediated signaling may have important therapeutic implications for a variety of neurological diseases, including stroke and Alzheimer's disease.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Antibodies; Apoptosis; Calcium Channels; Cardiovirus Infections; Caspase 3; Caspases; Cell Line, Tumor; Cell Survival; Humans; Hydrogen Peroxide; JNK Mitogen-Activated Protein Kinases; Membrane Microdomains; Mice; Mitogen-Activated Protein Kinases; Multiple Sclerosis; Myelin Proteins; Myelin Sheath; Nerve Degeneration; Oligodendroglia; Rats; Signal Transduction; Stem Cells; Tumor Necrosis Factor-alpha

Research has provided evidence about the role of excitotoxicity in the pathophysiology of sporadic amyotrophic lateral sclerosis and suggests that AMPA/kainate receptor activation contributes greatly in mediating glutamate injury to motor neurons. The recent finding of variable expression of metabotropic glutamate (mGlu) receptor subtypes in adult rat spinal cord has prompted us to investigate their contribution to the excitotoxic process. We report here that stimulation of mGlu receptors efficiently prevents motor neuron degeneration induced by kainate. The application of kainate to lumbar spinal cord slices from adult rats induced a massive degeneration of motor neurons which became shrunken, dark and TUNEL-positive. On the contrary, no significant neurotoxicity was observed after NMDA application. A blockade of ionotropic non-NMDA receptors by CNQX, and mGlu receptor stimulation, efficiently counteracted kainate-mediated cell death. Among the various agonists for mGlu receptors, we tested 3-hydroxyphenylglycine (3HPG), which selectively stimulates group I mGlu receptors. In addition, we tested 2-(carboxycyclopropyl)glycine (L-CCG-I) and 4-carboxy-3-hydroxyphenylglycine (4C3HPG), two selective agonists for group II receptors, as well as L-amino-4-phosphonobutyrate (L-AP4), a preferential agonist for group III. The results suggest that all three groups of mGlu receptors are involved in inhibiting excitotoxic phenomena mediated by kainate on spinal cord motor neurons. This was despite being localized differently and, possibly, activating different neuroprotective pathways.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Benzoates; Cell Survival; Choline O-Acetyltransferase; Cycloleucine; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Glycine; In Situ Nick-End Labeling; In Vitro Techniques; Kainic Acid; Male; Motor Neurons; N-Methylaspartate; Nerve Degeneration; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Receptors, Metabotropic Glutamate; Spinal Cord

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

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

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

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

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

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

Members of the nerve growth factor (NGF) family of neurotrophins bind to the second leucine-rich motif (LRM2) within the extracellular domains of their respective receptors (trkA, trkB, trkC). Small LRM2 peptides have been recently demonstrated to selectively bind the neurotrophins revealing similar complex binding characteristics as full-length receptors. We extend our recent findings, showing that the peptides (A and C) do not block nerve fiber outgrowth through high affinity trk receptors in a ganglia bioassay. Since the highest concentration of neurotrophins [NGF, brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3)] is found in the hippocampus, the peptides were injected into the 3rd ventricle of anesthetized adult rats. The (NGF binding) LRM2-A peptide, but not the (BDNF binding) LRM2-B or the (NT-3 binding) LRM2-C peptides, caused severe apoptotic neurodegeneration of hippocampal pyramidal CA1 neurons as revealed by cresyl violet staining and the TUNEL reaction. The degeneration was protected by intrahippocampal injection of NGF-beta and by the non-N-methyl-D-aspartate (NMDA) antagonist CNQX (6-cyano-7-nitroquinoxaline-2,3-dione), indicating a glutamatergic mechanism. In situ hybridization revealed that pyramidal CA1 neurons did not express trkA and p75 receptor mRNA in sham and LRM2-A-lesioned animals. It is concluded that the LRM2-A peptide represents a novel peptide with properties to induce apoptotic cell death of pyramidal CA1 neurons and may be useful as an experimental agent.

Topics: 2-Amino-5-phosphonovalerate; 6-Cyano-7-nitroquinoxaline-2,3-dione; Amino Acid Sequence; Analysis of Variance; Animals; Cell Death; Cerebral Ventricles; Excitatory Amino Acid Antagonists; Hippocampus; Injections, Intraventricular; Male; Microinjections; Molecular Sequence Data; Nerve Degeneration; Neuroprotective Agents; Peptide Fragments; Proto-Oncogene Proteins; Pyramidal Cells; Rats; Rats, Sprague-Dawley; Receptor Protein-Tyrosine Kinases; Receptor, trkA; Receptors, Nerve Growth Factor

Two non-protein amino acids of Lathyrus sativus, beta-(isoxazoline-5-on-2-yl)-alanine (BIA) and its higher homologue alpha-amino-gamma-(isoxazoline-5-on-2-yl)-alanine (ACI) were tested for excitotoxic potential. BIA (0.5-2.0 mM) but not ACI (2.0 mM) produced a concentration-dependent neurodegeneration in mouse cortical explants. The neuronal damage was prevented by the prior and simultaneous application of 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), indicating that it was mediated by non-N-methyl-D-aspartate type receptors. BIA (0.5-2.0 mM) activated CNQX-sensitive currents which were significantly smaller than those activated by 3-N-oxalyl-L-2,3-diaminopropanoic acid (beta-ODAP) or alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) in the majority of neurons. In a small number of cells, BIA (2 mM) produced currents which were similar in amplitude to those activated by beta-ODAP (50 microM). These results suggest that Lathyrus sativus plants engineered to block the synthesis of beta-ODAP may accumulate a neurotoxic precursor and therefore must be tested for the presence of both BIA and beta-ODAP.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Alanine; Aminobutyrates; Animals; Electrophysiology; Hippocampus; Isoxazoles; Lathyrism; Mice; Motor Cortex; Nerve Degeneration; Plants, Toxic

The distribution and density of non-NMDA receptors in spinal cord and motor cortex was compared in 10 cases of motor neuron disease (MND) and 8 neurologically normal controls by quantitative autoradiography using [3H]CNQX and [3H]kainate. In the motor cortex of MND cases, an increased density of [3H]kainate binding sites was observed which was most marked in the deep layers. No significant differences were observed in [3H]CNQX binding in the motor cortex between MND and control cases. In the spinal cord significantly increased densities of both [3H]CNQX and [3]kainate binding sites were found in the substantia gelatinosa and the intermediate grey matter in the MND group. The changes in [3H]kainate binding were observed only in the amyotrophic lateral sclerosis (ALS) subgroup of MND, while the changes in [3H]CNQX binding in the spinal cord were more marked in ALS compared to progressive muscular atrophy (PMA) cases. These findings provide evidence in support of a disturbance of glutamatergic neurotransmission in MND and suggest that there may be an increased excitatory drive to motor neurons via non-NMDA receptors. It is unclear at present whether the changes observed represent a compensatory response to loss of motor neurons in MND or a pathophysiological phenomenon contributing to motor neuron degeneration. Modulation of non-NMDA receptor activity may represent a possible target for therapeutic intervention in this disease.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Adult; Aged; Autoradiography; Female; Humans; Kainic Acid; Male; Middle Aged; Motor Cortex; Motor Neuron Disease; Nerve Degeneration; Receptors, AMPA; Receptors, Glutamate; Receptors, Kainic Acid; Spinal Cord

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

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

Murine neuronal and glial cell cultures exposed briefly to glutamate accumulated large amounts of 45Ca2+ from the extracellular medium during the exposure. Most of the accumulation likely reflected influx into neurons, as little accumulation was observed in similarly treated glial cultures. When the concentration of glutamate was varied between 10 and 1000 microM, or exposure duration was varied between 0 and 10 min, the amount of 45Ca2+ accumulation correlated closely with the amount of neuronal death 24 hr later. Both 45Ca2+ accumulation and cell death could be attenuated in a dose-dependent manner by the competitive NMDA antagonist D-aminophosphonovalerate or the noncompetitive antagonist dextrorphan, with IC50 values of approximately 100 microM and 15 microM, respectively. In contrast, neither 45Ca2+ accumulation nor cell death was blocked by the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)/kainate antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) in the presence of high glycine. With brief exposure, high concentrations of AMPA, kainate, or K+ produced much less death or 45Ca2+ accumulation than produced by glutamate, especially if 10 microM MK-801 was included in the exposure medium to block NMDA receptor activation. Kainate- or AMPA-induced 45Ca2+ accumulation or neuronal cell death was blocked with CNQX. However, high K(+)-triggered 45Ca2+ accumulation was only partially blocked with CNQX plus MK-801, consistent with mediation by voltage-gated Ca2+ channels. In addition to measuring the accumulation of 45Ca2+ occurring during agonist exposure, we also assessed accumulation during the 30 min immediately following completion of a 3-5 min exposure to 500 microM NMDA.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Calcium; Calcium Radioisotopes; Cells, Cultured; Cerebral Cortex; Glutamates; Glutamic Acid; Ibotenic Acid; N-Methylaspartate; Nerve Degeneration; Neurons; Quinoxalines; Receptors, Glutamate

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

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

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

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

Rat cerebellar granule cells, when subjected to a glucose-free environment for 4 h, developed extensive degeneration of neuronal cell bodies and their associated neurite network over the following 24 h. This neuronal damage was quantitated with a colorimetric assay using the metabolic dye 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide. Hypoglycemic neuronal injury could be markedly reduced by the presence of both competitive (3-(+/-)-2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid) and non-competitive (phencyclidine) N-methyl-D-aspartate receptor antagonists, but not by kainate/quisqualate preferring antagonists 6-cyano-7-nitroquinoxaline-2,3-dione and 6,7-dinitroquinoxaline-2,3-dione. Glucose deprivation neuronal injury was also reduced by adding glutamate-degrading enzymes to the incubation medium. Monosialoganglioside GM1, but not its asialo derivative (lacking sialic acid), was also effective in protecting against hypoglycemic neurodegeneration when included during the period of glucose deprivation. These results suggest that the neuronal injury to cerebellar granule cells resulting from glucose deprivation is mediated predominantly by activation of the N-methyl-D-aspartate type of excitatory amino acid receptor, perhaps through the action of endogenously released glutamate. Furthermore, the monosialoganglioside GM1, a member of a class of naturally occurring sialoglycosphingolipids, is able to attenuate this neuronal injury--as already observed for glutamate neurotoxicity and anoxic neuronal death in cerebellar granule cells. Gangliosides may thus prove to be of therapeutic utility in excitatory amino acid-associated neuropathologies.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Cerebellum; G(M1) Ganglioside; Gangliosides; Hypoglycemia; Nerve Degeneration; Neurons; Piperazines; Quinoxalines; Rats; Rats, Inbred Strains; Receptors, Amino Acid; Receptors, Cell Surface; Receptors, N-Methyl-D-Aspartate