Neuronal calcium channels.
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Journal Article↗
Biomedical subjects
D W Choi
Publications and source records attributed to D W Choi.
Glutamate neurotoxicity, calcium, and zinc.
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Animals↗
GABAergic neocortical neurons are resistant to NMDA receptor-mediated injury.
N-methyl-D-aspartate (NMDA) receptors are thought to mediate much of the central neuronal loss produced by certain neurologic insults, including hypoxia-ischemia, hypoglycemia, and trauma. Therefore, the specific vulnerability of GABAergic inhibitory neurons to NMDA receptor-mediated toxicity might be an important determinant of the potential for epileptogenesis following these insults. We have examined the fate of GABAergic cortical neurons in mouse cell cultured neuronal population) were identified either by immunoreactivity with antisera to GABA or by autoradiography following high-affinity uptake of 3H-GABA. Cultures exposed for 5 min to 20 to 750 microM NMDA showed NMDA concentration-dependent, widespread neuronal loss. However, GABAergic neurons were relatively spared, and thus represented an enhanced fraction of neuronal survivors. These observations suggest that GABAergic cortical neurons may possess some intrinsic resistance to NMDA receptor-mediated neurotoxicity, a property which might convey an anticonvulsant "inhibitory safety factor" to neocortex against certain forms of injury.
Animals↗
Delayed rescue of N-methyl-D-aspartate receptor-mediated neuronal injury in cortical culture.
The present study explored the neuroprotective efficacy of delayed manipulations performed after completion of an excitotoxic insult. Many cultured murine cortical neurons that would otherwise die after exposure to 500 microM N-methyl-D-aspartate (NMDA) or glutamate could be rescued by the late addition of NMDA antagonists to the bathing medium. D-2-Amino-5-phosphonovalerate (D-APV), MK-801 (5-methyl-10, 11-dihydro-5H-dibenzo[a,d]cyclohepten-5, 10-imine maleate) and dextrorphan all produced concentration-dependent neuroprotection, with EC50 about 10, 0.3 and 3 microM, respectively, and similar efficacy. The fraction of the doomed neuronal population that could be rescued depended on the time interval between washout of NMDA and antagonist addition, starting at a maximum of 40 to 80% with immediate addition, and decaying to zero after 30 min. D-APV only needed to be present for 30 min for near maximal protection. The ability of NMDA antagonists to rescue doomed neurons was not mimicked by several other drugs: 1 mM gamma-D-glutamyl-aminomethyl sulfonate, 1 mM L-glutamate diethyl ester, 10 microM 6-nitro-7-cyano-quinoxaline-2,3-dione, 1 mM secobarbital, 100 microM diphenylhydantoin, 10 microM nifedipine or 3 microM tetrodotoxin. The broad spectrum glutamate antagonist kynurenate had a protective action similar to that of D-APV, but when added to D-APV did not improve neuroprotection. Neurons could also be rescued by the delayed removal of extracellular calcium for 30 min after exposure to NMDA. In contrast, replacement of sodium with choline actually enhanced resultant neuronal damage.(ABSTRACT TRUNCATED AT 250 WORDS)
2-Amino-5-phosphonovalerate↗
Hypoxic neuronal injury in vitro depends on extracellular glutamine.
Hypoxic neuronal injury (HNI) in cortical cell cultures was enhanced in a concentration-dependent fashion by the presence of 500 microM to 2 mM (EC50 about 500 microM) glutamine in the medium, concentrations approximating those normally present in cerebrospinal fluid (CSF). Regardless of the glutamine concentration, glutamate receptor antagonists 2-amino-5-phosphonovalerate or dextrorphan could substantially reduce HNI. Thus, the availability of extracellular glutamine could be a determinant of hypoxic neuronal injury in vivo, most likely reflecting its importance in the synthesis of the neurotransmitter excitotoxins glutamate and aspartate.
Anaerobiosis↗
Opioids and non-opioid enantiomers selectively attenuate N-methyl-D-aspartate neurotoxicity on cortical neurons.
Addition of 1 microM-1 mM methadone to the bathing medium produced a concentration-dependent reduction in the neurotoxicity of exogenously applied N-methyl-D-aspartate (NMDA) in murine cortical cell culture (EC50 about 100 microM); the reduction persisted at intense NMDA exposure, consistent with non-competitive inhibition. Methadone also protected against exposure to quinolinate but not quisqualate or kainate. Concentrations (100 microM-3 mM) of several other opioids - morphine, fentanyl, codeine, meperidine, dextropropoxyphene, and naltrexone - were additionally found to produce concentration-dependent reductions in NMDA neurotoxicity. This novel neuron-protective effect of opioids was not mediated by conventional opioid receptors: the non-opioid enantiomer of methadone and morphine exhibited a potency equal to or greater than that of the opioid enantiomer, and 1 mM naloxone did not act as an antagonist. The possibility that opioids, or especially non-opioid enantiomers of opioids, might provide a useful therapeutic approach in diseases states involving NMDA receptor-mediated neurotoxicity, warrants further study.
Amino Acids↗
Tetrahydroaminoacridine selectively attenuates NMDA receptor-mediated neurotoxicity.
Addition of the acetylcholinesterase inhibitor 1,2,3,4-tetra-9-hydroaminoacridine (THA) at 1-3 mM markedly reduced the neuronal cell loss that otherwise followed brief exposure of murine cortical cell cultures to 500 microM N-methyl-D-aspartate (NMDA). This novel antagonism was selective for NMDA receptor-mediated toxicity, as it extended to glutamate toxicity but not to quisqualate toxicity, and was THA concentration-dependent between 100 microM and 3 mM, with IC50 approximately 500 microM. The antagonism was probably not due to enhancement of endogenous cholinergic action, as it was not mimicked by acetylcholine, carbachol, or bethanechol; rather, it likely reflected a recently described interaction of THA with the phencyclidine receptor. Exploration of structural specificity revealed some partial neuron-protection with high concentrations of other cholinesterase inhibitors--physostigmine, neostigmine, and edrophonium, but not the structurally related potassium channel blocker, 4-aminopyridine. Further examination of correlations between THA-like structure, and neuron-protective activity, may provide useful insights in the development of new antagonists of NMDA receptor-mediated neurotoxicity.
Aminoacridines↗
Beta-N-methylamino-L-alanine neurotoxicity: requirement for bicarbonate as a cofactor.
Ingestion of the excitotoxic cycad seed amino acid beta-N-methylamino-L-alanine may be responsible for the neuronal degeneration associated with Guam amyotrophic lateral sclerosis-parkinsonism-dementia in man. However, the basis for the central neurotoxicity of beta-N-methylamino-L-alanine has been unclear, as it lacks the omega acidic (or equivalent electronegative) moiety characteristic of other excitatory amino acids. beta-N-methylamino-L-alanine produced neurotoxic and neuroexcitatory effects in murine cortical cell cultures only when physiological concentrations of bicarbonate were available in the extracellular bathing medium. Bicarbonate may interact noncovalently with beta-N-methylamino-L-alanine to produce, in combination, a molecular configuration that activates glutamate receptors.
Amino Acids, Diamino↗
Adenosine reduces cortical neuronal injury induced by oxygen or glucose deprivation in vitro.
The endogenous neuromodulatory purine, adenosine, substantially attenuated neuronal degeneration when added to dissociated cortical cell cultures acutely deprived of either oxygen or glucose. The protective effect of adenosine, was concentration-dependent between 30 and 1000 microM (EC50 about 100 microM), and could be mimicked by the stable adenosine analogue N6-cyclohexyladenosine (10 microM). Unlike postsynaptic glutamate receptor antagonists, which also block these forms of neuronal injury, adenosine did not alter the neurotoxicity of exogenously applied glutamate.
Adenosine↗
Cultured striatal neurons containing NADPH-diaphorase or acetylcholinesterase are selectively resistant to injury by NMDA receptor agonists.
Cultured striatal neurons containing either NADPH-diaphorase or acetylcholinesterase were more resistant to injury by N-methyl-D-aspartate (NMDA) or quinolinate, than the general striatal neuronal population, although this resistance was not absolute and could be overcome by intense toxic exposure. Neurons containing NADPH-diaphorase, but not neurons containing acetylcholinesterase, also exhibited heightened vulnerability to injury by kainate. Given recent evidence that diaphorase- and cholinesterase-containing striatal neurons are selectively spared in Huntington's disease, our results strengthen the possibility that NMDA receptor-mediated neurotoxicity may participate in the pathogenesis of that disease.
Acetylcholinesterase↗
Morphinans attenuate cortical neuronal injury induced by glucose deprivation in vitro.
The non-narcotic dextrorotatory morphinan, dextrorphan, as well as its levorotatory opioid enantiomer, levorphanol, and its O-methyl derivative, dextromethorphan, have recently been shown to antagonize N-methyl-D-aspartate receptor-mediated neurotoxicity. Consistent with in vivo data suggesting that this neurotoxicity contributes to the neuronal damage associated with hypoglycemia, micromolar concentrations of these morphinans markedly attenuated the injury of cultured mouse cortical neurons produced by acute glucose deprivation. These observations lend specific support to the possibility that morphinan compounds may prove to have clinical therapeutic utility in hypoglycemic encephalopathy.
Animals↗
Dextrorphan and dextromethorphan, common antitussives, are antiepileptic and antagonize N-methyl-D-aspartate in brain slices.
The antitussive, dextromethorphan (DM), and its metabolite, dextrorphan (DX), were evaluated for antiepileptic properties in vitro. Interictal bursts and prolonged ictal epileptiform afterdischarges, induced by perfusion of guinea pig neocortical brain slices with Mg2+-free solution, were blocked by DX (1-250 microM) or DM (100 microM). Intracellular records showed that these agents blocked N-methyl-D-aspartate (NMDA)-induced depolarizations without altering intrinsic membrane properties. DX blocked NMDA but not quisqualate-evoked multi-unit excitatory responses. DM is a widely available, orally effective drug with low toxicity in antitussive doses, which has antiepileptic and NMDA-antagonist properties in vitro. Its toxicity and effectiveness as an anticonvulsant should be expeditiously examined in clinical trials.
2-Amino-5-phosphonovalerate↗
Dextromethorphan reduces neocortical ischemic neuronal damage in vivo.
The dextrorotatory morphinan dextromethorphan (DM), a clinically tested antagonist of the N-methyl-D-aspartate (NMDA) receptor-channel complex, was tested in an in vivo model of acute transient focal cerebral ischemia. Rabbits were randomly assigned to pretreatment with a 20 mg/kg i.v. bolus followed by 10 mg/kg/h of 0.4% DM in normal saline (NS), or with an equivalent volume of NS alone. They then underwent 1 h occlusion of the left internal carotid artery an anterior cerebral artery followed by 4 h of reperfusion. DM-treated animals showed a significant decrease in the percentage of severe neocortical ischemic neuronal damage (10.5%), as compared to NS-treated animals (49.6%).
Animals↗
Calcium-mediated neurotoxicity: relationship to specific channel types and role in ischemic damage.
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Animals↗
Zinc neurotoxicity in cortical cell culture.
Large amounts of zinc are endogenously present in synaptic vesicles of mammalian central excitatory boutons, and are likely released during synaptic activity; transient elevations in extracellular zinc concentration exceeding several hundred micromolar may accompany intense neuronal excitation. Exposure of mature cortical cell cultures, in mice, to similar concentrations of zinc for several minutes resulted in widespread neuronal injury; the extent of injury was dependent on both the concentration of zinc, and the length of exposure. Quantitative neuronal cell counts suggested an approximate neurotoxic ED50 of 600 microM for a 15 min zinc exposure, and 225 microM for an 18-24 h exposure. High zinc concentrations or long exposure times resulted in the addition of glial injury to the neuronal injury; this glial injury could also be demonstrated in neuron-free glial cell cultures, and hence likely represented a direct effect of zinc rather than a consequence of neuronal injury. Neurons in immature cultures were relatively resistant to zinc-induced injury, suggesting that neuronal vulnerability to zinc increases with maturation in vitro. An early event associated with toxic exposure to zinc was gross neuronal swelling. This swelling was dependent on the presence of extracellular sodium, and, interestingly, could be delayed by the continued presence of zinc itself. Zinc-induced neuronal cell loss, however, occurred even when both sodium and calcium were absent during the exposure to zinc. The present results provide direct evidence that zinc might be a relatively potent, rapidly acting neurotoxin, and somewhat less potent gliotoxin, in the mammalian central nervous system. We suggest that zinc should be included on the growing list of endogenous toxins which may be involved in the acute pathogenesis of central neuronal, and possibly glial, cell loss in some disease states.
Animals↗
Glutamate neurotoxicity and diseases of the nervous system.
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Animals↗
Phencyclidine receptor ligands attenuate cortical neuronal injury after N-methyl-D-aspartate exposure or hypoxia.
Phencyclidine (PCP) and related noncompetitive antagonists of N-methyl-D-aspartate (NMDA) receptor neuroexcitation were tested for their ability to attenuate either NMDA- or hypoxia-induced neuronal loss in dissociated cell cultures prepared from mouse neocortex. 10 microM PCP or MK-801 [(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate] selectively blocked the neurotoxicity produced by application of NMDA or the endogenous NMDA agonist quinolinate, without altering kainate or quisqualate neurotoxicity. Blockade of NMDA toxicity was concentration-dependent in the submicromolar and micromolar range, with a potency order (MK-801 greater than PCP greater than SKF 10,047 greater than pentazocine) consistent with primary mediation at PCP sites rather than at sigma sites. PCP reduced the toxicity of high NMDA concentrations, consistent with a noncompetitive mechanism of antagonism. PCP ligands also potently attenuated neuronal injury after exposure of the cultures to hypoxia. The effective concentrations and drug potency order for blockade of hypoxic injury were quantitatively similar to results obtained against the application of exogenous NMDA. These observations provide further support for the hypothesis that excessive activation of NMDA receptors may participate importantly in the pathogenesis of hypoxic brain injury.
Animals↗
Pharmacology of glutamate neurotoxicity in cortical cell culture: attenuation by NMDA antagonists.
The antagonist pharmacology of glutamate neurotoxicity was quantitatively examined in murine cortical cell cultures. Addition of 1-3 mM DL-2-amino-5-phosphonovalerate (APV), or its active isomer D-APV, acutely to the exposure solution selectively blocked the neuroexcitation and neuronal cell selectively blocked the neuroexcitation and neuronal cell loss produced by N-methyl-D-aspartate (NMDA), with relatively little effect on that produced by either kainate or quisqualate. As expected, this selective NMDA receptor blockade only partially reduced the neuroexcitation or acute neuronal swelling produced by the broad-spectrum agonist glutamate; surprisingly, however, this blockade was sufficient to reduce glutamate-induced neuronal cell loss markedly. Lower concentrations of APV or D-APV had much less protective effect, suggesting that the blockade of a large number of NMDA receptors was required to acutely antagonize glutamate neurotoxicity. This requirement may be caused by the amplification of small amounts of acute glutamate-induced injury by subsequent release of endogenous NMDA agonists from injured neurons, as the "late" addition of 10-1000 microM APV or D-APV (after termination of glutamate exposure) also reduced resultant neuronal damage. If APV or D-APV were present both during and after glutamate exposure, a summation dose-protection relationship was obtained, showing substantial protective efficacy at low micromolar antagonist concentrations. Screening of several other excitatory amino acid antagonists confirmed that the ability to antagonize glutamate neurotoxicity might correlate with ability to block NMDA-induced neuroexcitation: The reported NMDA antagonists ketamine and DL-2-amino-7-phosphono-heptanoate, as well as the broad-spectrum antagonist kynurenate, were all found to attenuate glutamate neurotoxicity substantially; whereas gamma-D-glutamylaminomethyl sulfonate and L-glutamate diethyl ester, compounds reported to block predominantly quisqualate or kainate receptors, did not affect glutamate neurotoxicity. The present study suggests that glutamate neurotoxicity may be predominantly mediated by the activation of the NMDA subclass of glutamate receptors--occurring both directly, during exposure to exogenous compound, and indirectly, due to the subsequent release of endogenous NMDA agonists. Given other studies linking NMDA receptors to channels with unusually high calcium permeability, this suggestion is consistent with previous data showing that glutamate neurotoxicity depends heavily on extracellular calcium.
2-Amino-5-phosphonovalerate↗