Role of glutamate receptors and voltage-dependent calcium and sodium channels in the extracellular glutamate/aspartate accumulation and subsequent neuronal injury induced by oxygen/glucose deprivation in cultured hippocampal neurons.
Ischemia is believed to induce neuronal damage by causing a sustained increase in the level of extracellular excitatory amino acids. In our study, we have examined the relationship between oxygen/glucose deprivation-induced changes in extracellular glutamate/aspartate level and subsequent neuronal injury by pharmacological manipulation of glutamate receptors and calcium and sodium channels. Cultured hippocampal neurons were exposed to combined deprivation of oxygen/glucose for 40 to 50 min. These cultures developed acute neuronal swelling and widespread neuronal degeneration over the next 20 hr. The extracellular levels of glutamate and aspartate at the end of the oxygen/glucose deprivation period were measured by high-performance liquid chromatography, and neuronal injury was assessed by lactate dehydrogenase efflux assay after subsequent aerobic incubation of the cells in normal medium for 20 hr. Both N-methyl-D-aspartate and non- N-methyl-D-aspartate receptor antagonists attenuated the extracellular level of glutamate/aspartate and the neuronal injury. L-type, N-type and P-type calcium channel blockers each significantly attenuated the neuronal injury, although the increase in the extracellular glutamate/aspartate was not significantly inhibited by any subtype-specific calcium channel blocker alone. A combination of calcium channel blockers of the three subtypes showed the most prominent neuroprotective effect and inhibited glutamate release. The sodium channel blocker tetrodotoxin also attenuated both glutamate efflux and neuronal injury. These observations suggest that the overactivation of glutamate receptors, calcium channels and sodium channels leads to excitotoxic neuronal injury through enhancing glutamate efflux into the extracellular space under the condition of oxygen/glucose deprivation.
Kimura
M
M
Eisai Tsukuba Research Laboratories, 5-1-3 Tokodai, Tsukuba, Ibaraki 300-2635, Japan.
Sawada
K
K
Miyagawa
T
T
Kuwada
M
M
Katayama
K
K
Nishizawa
Y
Y
D000818:Animals; D001224:Aspartic Acid / Q000187:drug effects* / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D015687:Cell Hypoxia; D002478:Cells, Cultured; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology*; D005947:Glucose / Q000172:deficiency / Q000494:pharmacology; D018698:Glutamic Acid / Q000187:drug effects* / Q000378:metabolism; D006624:Hippocampus / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D007511:Ischemia / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D009474:Neurons / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D010100:Oxygen / Q000009:adverse effects; D051381:Rats; D017208:Rats, Wistar; D017470:Receptors, Glutamate / Q000378:metabolism* / Q000502:physiology; D062687:Sodium Channel Agonists; D015222:Sodium Channels / Q000378:metabolism* / Q000502:physiology
D002121:Calcium Channel Blockers; D018690:Excitatory Amino Acid Agonists; D017470:Receptors, Glutamate; D062687:Sodium Channel Agonists; D015222:Sodium Channels; D001224:Aspartic Acid; D018698:Glutamic Acid; D007770:L-Lactate Dehydrogenase; D005947:Glucose; D010100:Oxygen