title: Involvement of Na+ and Ca2+ channel activation and resultant nitric oxide synthesis in glutamate-mediated hypoxic injury in rat cerebrocortical slices.
abstract: The role of Na+ and Ca2+ channels in glutamate-mediated hypoxic injury was investigated in slices of the rat cerebral cortex. Hypoxic injury was determined by mitochondrial reduction of 3-(4,5-dimethyl-2-thiazol)-2,5-diphenyltetrazolium bromide after exposure of brain slices to 30-min of hypoxia/glucose deprivation followed by 3-h of reoxygenation. Endogenous glutamate release was markedly elevated during hypoxia/glucose deprivation, but it returned almost to basal level during reoxygenation. Hypoxic injury was prevented by MK-801 or 6-cyano-7-nitroquinoxaline-2,3-dione. Combined treatment with omega-conotoxin GVIA, omega-agatoxin IVA, and tetrodotoxin reversed the hypoxic injury, although none of these agents alone or nifedipine was effective. Moreover, a novel Na+/Ca2+ channel blocker NS-7 [4-(4-fluorophenyl)-2-methyl-6-(5-piperidinopentyloxy) pyrimidine hydrochloride] significantly inhibited the hypoxic injury. Several inhibitors of nitric oxide synthase also blocked the hypoxic injury. Consistently, nitric oxide synthesis, as estimated from cyclic GMP formation in the extracellular fluids, was enhanced during hypoxia/glucose deprivation. NS-7 and other Na+ and Ca2+ channel blockers suppressed the enhancement of nitric oxide synthesis, although these compounds alone, or in combination, did not reduce hypoxic glutamate release. These findings suggest that hypoxic injury in rat cerebrocortical slices is triggered by glutamate and subsequent enhancement of nitric oxide synthesis through activation of both Na+ and Ca2+ channels. Thus, the simultaneous blockade of both Na+ channel as well as N-type and P/Q-type Ca2+ channels is required to sufficiently reverse the hypoxic injury.
authors.affiliation: Research Laboratories, Nippon Shinyaku Co., Ltd., Kyoto, Japan.
authors.forename: M
authors.initials: M
authors.lastname: Oka
authors.forename: Y
authors.initials: Y
authors.lastname: Itoh
authors.forename: Y
authors.initials: Y
authors.lastname: Ukai
mesh_terms: D018750:6-Cyano-7-nitroquinoxaline-2,3-dione / Q000494:pharmacology; D000818:Animals; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism*; D002540:Cerebral Cortex / Q000187:drug effects / Q000378:metabolism*; D016291:Dizocilpine Maleate / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D018698:Glutamic Acid / Q000378:metabolism*; D002534:Hypoxia, Brain / Q000378:metabolism*; D066298:In Vitro Techniques; D008297:Male; D009543:Nifedipine / Q000494:pharmacology; D009569:Nitric Oxide / Q000096:biosynthesis*; D011743:Pyrimidines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D015222:Sodium Channels / Q000187:drug effects / Q000378:metabolism*; D013779:Tetrodotoxin / Q000494:pharmacology; D020867:omega-Agatoxin IVA / Q000494:pharmacology; D020866:omega-Conotoxin GVIA / Q000494:pharmacology
chemical_list: C106144:4-(4-fluorophenyl)-2-methyl-6-(5-piperidinopentyloxy)pyrimidine hydrochloride; D015220:Calcium Channels; D018691:Excitatory Amino Acid Antagonists; D011743:Pyrimidines; D015222:Sodium Channels; D020867:omega-Agatoxin IVA; D009569:Nitric Oxide; D018698:Glutamic Acid; D013779:Tetrodotoxin; D016291:Dizocilpine Maleate; D018750:6-Cyano-7-nitroquinoxaline-2,3-dione; D020866:omega-Conotoxin GVIA; D009543:Nifedipine