Nitric oxide from neuronal nitric oxide synthase sensitises neurons to hypoxia-induced death via competitive inhibition of cytochrome oxidase.
Hypoxia/ischaemia is known to trigger neuronal death, but the role of neuronal nitric oxide synthase (nNOS) in this process is controversial. Nitric oxide (NO) inhibits cytochrome oxidase in competition with oxygen. We tested whether NO derived from nNOS synergises with hypoxia to induce neuronal death by inhibiting mitochondrial cytochrome oxidase. Sixteen hours of hypoxia (2% oxygen) plus deoxyglucose (an inhibitor of glycolysis) caused extensive, excitotoxic death of neurons in rat cerebellar granule cell cultures. Three different nNOS inhibitors (including the selective inhibitor N-4S-4-amino-5-2-aminoethyl-aminopentyl-N'-nitroguanidine) decreased this neuronal death by half, indicating a contribution of nNOS to hypoxic death. The selective nNOS inhibitor did not, however, block neuronal death induced either by added glutamate or by added azide (an uncompetitive inhibitor of cytochrome oxidase), indicating that nNOS does not act downstream of glutamate or cytochrome oxidase. Hypoxia plus deoxyglucose-induced glutamate release and neuronal depolarisation, and the nNOS inhibitor decreased this. Hypoxia inhibited cytochrome oxidase activity in the cultures, but a selective nNOS inhibitor prevented this inhibition, indicating NO from nNOS was inhibiting cytochrome oxidase in competition with oxygen. These data indicate that hypoxia synergises with NO from nNOS to induce neuronal death via cytochrome oxidase inhibition causing neuronal depolarisation. This mechanism might contribute to ischaemia/stroke-induced neuronal death in vivo.
Jekabsone
Aiste
A
Department of Biochemistry, University of Cambridge, Cambridge, UK.
Neher
Jonas J
JJ
Borutaite
Vilmante
V
Brown
Guy C
GC
D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003847:Deoxyglucose / Q000633:toxicity; D003576:Electron Transport Complex IV / Q000037:antagonists & inhibitors* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D018698:Glutamic Acid / Q000378:metabolism; D006162:Guanylate Cyclase / Q000378:metabolism; D002534:Hypoxia, Brain / Q000378:metabolism*; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D009569:Nitric Oxide / Q000494:pharmacology / Q000502:physiology*; D052248:Nitric Oxide Synthase Type I / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar
D004791:Enzyme Inhibitors; D009569:Nitric Oxide; D018698:Glutamic Acid; D003847:Deoxyglucose; D052248:Nitric Oxide Synthase Type I; D003576:Electron Transport Complex IV; D006162:Guanylate Cyclase