title: Depolarization of mitochondria in neurons promotes activation of nitric oxide synthase and generation of nitric oxide.
abstract: The diverse signaling events following mitochondrial depolarization in neurons are not clear. We examined for the first time the effects of mitochondrial depolarization on mitochondrial function, intracellular calcium, neuronal nitric oxide synthase (nNOS) activation, and nitric oxide (NO) production in cultured neurons and perivascular nerves. Cultured rat primary cortical neurons were studied on 7-10 days in vitro, and endothelium-denuded cerebral arteries of adult Sprague-Dawley rats were studied ex vivo. Diazoxide and BMS-191095 (BMS), activators of mitochondrial KATP channels, depolarized mitochondria in cultured neurons and increased cytosolic calcium levels. However, the mitochondrial oxygen consumption rate was unaffected by mitochondrial depolarization. In addition, diazoxide and BMS not only increased the nNOS phosphorylation at positive regulatory serine 1417 but also decreased nNOS phosphorylation at negative regulatory serine 847. Furthermore, diazoxide and BMS increased NO production in cultured neurons measured with both fluorescence microscopy and electron spin resonance spectroscopy, which was sensitive to inhibition by the selective nNOS inhibitor 7-nitroindazole (7-NI). Diazoxide also protected cultured neurons against oxygen-glucose deprivation, which was blocked by NOS inhibition and rescued by NO donors. Finally, BMS induced vasodilation of endothelium denuded, freshly isolated cerebral arteries that was diminished by 7-NI and tetrodotoxin. Thus pharmacological depolarization of mitochondria promotes activation of nNOS leading to generation of NO in cultured neurons and endothelium-denuded arteries. Mitochondrial-induced NO production leads to increased cellular resistance to lethal stress by cultured neurons and to vasodilation of denuded cerebral arteries.
authors.affiliation: Department of Pharmacology, Tulane University School of Medicine, Tulane University, New Orleans, Louisiana pkatakam@tulane.edu.
authors.forename: Prasad V G
authors.initials: PV
authors.lastname: Katakam
authors.affiliation: Department of Pharmacology, Tulane University School of Medicine, Tulane University, New Orleans, Louisiana.
authors.forename: Somhrita
authors.initials: S
authors.lastname: Dutta
authors.affiliation: Department of Pharmacology, Tulane University School of Medicine, Tulane University, New Orleans, Louisiana.
authors.forename: Venkata N
authors.initials: VN
authors.lastname: Sure
authors.affiliation: Department of Pharmacology, Tulane University School of Medicine, Tulane University, New Orleans, Louisiana.
authors.forename: Samuel M
authors.initials: SM
authors.lastname: Grovenburg
authors.affiliation: Department of Pharmacology, Tulane University School of Medicine, Tulane University, New Orleans, Louisiana.
authors.forename: Angellica O
authors.initials: AO
authors.lastname: Gordon
authors.affiliation: Department of Pharmacology, Tulane University School of Medicine, Tulane University, New Orleans, Louisiana.
authors.forename: Nicholas R
authors.initials: NR
authors.lastname: Peterson
authors.affiliation: Department of Pharmacology, Tulane University School of Medicine, Tulane University, New Orleans, Louisiana.
authors.forename: Ibolya
authors.initials: I
authors.lastname: Rutkai
authors.affiliation: Department of Pharmacology, Tulane University School of Medicine, Tulane University, New Orleans, Louisiana.
authors.forename: David W
authors.initials: DW
authors.lastname: Busija
mesh_terms: D000818:Animals; D001578:Benzopyrans / Q000494:pharmacology; D002478:Cells, Cultured; D002536:Cerebral Arteries / Q000187:drug effects / Q000294:innervation / Q000378:metabolism*; D003981:Diazoxide / Q000494:pharmacology; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology; D007191:Indazoles / Q000494:pharmacology; D008297:Male; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology*; D026602:Nitrergic Neurons / Q000187:drug effects / Q000201:enzymology*; D009569:Nitric Oxide / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019899:Paracrine Communication / Q000187:drug effects; D010766:Phosphorylation; D015221:Potassium Channels / Q000819:agonists / Q000378:metabolism; D061251:Primary Cell Culture; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism; D012694:Serine; D015398:Signal Transduction; D014664:Vasodilation / Q000187:drug effects
chemical_list: C428117:BMS 191095; D001578:Benzopyrans; D004791:Enzyme Inhibitors; D007093:Imidazoles; D007191:Indazoles; D015221:Potassium Channels; D017382:Reactive Oxygen Species; C117780:mitochondrial K(ATP) channel; D009569:Nitric Oxide; D012694:Serine; D052248:Nitric Oxide Synthase Type I; C496322:Nos1 protein, rat; D003981:Diazoxide; C080122:7-nitroindazole
keywords: BMS-191095; diazoxide; membrane potential; mitochondrial ATP-sensitive potassium channels; superoxide anion