Neurotoxic nitric oxide rapidly depolarizes and permeabilizes mitochondria by dynamically opening the mitochondrial transition pore.
Exposure of SH-SY5Y neuroblastoma or rat cortical neurons to diethylenetriamine-NO (DETA-NO) rapidly depolarized mitochondria. In SH-SY5Y DETA-NO activated caspase 3 and produced cell death. Mitochondrial depolarization in SH-SY5Y was visualized both with JC-1 accumulation and as dequenching of calcein fluorescence in mitochondria initially loaded with calcein-AM and tetramethylrhodamine methyl ester (TMRM). Calcein/TMRM-visualized mitochondrial depolarization was prevented by cyclosporin A (CsA) or approximately two-fold increased levels of BclXL protein. Dynamic imaging of mitochondrial potential (Deltapsi M) with TMRM showed that DETA-NO induced cycles of mitochondrial depolarization/repolarization ("flickering"). Fifteen-30 min of DETA-NO exposure caused high-frequency flickering with small peak size; 2 h of DETA-NO produced large peaks with prolonged depolarization. NO-induced flickering but not that from Bax was blocked by the calcium uniporter antagonist Ru360. Our findings show rapid-onset, dynamic regulation of Deltapsi M by NO, implying that neuroprotective therapies for brain ischemia target cell death processes downstream of effects of NO on mitochondria.
Kindler
Dean D
DD
Center for the Study of Neurodegenerative Diseases and Department of Neurology, University of Virginia, Charlottesville, VA 22908, USA.
Thiffault
Christine
C
Solenski
Nina J
NJ
Dennis
Jameel
J
Kostecki
Vanessa
V
Jenkins
Russell
R
Keeney
Paula M
PM
Bennett
James P
JP
D000818:Animals; D002545:Brain Ischemia / Q000188:drug therapy / Q000378:metabolism* / Q000503:physiopathology; D015220:Calcium Channels; D002135:Calcium-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D053148:Caspase 3; D020169:Caspases / Q000187:drug effects / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects / Q000502:physiology*; D002463:Cell Membrane Permeability / Q000187:drug effects / Q000502:physiology; D016572:Cyclosporine / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005456:Fluorescent Dyes; D006801:Humans; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism; D007473:Ion Channels / Q000187:drug effects / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D009410:Nerve Degeneration / Q000188:drug therapy / Q000378:metabolism* / Q000503:physiopathology; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism* / Q000633:toxicity; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D051381:Rats; D012235:Rhodamines / Q000494:pharmacology; D017975:Ruthenium Compounds / Q000494:pharmacology; D014226:Triazenes / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D051020:bcl-X Protein
C105351:1-hydroxy-2-oxo-3,3-bis(2-aminoethyl)-1-triazene; C494811:BCL2L1 protein, human; C494813:Bcl2l1 protein, rat; D015220:Calcium Channels; D002135:Calcium-Binding Proteins; D005456:Fluorescent Dyes; D007473:Ion Channels; D019253:Proto-Oncogene Proteins c-bcl-2; D012235:Rhodamines; C112020:Ru 360; D017975:Ruthenium Compounds; D014226:Triazenes; D051020:bcl-X Protein; C034703:mitochondrial calcium uniporter; C401833:tetramethylrhodamine methyl ester; D009569:Nitric Oxide; D016572:Cyclosporine; C505407:CASP3 protein, human; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases