Nitric oxide induces oxidative stress and apoptosis in neuronal cells

Within the central nervous system and under normal conditions, nitric oxide (NO) is an important physiological signaling molecule. When produced in la rge excess, NO also displays neurotoxicity. In our previous report, we have demonstrated that the exposure of neuronal cells to NO donors induced apop totic cell death, while pretreatment with free radical scavengers L-ascorbi c acid 2-[3,4-dihydro-2,5,7,8-tetramethyl-2-(4,8,12-trimethyltridecyl)-2H-1 -benzopyran-6-yl-hydrogen, phosphate] potassium salt (EPC-KZ) or superoxide dismutase attenuated apoptosis effectively, suggesting that reactive oxyge n species (ROS) may be involved in the cascade of events leading to apoptos is. In the present investigation, we directly studied the kinetic generatio n of ROS in NO-treated neuronal cells by flow cytometry using 2',7'-dichlor o-fluorescein diacetate and dihydrorhodamine 123 as odor-sensitive fluoresc ence probes. The results indicated that exposure of cerebellar granule cell s to the NO donor S-nitroso-N-acetylpenicillamine (SNAP) induced oxidative stress, which was characterized by the accumulation of cytosolic and mitoch ondrial ROS, the increase in the extracellular hydrogen peroxide level, and the formation of lipid peroxidation products. SNAP treatment also induced apoptotic cell death as confirmed by the formation of cytosolic mono- and o ligonucleosomes. Pretreating cells with the novel antioxidant EPC-KI effect ively prevented oxidative stress induced by SNAP, and attenuated cells from apoptosis. (C) 2000 Elsevier Science B.V. All rights reserved.