Two distinct mechanisms of nitric oxide-mediated neuronal cell death show thiol dependency

To better understand the mechanism(s) underlying nitricoxide (· NO)-mediated toxicity, in the presence and absenceof concomitant oxidant exposure, postmitotic terminally differentiatedNT2N cells, which are incapable of producing · NO, wereexposed to PAPA-NONOate (PAPA/NO) and 3-morpholinosydnonimine (SIN-1).Exposure to SIN-1, which generated peroxynitrite in the range of25-750 nM/min, produced a concentration- and time-dependentdelayed cell death. In contrast, a critical threshold concentration(>440 nM/min) was required for · NO to produce significantcell injury. Examination of cells by electron microscopy shows alargely necrotic injury after peroxynitrite exposure but mainlyapoptotic-like morphology after · NO exposure. Cellularlevels of reduced thiols correlated with cell death, and pretreatmentwith N-acetylcysteine (NAC) fully protected from cell death ineither PAPA/NO or SIN-1 exposure. NAC given within the first 3 hposttreatment further delayed cell death and increased theintracellular thiol level in SIN-1 but not · NO-exposedcells. Cell injury from · NO was independent of cGMP,caspases, and superoxide or peroxynitrite formation. Overall, exposureof non-· NO-producing cells to · NO orperoxynitrite results in delayed cell death, which, although occurringby different mechanisms, appears to be mediated by the loss ofintracellular redox balance.