Manganese-induced rat pheochromocytoma (PC12) cell death is independent ofcaspase activation

Manganese (Mn) is an essential mineral that at high concentrations can prod uce an irreversible syndrome resembling Parkinson's disease. To examine the mechanism by which Mn elicits its toxic response, we have selected the rat pheochromocytoma cells (PC12) as our model system because it possesses muc h of the biochemical machinery associated with dopaminergic neurons. Win-in duced PC12 cell death is both time and concentration dependent with approxi mately 50% cell survival at 48 hr in the presence of 0.3 mM Mn. To determin e whether oxidative stress contributed to cytotoxicity induced by Mn, lipid peroxidation was assessed in Mn-treated in PC12 cells. The highly sensitiv e HPLC assay that measures the lipid peroxide product, 9-HODE, was used and results of these experiments demonstrate there was no increase in the lipi d peroxidation in cells exposed to 0.3 mM Mn for 24 hr. Mn was found to sti mulate the activation of the apoptotic marker proteins, p38 and caspase-3 w ithin the first 24 hr of treatment. The selective inhibitor of caspase-3, D EVD-CHO, and the nonselective caspase inhibitor, Z-VAD-FMK, however, fail t o prevent Mn-induced PC12 cell death. Studies were performed to determine t he role of mitochondria in initiating or supporting Mn cytotoxicity, becaus e Mn has been reported to cause changes in membrane permeability. Mn caused a decrease in ATP levels in PC12 cells in both a time and concentration de pendent manner. We hypothesize that both apoptosis and necrosis contribute to PC12 cell death although the necrotic events prevail even when the apopt otic signaling is inhibited. (C) 2000 Wiley-Liss, Inc.