Diazoxide triggers cardioprotection against apoptosis induced by oxidative stress

Although mitochondrial ATP-sensitive potassium (mitoK(ATP)) channels have been reported to reduce the extent of apoptosis, the critical timing of mitoK(ATP) channel opening required to protect myocytes against apoptosis remains unclear. In the present study, we examined whether the mitoK(ATP) channel serves as a trigger of cardioprotection against apoptosis induced by oxidative stress. Apoptosis of cultured neonatal rat cardiomyocytes was determined by flow cytometry (light scatter and propidium iodide/annexin V-FITC fluorescence) and by nuclear staining with Hoechst 33342. Mitochondrial membrane potential (DeltaPsi) was measured by flow cytometry of cells stained with rhodamine-123 (Rh-123). Exposure to H2O2 (500 muM) induced apoptosis, and the percentage of apoptotic cells increased progressively and peaked at 2 h. This H2O2-induced apoptosis was associated with the loss of DeltaPsi, and the time course of decrease in Rh-123 fluorescence paralleled that of apoptosis. Pretreatment of cardiomyocytes with diazoxide (100 muM), a putative mitoK(ATP) channel opener, for 30 min before exposure to H2O2 elicited transient and mild depolarization of DeltaPsi and consequently suppressed both apoptosis and DeltaPsi loss after 2-h exposure to H2O2. These protective effects of diazoxide were abrogated by the mitoK(ATP) channel blocker 5-hydroxydecanoate (500 muM) but not by the sarcolemmal K-ATP channel blocker HMR-1098 (30 muM). Our results suggest for the first time that diazoxide-induced opening of mitoK(ATP) channels triggers cardioprotection against apoptosis induced by oxidative stress in rat cardiomyocytes.