Mitochondrial ATP-sensitive potassium channels attenuate matrix Ca2+ overload during simulated ischemia and reperfusion -: Possible mechanism of cardioprotection

Mitochondrial ATP-sensitive potassium (mitoK(ATP)) channels play a key role in ischemic preconditioning of the heart. However, the mechanism of cardioprotection remains controversial. We measured rhod-2 fluorescence in adult rabbit ventricular cardiomyocytes as an index of mitochondrial matrix Ca2+ concentration ([Ca2+](m)), using time-lapse confocal microscopy. To simulate ischemia and reperfusion (I/R), cells were exposed to metabolic inhibition (50 minutes) followed by washout with control solution. Rhod-2 fluorescence gradually increased during simulated ischemia and rose even further with reperfusion. The mitoK(ATP) channel opener diazoxide attenuated the accumulation of [Ca2+](m) during simulated I/R (EC50= 18 mu mol/L). These effects of diazoxide were blocked by the mitoK(ATP) channel antagonist 5-hydroxydecanoate (5HD). In contrast, inhibitors of the mitochondrial permeability transition (MPT), cyclosporin A and bongkrekic acid, did not alter [Ca2+](m) accumulation during ischemia, but markedly suppressed the surge in rhod-2 fluorescence during reperfusion. Measurements of mitochondrial membrane potential, Delta Psi (m), permeabilized myocytes revealed that diazoxide depolarized Delta Psi (m) (by 12% at 10 mu mol/L, P <0.01) in a 5HD-inhibitable manner. Our data support the hypothesis that attenuation of mitochondrial Ca2+ overload, as a consequence of partial mitochondrial membrane depolarization by mitoK(ATP) channels, underlies cardioprotection. Furthermore, mitoK(ATP) channels and the MPT differentially affect mitochondrial calcium homeostasis: mitoK(ATP) channels suppress calcium accumulation during, I/R, while the MPT comes into play only upon reperfusion.