Background: The large-conductance Ca2+-activated K+ (BK(Ca)) channel in the cardiac inner mitochondrial membrane (mitoK(Ca) channel) has been shown to protect the heart against ischemic injury. However, questions about the cardioprotective mechanism and the kinase-mediated regulation of mitoK(Ca) channels remain to be answered.
Methods and results: Flavoprotein fluorescence in guinea pig ventricular myocytes was measured to assay mitoK(Ca) channel activity. The mitochondrial Ca2+ concentration ([Ca2+]m) and membrane potential (DeltaPsi(m)) were measured by loading cells with rhod-2 and JC-1, respectively. Cell death was assessed by trypan blue permeability. The BK(Ca) channel opener NS1619 reversibly increased the flavoprotein oxidation in a concentration-dependent manner. NS1619 (30 micromol/L) attenuated the ouabain (1 mmol/L)-induced elevation of [Ca2+]m with accompanying depolarization of DeltaPsi(m). These effects of NS1619 were completely antagonized by the BK(Ca) channel blocker paxilline (2 micromol/L) but not by the mitochondrial ATP-sensitive K+ (mitoK(ATP)) channel blocker 5-hydroxydecanoate (500 micromol/L). Paxilline, however, failed to block the oxidative effect of diazoxide (100 micromol/L), a mitoK(ATP) channel opener. The combined application of submaximally effective concentrations of NS1619 (10 micromol/L) and diazoxide (30 micromol/L) produced additive effects. NS1619 (30 micromol/L) blunted the rate of cell death during exposure to ouabain; this cardioprotective effect was prevented by paxilline. Activation of cAMP-dependent protein kinase by 8-bromoadenosine 3'5'-cyclic monophosphate (0.5 mmol/L) and forskolin (10 micromol/L) potentiated the NS1619-induced flavoprotein oxidation.
Conclusions: Opening of mitoK(Ca) channels, which is modulated by cAMP-dependent protein kinase, depolarizes the DeltaPsi(m) and attenuates the mitochondrial Ca2+ overload. Our study further indicates that mitoK(Ca) channel activation confers cardioprotection in a manner similar to but independent of mitoK(ATP) channel activation.