Distinct myoprotective roles of cardiac sarcolemmal and mitochondrial KATP channels during metabolic inhibition and recovery

The protective roles of sarcolemmal (sarc) and mitochondrial (mito) K-ATP channels are unclear despite their apparent importance to ischemic preconditioning. We examined these roles by monitoring intracellular calcium ([Ca](int)), using fura-2 and fluo-3, in enzymatically isolated rat right ventricular myocytes. Myocyte mortality, estimated using a trypan blue assay, changed approximately in parallel with changes in [Ca](int). Chemically induced hypoxia (CIH), induced by application of cyanide and 2-deoxy-glucose, caused a steady rise in [Ca](int). Calcium increased more rapidly on 'reoxygenation' by return to control solutions. The protein kinase C (PKC) activator PMA abolished both phases of calcium increase. The mitoK(ATP) channel-selective blocker 5-hydroxydecanoate partially prevented the PMA-induced protection during CIH, but not during reoxygenation. In contrast, HMR 1098, a sarcK(ATP) channel-selective blocker, abolished protection only during the reoxygenation. Adenosine (A(1)) receptor activation prevented or reduced increases in [Ca](int) and improved cell viability via a PKC and mito/sarcK(ATP) channel-dependent mechanism. PKC-dependent protection against cytoplasmic calcium increases was also observed in a human cell line (tsA201) transiently expressing sarcK(ATP) channels. Protection was abolished only during the reoxygenation phase by the amino acid substitution (T180A) in the pore-forming Kir6.2 subunit, a mutation previously shown to prevent PKC-dependent modulation. Our data suggest that sarc and mitoK(ATP) channel populations play distinct protective roles, triggered by PKC and/or adenosine, during chemically induced hypoxia/reoxygenation.