A mitochondrial uncoupler increases K_Ca currents but decreases K_v currents in pulmonary artery myocytes

Intracellular free Ca²⁺ concentration ([Ca^2-]_i) and ATP play important roles in the regulation of K^- channels in pulmonary artery (PA) myocytes. Previous studies have demonstrated that hypoxia and the metabolic inhibitor, 2-deoxy-D-glucose, decrease voltage-gated K^- (K_v) currents [I_k(v)] and thereby depolarize PA myocytes; these effects lead to a rise in [Ca²⁺]_i. Here, we used carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP), a protonophore that uncouples mitochondrial respiration from ATP production, to test whether the inhibition of oxidative phosphorylation affects K^- channel activities in rat PA myocytes. Patch-clamp and fluorescentimaging microscopy techniques were used to measure K⁺ currents (I_k) and [Ca^2-]_i, respectively. FCCP (3-5 μM) reversibly raised [Ca²⁺]_i in the presence and absence of external Ca²⁺. This effect was prevented by pretreating the cells with the membrane-permeable Ca²⁺ chelator, 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid-acetoxymethyl ester (BAPTA-AM). This suggests that much of the FCCP-evoked rise in [Ca²⁺]_i was due to Ca²⁺ release from intracellular stores. Brief exposure to FCCP (∼2 min) reversibly enhanced I_K. This augmentation was not influenced by glibenclamide, an ATP-sensitive K⁺ channel blocker, but was eliminated by pretreatment with BAPTA-AM. This implies that the FCCP-evoked rise in [Ca²⁺]; activated Ca²⁺-activated K⁺ (K_ca) channels. Furthermore, in BAPTA-treated cells, longer application (≥6 min) of FCCP reversibly decreased I_K(V) in PA cells bathed in Ca^2--free solution. These results demonstrate that FCCP affects K_ca and K_V channels by different mechanisms. FCCP increases I_K(Ca) by raising [Ca²⁺]_i primarily as a result of Ca²⁺ release, but decreases I_K(V), by a Ca²⁺-independent mechanism, presumably the inhibition of oxidative ATP production.