Inhibition of cytochrome P-450 reduces voltage-gated K+ currents in pulmonary arterial myocytes

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Abstract
Cytochrome P-450 (P-450) is a NADPH-requiring and O2-dependent monooxygenase system. It is present in lung and has been postulated to act as an O2 sensor in hypoxic pulmonary vasoconstriction. To determine whether P-450 is involved in the regulation of voltage-gated K+ (KV) channel activity in pulmonary artery (PA) myocytes, we used the whole cell patch-clamp technique to evaluate the effects of P-450 inhibitors on KV channel currents (IKV) and membrane potential (Em). Bath application of the P-450 inhibitors clotrimazole, miconazole, and 1-aminobenzotriazole (1-ABT) significantly and reversibly inhibited steady-state IKV (IKss) and depolarized PA cells bathed in either Ca(2+)-containing (1.8 mM) or Ca(2+)-free [0.5-1 mM ethylene glycol-bis(beta-aminoethyl ether)-N,N,N'N'-tetraacetic acid present] bath solution. Clotrimazole (1 microM), miconazole (10 microM), and 1-ABT (1 mM) reversibly reduced IKss, elicited by a test potential of +80 mV, by 40, 70, and 31%, respectively. Pretreatment of PA smooth muscle cells with 10 mM 4-aminopyridine (4-AP) prevented the subsequent inhibitory effect of clotrimazole on IKV. However, pretreatment of the cells with 1 mM tetraethylammonium negligibly altered the effects of miconazole on IKV and Em. In current-clamp (I = 0) measurements, clotrimazole depolarized PA myocytes by 9 and 11 mV during perfusion with Ca(2+)-containing and Ca(2+)-free bath solution, respectively. 1-ABT also caused a 9-mV depolarization in PA myocytes bathed in Ca(2+)-free solution. These effects are similar to those induced by hypoxia, reduced glutathione, and 4-AP. Clotrimazole also decreased IKV and depolarized mesenteric arterial myocytes. These data raise the possibility that the P-450 system, due to its influence on IKV and sensitivity to O2 tension and NADPH, may play a role in linking the regulation of pulmonary vascular tone to the alteration of cellular redox status through a common pathway of KV channel activity.