Clotrimazole inhibits the recombinant human cardiac L-type Ca2+ channel α1C subunit

1 Clotrimazole (CLT) is an antimycotic agent with a potential role in the treatment of cancer. Whole-cell patch clamp recordings and Fura-2 AM fluorescence measurements were used to investigate the inhibition by CLT of recombinant human cardiac L-type Ca2+ channel alpha(1C) subunits, stably expressed in human embryonic kidney (HEK 293) cells. 2 CLT (100 nmol 1(-1) to 25 mu mol 1(-1)) reduced Ca2+ channel currents in a concentration-dependent manner. Inhibition was neither use- or voltage-dependent. The effects of CLT were rapid and maximal effects were attained within 3 min. Application of CLT also caused an acceleration of apparent Ca2+ channel current inactivation. 3 Basal current density and the degree of inhibition due to CLT were not significantly altered by pretreating cells with 3 mmol 1(-1) 1-aminobenzotriazole for 1 h, or by dialysing cells for 10 min with 2 mmol 1(-1) alpha-napthoflavone via the patch pipette, suggesting that the inhibitory action of CLT was not due to inhibition of cytochrome P-450. 4 CLT (10 mu mol 1(-1)) did not influence [Ca2+](i), as determined by Fura-2 AM fluorescence measurements. 5 Dialysing cells for 10 min with the non-specific serine/threonine kinase inhibitor H-7 (10 mu mol 1(-1)) was without effect on basal current density or on the inhibitory response to 10 mu mol 1(-1) CLT, indicating that CLT is not acting via an indirect effect on these kinases. 6 These data suggest that CLT exerts a direct blocking effect on the alpha(1C) subunit at therapeutic concentrations. This effect may explain the abbreviation of the action potential duration by CLT observed in cardiac myocytes.