Potentiation of P1075-induced K+ channel opening by stimulation of adenylate cyclase in rat isolated aorta.

Abstract

1. The effects of analogues and stimulators of cyclic AMP on the 86Rb+ efflux-stimulating and binding properties of P1075, an opener of ATP-dependent potassium channels, were studied in rat aortic rings. The increase in 86Rb+ efflux stimulated by P1075 was taken as a qualitative measure of K+ channel opening. 2. Forskolin, a direct activator of adenylate cyclase, isobutylmethylxanthine (IBMX), a phosphodiesterase inhibitor, and dibutyryl-cyclic AMP (db-cyclic AMP), a membrane permeant cyclic AMP-analogue, relaxed rat aortic rings contracted by noradrenaline with EC50 values of 0.06, 2 and 10 microM, respectively. 3. Forskolin, IBMX and db-cyclic AMP produced concentration-dependent increases of the 86Rb+ efflux induced by P1075 (50 nM) by up to twofold with EC50 values of about 0.1, 1.7 and 81 microM. At these concentrations the agents had little effect on the basal rate of 86Rb+ efflux. 4. The 86Rb+ efflux produced by P1075 in the presence of the cyclic AMP stimulators was inhibited by glibenclamide, a blocker of ATP-sensitive potassium channels. 5. IBMX (100 microM) induced a leftward shift of the concentration-86Rb+ efflux curve of P1075 without increasing the maximum. The enhancements of P1075-stimulated 86Rb+ efflux produced by combinations of forskolin and IBMX were either additive or less than additive. 6. The protein kinase A inhibitor, H-89, inhibited P1075-stimulated 86Rb+ efflux in the presence of IBMX significantly more than in the absence of IBMX, suggesting that the effect of increased cyclic AMP levels is mediated by protein kinase A. 7. At high concentrations, forskolin and IBMX slightly increased basal 86Rb+ efflux and inhibited the tracer efflux induced by P1075.8. Binding of [3H]-P1075 to rat aortic rings was either unaffected or inhibited by forskolin, IBMX and db-cyclic AMP.9. This study shows that moderate stimulation of the cyclic AMP system potentiates the K+ channel opening effect of P1075 by activation of protein kinase A. The fact that binding of [3H]-P1075 remains unchanged or is diminished favours the hypothesis that the K'channel openers activate ATP-dependent K+ channels by an indirect mechanism.