Inhibitors of ATP-sensitive potassium channels in guinea pig isolated ischemic hearts

Abstract

During heart ischemia, ATP-sensitive potassium channels in the sarcolemmal membrane (sarcK_ATP) open and cause shortening of the action potential duration. This creates heterogeneity of repolarization, being responsible for the development of re-entry arrhythmias and sudden cardiac death. Therefore, the aim is to develop selective blockers of the cardiac sarcK_ATP channel. In the present study we established an in vitro model and classified 5 K_ATP channel inhibitors with respect to their potency and selectivity between cardiomyocytes and the coronary vasculature and compared the results with inhibition of Kir6.2/SUR2A channels expressed in HEK293 cells, recorded with the Rb⁺-efflux methods. We used Langendorff-perfused guinea pig hearts, where low-flow ischemia plus hypoxia was performed by reducing the coronary flow (CF) to 1.2 ml/min and by gassing the perfusion solution with N₂ instead of O₂. Throughout the experiment, the monophasic action potential duration at 90% repolarization (MAPD₉₀) was recorded. In separate experiments, high-flow hypoxia was produced by oxygen reduction in the perfusate from 95% to 20%, which caused an increase in the coronary flow. Under normoxic conditions, the substances glibenclamide, repaglinide, meglitinide, HMR 1402 and HMR 1098 (1 μM each) reduced the CF by 34%, 38%, 19%, 12% and 5%, respectively. The hypoxia-induced increase in CF was inhibited by the compounds half-maximally at 25 nM, approximately 200 nM, 600 nM, approximately 9 μM and >100 μM, respectively. In control experiments after 5 min low-flow ischemia plus hypoxia, the MAPD₉₀ shortened from 121±2 to 99±2 ms (n=29). This shortening was half-maximally inhibited by the substances at concentrations of 95 nM, 74 nM, 400 nM, 110 nM and 550 nM, respectively. In HEK293 cells the Rb⁺-efflux through KIR6.2/SUR2A channels was inhibited by the compounds with IC₅₀ values of 21 nM, 67 nM, 205 nM, 60 nM and 181 nM, respectively. In summary, the present data demonstrate that the sulfonylurea glibenclamide, and the carbamoylbenzoic acid derivatives repaglinide and meglitinide are unselective blockers of K_ATP channels in cardiac cells and in the cardiac vascular system, whereas the sulfonylthioureas HMR 1402, and especially HMR 1098 selectively blocked the cardiac sarcK_ATP channel. Blockade of Kir6.2/SUR2A channels in HEK293 cells occurred with comparable efficacy as in the cardiac tissue, indicating that the expression system is suited for screening for novel inhibitors.