Pharmacological Evidence that Cromakalim Inhibits $Ca^{2+}$ Release from Intracellular Stores in Porcine Coronary Artery

In the present study, it was aimed to further indentify the intracellular action mechansm of cromakalim and levcromakalim in the porcine coronary artery. In intact porcine coronary arterial strips loaded with fura-2/AM, acetylcholine caused an increase in intracellular free $Ca^{2+}$ $([Ca^{2+}]_i)$ in association with a contraction in a concentration-dependent manner. Cromakalim (1 ${\mu}M$) caused a reduction in acetylcholine-induced increased $[Ca^{2+}]_i$ not only in the mormal physiological salt solution (PSS) but also in $Ca^{2+}$-free PSS (containing 1 mM EGTA). In the skinned strips prepared by exposure of tissue to 20 .${\mu}M$ B-escin, inositol 1,4,5-trisphosphate ($IP_3$) evoked an increase in $[Ca^{2+}]_i$, but it was without effect on the intact strips. The $IP_3$-induced increase in $[Ca^{2+}]_i$ was inhibited by cromakalim by 78% and levcromakalim by 59% (1 .${\mu}M$, each). Pretreatment with glibenclamide (a blocker of ATP-sensitive $K^+$ channels, 10 .${\mu}M$) and apamin (a blocker of small conductance $Ca^{2+}$-activated $K^+$ channels, 1 .${\mu}M$) strongly blocked the effect of cromakalim and levcromakalim. However, charybdotoxin (a blocker of large conductance $Ca^{2+}$-activated $K^+$ channels, 1 .${\mu}M$) was without effect. In addition, cromakalim inhibited the $GTP{\gamma}S$ (100 .${\mu}M$, non-hydrolysable analogue of GTP)-induced increase in $[Ca^{2+}]_i$. Based on these results, it is suggested that cromakalim and levcromakalim exert a potent vasorelaxation, in part, by acting on the $K^+$ channels of the intracellular sites (e.g., sarcoplasmic reticulum membrane), thereby, resulting in decrease in release of $Ca^{2+}$ from the intracellular storage site.