Ca2+-transients induced by K+ channel openers in isolated coronary capillaries

To study the role of endothelial ATP-sensitive K+ channels in the regulation of vascular tone we examined the intracellular calcium concentration ([Ca2+]i) in coronary capillaries consisting only of endothelial cells. Coronary capillary fragments were isolated enzymatically from the guinea-pig heart and [Ca2+]i was determined by microfluorometry of fura-2 loaded cells. Low concentrations of the K+ channel opener diazoxide, which caused pronounced glibenclamide-sensitive hyperpolarization in capillaries, induced a rapid, transient rise in [Ca2+]i followed by a sustained elevation of [Ca2+]i (19 of 40 experiments). [Ca2+]i in the endothelial cells increased from 32 +/- 7 nM at rest to 66 +/- 11 nM at the peak (n = 19). One third of the [Ca2+]i-transients showed irregular oscillations of [Ca2+]i. No significant difference in the [Ca2+]i-response induced by 100 nM or 1 muM diazoxide was found. Similar results were obtained with the K+ channel opener rilmakalim. Simultaneous measurements of the membrane potential and [Ca2+]i with fluorometric methods indicated that the hyperpolarization but not the [Ca2+]i-transient could be repeatedly induced in a single capillary by the K+ channel openers. Electrophysiological recordings of the membrane potential using the "perforated patch" method (n = 4), showed that rilmakalim (1 muM) induced hyperpolarization of capillaries towards the K+ equilibrium potential, confirming our fluorometric measurements. In conclusion, for the first time, these data indicate that K+ channel openers induce [Ca2+]i-transients in microvascular endothelial cells. This raises the possibility that these drugs not only act as synthetic vasoactive factors via hyperpolarizing smooth muscle cells but also via NO release of microvascular endothelial cells. Interestingly, only 100 nM diazoxide was sufficient for a maximal response, suggesting the expression of a new type of KATP-channel in coronary capillaries characterised by high sensitivity to diazoxide.