Endothelin-1 regulates cytosolic and nuclear Ca2+ in human endocardial endothelium

Endocardial endothelium is far less studied than its systemic and pulmonary counterparts and its role in cardiac function is not well known. In the study reported here, we succeeded in isolating and culturing endocardial endothelial cells (EECs) from 21-week-old human fetal heart and verified whether endothelin-1 (ET-1) receptors are present on these cells and whether their activation regulates cytosolic ([Ca](c)) and nuclear calcium ([Ca](n)). Using fluo-3 Ca2+ measurement and three-dimension confocal microscopy techniques, superfusion of fetal human EECs from right and left ventricles with increasing concentrations of ET-1 induced a dose-dependent sustained increase of free cytosolic and nuclear Ca2+ levels with an EC50 near 10(-10) M and 10(-11) M. respectively. The sustained increase of cytosolic and nuclear Ca2+ by ET-1 in both EEC preparations was completely blocked by the calcium chelator ethylene glycol-his (beta -aminoethylether)-N,N,N',N'-tetra-acetic acid (EGTA) but was insensitive to the L-type Ca2+ channel blocker, nifedipine. The effect of ET-1 was prevented by the ETA-receptor antagonist BQ 123. However the ETB-receptor antagonist BQ 788 had no effect. Our results suggest that ET-1 receptors are present in human EECs and that their stimulation induces sustained Ca2+ influx through ETA-receptor stimulation of a nifedipine-insensitive Ca2+ channel, probably the R-type Ca2+ channel. The presence of Ca2+-dependent responses to ET-1 supports a possible modulatory role of the endocardial endothelium in myocardial functions.