17-BETA-ESTRADIOL INCREASES INTRACELLULAR FREE CALCIUM CONCENTRATIONSOF HUMAN VASCULAR ENDOTHELIAL-CELLS AND MODULATES ITS RESPONSES TO ACETYLCHOLINE

In this study, we have investigated the effect of (17)beta-estradiol ( E2) on intracellular free calcium concentrations ([Ca2+](i)) in human umbilical vein endothelial cells (HUVEC) using fura-2 fluorescence, E2 at concentrations of 1nM-1 mu M was added subsequently to HUVEC cultu res which were either deprived of estrogens or preincubated with E2 (1 00 nM) for 23 hours, In both groups of cultures, E2 stimulated signifi cant increases in [Ca2+](i) in a dose-dependent manner, The effects we re more prominent in E2-deprived cells, Preincubation of cells with ta moxifen or the presence of it in the buffer during the experiments did not inhibit the response of the cells to E2, Experiments performed in Ca2+ free/EGTA buffer yielded transient increases in [Ca2+](i) sugges ting release of Ca2+ from intracellular stores was responsible for the initial peak, while sustained elevations were supported by Ca2+ influ x from the extracellular space, Addition of La3+ abolished the sustain ed [Ca2+](i) elevations, Carbachol (CCh) (1nM, 100 nM) did not induce changes in [Ca2+](i) of estrogen-deprived cells but produced significa nt increases in [Ca2+](i) of the same cells after incubation with E2 f or 30 minutes, The cultures which were preincubated with E2 for 24 hou rs responded to carbachol directly, The results of our study indicate that E2 may modulate the functions of endothelial cells after only a b rief exposure and also may be necessary for the response to acetylchol ine especially at low concentrations.