A patch-clamp study of the Ca2+ mobilization from internal stores in bovine aortic endothelial cells. II. Effects of thapsigargin on the cellular Ca2+ homeostasis.

Abstract

Evidence was provided, in the preceding paper (Thuringer & Sauvé, 1992), that the external Ca(2+)-dependent phase of the Ca2+ signals evoked by bradykinin (BK) or caffeine in bovine aortic endothelial cells (BAE), differ in their respective sensitivity to procaine. To examine whether the emptying of the InsP3-sensitive Ca2+ store is the signal for activating the agonist-evoked Ca2+ entry, we have investigated the effects of thapsigargin (TSG), a known inhibitor of the microsomal Ca(2+)-ATPase activity in a variety of cell types, via the activity of calcium-activated potassium channels [K(Ca2+) channels]. In cell-attached experiments, the external application of TSG caused a sustained or oscillatory activation of K(Ca2+) channels depending on both the cells and doses tested. The TSG-evoked channel activity could be reversibly blocked by removing extracellular Ca2+, and strongly decreased by adding 10 mM procaine to the bath medium. In Ca(2+)-free external conditions, TSG did not promote an apparent Ca2+ discharge from internal stores but prevented in a dose- and time-dependent manner the subsequent agonist-evoked channel activity related to the release of internally sequestered Ca2+. These results confirm that TSG and BK release Ca2+ from the same internal stores but with different kinetics. Because the channel response to caffeine was found to be poorly sensitive to procaine, in contrast to that evoked by BK and TSG, it may be concluded that both BK and TSG activate the same Ca2+ entry pathway. Therefore, the emptying of the InsP3-sensitive Ca2+ store is likely to be the main signal for activating the agonist-evoked Ca2+ entry in BAE cells.