Progesterone metabolites rapidly stimulate calcium influx in human platelets by a src-dependent pathway

The effects of several steroids and their metabolites were examined for their ability to rapidly alter intracellular free calcium ([Ca2+](i)) in the anucleate human platelet. Earlier studies suggested that steroids had direct and rapid non-genomic effects to alter platelet physiology. The rationale for performing this study was to investigate the signal transduction events being activated by steroids. Super-physiologic concentrations (1.0-10.0 mu M) of beta-estradiol and several estradiol. metabolites and analogs potentiated (approximately twofold) the action of thrombin to elevate [Ca2+](i) in platelets, whereas 10.0 mu M progesterone inhibited the action of thrombin by 10-15%. Progesterone and p-estradiol by themselves did not affect [Ca2+](i).. Progesterone metabolites can achieve high blood concentrations. Some progesterone metabolites, particularly those in the p-conformation, were potent stimulators of Ca2+ influx and intracellular Ca2+ mobilization in platelets. They activated phospholipase C because their ability to increase [Ca2+](i) was inhibited by the phospholipase C inhibitor U-73122. The ability of pregnanediol and collagen to increase [Ca2+](i) was inhibited by the src tyrosine kinase inhibitor PP1, whereas the actions of thrombin and thapsigargin to increase [Ca2+](i) were not affected by PP1. The effects of progesterone metabolites to increase [Ca2+](i) were observed with concentrations as low as 0.1 mu M. Pregnanolone synergized with thrombin to increase [Ca2+](i). it is hypothesized that human platelets possess receptors for progesterone metabolites. These receptors when stimulated will activate platelets by causing a rapid increase in [Ca2+](i). Pregnanolone, isopregnanediol and pregnanediol were the most effective stimulators of this newly identified src-dependent signal transduction system in platelets. Progesterone metabolites may regulate platelet aggregation and hence thrombosis in vivo. (C) 2008 Elsevier Inc. All rights reserved.