Phosphoinositides are required for store-mediated calcium entry in human platelets

We have recently observed that small GTP-binding proteins are important for mediation of store-mediated Ca2+ entry in human platelets through the reor ganization of the actin cytoskeleton. Because it has been shown in platelet s and other cells that small CTP-binding proteins regulate the activity of phosphatidylinositol S-kinase and phosphatidylinositol 4-kinase, whose prod ucts, phosphoinositides, play a key role in the reorganization of the actin cytoskeleton, we have investigated the role of these lipid kinases in stor e mediated Ca2+ entry. Treatment of platelets with LY294002, an inhibitor o f phosphatidylinositol 3- and phosphatidylinositol I-kinases, resulted in a concentration-dependent inhibition of Ca2+ entry stimulated by thapsigargi n or the physiological agonist, thrombin. In addition, wortmannin, another inhibitor of these kinases, which is structurally unrelated to LY294002, si gnificantly reduced store-mediated Ca2+ entry. The inhibitory effect of LY2 94002 was not mediated either by blockage of Ca2+ channels or by modificati on of membrane potential. LY294002 inhibited actin polymerization stimulate d by thrombin or thapsigargin. These results indicate that both phosphatidy linositol 3-kinase and phosphatidylinositol 4-kinase are required for activ ation of store-mediated Ca2+ entry in human platelets and that the mechanis m could involve the reorganization of the actin cytoskeleton.