EFFECTS OF THE PRIOR ACTIVATION OF PROTEIN-KINASE-C ON HUMAN PLATELETACTIVATION-INDUCED BY THROMBIN

Effects of the prior activation of protein kinase C (PKC) on the respo nses induced by thrombin were studied using human blood platelets, in which PKC is abundantly expressed. At a concentration of 25 nM, 12-O-t etradecanoyl-phorbol 13-acetate (TPA) induced little aggregation or re lease by itself but selectively elicited PKC activation in a time-depe ndent manner, as monitored by 47-kDa protein phosphorylation. Increase s in the intracellular Ca2+ concentration of human platelets caused by thrombin, at any concentration, were markedly inhibited by the prior addition of 25 nM TPA in a time-dependent manner. However, the effects of TPA on platelet aggregation and secretion induced by thrombin vari ed, depending upon the agonist concentration; the PKC activator marked ly enhanced the aggregation and secretion induced by lower concentrati ons of thrombin but had a tendency to weakly inhibit those induced by higher concentrations of thrombin. TPA enhanced the ionomycin effect u pon aggregation and release without obvious effects on the ionophore-i nduced Ca2+ mobilization, suggesting that PKC potentiates platelet fun ction by increasing intracellular sensitivity to Ca2+. There was a goo d time-dependent correlation of the TPA effects among the three parame ters, namely, the phosphorylation of the 47-kDa protein, inhibition of the [Ca2+]i increase induced by thrombin, and enhanced ionomycin effe cts in aggregation and release. It is most likely that the Final respo nses of agonist-activated platelets depend upon the balance of two fac tors induced by PKC, inhibition of Ca2+ signals and enhancement of int racellular sensitivity to Ca2+.