Phorbol esters modulate thrombin-operated calcium mobilisation and dense granule release in human platelets

Abstract

Human α-thrombin-induced elevation of cytosolic free calcium ([Ca 2+] i) and dense granule release was examined in platelets preincubated with either activators or an inhibitor of protein kinase C. 12- O-Tetradecanoylphorbol 13-acetate (TPA) or two 12-deoxy analogues of TPA, when added alone to platelets, did not elevate [Ca 2+] i, as monitored by quin2 fluorescence, though small amounts of dense granule release were detected. Preincubation of the platelets with either TPA or 12-deoxyphorbol 13-phenylacetate, but not the parent, 4-β-phorbol, produced a dose-dependent inhibition of the elevation of [Ca 2+] i and 5-hydroxytryptamine release induced by human α-thrombin. Furthermore, this phorbol ester-mediated inhibition of human α-thrombin-induced activation could be prevented by H7 (1-[5-isoquinolinesulphonyl]-2-methylpiperazine), the recently described inhibitor of protein kinase C. These results suggest a role for protein kinase C as a modulator of receptor-operated calcium fluxes in human platelets.