Cyclic nucleotides modulate store-mediated calcium entry through the activation of protein-tyrosine phosphatases and altered actin polymerization in human platelets

Agonists elevate the cytosolic calcium concentration in human platelets via a receptor-operated mechanism, involving both Ca2+ release from intracellular stores and subsequent Ca2+ entry, which can be inhibited by platelet inhibitors, such as prostaglandin E-1 and nitroprusside which elevate cAMP and cGMP, respectively. In the present study we investigated the mechanisms by which cAMP and cGMP modulate store-mediated Ca2+ entry. Both prostaglandin E-1 and sodium nitroprusside inhibited thapsigargin-evoked store-mediated Ca2+ entry and actin polymerization. However, addition of these agents after induction of store-mediated Ca2+ entry did not affect either Ca2+ entry or actin polymerization. Furthermore, prostaglandin E-1 and sodium nitroprusside dramatically inhibited the tyrosine phosphorylation induced by depletion of the internal Ca2+ stores or agonist stimulation without affecting the activation of Ras or the Ras-activated phosphatidylinositol 3-kinase or extracellular signal-related kinase (ERK) pathways. Inhibition of cyclic nucleotide-dependent protein kinases prevented inhibition of agonist-evoked Ca2+ release but it did not have any effect on the inhibition of Ca2+ entry or actin polymerization. Phenylarsine oxide and vanadate, inhibitors of protein-tyrosine phosphatases prevented the inhibitory effects of the cGMP and cAMP elevating agents on Ca2+ entry and actin polymerization. These results suggest that Ca2+ entry in human platelets is directly down-regulated by cGMP and cAMP by a mechanism involving the inhibition of cytoskeletal reorganization via the activation of protein tyrosine phosphatases.