HEMOSTASIS, THROMBOSIS, AND VASCULAR BIOLOGY Reciprocal cross-talk between P2Y1 and P2Y12 receptors at the level of calcium signaling in human platelets

Adenosine diphosphate (ADP), an important platelet agonist, acts through 2 Gprotein–coupled receptors (GPCRs), P2Y1 and P2Y12, which signal through Gq and Gi, respectively. There is increasing evidence for cross-talk between signaling pathways downstream of GPCRs and here we demonstrate cross-talk between these 2 ADP receptors in human platelets. We show that P2Y12 contributes to platelet signaling by potentiating the P2Y1-induced calcium response. This potentiation is mediated by 2 mechanisms: inhibition of adenylate cyclase and activation of phosphatidylinositol 3 (PI 3)–kinase. Furthermore, the Src family kinase inhibitor PP1 selectively potentiates the contribution to the calcium response by P2Y12, although inhibition of adenylate cyclase by P2Y12 is unaffected. Using PP1 in combination with the inhibitor of PI 3-kinase LY294002, we show that Src negatively regulates the PI 3-kinase–mediated component of the P2Y12 calcium response. Finally, we were able to show that Src kinase is activated through P2Y1 but not P2Y12. Taken together, we present evidence for a complex signaling interplay between P2Y1 and P2Y12, where P2Y12 is able to positively regulate P2Y1 action and P2Y1 negatively regulates this action of P2Y12. It is likely that this interplay between receptors plays an important role in maintaining the delicate balance between platelet activation and inhibition during normal hemostasis. (Blood. 2004; 104:1745-1752)