ADP receptor P2Y12 is the capstone of the cross-talk between Ca2+ mobilization pathways dependent on Ca2+ ATPases sarcoplasmic/endoplasmic reticulum type 3 and type 2b in platelets.

Background: Blood platelet Ca2+ stores are regulated by 2 Ca2+-ATPases (SERCA2b and SERCA3). On thrombin stimulation, nicotinic acid adenosine dinucleotide phosphate mobilizes SERCA3-dependent stores, inducing early adenosine 5'-diphosphate (ADP) secretion, potentiating later SERCA2b-dependent secretion. Objectives: The aim of this study was to identify which ADP P2 purinergic receptor (P2Y1 and/or P2Y12) is(are) involved in the amplification of platelet secretion dependent on the SERCA3-dependent Ca2+ mobilization pathway (SERCA3 stores mobilization) as triggered by low concentration of thrombin. Methods: The study used the pharmacologic antagonists MRS2719 and AR-C69931MX, of the P2Y1 and P2Y12, respectively, as well as Serca3 -/- mice and mice exhibiting platelet lineage-specific inactivation of the P2Y1 or P2Y12 genes. Results: We found that in mouse platelets, pharmacological blockade or gene inactivation of P2Y12 but not of P2Y1 led to a marked inhibition of ADP secretion after platelet stimulation with low concentration of thrombin. Likewise, in human platelets, pharmacological inhibition of P2Y12 but not of P2Y1 alters amplification of thrombin-elicited secretion through SERCA2b stores mobilization. Finally, we show that early SERCA3 stores secretion of ADP is a dense granule secretion, based on parallel adenosine triphosphate and serotonin early secretion. Furthermore, early secretion involves a single granule, based on the amount of adenosine triphosphate released. Conclusion: Altogether, these results show that at low concentrations of thrombin, SERCA3- and SERCA2b-dependent Ca2+ mobilization pathways cross-talk via ADP and activation of the P2Y12, and not the P2Y1 ADP receptor. The relevance in hemostasis of the coupling of the SERCA3 and the SERCA2b pathways is reviewed.