Dual P2Y 12 receptor signaling in thrombin-stimulated platelets--involvement of phosphoinositide 3-kinase beta but not gamma isoform in Ca2+ mobilization and procoagulant activity.
During thrombus formation, thrombin, which is abundantly present at sites of vascular injury, activates platelets in part via autocrine-produced ADP. We investigated the signaling pathways by which thrombin and ADP in synergy induced platelet Ca(2+) elevation and procoagulant activity, and we monitored the consequences for the coagulation process. Even at high thrombin concentration, autocrine and added ADP enhanced and prolonged Ca(2+) depletion from internal stores via stimulation of the P2Y(12) receptors. This P2Y(12)-dependent effect was mediated via two distinct signaling pathways. The first is enhanced Ca(2+) mobilization by the inositol 1,4,5-trisphosphate receptors due to inhibition of protein kinase A. The second pathway concerns prolonged activation of phosphoinositide 3-kinase (PI3-K) and phospholipase C. Experiments with phosphoinositide 3-kinase isoform-selective inhibitors and p110gamma deficient platelets demonstrated that the phosphoinositide 3-kinase beta and not the phosphoinositide 3-kinase gamma isoform is responsible for the prolonged Ca(2+) response and for the subsequent increases in procoagulant activity and coagulation. Taken together, these results demonstrate a dual P2Y(12)-dependent signaling mechanism, which increases the platelet-activating effect of thrombin by prolongation of Ca(2+) elevation, thereby facilitating the coagulation process.
van der Meijden
Paola E J
PE
Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, the Netherlands.
Schoenwaelder
Simone M
SM
Feijge
Marion A H
MA
Cosemans
Judith M E M
JM
Munnix
Imke C A
IC
Wetzker
Reinhard
R
Heller
Regine
R
Jackson
Shaun P
SP
Heemskerk
Johan W M
JW
D001792:Blood Platelets / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D003029:Coagulants / Q000494:pharmacology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D066298:In Vitro Techniques; D053496:Inositol 1,4,5-Trisphosphate Receptors / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D058925:Receptors, Purinergic P2Y12; D015398:Signal Transduction; D013917:Thrombin / Q000494:pharmacology*
D003029:Coagulants; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007527:Isoenzymes; C548576:P2RY12 protein, human; D018048:Receptors, Purinergic P2; D058925:Receptors, Purinergic P2Y12; D019869:Phosphatidylinositol 3-Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D013917:Thrombin; D002118:Calcium