title: Signalling components underlying platelet aggregation to a Ca2+ ionophore and a phorbol ester.
abstract: Although it is well established that G(q)- and G(i)-coupled receptors can combine to mediate platelet aggregation, the signalling events underlying the synergy are not fully characterised. This study has used the calcium ionophore, A23187, and phorbol ester, PMA, to investigate this question. We show that aggregation to submaximal but not maximal concentrations of ionophore is partially inhibited by antagonism of the P2Y(12) ADP receptor or PKC blockade. However, a full aggregation response can be restored under these conditions by addition of PMA or ADP. Studies using PI 3-kinase inhibitors demonstrate that this is the second messenger pathway that restores aggregation by the G(i)-coupled receptor in the presence of PKC blockade. However, under normal circumstances, PI 3-kinase activity is not a major requirement for aggregation to the ionophore. PMA stimulates a weak aggregation which takes several minutes to reach a maximum. Threshold concentrations of PMA and a G(i)-coupled receptor agonist when added alone show no effect on aggregation, but when combined induce aggregation responses. This study demonstrates that calcium and PKC interact synergistically with a G(i)-coupled receptor agonist to mediate aggregation, and also with each other. Activation of G(i) supports aggregation in part through the PI 3-kinase pathway. High concentration of ionophore on their own can induce aggregation independent of PKC and activation of G(i). Multiple signalling pathways mediate platelet aggregation and their relative importance depends on experimental conditions.
authors.affiliation: Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, UK. margaret@bioch.ox.ac.uk
authors.forename: M J
authors.initials: MJ
authors.lastname: Stafford
authors.forename: B T
authors.initials: BT
authors.lastname: Atkinson
authors.forename: S P
authors.initials: SP
authors.lastname: Watson
authors.forename: C J
authors.initials: CJ
authors.lastname: Pears
mesh_terms: D000244:Adenosine Diphosphate / Q000378:metabolism; D000730:Androstadienes / Q000037:antagonists & inhibitors; D000001:Calcimycin / Q000494:pharmacology*; D020013:Calcium Signaling / Q000187:drug effects*; D002867:Chromones / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D004789:Enzyme Activation; D006801:Humans; D007211:Indoles / Q000378:metabolism*; D007476:Ionophores / Q000494:pharmacology*; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000494:pharmacology; D010974:Platelet Aggregation / Q000187:drug effects*; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism; D011493:Protein Kinase C / Q000187:drug effects*; D012701:Serotonin / Q000378:metabolism*; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D013931:Thromboxanes / Q000378:metabolism; D010738:Type C Phospholipases / Q000502:physiology*; D000077191:Wortmannin
chemical_list: D000730:Androstadienes; D002867:Chromones; D007211:Indoles; D007476:Ionophores; D009025:Morpholines; D013931:Thromboxanes; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D012701:Serotonin; D000001:Calcimycin; D004533:Egtazic Acid; D000244:Adenosine Diphosphate; D011451:Prostaglandin-Endoperoxide Synthases; D019869:Phosphatidylinositol 3-Kinases; D011493:Protein Kinase C; D010738:Type C Phospholipases; D013755:Tetradecanoylphorbol Acetate; C064758:Ro 31-8220; D000077191:Wortmannin