Second messengers in platelet aggregation evoked by serotonin and A23187, a calcium ionophore

We investigated the combined effect of 5-hydroxytryptamine (5-HT, serotonin ) and calcium ionophore (A23187) on human platelet aggregation. Aggregation , monitored at 37 degreesC using a Dual-channel Lumi-aggregometer, was reco rded for 5 min after challenge by a change in light transmission as a funct ion of time. 5-HT (2-200 muM) alone did not cause platelet aggregation, but markedly potentiated A23187 (low dose) induced aggregation. Inhibitory con centration (IC50) values for a number of compounds were calculated as means SEM from dose-response determinations. Synergism between 5-HT (2-5 muM) an d A23187 (0.5-2 muM) was inhibited by 5-HT receptor blockers, methysergide (IC50 = 18 muM) and cyproheptadine (IC50 = 20 muM), and calcium channel blo ckers (verapamil and diltiazem, IC50 = 20 muM and 40 muM respectively). Int erpretation of the effects of these blockers is complicated by their lack o f specificity. Similarly, U73122, an inhibitor of phospholipase C (PLC), bl ocked the synergistic effect at an IC50 value of 9.2 muM. Wortmannin, a pho sphatidylinositide 3-kinase (PI 3-K) inhibitor, also blocked the response ( IC50 = 2.6 muM). However, neither genistein, a tyrosine-specific protein ki nase inhibitor, nor chelerythrine, a protein kinase C inhibitor, affected a ggregation at concentrations up to 10 muM. We conclude that the synergistic interaction between 5-HT and ionophore may be mediated by activation of PL C/Ca2+ and PI 3-kinase signalling pathways, but definitive proof will requi re other enzyme inhibitors with greater specificity. (C) 2001 Elsevier Scie nce Inc. Ali rights reserved.