Evidence for separate effects of U73122 on phospholipase C and calcium channels in human platelets

Abstract

U73122 ({1-[6-((17β-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)exyl]-1H-pyrrole-2,5-dione}) is generally used as a selective inhibitor of phospholipase C (PLC) and the related rise in cytosolic Ca 2+. Recently, by using hepatocytes, it was suggested that its action sites are different for PLC activation and increase in Ca 2+ concentration. To verify whether U73122 has different sites for inhibiting PLC activation and calcium responses in human platelets, aggregation, Mn 2+ influx, cytosolic Ca 2+ increase and PLC activation were studied in response to thrombin and the synthetic agonist of the thromboxane receptor U46619 (9,11-dideoxy-9α,11α-methanoepoxyprostaglandin F 2α). With both agonists, U73122 inhibited aggregation, Mn 2+ influx and the enhancement of cytosolic calcium at concentrations of 2 μM or lower, while 10 μM was necessary to inhibit PLC activation. Our results suggested that U73122 is much more active in antagonizing Ca 2+ channels, both the intracellular ones, which are activated by formation of inositol 1,4,5 P 3 and those present on plasma membrane, than in reducing the activation of PLC.