THROMBIN-STIMULATED PHOSPHOLIPASE-C ACTIVITY IS INHIBITED WITHOUT VISIBLE DELAY BY A RAPID INCREASE IN THE CYCLIC-GMP LEVELS INDUCED BY SODIUM-NITROPRUSSIDE

Different drugs that elevate the cGMP levels inhibit the agonist-induc ed platelet activation. The mechanisms of action of cGMP probably incl ude inhibition of both phospholipase C and the increase in intracellul ar Ca2+ concentration, and these effects seem to be mediated by cGMP-d ependent protein kinases. However, in most studies, cells were preincu bated with nitrovasodilators before stimulation. The effect of the pre incubation with sodium nitroprusside before stimulation or the simulta neous addition of sodium nitroprusside and thrombin has been compared. The simultaneous addition of sodium nitroprusside and thrombin was ab le to inhibit without any significant delay the platelet aggregation. This rapid effect was correlated with an inhibition of both the maximu m increase in intracellular Ca2+ concentration and the phospholipase C activity. Also, the simultaneous addition of sodium nitroprusside and thrombin clearly accelerated the decline in the Ca2+ signal, which wa s not observed in platelets preincubated with sodium nitroprusside. Th e rapid inhibition induced by sodium nitroprusside was correlated with a rapid and significant increase in the cGMP levels and reversed when platelets were pretreated with methylene blue. The inhibitor of cAMP- dependent protein kinase Rp-8-(4-chlorophenylthio)-adenosine-3',5'-cyc lic monophosphorothioate was able to abolish nearly completely the inh ibitory effect induced by sodium nitroprusside independent of the prot ocol used. Thus, the rapid inhibition induced by sodium nitroprusside seems to be induced by a rapid phosphorylation-dependent mechanism. In addition, both cGMP- and cAMP-dependent protein kinase seem to be inv olved; however, the cAMP-dependent protein kinase seems to be more imp ortant.