SYNERGISTIC INTERACTION OF ADENYLATE-CYCLASE ACTIVATORS AND NITRIC-OXIDE DONOR SIN-1 ON PLATELET CYCLIC-AMP

The molecular mechanism of the synergistic platelet inhibition by acti vators of adenylate cyclase and guanylate cyclase in human platelets w as investigated. The adenylate cyclase activators iloprost and prostag landin E(1) and the guanylate cyclase activator 3-morpholino-sydnonimi ne (SIN-1) dose-dependently inhibited thrombin-induced aggregation of washed human platelets. Furthermore, SIN-1 at a concentration inhibiti ng platelet aggregation by only 10% shifted the IC50 values of ilopros t and prostaglandin E(1) by one order of magnitude to the left, indica ting a synergistic action of adenylate cyclase and guanylate cyclase a ctivators. Iloprost and prostaglandin E(1) dose-dependently elevated p latelet cAMP without a significant influence on cGMP. In contrast, the platelet cGMP level was dose-dependently elevated by SIN-1. In additi on, SIN-1 markedly increased cAMP level induced by low concentrations of adenylate cyclase activators (0.1-0.3 nM iloprost or 10-150 nM pros taglandin E(1)). In contrast, the rise in cAMP induced by higher adeny late cyclase activator concentrations (3 nM iloprost or 30 mu M prosta glandin E(1)) was significantly reduced in the presence of SIN-1. The same biphasic mode of action of SIN-1 was observed with forskolin, an adenylate cyclase stimulator acting receptor independently, indicating a prostacyclin-receptor independent mechanism. The cAMP elevating eff ect of SIN-1 in the presence of low prostanoid concentrations was comp letely abolished by piroximone, a selective inhibitor of phosphodieste rase type III. Therefore, the inhibition of phosphodiesterase III by c GMP seems to be the mechanism for the elevation of cAMP levels by SIN- 1 in the presence of low concentration of adenylate cyclase activators in human platelets. In contrast, the mechanism by which SIN-1 reduces platelet cAMP levels induced by higher concentrations of prostaglandi n E(1), iloprost, and forskolin, remains to be clarified.