| Since the discovery that nitric oxide (NO) mediates smooth muscle relaxation and blood vessel dilation, studies on cellular functions of NO have received much attention from medical researchers. Significant attention has been given to the effects of NO on the function of platelets, blood cells critical for thrombus formation. It has been a textbook concept in the past 30 years that NO inhibits platelet function. This concept was mainly based on the observation that high concentrations of NO donors inhibit platelet activation. However, platelet agonists are known to stimulate low level NO production. This thesis investigated the mechanisms of endogenous NO production and its role in platelet activation. By using platelets isolated from genetically manipulated mice, we show that platelet NO syntheses (NOS), both endothelial NOS (eNOS) and inducible NOS (iNOS), mediate platelet-agonist induced NO production. Agonist-stimulated NO production occurs almost immediately and is not affected by an inhibitor of protein synthesis, indicating that it is post-translationaly regulated. These data suggest a novel mechanism for the regulation of iNOS activation, independently of the previously known mechanisms through the regulation of its expression. The studies on isolated human platelets, using both NOS inhibitors and activators, and studies on genetically manipulated mouse platelets, show that platelet NOS plays a stimulatory role in platelet activation induced by low doses of platelet agonists. Furthermore, we show that eNOS promotes thrombosis in vivo. The stimulatory role of NOS is mediated by soluble guanylyl cyclase (sGC), and results from a cyclic guanosine monophosphate (cGMP)-dependent stimulation of platelet granule secretion. NO-cGMP-dependent stimulation of platelet secretion serves to amplify platelet activation and stabilize platelet aggregates. Our data also suggest that NO plays a biphasic role in platelet activation: low NO concentrations, such as those endogenously produced, play a stimulatory role, while high concentrations exhibit an inhibitory role. |
