Bridge Grant: cGMP regulates platelet function and thrombosis

Platelets are essential for physiological process of hemostasis, but can also contribute to development of pathologic thrombus formation. At sites of vascular injury, exposure of platelets to soluble agonists or subendothelial adhesive proteins triggers platelet activation, leading to thrombus formation. However, under physiological conditions, this process is tightly regulated, limiting platelet activation to ensure that the response to injury halts bleeding without causing vascular occlusion. The secondary messenger, cGMP, plays an important role in regulating platelet activation. In platelets, cGMP is believed to be synthesized predominantly by soluble guanylyl cyclase (sGC), which can be activated by platelet agonists and nitric oxide (NO). Surprisingly, agonist-induced cGMP production was not abolished in the platelets lacking sGC. We further identified a membrane-bound guanylyl cyclase, the natriuretic peptide receptor-A (NPRA), in platelets. Because NO donors inhibit platelet activation, the prevailing concept is that cGMP inhibits platelet function. However, our previously published work suggests a biphasic platelet response with initial cGMP production stimulating platelet activation and subsequent generation inhibiting function. Based on these novel observations, we propose that cGMP serves as critical switch to regulate platelet function and that the initially low levels generated by physiological agonists promote platelet secretion and activation. In this application, we will test our hypothesis by determining contribution of NPRA to platelet activation and thrombosis. Following the initial response, spatial changes in the localization of cGMP production triggers subsequent pathways to inhibit platelet recruitment and limit thrombus overgrowth. This hypothesis is consistent with observations of spatial regulation in cGMP signaling and that cGMP-dependent pathways regulate neutrophil degranulation and neuron cell exocytosis in a biphasic manner. We have generated an innovative set of reagents that uniquely position us to complete the proposed studies. Given the increasing clinical use of drugs that target cGMPdependent pathways, our findings could have important ramifications by providing novel insights into how platelet activation is regulated that could be used to guide the development of new antithrombotic drugs.