The relationship of cyclic AMP to platelet function has been examined directly by assay of the platelet content of the nucleotide and platelet aggregation. Platelet aggregation is favored by an decrease in platelet cyclic AMP and inhibited by an increase in cyclic AMP. The platelet aggregation is accompanied by reduction in cyclic AMP. beta adrenergic substances increase platelet cyclic AMP in vivo and simultaneously block platelet aggregation. Thrombin selectively causes human platelets to form and release PGE. The amount formed was proportional to the amount of thrombin added and high enough to influence platelet aggregation in vitro. Platelet prostaglandin formation by washed platelets and platelet-rich plasma was not proportional to the degree of platelet aggregation induced by stirring. We were unable to detect PGE-formation in response to ADP. Thrombin-induced synthesis of PGE is inhibited by ADP. Aspirin selectively inhibits PGE-production in human platelets. This reaction does not depend on the extend of platelet aggregation. These studies suggest that cyclic AMP is the key to regulation of platelet aggregation and that PGE, formed by platelets, through its effects on cyclic AMP may play a role in regulating platelet aggregation.