Thrombin plays an important role in promoting arterial thrombosis by platelet activation and by catalyzing fibrin formation. Use of thrombin inhibitors that block both the platelet-activating and fibrin formation properties of thrombin are associated with hemostasis. This problem might be overcome by developing agents that block only the platelet-activating property of thrombin. Because the platelet-activating property of thrombin is mediated by the thrombin receptor, antagonists of the thrombin receptor might be efficacious and potentially safer with regard to bleeding complications. We investigated whether a peptide ligand (AFLARAA) of the thrombin receptor that blocked alpha-thrombin and thrombin receptor activating peptide-induced platelet aggregation could inhibit thrombosis. A partially occlusive thrombus was generated by application of electric current in rabbit carotid artery. In control animals, the artery was completely occluded within 42+/-12 min after the current was discontinued. When the thrombin receptor activating peptide antagonist was given (100 micromol/kg as an IV bolus followed by 900 micromol/kg infusion for a period of 180 min) starting at the time the current was stopped, blood flow remained patent throughout the infusion period and for an additional 60 min after the infusion was stopped. The antithrombotic effect of the antagonist peptide was not associated with increased bleeding tendency, as judged by the amount of blood adsorbed by a gauze pad placed in a surgical incision extending to the muscle tissue and by a standard template bleeding time. These results indicate that thrombin receptor antagonist peptides can be used as antithrombotic agents.