Platelet membrane glycoprotein V ( GPV ) is hydrolyzed during thrombin-induced platelet aggregation. The present studies were undertaken to determine whether cleavage of GPV was a direct result of thrombin action or was secondary to platelet activation. Acetylsalicylic acid or vincristine had little or no effect on platelet activation or cleavage of GPV by thrombin. Prostacyclin or antimycin A and 2-deoxyglucose inhibited platelet activation but had no effect on hydrolysis of GPV . These findings suggest that the hydrolysis of GPV occurs independently of the biochemical and morphological changes associated with platelet activation and is therefore a direct effect of thrombin.