To examine the temporal effects of plasmin generated in vivo on platelet function, we infused tissue-type plasminogen activator (t-PA) in rabbits over 3 hours and measured ex vivo platelet aggregation. We noted an initial increase in the aggregation response to ADP occurring 30 minutes after the start of infusion. This enhanced response was short-lived and by 180 minutes was reduced, compared with pretreatment levels. Baseline aggregation response was restored by 240 minutes. This pattern of aggregation response to t-PA infusion was also seen with thrombin as the agonist. Coinfusion of either prostaglandin I2 or prostaglandin E1 abolished the initial hyperaggregable phase induced by t-PA; the hypoaggregable phase occurred earlier (after 60 minutes) and persisted throughout the 1-hour recovery period. Similarly, streptokinase infused for 1 hour also increased platelet aggregation at early times and then reduced aggregation responses after the first hour. Plasma plasmin activity increased as expected with t-PA infusion alone, peaking at 30 minutes and returning to baseline by the first hour. Interestingly, prostaglandin E1 blunted the rise in plasma plasmin activity. This same dose of prostaglandin E1 or prostaglandin I2 used alone did not appreciably alter platelet function at any time during the experiment. Our data show that therapeutic doses of t-PA or streptokinase produce a biphasic effect on platelet aggregation response in the rabbit. Coinfusion of either of the antiplatelet agents, prostaglandin E1 or prostaglandin I2, abolishes the hyperaggregable phase and prolongs the inhibitory effects on platelet aggregation produced by t-PA. These data suggest that the effects of thrombolytic agents on platelet function are complex and can be modulated by antiplatelet prostaglandins.