This study compared the responses of canine and human platelets to various aggregating agonists in the presence or absence of extracellular ATP and ATP analogues. Canine and human platelets were approximately equally reactive with ADP or collagen while the canine platelets were about 10 fold more sensitive to thrombin. Canine platelets were insensitive to the thromboxane mimetic U46619 but were synergistically aggregated by a mixture of ADP and U46619. Human platelets were very sensitive to U46619. Aggregations of human platelets with all of the above agonists were inhibited by extracellular ATP; beta, gamma methylene ATP (beta gamma ATP) and benzoyl ATP (BzATP) with a rank order suggestive of an interaction with P2x-like purinoceptors which support our previous findings. The comparable aggregations of canine platelets were likewise inhibited by ATP and its analogues but with a rank order suggestive of an interaction with P2y-like purinoceptors. ATP inhibited U46619- and ADP-induced aggregation of human platelets and ADP-induced aggregation of canine platelets, presumably, in part, due to competition for the ADP P2T receptor. However, when U46619 was added to either ATP or ATP analogue-inhibited ADP-treated canine platelets, the inhibition was nullified. Furthermore, we demonstrated, for the first time, that the canine thromboxane receptor becomes reactive to U46619 alone after incubation at room temperature for 3.5-5 hrs while human platelets become inactive under similar conditions. The implication of these studies is that there are significant differences in the canine and human platelet thromboxane and purine receptors. The future characterization of these differences and the mechanism by which they function should further our understanding of the impact of extracellular ATP on hemostasis and thrombosis.