The supernatant obtained from platelet-suspension incubated with thrombin caused contractions in both intact and endothelium-removed canine basilar arteries. Cyproheptadine (5 x 10(-7) M), which reduced the serotonin-induced contraction to about 20%, attenuated the supernatant-induced contraction to 51% in intact arteries and to 24% in endothelium-removed arteries. In both absence and presence of cyproheptadine, the supernatant-induced contraction of intact arteries was significantly larger than that of endothelium-removed arteries. The cyproheptadine-resistant contraction by the supernatant of intact arteries was attenuated by aspirin, OKY-046 (a thromboxane A2 synthetase inhibitor) and ONO-3708 (a thromboxane A2 antagonist), whereas such contraction of endothelium-removed arteries was not affected by these agents. The concentrations of serotonin, ATP and ADP in the supernatant used here were approximately 3, 67 and 44 micrograms/ml, respectively. The mixed solution of serotonin, ATP and ADP, at the same concentrations as the supernatant, also induced cyproheptadine-resistant, endothelium-dependent contractions in intact arteries. The contraction induced by the mixed solution in intact arteries was also attenuated by aspirin, OKY-046 and ONO-3708. The present experiments indicate that the supernatant causes endothelium-dependent contraction in canine cerebral arteries and that platelet-derived ATP and ADP contribute to this contraction.