To examine the effect of platelets on the tone of quiescent vessels, rat aortic rings with intact endothelium and others with denuded endothelium were exposed to platelets in an organ bath. Suspension of washed platelets induced a mild but consistent contraction of rings with intact endothelium. The subsequent contractile response of these rings to norepinephrine (NE) was also potentiated. Platelets caused a marked contraction of quiescent rings without intact endothelium and potentiated the subsequent contractile effects of NE. The procontractile effect of platelets in rat aortic rings with and without intact endothelium was not modulated by the selective thromboxane A2 (TXA2)/endoperoxide receptor antagonist SQ29,548 or the cyclooxygenase inhibitor indomethacin, but was completely blocked by serotonin (S2) receptor antagonist LY53,857, suggesting that release of serotonin from aggregating platelets is perhaps more important than release of TXA2 in the procontractile effects of platelets on quiescent rat aortic rings. To examine the role of vascular endothelium in the procontractile effect of platelets further, we treated aortic rings with intact endothelium with the inhibitor of endothelium-derived relaxing factor (EDRF) NG-monomethyl-L-arginine (L-NMMA) or the adenine nucleotide scavenger apyrase. In rings treated with L-NMMA or apyrase, the contractile effect of platelets was enhanced (p < 0.01) and mimicked that in rings without intact endothelium. These observations indicate that although basal release of EDRF modulates the contractile effect of platelets, the dominant effect of platelets on quiescent rat aortic rings with intact endothelium is procontractile and this effect is mediated exclusively by release of serotonin.