We performed an in vitro study to assess injury to vascular endothelial cells by platelets. Cultured endothelial cells isolated from fetal bovine aorta were used. Addition of human platelets, activated by collagen or lysed by sonication, to the culture dish resulted in dose- and time-dependent damage to the cells as estimated by [3H]adenine release. Analysis of [3H]adenine nucleotides by thin-layer chromatography on PEI-cellulose revealed decreased intracellular ATP content in the cells treated with platelet lysate. The medium contained AMP and adenosine, the latter increasing following the treatment of the cells. Of the substances released by the activated platelets, thromboxane A2 (TxA2) and serotonin caused cell damage. Platelet-derived growth factor (PDGF), however, did not damage the endothelial cells up to a concentration of 200 ng/ml. Pretreatment of the cells with methysergide (10(-6) M) or ONO 3708 (10(-5) M), a TxA2 antagonist, only partially prevented the damage, while ZK 36374 (10(-6) M), a prostacyclin analog, and 3-isobutyl-1-methylxanthine (IBMX; 10(-3) M), a phosphodiesterase inhibitor, potently inhibited injury. We conclude that the substances released from activated platelets may injure endothelial cells in an additive or synergistic manner and that agents which produce effects that elevate cyclic AMP levels may protect the cells from damage induced by the platelets.