Although endothelial cell (EC)-dependent relaxation has been described in most mammals, there have been no detailed reports of its existence in humans. Consequently, we evaluated human pulmonary artery segments taken from uninvolved regions of resected lung from 11 patients. EC removal did not significantly alter relaxation to vasoactive intestinal peptide (VIP). However, relaxation to acetylcholine (ACh) was observed only in segments with EC. Preincubation with either 1 microM propranolol or 10 microM indomethacin failed to block relaxation, but the addition of either 30 microM quinacrine hydrochloride or 100 microM nordihydroguaiaretic acid prevented it entirely. EC-dependent relaxation to ATP was also demonstrated. These data demonstrate that EC-dependent relaxation occurs in human pulmonary arteries. Neither beta-adrenergic pathways nor prostaglandin intermediaries are utilized. An oxidized breakdown product of arachidonic or some other fatty acid from EC phospholipid appears to be involved. These data suggest that interactions between endothelium and smooth muscle may be important in modulating tone in the vessel wall and that damage to the endothelium may play a role in the development of pulmonary hypertension in humans.