We examined the effects of flow on lymphatic endothelial cells by using conventional cascade preparations of isolated coronary arteries without intact endothelium. The pressurized thoracic ducts were intraluminally perfused at a constant flow rate ranging from 0.5 to 2.0 ml/min. A linear relationship was observed between the flow rate and the normalized amount of relaxing substance(s) released from the lymphatic endothelial cells. Thus the flow rate of 2.0 ml/min produced approximately 39% of sodium nitroprusside (SNP)-produced maximal relaxation in the cascade arterial rings. The acetylcholine (ACh, 10(-5) M)- and flow-induced relaxations of the cascade arterial rings were completely reduced by the mechanical rubbing of lymphatic endothelial cells in the pressurized lymph vessels. Pretreatment with 5 x 10(-5) M N(G)-nitro-L-arginine methyl ester (L-NAME) on the lymphatic endothelial cells caused a significant reduction of the ACh- and flow-induced vasodilations of the cascade arterial rings. Pretreatment with 10(-5) M indomethacin on the lymphatic endothelial cells produced no significant effect on the ACh- and flow-induced vasodilations. These findings suggest that lymphatic endothelial cells of canine thoracic ducts can produce and release endogenous nitric oxide by stimulation of flow (approximately 2.0 ml/min).