Retinal relaxing factor (RRF) is suggested to be released from the retina and to contribute in the maintenance of retinal arterial tone. Herein, we aimed to clarify the effects of retinal tissue in isolated bovine retinal arteries in comparison with choroidal tissue and to evaluate the possible role of endothelium and potassium channels. In parallel, the effects of palmitic acid methyl ester (PAME), a putative vasodilator proposed to be released from the retina, was also examined. A piece of bovine retinal or choroidal tissue was placed within a close proximity on top of retinal arteries mounted in a wire myograph and precontracted with noradrenaline, prostaglandin F(2α), endothelin-1, thromboxane A(2) mimetic, U46619 or potassium (K(+)). To elucidate possible mechanisms in the effects of retinal tissue, retinal arteries were either deendothelized or incubated with inhibitors of endothelial vasodilators, i.e. nitric oxide (NO) and prostaglandins, or K(+) channels. Unlike the choroid, retinal tissue produced rapid, biphasic and complete relaxations in isolated bovine retinal arteries precontracted with various spasmogens acting on distinct receptors. Endothelium removal or preincubation of retinal arteries with inhibitors of NO synthase; L-NOARG (10(-4)M), guanylate cyclase; ODQ (10(-5)M) and cyclooxygenase; indomethacin (10(-5)M), did not cause a significant difference in the relaxation profile. Additionally, retinal relaxations remained unchanged in the presence of respective inhibitors of ATP-sensitive (K(ATP)) (glibenclamide, 10(-5)M), voltage-dependent (K(V)) (4-aminopyridine, 2×10(-3)M), and calcium-activated (K(Ca)) (tetraethylammonium 10mM; charybdotoxin, 10(-7)M; and apamin, 5×10(-7)M) K(+) channels. Thus, our results provide novel evidence regarding the biphasic relaxing profile of RRF in the retinal artery which was unrelated to endothelium and K(+) channels (K(ATP), K(V) and K(Ca)). Interestingly, PAME (10(-14)-10(-5)M) did not provoke a relaxation in bovine retinal artery suggesting no association with RRF.