The effects of a blockade of the action or synthesis of endothelium-derived nitric oxide (EDRF) on vascular resistance and reactivity, platelet cGMP and tissue oxygenation were studied. Experiments were performed in isolated perfused rabbit hearts as well as in rabbit hindlimbs in vivo. In isolated hearts, perfusion with hemoglobin (6 μM) or N^G-nitro-L-arginine (30 μM) significantly increased vascular resistance. The cGMP level in platelets passing through the coronary bed was found to be more than 50% lower than with intact EDRF production. EDRF inhibition also resulted in a reduced peak reactive hyperemia, an enhanced reactive vasoconstriction after a rapid increase in perfusion pressure (myogenic response), and in abolition of flow-dependent dilation of coronary resistance vessels. In rabbit hindlimbs, local blockade of EDRF-mediated dilations by gossypol resulted also in an increased vascular resistance and abolition of the increase in platelet cGMP induced by intraarterial infusion of acetylcholine. In addition, the oxygen uptake of the hindlimb (−46%) and the skeletal muscle pO₂ were significantly reduced. It is concluded that continuously released EDRF has a functional role in maintaining adequate tissue perfusion and oxygen supply. Furthermore, the adaption of the vascular bed to rapid changes in flow and pressure is impaired after inhibition of EDRF.