Changes in perfusion pressure in response to graded doses of prostaglandin F 2 alpha (PGF 2 alpha) were measured during both forward and retrograde perfusion of isolated cat lungs perfused at a constant flow rate. During forward perfusion, PGF 2 alpha produced a dose-dependent increase in pulmonary artery pressure and a decrease in lung fluid volume. During retrograde perfusion, PGF 2 alpha also produced a dose-dependent increase in perfusion pressure; however, the dose required was fivefold greater than that needed to produce an identical change in pressure during forward perfusion. In addition, during retrograde perfusion, the lung fluid volume increased in response to PGF 2 alpha. These results suggest that the major site of activity of PGF 2 alpha is on the arterial side of the pulmonary vascular bed and that inactivation of PGF 2 alpha by the lung occurs primarily distal to this arterial site of vasomotion. The changes in perfusion pressure in response to PGF 2 alpha were markedly dependent on pH and oxygen tension (Po-2), being abolished by severe alkalosis and potentiated by both acidosis and hypoxia. In contrast, neither serotonin nor norepinephrine exhibited such a pH or Po-2 dependency. Since the ratio of the forward response to the retrograde response was not decreased by alterations of pH or Po-2, their influence on the responses appears to be through interaction at the site of vascular activity rather than through alteration of the rate of prostaglandin inactivation.