The renal glycoprotein hormone erythropoietin (Epo) interacts with erythrocytic progenitors to stimulate their proliferation and differentiation in the bone marrow. The renal O2-sensing mechanism in the control of the synthesis of Epo is still poorly understood. Therefore, the capacity of isolated rat kidneys to produce Epo during hypoxic and anemic perfusion was studied. The kidneys were perfused at a constant perfusion pressure of 100 mm Hg with a substrate-enriched Krebs-Henseleit solution containing 60 g/liter BSA and freshly drawn human erythrocytes. Epo was measured by RIA. When the kidneys were perfused at an arterial pO2 of 720 or 150 mm Hg (hematocrit, 5%), Epo production was very low (0.1-0.2 U/g kidney within 3 h of perfusion). When the arterial pO2 was lowered to 35 or 20 mm Hg, Epo production increased to 0.4 and 0.9 U/g kidney, respectively. The release of Epo during hypoxic perfusion (pO2 35 and 20 mm Hg) was little affected by changes in the hematocrit, i.e. the O2-carrying capacity of the perfusion medium over a wide range (0-40%). These results indicate that the production of Epo in the isolated perfused kidney depends on the availability of O2 and can be modulated by changes in the arterial pO2.