The renal origin of natriuretic factor (NF) of high molecular weight (MW 45,000) has been previously demonstrated. Its role in the control of sodium excretion by the kidney has been strongly suggested. The aim of the present work was to study the relationship between NF and prostaglandins, which are known to modify the renal regulation of sodium. (1) The activity of NF, obtained by ultrafiltration and gel filtration from pooled plasma of salt-loaded rats, was tested on an isolated rat kidney perfused by a cell-free medium. A significant natriuresis was observed in the kidneys isolated either from normal rats or from indomethacin-pretreated rats after injection of NF. Moreover, a dose-response curve was obtained with increasing amounts of NF. (2) The ability of isolated rat kidney to produce NF was studied in two series of salt-deprived rats, the second series being pretreated with indomethacin. The production of NF by isolated kidneys was induced by addition of saline to the fluid perfusing kidneys isolated from normal or indomethacin-pretreated rats fed a salt-depleted diet. Before addition of saline, no natriuretic activity was detectable. After the addition of 0.9% NaCl, NF was only found in the perfusate from kidneys isolated from normal rats. Perfusate extracts from indomethacin-pretreated kidneys did not induce any significant rise in natriuresis. (3) Kidneys isolated from rats previously fed a free fatty acid-deficient diet and thus prostaglandin-depleted were unable to produce NF even if they were salt-loaded. Intraarterial perfusion of prostaglandin E2 restored their ability to release or secrete NF. It is concluded that the release or the synthesis of NF is only possible in the presence of prostaglandins. Its action is not influenced by prostaglandin inhibition or deficiency.