Omega- and omega-1 hydroxylations are the major pathways by which arachidonic acid is metabolized in cortical and outer medullary microsomes of rat and rabbit kidneys. It is a cytochrome P450-dependent oxidation leading to the formation of 20-hydroxy- and 19-hydroxyeicosatetraenoic acids. In this study, we compared the effects of the synthetically prepared omega- and omega-1 metabolites of arachidonic acid on the activity of the renal Na+-K+-ATPase partially purified from rat renal cortical microsomes. 19(S)-hydroxyeicosatetraenoic acid caused a dose related stimulation of Na+-K+-ATPase activity with an EC50 of 3 x 10(-7) M. In contrast, neither 19(R)-hydroxyeicosatetraenoic acid, 20-hydroxyeicosatetraenoic acid nor arachidonic acid at 10(-6) M had any effect on Na+-K+-ATPase activity. In the same preparation, ouabain at 10(-3) M and 12(R)-hydroxyeicosatetraenoic acid at 10(-6) M inhibited the enzyme activity by 75% and 60%, respectively. We conclude that 19(S)-hydroxyeicosatetraenoic acid is a specific stimulator of renal Na+-K+-ATPase. Therefore, the formation of 19(S)-hydroxyeicosatetraenoic acid by renal cortical cytochrome P450 omega-1-hydroxylase may contribute to the regulation of renal function by regulating Na+-K+-ATPase which is essential for transtubular transport processes.