The effects of eicosapentaenoic (EPA, 20:5n-3) and docosahexaenoic acids (DHA, 22:6n-3) on the phospholipase $A_2$ ($PLA_2$)-mediated release of arachidonic acid (AA, 20:4n-6) were studied in kidney microsomes from rats fed diets containing sunflower oil (SO) or fish oil (FO) concentrate for 11 months. The amounts of AA released by the endogenous $PLA_2$ enzyme were significantly lower by 38% in the FO, compared to the SO-fed rats (23.2 nmol versus 60.7 nmol AA released/mg protein/h in the FO- and SO-treated groups, respectively). The FO-derived microsomes released less linoleic acid (LA, 18:2n-6) and adrenic acid (22:4n-6), but larger amounts of the n-3 fatty acids, including EPA, DHA, docosapentaenoic acid (DPA, 22:5n-3), and 20:4n-3 than the SO-derived microsomes. A similar replacement of the AA and adrenic acid with the n-3 fatty acids including EPA and DHA was also observed in the microsomal phospholipid fraction from the FO-fed rats relative to the SO-treated group. The results suggest that the $PLA_2$-mediated release of AA is reduced and that of EPA is increased in compensation for AA decline in kidney microsomes from FO-fed rats (0.7 nmol EPA/mg protein/h versus 22.7 nmol EPA/mg protein/h for the SO and FO-treated groups). Replacement of the n-6 with n-3 fatty acids may explain the reduced synthesis of the AA-derived prostaglandins and the concomitant rise in the EPA-derived prostaglandins observed in kidneys of FO-treated rats.