The effects of a moderate dietary intake of n-3 polyunsaturated fatty acids were determined on the eicosanoid synthesis by rat peritoneal macrophages: the availability of arachidonic acid (20:4 (n-6)) and eicosapentaenoic acid (20:5 (n-3] in the membrane phospholipids of peritoneal macrophages and splenic lymphocytes and the synthesis of the main eicosanoids were evaluated in parallel. The n-6/n-3 ratio was decreased from 13.5 in the control diet rich in 18:2(n-6) to about 2 by an addition of either linseed oil providing 18:3(n-3) (linseed oil diet) or fish oil providing 20:5(n-3) and 22:6(n-3) (fish oil diet). The dietary linoleic acid content was at least 3.5% of energy in each diet. In peritoneal macrophage and splenic lymphocyte membrane phospholipids, arachidonic acid content was significantly decreased with the linseed oil and fish oil diets. 20:5(n-3) content was significantly raised up in the linseed oil group (3-fold in macrophage phospholipids) and more strikely in the fish oil group (7-fold). In response to a stimulation by the calcium ionophore A23187, peritoneal macrophages released 6-ketoPGF1 alpha, TXB2, PGE2 and LTB4 and/or 5, but no or few peptidoleukotrienes. The linseed oil did not affect significantly the synthesis of these eicosanoids, except for LTB5, which was increased 2-fold. In contrast, the fish oil diet led to significant decreases in the productions of 6-ketoPGF1 alpha and PGE2 (-50%) and LTB4 (-40%) and to a 10-fold increase in the release of LTB5. TXB2 was also decreased (-35%), but not significantly. These results and the parallel evolutions of the two ratios 20:4(n-6)/20:5(n-3) and LTB4/LTB5 showed that eicosanoid synthesis is greatly dependent on the relative availability of the two substrates in competition.