The aim of the present work was to elucidate the role of cytosolic calcium ions, [Ca2+]i, in the control of arachidonic acid release and metabolism. [Ca2+]i was measured in resident peritoneal rat macrophages loaded with Fura2, and compared with the release of leukotriene B4(LTB4) and prostaglandin l2 (PGL2, assayed through its hydrolysis product 6-keto-PGF1 alpha). The calcium ionophore A 23187 stimulated both an increase in [Ca2+]i and the release of LTB4 and 6-keto-PGF1 alpha. On the contrary, zymosan and opsonized zymosan, while stimulating eicosanoid release to an extent only slightly lower than A 23187, did not affect [Ca2+]i. Lipopolysaccharide stimulated 6-keto-PGF1 alpha, but not LTB4, release, without affecting [Ca2+]i. In parallel experiments, macrophages were prelabelled with [3H]arachidonic acid and the release of total 3H-products was assayed and taken as an index of phospholipase activity. A 23187, zymosan and opsonized zymosan increased the release of 3H-products in the presence of Ca2+. When extracellular Ca2+ was removed, the ionophore-induced 3H-products release was greatly blunted, while the release induced by zymosan was actually augmented. Our data indicate that a generalized [Ca2+]i increase is not necessary for arachidonic acid release and metabolism in rat peritoneal macrophages.