V. Kaever 1, K. Wessel, H.H. Radeke and K. Resch Institute of Molecular Pharmacology, Department of Pharmacology and Toxicology, Medical School Hannover, Konstanty-Gutschow-Str. 8, D-3000 Hannover 61 Introduction Prostaglandins (PG) and leukotrienes (LT) play important roles in many physiological and patho- physiological situations, including inflammatory and allergic diseases. Tissue-bound macrophages are a major source of different eicosanoids in re- sponse to various soluble as well as particulate stimuli [1]. The basal as well as the stimulus-depen- dent formation of PG, LT, and hydroxyeicosate- traenoic acids is regulated in a complex manner by several enzymes and cofactors. We have shown previously that in resident mouse peritoneal macrophages the activation of the calcium- and phospholipid-dependent protein kinase C (PKC) leads to an increase in free arachidonic acid (AA) available for PG synthesis within minutes [2]. We further investigated in more detail by which factors the ratio of PG and LT synthesis may be controlled within this cell type. In this report we demonstrate the effects of calcium channel antagonists and calmodulin antagonists on macrophage eicosanoid synthesis taking into account the dominant role of calcium in the regulation of AA liberation and metabolism. Materials and methods Resident macrophages were harvested by peri- toneal lavage of untreated DBA/2 mice and main- tained in serumfree medium in 96-well plates at a 1 Author for correspondence. cell density of 5 x 105 cells/ml [3]. After a preincu- bation time of 2 hrs non-adherent cells were re- moved by washing and the adherent macrophages (about 5 x 104 cells/well in 200 ~tl medium) were used in the individual experiments. Eicosanoids were determined from the cellular supernatants by specific enzyme-linked immunosorbent assay us- ing monoclonal anti PGE2 or LTC4 antibodies (kindly provided by Prof. K. Brune, Erlangen, FRG) as previously described [4]. PGE 2 was ob- tained from Paesel, Frankfurt, FRG, and N-(6- aminohexyl)- 5-chloro-l-naphthalene sulfonamide (W-7) was from Sigma, Deisenhofen, FRG. LTC4 was kindly provided by Hoechst, Frankfurt, FRG, and 1,3-dihydro-l-[1-[(4-methyl-4H,6H-pyrrolo [1,2-@[4,1] benzoxazepin-4-yl)methyl]-4-piperi- dinyl]-2H-benzimidazol-2-one maleate (CGS9343B) was kindly provided by Ciba-Geigy, Summit, USA. Data are means of duplicate cell incubations and quadruplicate eicosanoid determinations. Results and discussion One important cofactor for PKC, different phos- pholipases, and AA 5-1ipoxygenase activities is the intracellular free calcium. Many physiological stimuli induce eicosanoid synthesis by an increase of phosphoinositide turnover and thus lead to acti- vation of PKC and an enhanced calcium level. In a model system this can also be achieved by the combined action of direct PKC activators and cal- cium ionophores [31.