In rat basophilic leukemia cells (2 H3-RBL) stimulated with the calcium ionophore A23187, a rapid build-up of PAF-acether was observed within 5 minutes. Thereafter, a slow and complete catabolism was observed within the next 55 minutes. Accumulation of PAF-acether required calcium in the medium and was increased in the presence of acetyl-CoA. Phenyl methyl sulfonyl fluoride, a serine hydrolase inhibitor active on 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine acetyl hydrolase also produced an increased accumulation of PAF-acether in ionophore-stimulated cells. Quinacrine, a non specific inhibitor of phospholipase A2 impaired PAF-acether formation in a dose-dependent manner. The time-course of PAF-acether formation was compared with the ionophore-induced release of arachidonic acid from these cells.