The effects of a selective phosphodiesterase (PDE) type IV inhibitor, rolipram, on activation of neutrophil phospholipases in response to the chemotactic peptide formyl-methiony-leucyl-phenylalanine (fMLP) were investigated. fMLP caused liberation of arachidonic acid, a precursor of eicosanoids and in the presence of 0.3% butanol, production of phosphatidylbutanol, an indicator of phospholipase D activation. Rolipram inhibited arachidonic acid release and phosphatidylbutanol formation. The inhibition was considered to be mediated through a cAMP-dependent mechanism, probably protein kinase A, because selective inhibitors for protein kinase A, H-8 or H-89 overcame the action of rolipram. The concentration-dependent inhibitory profile for phospholipase D activation was similar to that for lysosomal enzyme release, providing additional evidence for the functional link of these two events. In contrast, rolipram was without effect on fMLP-induced inositol trisphosphate production. These results indicate that this selective PDE IV inhibitor had no effect on phosphatidylinositol-specific phospholipase C activation but effectively prevented activation of phospholipases A2 and D coupled to arachidonic acid liberation and lysosomal enzyme release, respectively.