Signal transduction of interleukin-8 (IL-8) was analyzed in neutrophils, and compared with the well known neutrophil activator N-formyl peptide. Stimulation of human neutrophils with IL-8 induced a rapid polymerization of actin as detected by 7-nitrobenz-2-oxa-1,3-diazol-(NBD)-phallacidin staining of f-actin and reduction of monitored right-angle light scatter. Actin polymerization peaked within 10 seconds after the addition of IL-8 and was short-lived as compared to N-formyl peptide-induced stimulation. Analysis of phospholipids by thin-layer chromatography and analysis of deacylation products of lipid extracts by high-pressure liquid chromatography (HPLC) showed that IL-8 triggered a rapid rise of [32P]phosphatidyl-inositol(3,4,5)trisphosphate (PtdInsP3) followed by a slower increase of [32P]phosphatidylinositol(3,4)bisphosphate (PtdIns-3,4-P2) along with a rapid decrease of [32P]phosphatidylinositol(4,5)bisphosphate (PtdIns-4,5-P2). Changes in polyphosphoinositide metabolism were more moderate and transient than those obtained by N-formyl peptide. Moreover, [32P]phosphatidic acid (PA) production stimulated by IL-8 was minimal and transient as compared to the response activated by N-formyl peptide. Both IL-8 and N-formyl peptide induced Ca++ mobilization from intracellular stores, but IL-8 in contrast to N-formyl peptide failed to trigger the secondary influx of Ca++ from the extracellular medium. In summary, IL-8 and N-formyl peptide stimulated similar and distinct patterns of intracellular activation steps. This study indicates that IL-8 is a potent activator of intracellular events presumably required for chemotaxis, but a relatively weak activator for events associated with superoxide anion generation and proinflammatory activity.