Over the last few years, several studies showing that production of superoxide by neutrophils in response to chemotactic factors such as FMLP is enhanced after preincubation of the cells with granulocyte-macrophage (GM)-CSF or TNF-alpha have been published. Subsequent reports have indicated that this effect of the cytokines may be mediated by modulation of the number and/or affinity of surface receptors for FMLP. In the present study we have investigated the effect of preincubation with GM-CSF and TNF-alpha on the oxidative burst induced by sodium fluoride and guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S)-agents which directly activate guanine-nucleotide binding proteins in neutrophils. Pretreatment of neutrophils with either GM-CSF or TNF-alpha dose-dependently enhanced the production of superoxide induced by NaF, as determined by the superoxide dismutase-inhibitable reduction of ferricytochrome c. Furthermore, preincubation of neutrophils with these cytokines enhanced the production of hydrogen peroxide induced by GTP gamma S in electroporated neutrophils. Because both NaF and GTP gamma S directly activate G proteins independently of external receptor-G protein interaction, these results imply that both GM-CSF and TNF-alpha alter the neutrophil signal transduction pathway in response to subsequent agonists independently of a modulation in the expression of the cell surface receptors for such agonists.