The effect of recombinant human tumor necrosis factor-alpha (TNF) on voltage-gated membrane currents of cultured neurons derived from embryonic rat cerebral cortex was studied using the whole-cell patch-clamp technique. Treatment of neurons with TNF resulted in an increase in outward potassium current density, dependent upon the concentration of TNF and the incubation time, without affecting other membrane currents such as barium and N-methyl-D-aspartate (NMDA). Long exposures (12-48 hr) to TNF (10-100 ng/ml) increased transient outward potassium current (A-current) density without affecting the parameters of activation and inactivation of the current. Prolonged exposures to TNF diminished its increasing effect on the A-current. Since the increase of A-current density induced by TNF is inhibited by both the anti-TNF receptor antibody and cycloheximide treatment, the effect of TNF might be mediated through receptors and by de novo synthesis of the channel protein itself and/or modulating proteins associated with the channel activities. Results indicate that phosphatidylcholine-specific phospholipase C and protein kinase C, but not ceramide, are involved in the signal transduction. In toxicological experiments, TNF had no neurotoxicity. Moreover, a 12 hr pretreatment of TNF protected neurons against NMDA-induced neurotoxicity. This protective effect of TNF was cancelled by 4-aminopyridine, an A-current blocker, suggesting that the increase of A-current densities induced by TNF contributes to the neuroprotection.