Neurotrophins activate multiple signaling pathways in neurons. However, the precise roles of these signaling molecules in cell survival are not well understood. In this report, we show that nerve growth factor (NGF) activates the transcription factors NF-κB and AP-1 in cultured sympathetic neurons. Activated NF-κB complexes were shown to consist of heterodimers of p50 and Rel proteins (RelA, as well as c-Rel), and NF-κB activation was found to occur independently ofde novo protein synthesis but in a manner that required the action of the proteasome complex. Treatment with the NF-κB inhibitory peptide SN50 in the continuous presence of NGF resulted in dose-dependent induction of cell death. Under the conditions used, SN50 was shown to selectively inhibit NF-κB activation but not the activation of other cellular transcription factors such as AP-1 and cAMP response element-binding protein. Cells treated with SN50 exhibited morphological and biochemical hallmarks of apoptosis, and the kinetics of cell killing were accelerated relative to death induced by NGF withdrawal. Finally, experiments were conducted to test directly whether NF-κB could act as a survival factor for NGF-deprived neurons. Microinjection of cells with an expression plasmid encoding NF-κB (c-Rel) resulted in enhanced neuronal survival after withdrawal of NGF, whereas cells that were transfected with a vector encoding a mutated derivative of c-Rel lacking the transactivation domain underwent cell death to the same extent as control cells. Together, these findings suggest that the activation of NF-κB/Rel transcription factors may contribute to the survival of NGF-dependent sympathetic neurons.