Neuronal polarity development begins by the outgrowth of the axon and the formation of the axon initial segment which acts as a diffusion barrier and it is the place of action potential generation. The mechanisms controlling this development are largely unknown. We describe a role for IkappaB alpha, the NFkappaB inhibitor, in the initial stages of axon outgrowth and the development of the axon initial segment. In cultured hippocampal neurons, inhibition of IkappaB alpha phosphorylation by IkappaB kinases (IKKs) impedes axon outgrowth. Moreover, the absence of IkappaB alpha phosphorylation, in the next stages of axon development, impairs the localization of structural and functional proteins at the axon initial segment, such as ankyrin G and voltage gated sodium channels. These results demonstrate a new role for proteins of the NFkappaB pathway in the acquisition of neuronal polarity and its involvement in the development of the axon initial segment.