Signals activating the kinases that phosphorylate neurofilament proteins in the axon remain unknown. In a previous study, we have demonstrated that a constitutively active form of MEK1 activates Erk1 and Erk2 kinases, which phosphorylate co-transfected NF-M in NIH 3T3 cells. In this study, we report the activation of endogenous Erk1 and Erk2 by membrane depolarization and calcium influx through L-type calcium channels, which resulted in phosphorylation of the NF-M tail domain in PC12 cells. This phosphorylation was inhibited in the presence of nifedipine, an L-type calcium channel inhibitor, and PD98059, a specific MEK1 inhibitor. Our data suggest a mechanism linking calcium influx through voltage-gated calcium channels with the MAP kinase pathway and NF-M tail domain phosphorylation in cell body and neurite. These findings may provide significant new insights into mechanisms involved in some neurological diseases.