The role of protein kinase C (PKC) isoforms in the neural cell adhesion molecule (NCAM)‐mediated neurite outgrowth was tested using a co‐culture system consisting of fibroblasts with or without NCAM expression upon which either primary cerebellar granular neurones (CGN) or pheochromocytoma (PC12‐E2) cells were grown. The latter transiently expressed various PKC isoforms and domains derived from selected PKCs. PKC inhibitors of various specificity inhibited NCAM‐stimulated neuritogenesis from CGN, indicating that PKC is involved in this process. Moreover, stimulation by the NCAM‐mimetic peptide, C3d, elicited phosphorylation of PKC in CGN. Expression of kinase‐deficient forms of PKCα, βI and βII blocked NCAM‐mediated neurite extension, but had no effect on nerve growth factor (NGF)‐mediated neurite outgrowth. Expression of two PKCɛ constructs: (i) a fragment from PKCɛ encompassing the pseudosubstrate, the C1a domain (including the actin‐binding site, ABS), and parts of the V3 region, or (ii) the PKCɛ‐specific ABS blocked NCAM‐mediated neurite extension in both cases. These two constructs also partially inhibited NGF‐stimulated neuritogenesis indicating that PKCɛ is a positive regulator of both NCAM‐ and NGF‐mediated differentiation. We suggest that PKCɛ is a common downstream mediator for several neuritogenic factors, whereas one or more conventional PKCs are specifically involved in NCAM‐stimulated neurite outgrowth.