In this work, the differential expression of a chemical marker, the alpha-isoform of the calcium/calmodulin-dependent protein kinase II (CaM-Kinase II alpha) and the development of the spinal cord projection were used to determine in vivo the embryonic stages at which different aspects of the phenotype of neocortical cells are specified. We first performed a quantitative, immunocytochemical study on the levels of CaM-Kinase II alpha expression in the frontal, parietal and occipital cortical areas of control adult rats. We found that the levels of expression of CaM-Kinase II alpha were larger in the frontal and parietal areas than in the occipital areas. In addition, all layer V neurons identified as projecting to the spinal cord were CaM-Kinase II alpha immunopositive. We then grafted embryonic day (E) 12 or 14 cells from the presumptive frontal or occipital cortex of donor fetuses into the frontal or occipital cortex of newborn hosts. Cortical cells grafted at E12 differentiate neurons with molecular (CaM-Kinase II alpha) and connectivity (spinal cord projection) phenotypes appropriate to the cortical area where they complete their development whereas cells taken at E14 differentiate neurons with molecular and connectivity phenotypes appropriate to their cortical locus of origin. These findings suggest that E12 progenitors destined to generate layer V neurons are multipotent. The final phenotype of their progeny depends on regionalizing signals expressed in the environment. Later in corticogenesis, committed progenitors become unable to respond to regionalizing signals and generate neurons whose phenotype is appropriate to the initial cortical position of the precursor.