Using immunocytochemistry, we have compared the distribution of neurocan and phosphacan in the developing central nervous system. At embryonic day 13 (E13), phosphacan surrounds the radially oriented neuroepithelial cells of the telencephalon, whereas neurocan staining of brain parenchyma is very weak. By E16-19, strong staining of both neurocan and phosphacan is seen in the marginal zone and subplate of the neocortex, and phosphacan is present in the ventricular zone and also has a diffuse distribution in other brain areas. Phosphacan is also widely distributed in embryonic spinal cord, where it is strongly expressed throughout the gray and white matter, in the dorsal and ventral nerve roots, and in the roof plate at E13, when neurocan immunoreactivity is seen only in the mesenchyme of the future spinal canal. Neurocan first begins to appear in the spinal cord at E16-19, in the region of ventral motor neurons. In early postnatal and adult cerebellum, neurocan immunoreactivity is seen in the prospective white matter and in the granule cell, Purkinje cell, and molecular layers, whereas phosphacan immunoreactivity is associated with Bergmann glial fibers in the molecular layer and their cell bodies (the Golgi epithelial cells) below the Purkinje cells. These immunocytochemical results demonstrate that the expression of neurocan and phosphacan follow different developmental time courses not only in postnatal brain (as previously demonstrated by radioimmunoassay) but also in the embryonic central nervous system. The specific localization and different temporal expression patterns of these two proteoglycans are consistent with other evidence indicating that they have overlapping or complementary roles in axon guidance, cell interactions, and neurite outgrowth during nervous tissue histogenesis.