The localization of aggrecan and mRNA splice variants of versican in the developing rat central nervous system has been examined by using specific polyclonal antibodies to the nonhomologous glycosaminoglycan attachment regions of these hyaluronan-binding chondroitin sulfate proteoglycans. At embryonic day 16 (E16), aggrecan and versican splice variants containing either or both the α-and β-domains are present in the marginal zone and subplate of the cerebral cortex and in the amygdala, internal capsule, and the optic and lateral olfactory tracts. There is strong staining of versican but not of aggrecan in the hippocampus and dentate gyrus by E19, whereas both aggrecan and α-versican are present in the fimbria. At E19, aggrecan is seen throughout the cerebral cortex, whereas the distribution of versican is considerably more limited, being confined essentially to the marginal zone and subplate. At 1 week postnatal, both aggrecan and versican are present in the prospective white matter and in the molecular and granule cell layers of the cerebellum, but neither proteoglycan is seen in the external granule cell layer. α- but not β-versican staining is seen in Purkinje cells, and aggrecan staining of Purkinje cells is also rather minimal. In the spinal cord at E13, aggrecan is present in the dorsal root entry zone, ventral funiculus, mantle layer, and floor plate, as well as in the dorsal root ganglia and ventral roots. However, α-versican is confined to the dorsal root entry zone and the ependyma surrounding the spinal canal, and β-versican is not present in spinal cord parenchyma at this developmental stage, being limited to the surrounding connective tissue. By E19, there are significant amounts of all three proteoglycans in the spinal cord. Aggrecan staining is most intense in the lateral funiculus and the fasciculi gracilis and cuneatus, where α-versican staining is also strong. In contrast, β-versican is seen predominantly in the motor columns. Differences in the localization and temporal expression patterns of these chondroitin sulfate proteoglycans suggest that, like neurocan and phosphacan, they have partially complementary roles during central nervous system development. Development Dynamics 227:143–149, 2003. © 2003 Wiley-Liss, Inc.