Exostosin tumor-like 3 (EXTL3) is a glycosyltransferase involved in heparan sulfate (HS) biosynthesis. HS proteoglycans are critically involved in different steps during brain development. The present in situ hybridization in mice revealed wide EXTL3 expression at different grades in the central and peripheral nervous system components including the neural retina and neural crest-derived structures at embryonic days (E) 11.5, E12.5, E14.5, and E16.5. In the neopallial cortex, an intense EXTL3 expression was observed in the neuroepithelial cells lining the ventricular zone at E11.5 and E12.5. The signal decreased at E14.5 and was further downregulated at E16.5 in the ventricular zone. The pioneer neurons of the preplate at E12.5 differentially expressed the gene. Heavily stained among weakly or negatively stained neurons were observed. At E14.5, the cortical plate cells were moderately and homogeneously stained. In contrast, at E16.5, an upregulated and differential expression pattern was detected. The labeling pattern at E16.5 subdivided the cortical plate cells into a large number of heavily, a moderate number of less intensely, and some negatively stained cell populations. Interestingly, the distinct expression pattern displayed by the three main cell types of the adult cerebral cortex was similar to that of the late corticogenesis stage (E16.5). In the adult, the strongest expression was observed in the pyramidal neurons. The granule-type neurons showed less intense staining while the glia cells were devoid of signals. Our data revealed that EXTL3 expression is developmentally regulated in the mouse nervous system and suggested that it differentially contributes to brain development and corticogenesis.