The glycosphingolipid (GSL) composition of cells changes dramatically during cellular differentiation. Nerve growth factor (NGF) or forskolin (FRK) are known to induce cellular differentiation including process formation in PC12 pheochromocytoma cells. In this respect, we present the NGF/FRK‐dependent regulation of glycosyltransferase activities and the corresponding GSL expression in PC12D cells. After treatment of PC12D cells with NGF or FRK, the cell processes, including varicoses and growth cones, became strongly immunoreactive with an antibody against a unique globo‐series neutral GSL, Galα1‐3Galα1‐4Galβ1‐4Glcβ1‐1′Cer (GalGb3), and the activity of GalGb3‐synthase increased significantly. Other glycosyltransferase activities, including GM1 containing blood group B determinant (BGM1)‐, GM3‐, GD1a‐, and GM2‐synthases, also increased significantly upon NGF treatment, but the immunoreactivity against BGM1 did not show any appreciable change. For the parent PC12 cells, NGF/FRK treatment significantly increased the percentage of anti‐GalGb3 positive cells and induced some immunoreactive cell processes. Because the parent PC12 cells do not express appreciable amounts of GalGb3, and because PC12D cells are considered to be more differentiated than the parent PC12 cells, the expression of GalGb3 and the increase of GalGb3‐synthase activity may be closely related to the cellular differentiation process in this cell line.