Viruses utilize normal cell surface structures as attachment sites. Interaction of viral components with these structures may alter target cell growth. In the present study, the expression and function of the cell surface receptor for reovirus type 3 (Reo3R) was studied in neonatal rat optic nerve glial cultures. The Reo3R is expressed by mature oligodendrocytes and astrocytes but not by O-2A progenitor cells. It appears at an early stage of oligodendrocyte development, coincident with the O4 marker but prior to galactocerebroside or myelin basic protein. Anti-Reo3R antibodies stimulate the expression of galactocerebroside by developing oligodendrocytes. Divalent Reo3R-binding peptides are similarly active. Maximal stimulation of galactocerebroside expression occurs with treatment as short as 4 hr, consistent with a receptor-mediated process. Cell surface structures used as an attachment site by reovirus type 3 may also play a role in the regulation of oligodendrocyte differentiation.