We have analyzed the expression of the oligodendrocyte-derived extra-cellular matrix molecule janusin (previously termed J1-160/180) in the retina and optic nerve of developing and adult mice using indirect light and electron microscopic immunocytochemistry, immunoblot analysis, and enzyme-linked immunosorbent assay. In the optic nerve, janusin is not detectable in neonatal and only weakly detectable in 7-day-old animals. Expression is at a peak in 2- or 3-week-old animals and subsequently decreases with increasing age. In the retina, expression increases until the third postnatal week and then remains at a constant level. In immunocytochemical investigations at the light microscopic level, janusin was found in the myelinated regions of the nerve with spots of increased immunoreactivity possibly corresponding to an accumulation of the molecule at the nodes of Ranvier. At the electron microscopic level, contact sites between unmyelinated axons, between axons and glial cells, and between axons and processes of myelinating oligodendrocytes were immunoreactive. Cell surfaces of astrocytes at the periphery of the nerve and forming the glial-limiting membrane, in contrast, were only weakly immunopositive or negative. In cell cultures of young postnatal mouse or rat optic nerves, oligodendrocytes and type-2 astrocytes, but not type-1 astrocytes were stained by janusin antibodies. In the oligodendrocyte-free retina, janusin was detectable in association with neuronal cell surfaces, but not with cell surfaces of Müller cells or retinal astrocytes. Our observations indicate that expression of janusin in the optic nerve and in the retina is developmentally differentially regulated and that other cell types, in addition to oligodendrocytes, express the molecule. Since the time course of janusin expression in the optic nerve coincides with the appearance of oligodendrocytes and myelin and since janusin is associated with cell surfaces of oligodendrocytes and outer aspects of myelin sheaths and is concentrated at nodes of Ranvier, we suggest that janusin is functionally involved in the process of myelination.