The properties of glial cells in lesioned nerves contribute quite substantially to success or failure of axon regeneration in the CNS. Goldfish retinal axons regenerate after optic nerve lesion (ONS) and express the L1-like cell adhesion protein E587 antigen on their surfaces. Goldfish oligodendrocytes in vitro also produce E587 antigen and promote growth of both fish and rat retinal axons. To determine whether glial cells in vivo synthesize E587 antigen, in situ hybridizations with E587 antisense cRNA probes and light- and electron microscopic E587 immunostainings were carried out. After lesion, the goldfish optic nerve/tract contained glial cells expressing E587 mRNA, which were few in number at 6 days after ONS, increased over the following week and declined in number thereafter. Also, E587-immunopositive elongated cells with ultrastructural characteristics of oligodendrocytes were found. Thus, glial cells synthesize E587 antigen in spatiotemporal correlation with retinal axon regeneration. To determine the functional contribution of E587 antigen, axon-oligodendrocyte interactions were monitored in co-culture assays in the presence of Fab fragments of a polyclonal E587 antiserum. E587 Fabs in axon-glia co-cultures prevented the normal tight adhesion of goldfish retinal growth cones to oligodendrocytes and blocked the preferential growth of fish and rat retinal axons on the oligodendrocyte surfaces. The ability of glia in the goldfish visual pathway to upregulate the expression of E587 antigen and the growth supportive effect of oligodendrocyte-associated E587 antigen in vitro suggests that this L1-like adhesion protein promotes retinal axon regeneration in the goldfish CNS.