Optic nerve fibers run parallel from the retina as far as the optic tract in fish, then suddenly criss-cross into a new pattern matching the tectal map. This change coincides with a unique demarcation between two astroglial territories in the retinotectal pathway, located where the optic chiasm occurs in other vertebrates, which we defined using antibodies directed against intermediate filaments (IF). We found that astroglia in optic nerve territory express an Mr 56,000 IF polypeptide, band 3, which we identify as the fish equivalent of vimentin in mammals. These astrocytic cells lack glial fibrillary acidic protein (GFAP; cf. Dahl and Bignami, 1973). Conversely, glia in brain territory, that is, in the optic tract and elsewhere in the CNS, lack the fish vimentin, but express GFAP. By electron microscopy, we obtained evidence that new retinal axons extend swiftly through the growing optic nerve, where they are tightly shepherded into a narrow track by newly differentiating glial cells, positive for the fish vimentin. In the GFAP-positive glial territory of the optic tract, by contrast, growing axons are slowed down and probably branch. We suggest that this allows them to fasciculate accurately with older fibers and thereby propagate a tectotopic pattern established by pioneer axons in the embryo.