During the development of the Drosophila visual system, photoreceptor (retinal) axons (R axons) project retino-topically to their targets in the optic lobes. The establishment of this precise pattern of connections does not depend on interactions between adjacent axon bundles, suggesting that R axons rely on environmental signals for proper pathfinding. Glial cells that are located along the R-axon trajectory are likely candidates to provide guidance cues for R-axon navigation. This study defines the origin of lamina glia (L glia), and demonstrates that L glia migrate into the lamina over a considerable distance. Glia are located in positions at which the R axons make critical growth choices. In the absence of cues from the eye, several classes of glia migrate to their final positions within the optic lobe anlage and begin to differentiate. Our results are consistent with a role for the glia in providing guidance cues to the R axons.