Eph/ephrin signaling is crucial for organizing retinotopic maps in vertebrates. Unlike other EphAs, which are expressed in the embryonic ventral retina, EphA4 is found in the retinal ganglion cell (RGC) layer at perinatal stages, and its role in mammalian visual system development remains unclear. Using classic in vitro stripe assays, we demonstrate that, while RGC axons are repelled by ephrinB2, they grow on ephrinB1 stripes through EphA4-mediated adhesion. In vivo, retinal axons from EphA4-deficient mice from either sex show impaired arborization in the medial, but not lateral, regions of the superior colliculus that express ephrinB1. Gain-of-function experiments further reveal that ephrinB1-mediated adhesion depends on EphA4 tyrosine kinase activity but it is independent of its sterile alpha motif. Together, our findings suggest that EphA4/ephrinB1 forward signaling likely facilitates adhesion between retinal axon terminals and cells in the medial colliculus, contributing to the establishment of proper connectivity within the visual system.