The retina sends spatially ordered visual information to the superior colliculus (SC) directly and indirectly via the thalamus and primary visual cortex (V1). Gradients of Ephs and ephrins are present in all of these regions, and have been shown to be involved in establishing topography of at least some of these interconnected visual pathways. Studies in ephrin-A knockout mice show that abnormal retinotectal termination zones (TZs) are present in a majority of mice lacking (-/-) ephrin-A2 (57%), and ephrin-A2 and -A5 (89%). A similar but seemingly less disordered pattern is detected in the retina-to-dorsal lateral geniculate nucleus (dLGN) and dLGN-to-V1 projections. Here we analyse the dLGN-to-V1 and V1-to-SC projections in ephrin-A(-/-) mice to determine the extent to which topographic errors are transmitted across synaptic relays. Fluorescent tracers were injected into V1 of wild-type (WT), ephrin-A2(-/-) or ephrin-A2A5(-/-) mice. We examined the number, location and size of anterograde TZs in SC, and mapped the distribution of retrogradely labelled neurons in dLGN. Compared with WT and ephrin-A2(-/-) mice, the volume of individual TZs in the SC was smaller in ephrin-A2A5(-/-) mice (P = 0.002). Single V1 injections labelled two foci of dLGN neurons in 70%, and two SC TZs in 80% of ephrin-A2A5(-/-) mice. Abnormalities in one or other of the projections were detected in 10% of ephrin-A2(-/-) mice. Importantly, there was no consistent correspondence between the organization of geniculocortical and corticotectal projections in either genotype, suggesting a role for ephrin-As in maintaining topographic organization in register across multiple interconnected central visual pathways.