The avian retinotectofugal pathway has been suggested to be homologous to the mammalian retinotectofugal pathway. The projection of the nucleus rotundus upon the ectostriatum is equivalent to that of the pulvinar nucleus upon the extrastriate cortex in mammals. In this system, the optic tectum relays retinal input to the nucleus rotundus, which then ascends to the ectostriatum of the telencephalon. Given the fact that the chick retinotectofugal system becomes mature early during development, the present study attempted to investigate the timecourse of neuronal connections of the embryonic rotundoectostriatal pathway. We used multiple injections of cholera toxin B subunit (CTb) in the ectostriatum of chick embryos to retrogradely trace projections to the nucleus rotundus. We found CTb-labeled neurons in the nucleus rotundus at embryonic day 7.5–8. By embryonic day 8–8.5, increased numbers of CTb-labeled neurons were seen in the nucleus rotundus. It was noted that the time of this initial connection between the nucleus rotundus and the ectostriatum is nearly synchronous with that of the retinotectal and tectorotundal pathways, respectively (Crossland et al., 1975; Thanos & Bonhoeffer, 1987; Wu et al., 2000). These findings, combined with the present study, suggest that the retinotectofugal system becomes established, at least at a structural level, by embryonic day E8.