In order to study the relationship between retinal projections and immunohistochemically identified neurotransmitter systems in the primary visual centers of the brain in lizards, intraocular injections of horseradish peroxidase were combined with immunohistochemistry. Antibodies raised against six substances were applied: choline acetyltransferase (ChAT), serotonin (5-HT), tyrosine hydroxylase (TH), dopamine (DA), substance P (SP), and leu-enkephalin (LENK). In the primary visual centers of the lizards Gekko gecko and Gallotia galloti, notable overlap was observed between retinofugal fibers with: 1) ChAT-immunoreactive fibers in almost all primary visual centers; 2) 5-HT-immunoreactive fibers in the ventral lateral geniculate body and the basal optic nucleus; 3) TH-immunoreactive fibers in the nucleus ovalis and the dorsal lateral geniculate body; 4) SP- and LENK-immunoreactive fibers in the perirotundal belt; and 5) TH- and SP-immunoreactive fibers in the pretectal posterodorsal nucleus. The latter nucleus also contains dopaminergic cell bodies that lie outside the retinal target area but have dendrites extending into it. Several differences were noted in the distribution of 5-HT-, TH-, DA-, and LENK-immunoreactive fibers in the tectum of the midbrain in the two species studied. Distinct laminae of 5-HT-immunoreactive fibers (layer 9) and TH- and DA-immunoreactive fibers (layers 9 and 11) are present in G. gecko but absent or, at least, less distinct in G. galloti. On the contrary, the optic layers in the tectum of G. galloti show a rather dense plexus of LENK immunoreactive fibers, whereas the corresponding layers in G. gecko are devoid of LENK-immunoreactivity. Since only a very few ChAT immunoreactive fibers were observed in the optic nerve of G. galloti, most of the observed immunoreactive fibers in the primary visual centers are considered to have an extraretinal origin. Putative sources of the cholinergic, the monoaminergic, and the peptidergic innervation of the primary visual centers in reptiles include the isthmic nucleus, the raphe nuclei, the substantia nigra and the nucleus of the posterior commissure. as reported in other amniotes.