The aim of the present study is to provide a complete description of the distribution of choline acetyltransferase (ChAT) immunoreactivity (i) in the brain of the lizard Gallotia galloti, on the basis of two different primary antisera: rat anti-ChAT and rabbit anti-chicken ChAT. Considering that the brain is a segmented structure, we have analysed our data with respect to transverse segmental domains (or neuromeres), which have been previously described by several authors in the brain of vertebrates. In the telencephalon, ChATi neurons are seen in the cortex, anterior dorsal ventricular ridge, basal ganglia, diagonal band, and bed nucleus of the stria terminalis. Further caudally, ChATi cell bodies are located in the preoptic area, hypothalamus, habenula, isthmus, and all motor efferent centers of the brainstem and spinal cord. Plexuses of ChATi fibers are observed in the areas containing cholinergic cell bodies. In addition, distinct plexuses are found in the cortex, the posterior dorsal ventricular ridge, the neuropiles of all primary visual centers of the diencephalon and mesencephalon, and several non-visual nuclei of the brainstem. The distribution of ChAT immunoreactivity in the brain of G. galloti resembles in many respects that of other vertebrates, and differences are mainly observed in the pretectum and midbrain tectum. Transverse segmental domains were identified in the brainstem and forebrain of Gallotia when the cranial nerve roots and fiber tracts were used as a reference, and most cranial motor nuclei were found to occupy the same segmental positions as have been reported in the chick.