Microiontophoresis of acetylcholine (ACh), atropine, or curare was accompanied by single-unit analysis of visual neuronal responses in the optic tectum of Rana nigromaculata. In 71% of the units tested, ACh enhanced responses to visual stimuli, while atropine suppressed visual responses. Twenty-two percent of the remainder showed no ACh- or atrophin-induced effects and were recorded within 125 microns of the pial surface. The majority of ACh-enhanced, atropine-suppressed units were recorded from visually responsive units localized in the postsynaptic cellular layer 8. The iontophoretic effects of curare were considerably more varied, with approximately equal numbers of units showing an increase (33%), a decrease (35%) or no change (32%) in visually activated responses. The specific effects of curare were also related to the depth from which unit recordings were obtained. These findings indicate that ACh functions as a modulatory neurotransmitter in the frog optic tectum, with a predominantly muscarinic mode of action at postsynaptic levels.