The effects of a specific presynaptic cholinergic antagonist, toosendanin, on memory formation following a passive avoidance training experience in day-old chicks was investigated. Bilateral injection of toosendanin into the neostriatal/hyperstriatal region of the chick forebrain produced memory impairment in a dose-dependent manner. Retention deficits were apparent from 20 min following training in chicks treated with toosendanin, regardless of the injection time relative to training. Chicks that received injections of the drug at corresponding times prior to retention tests showed normal retention levels, suggesting that toosendanin has no effect on performance and memory retrieval. These results indicate an involvement of cholinergic transmission during an early stage of memory formation.