In the chicken retina, choline acetyltransferase-like immunoreactivity (ChAT-LI) defines three populations of cholinergic amacrine cells and two terminal bands in the inner plexiform layer (IPL). Acetylcholinesterase (AChE) histochemistry defines two prominent bands within the IPL which corresponded to those containing ChAT. Other AChE-positive bands in the IPL are not associated with cholinergic transmission sites. Cholinergic cell bodies contain AChE, but the most intensely AChE-positive cells do not appear to be cholinergic. AChE histochemistry may be used to define the major cholinergic synaptic sites in the IPL, and may be a useful marker of IPL lamination.