Frog retinal ganglion cells projecting to the tectum layer F release acetylcholine as co-mediator

► Co-release of acetylcholine from frog retinotectal terminals was investigated. ► Application of KYNA or CNQX reduced retinotectal transmission to subthreshold level. ► The reduction had no effect on the phasic potentiation of retinotectal transmission. ► This indicated that source of acetylcholine can only be in retinotectal terminals.