Abstract We studied the effects of aging on the [ 3H]-choline uptake, acetylation, [ 3H]-ACh release and muscarinic modulation of [ 3H]-ACh release in cortical synaptosomes prepared from Fischer 344 male rats. Our results indicate that 6 and 24 month old rats take up and acetylate [ 3H]-choline to a similar extent, but that the older animals release significantly less [ 3H]-ACh in response to K +-depolarization than the young adults do. This difference in K +-induced release is not due to a difference in presynaptic muscarinic receptor inhibitory activity since the older animals appear to be, if anything, slightly less sensitive to oxotremorine than the younger animals are. Atropine (1 μM) had no effect on ACh-release but blocked oxotremorine-induced modulation. Our results suggest that acetylcholine release is decreased in synaptosomes prepared from old rats although the presynaptic muscarinic regulation of release is functional. Thus, muscarinic receptor-mediated release-modulation is a potential site for pharmacologically altering ACh release.