The significance of age-dependent changes in acetylcholinesterase (AChE) activity level is poorly understood. Reported here is one approach to understand AChE's function as it relates to age: to investigate how inhibition of AChE affects behavior in freely moving Aplysia of two age groups, mature and old. The siphon/gill withdrawal reflex (S/GWR) and gill pumping movement (GPM) were examined to assay the effects of AChE inhibition by BW284c51, a specific and reversible AChE inhibitor. In mature Aplysia AChE inhibition by 2 microM and 5 microM of BW284c51 resulted in a significantly shortened S/GWR duration, and in suppression of habituation and dishabituation. In old animals, AChE inhibition by 2 microM of BW284c51 did not affect S/GWR and only dishabituation was suppressed by inhibition by 5 microM of BW284c51. AChE inhibition reduced the GPM rate significantly only in mature animals. AChE inhibition did not alter the decrement in GPM rate which is regularly observed in both age groups during repetitive exposure to acidified sea water. Thus both S/GWR and GPM were affected by AChE inhibition, and a significant age effect on the two behaviors was observed. Comparisons of the results of AChE inhibition which would elevate acetylcholine (ACh) levels with those of carbachol administration revealed that AChE inhibition affects both cholinergic and non-cholinergic mechanisms underlying the two behaviors.