Roots of Plumbago zeylanica have been reported to improve learning and memory of mice, but the effect of plumbagin on learning and memory of mice and the possible mechanisms for its effect on memory have not been explored till date. So the present study was designed to evaluate the effect of plumbagin on learning and memory of Swiss young male albino mice. Plumbagin (4, 8, 16 mg/kg, p.o.) and physostigmine (0.1 mg/kg,i.p.) per se were administered for 15 successive days to separate groups of mice. Behavioral parameters of learning and memory were recorded using Morris water maze. Acetylcholinesterase activity was estimated in brain of mice. Effect of plumbagin on scopolamine (0.4 mg/kg, i.p.) and diazepam (1 mg/kg, i.p.)-induced amnesia was also investigated. Locomotor activities of mice were also recorded. Plumbagin (8 and 16 mg/kg) and physostigmine significantly improved learning and memory of mice, as indicated by decrease in escape latency during training and increase in time spent in target quadrant of Morris water maze during retrieval. The drugs did not show any significant effect on locomotor activities of mice. Memory improving activity of plumbagin (16 mg/kg) was equivalent to physostigmine. Plumbagin significantly reversed scopolamine- and diazepam- induced amnesia in mice. Plumbagin and physostigmine also significantly reduced brain acetylcholinesterase activity of mice. In conclusion, plumbagin significantly improved memory of mice possibly through inhibition of brain acetylcholinesterase activity and through involvement of GABA-benzodiazepine pathway. Thus, plumbagin may be explored further for management of cognitive dysfunction.