The alcoholic leaf extract of Ocimum sanctum (OS, Tulsi) was tested for analgesic activity in mice. In the glacial acetic acid (GAA)-induced writhing test, OS (50, 100 mg/kg, i.p.; and 50, 100, 200 mg/kg, p.o.) reduced the number of writhes. OS (50, 100 mg/kg, i.p.) also increased the tail withdrawal latency in mice. Naloxone (1 mg/kg, i.p.), an opioid antagonist, and DSP-4 (50 mg/kg, i.p.), a central noradrenaline depletor, attenuated the analgesic effect of OS in both the experimental models, whereas, PCPA (300 mg/kg, i.p.), a serotonin synthesis inhibitor, potentiated the action of OS on tail flick response in mice. The results of our study suggest that the analgesic action of OS is exerted both centrally as well as peripherally and involves an interplay between various neurotransmitter systems.