The effect of THIP, a direct-acting gamma-aminobutyric acid (GABA) receptor agonist, on the antinociceptive response to a variety of agents was examined using the mouse tail-immersion assay. Alone, THIP produced an antinociceptive response in smaller doses (5 mg/kg) but was ineffective at doses exceeding 10 mg/kg. Treatment with THIP (15 mg/kg) was found to block the antinociceptive response to an inhibitor of the uptake of GABA, an inhibitor of GABA-transaminase, a direct-acting GABA receptor agonist and to a cholinesterase inhibitor. In contrast, THIP had no effect on the antinociceptive responses to morphine, clonidine or oxotremorine. The results indicate that large doses of THIP reduce cholinergic activity in a pathway important for mediating the antinociceptive action of GABAergic drugs and physostigmine.