The anxiolytic effect of the alcoholic extract of ting-chih-wan (TCWa) was studied using the black and white test (BWT) and the elevated plus-maze (EPM). We further demonstrated the anxiolytic mechanism of TCWa by combining with diazepam (DIZ), serotonin (5-HT) agonists or antagonists, and measuring the levels of monoamines and its metabolites in the brain stem and cortex. In the BWT, TCWa (0.1-1.0 g/kg, p.o.) increased the time spent in the white chamber and total change between the two chambers, and decreased the time spent in the black chamber. TCWa (0.1-0.5 g/kg, p.o.) increased the arm entries and the time spent on the open arms, and decreased the arm entries and the time spent on the closed arms in the EPM. On the other hand, TCWa (1.0 g/kg, p.o.) decreased horizontal activity and prolonged pentobarbital-induced sleeping times. TCWa (0.1, 0.5 g/kg) decreased the levels of norepinephrine (NE), dopamine (DA), 5-HT and 5-hydroxy-indoleacetic acid (5-HIAA) and increased the levels of vanillylmandelic acid (VMA) and homovanillic acid (HVA) in the brain stem. TCWa (0.1 and 0.5 g/kg) decreased the levels of NE, DA and increased the levels of VMA and HVA in the cortex. TCWa also attenuated the anxiogenic effect of 5-hydroxytryptophan (5-HTP) and enhanced the anxiolytic effect of 9p-chlorophenylalanine (PCPA), buspirone (BUS) and ritanserin (RIT) in the EPM. From these results, TCWa at 0.1 and 0.5 g/kg possessed an anxiolytic effect T heanxiolytic mechanisms of TCWa might be due to decreased catecholaminergic activity caused by the increase in the turnover rate of catecholamines in the brain and decreased concentrations of 5-HT in the brain stemvia activating somatodendritic 5-HT1A autoreceptors and inhibiting postsynaptic 5-HT receptors.