The aim of this study was to identify the effects of 85% methanolic extract of Morus alba leaves (EMA), which is a traditional herb, in mice. The effects of EMA on the anxiolytic-like behaviour were studied using the elevated plus maze (EPM) and hole-board test. To elucidate the mode of action of the anxiolytic-like effects of EMA, the mice were subjected to the co-administration of EMA (200 mg/kg, per os (p.o.)) and either antagonist. EMA (at 200 or 400 mg/kg) significantly increased the percentages of time-spent in the open arms and entries into the open arms of the EPM versus vehicle-treated control group (p<0.05). Moreover, in the hole-board test, EMA (200 and 400 mg/kg) significantly increased the number of head-dips versus vehicle-treated control group (p<0.05). However, there were no changes in the locomotor activity and myorelaxant effects in any group compared with the vehicle-treated control group. In addition, the anxiolytic-like effects of EMA were abolished by thioperamide (10 mg/kg, intraperitoneally (i.p.)), which is a histamine H3 receptor antagonist. Moreover, results from reverse transcription polymerase chain reaction (RT-PCR) also revealed that the amygdalal histidine decarboxylase mRNA expression levels in EMA (200 mg/kg)-treated group were significantly higher than those in the vehicle-treated controls (p<0.05). These results suggest that EMA might prove to be an effective anxiolytic agent and that EMA acts via the histaminergic system in central nerve system.