Abstract: Rationale: Comorbidity of depression and anxiety is one of the most common mental disorders in practice. Few experimental procedures, however, have focused on identifying mixed depression- and anxiety-like behaviors in animal models. Objectives: To develop a novel testing paradigm, the so-called Liang’s contextual-stress Box (briefly, Liang’s Box), for evaluating depression- and/or anxiety-like behaviors in mice. Methods: Liang’s Box consisted of a central area (CA) with three peripheral arms (PA). Mice were placed individually in the end of a peripheral arm facing to the central area and were allowed to travel around Liang's Box. During a test period, behavioral parameters were recorded and collected by a video-tracking system named SMART 3.0 (Panlab, Spain), including (1) Latency to CA, (2) Time spent in CA (CA-time), (3) Distance traveled in CA (CA-distance), (4) Distance traveled in PA (PA-distance), (5) Transition number between CA and PA, (6) Global activity, (7) CA-time/cm2, (8) PA-time/cm2, (9) CA-distance/cm2, and (10) PA-distance/cm2. In order to assess the reliability and validity of Liang’s Box and get baseline information, we undertook a series of experiment, including verifying consistency and stability in the various strains of mice (KM, ICR, and C57BL/6J mice), imposing high illumination, restraint and tail suspension stress, pre-treating with an anxiolytic (diazepam, buspirone) or antidepressant (desipramine, mianserin).
Results: (1) There were not remarkable differences in all the parameters among KM, ICR, and C57BL/6J mice; (2) High illumination, restraint and tail suspension significantly increased the latency to CA and decreased the CA-time, CA-distance and transition number. However, these three types of stresses evoked different effect on global activity. High illumination increased activity, but restraint stress had no significant influence. In contrast with high illumination, tail suspension stress markedly reduced activity in mice; (3) The anxiolytics diazepam (0.25mg•kg-1 and 0.5mg•kg-1, i.p.) and buspirone (0.5mg•kg-1 and 1.0mg•kg-1, i.p.) significantly decreased latency to CA and transition number, and dose-dependently increased the CA-time. However, they did not affect CA- or PA-distance, or global activity; (4) The antidepressants desipramine (10.0mg•kg-1 and 30.0mg•kg-1, i.p.) and mianserin (0.25mg•kg-1 and 0.5mg•kg-1, i.p.) significantly decreased latency to CA, and dose-dependently enhanced CA-time, CA-distance and transition number, while neither affected PA-distance or global activity. Conclusions: Our data suggest that Liang’s Box is a useful experimental instrument and method for screening and assessing potential antidepressants and/or anxiolytics. Moreover, depression- and anxiety-like behaviors induced by Liang’s Box in mice may be a new hybrid animal model to investigate the pathogenic mechanisms underlying the comorbidity of anxiety and depression.

Key words: Depression, Anxiety, Stress, Animal model, Comorbidity