Young (10 + 2 weeks; n=20) male Swiss albino mice weighing 25-30 g inbred and maintained in the animal house of Department of Zoology, Banaras Hindu University, India were used. Mice were housed 5 per cage in a temperature-, humidity-, and light-controlled (12:12-hr light/dark cycle) animal room, and provided with food and water ad libitum. Mice were divided in control and experimental groups (n=10 each). All animal experiments were carried out in accordance with the principles and procedures outlined in the guidelines of animal ethical committee of Banaras Hindu University.
2.2 CUMS procedure
The CUMS procedure employed was a modification of published literatures [5, 25, 26]. Experimental mice underwent CUMS exposure for 14 consecutive days as outlined in Fig. 1 and stressors were given in a random order to ensure unpredictability. Mice were subjected each day to at least one stressor that was randomly chosen from 9 different stressors. For restraint stress, mice were placed in clear plastic restraint bags with opening in one corner allowing free respiration but restricting any movement. The predator sense experiment was carried out by placing experimental mice with a Sprague-Dawley rat individually and care was taken to avoid any physical injury. Intermittent light exposure was given for 8-hr and consisted of successive light and dark intervals of 2-hr duration. For social isolation, mice were individually housed in separate cages overnight and regrouped the following morning. After each stressor, sufficient interval of at least 2-hr was given to rest, following which they were placed in clean cages with fresh bedding. To maximise the resemblance to a psychological stress, painful stressors or food and water deprivation were not included in the protocol. Control mice did not receive any kind of stressor and were housed in normal conditions.
Body weight was measured at the onset of CUMS procedure and subsequently at the interval of 7 days. After 2 weeks of CUMS exposure, mice were subjected to the following behavioural tests: elevated plus maze (EPM), open field test (OFT), forced swim test (FST) and sucrose preference test.
2.3 Behavioural Tests
Tests were performed from 15th day following CUMS paradigm and mice were placed in the testing room about 30 min before the assays. All tests were conducted between 8:00 and 12:00-hr and after each test mice were temporarily placed in a cage and eventually regrouped to their cage after every mouse was tested. At least 24-hr time interval was maintained between successive behavioural tests. Tests were recorded with a video camera for off-line analysis.
2.3.1 Elevated plus-maze
Twenty-four hours after the last stress, control and CUMS mice were subjected to elevated plus-maze test . The apparatus was elevated 50 cm from the ground and consisted of two open arms (30 x 5 cm, surrounded by 0.25 cm high wall) and two closed arms (30 x 5 cm, surrounded by opaque 15 cm high wall) emerging from a central...