Fig 1.
Schematic representation of the experimental procedure used to test the effects of housing condition on predator exposure induced anxiety.
After housed in a standard condition (SC; for 7 days), mice were randomly separated to short-term at standard condition (STSC) or long-term at standard condition (LTSC) or enriched condition (EC) and kept in the animal facility until to complete all behavior.
Fig 2.
Effect of environmental enrichment on the behavioral responses of mice that were exposed to the predator.
The exposure chamber is an area for mice to explore and investigate about the predator but the mesh partition prevents interactions. The behavioral activities of mice during exposure to a rat were shown. The columns in the graph represent duration of mice activity in respective region during predator exposure session. Data were shown as mean ± SEM, * indicates significant difference (*P < 0.05; **P < 0.01; ***P < 0.001). The letters indicates groups were compared for statistical analysis (a: STSC verses LTSC; b LTSC verses EC; c: STSC verses EC). Raw data is provided in S1 Data.
Table 1.
Elevated plus-maze test profile of male field mouse Mus booduga.
Fig 3.
Behavioural profile of mice in light/dark-box test after housed in environmental enrichment exposed to predator.
(a) EC made more entries to light chamber after seven hours and seven days of predator exposure and exert anxiolytic effect compared to STSC and LTSC mice. (b) EC mice spent more time in light zone compared to STSC and LTSC mice. (c) EC mice exhibit few risk assessment behaviour than other groups of mice. Data are presented as means ± SEM, * indicates significant difference (***P < 0.001; **P < 0.01; *P < 0.05), respect to comparison between groups a = STSC versus LTSC: b = LTSC verses EC: c = STSC verses EC. Raw data is provided in S1 Data.
Fig 4.
Effect of environmental enrichment on activation of SERT and 5-HT level in mice exposed to predator.
(a) Representative western blots shows the level of SERT (70 kDa) and β-actin (43 kDa). (b) There was a significant increase in 5-HT level after exposure to predator in STSC, LTSC and EC mice. (C) SERT protein level was significantly increased in EC following predator exposure than STSC and LTSC mice. Data were shown as mean ± SEM, * indicates significant difference (*P < 0.05; **P < 0.01; ***P < 0.001), respect to comparison between groups (a = STSC verses LTSC; b = LTSC verses EC; c = STSC verses EC). Raw data is provided in S2 Data.
Fig 5.
Effect of environmental enrichment on activation of 5-HT1A and 5-HT2C receptors in mice exposed to predator.
(a) 5-HT1A receptor mRNA level significantly increased in STSC and EC mice, an effect were not noticed in LTSC mice. (b) 5-HT2C receptor mRNA level was significantly altered in STSC, EC mice but it was significantly high in LTSC mice. Data were shown as mean ± SEM, * indicates significant difference (*P < 0.05; **P < 0.01; ***P < 0.001), respect to comparison between groups (a = STSC verses LTSC; b = LTSC verses EC; c = STSC verses EC). Raw data is provided in S2 Data.
Fig 6.
Effect environmental enrichment on activation of CaMKII/CREB in mice exposed to predator.
(a) Representative western blots showing the activation (measured as phosphorylated level) of CaMKII (50 kDa) and CREB (43 kDa) after exposed to predator. Phosporylation of (b) CaMKII, (c) CREB increased after predator exposure in STSC, LTSC and EC mice The fold variation shown as mean ± SEM. * Indicates significant difference (*P < 0.05; **P < 0.01; ***P < 0.001) respect to comparison between groups (a = STSC verses LTSC; b = LTSC verses EC; c = STSC verses EC). Raw data is provided in S2 Data.
Fig 7.
Effect of environmental enrichment on chromatin modifications (alternation in histone acetylation and deacetylation) in mice exposed to predator.
(a) Representative western blots shows the levels of H3 (17 kDa), H4 (11 kDa), HDAC-1(60 kDa) and HDAC-2(60 kDa) and β-actin (43 kDa). (b) H3 (c) H4 acetylation was significantly increased in STSC and EC mice but not in LTSC mice. While, histone deactylase (d) HDAC1 (e) HDAC2 significantly altered in STSC and LTSC mice but not in EC mice. The fold variation shown as mean ± SEM. * Indicates significant difference (*P < 0.05; **P < 0.01; ***P < 0.001) respect to comparison between groups (a = STSC verses LTSC; b = LTSC verses EC; c = STSC verses EC). Raw data is provided in S2 Data.
Fig 8.
Effect of environmental enrichment on anxiolytic/anxiety related markers in neuropeptide—Y (NPY) ergic system and BDNF in mice exposed to predator.
Real-time qPCR showing increase in (a) NPY and (b) NPY Y1 receptor mRNA level in STSC, EC mice but not in LTSC mice. Whereas, NPY Y2 receptor (c) mRNA increased only in LTSC mice but not in STSC and EC mice. (e) BDNF (exon IV) mRNA level increased in STSC and EC mice but not in LTSC mice. The fold variation shown as mean ± SEM. * Indicates significant difference (*P < 0.05; **P < 0.01; ***P < 0.001) respect to comparison between groups (a = STSC verses LTSC; b = LTSC verses EC; c = STSC verses EC). Raw data is provided in S2 Data.