Update in the methodology of the chronic stress paradigm: internal control matters

Abstract

To date, the reliability of induction of a depressive-like state using chronic stress models is confronted by many methodological limitations. We believe that the modifications to the stress paradigm in mice proposed herein allow some of these limitations to be overcome. Here, we discuss a variant of the standard stress paradigm, which results in anhedonia. This anhedonic state was defined by a decrease in sucrose preference that was not exhibited by all animals. As such, we propose the use of non-anhedonic, stressed mice as an internal control in experimental mouse models of depression. The application of an internal control for the effects of stress, along with optimized behavioural testing, can enable the analysis of biological correlates of stress-induced anhedonia versus the consequences of stress alone in a chronic-stress depression model. This is illustrated, for instance, by distinct physiological and molecular profiles in anhedonic and non-anhedonic groups subjected to stress. These results argue for the use of a subgroup of individuals who are negative for the induction of a depressive phenotype during experimental paradigms of depression as an internal control, for more refined modeling of this disorder in animals.

Figure 1

Chronic stress results in decreased sucrose preference in a subgroup of mice. After initial increase in sucrose preference on the 3rd week of stress, by the termination of a 4-week stress, a cohort of mice display a prominent decrease in sucrose preference. Mice subjected to chronic stress were split into anhedonic and non-anhedonic subgroups according to the criterion of 65% preference for sucrose solution (see the text), Reproduction of this material is permitted by Macmillan Publishers Ltd.

Figure 2

Differential stress-induced changes in the sucrose test parameters in anhedonic and non-anhedonic mice. (A) Sucrose preference in the anhedonic group is significantly lower than in non-anhedonic and control mice after 2.5, 3.5 and 4 weeks of stress. (B) Sucrose intake in the anhedonic group is significantly increased after 2.5 weeks of stress and significantly decreased after 3.5 weeks of stress (vs. non-anhedonic group) and after 4 weeks of stress (vs. control and non-anhedonic group). Non-anhedonic mice show elevated sucrose intake after 2.5 and 3.5 weeks of stress. (C) Water intake is elevated in the anhedonic animals after 2.5 - 4 weeks of stress (vs. control and non-anhedonic group). In the non-anhedonic group; water intake is increased after 2.5 weeks of stress as compared to control. (D) Total liquid intake is elevated both in the anhedonic and in non-anhedonic animals after 2.5 and 3.5 weeks of stress (vs. control group). After 2.5 weeks, anhedonic mice show significantly higher total liquid intake than non-anhedonic mice. Parameters of the sucrose test are expressed as a percentage of the mean values of the control group, and compared between anhedonic (dashed line) and non-anhedonic (plain line) groups during a 4-week stress procedure as mean ± (SEM) (*p < 0.05 vs. control group; #p < 0.05 vs. non-anhedonic group; Mann-Whitney).

Figure 3

Dynamics of the sucrose test parameters recovery in anhedonic and non-anhedonic mice. Parameters of the sucrose test were expressed as a percentage of mean values of the control group, and compared between anhedonic (dashed line) and non-anhedonic (plain line) groups during a 4-week stress procedure (*p < 0.05 vs. control group; # p < 0.05 vs. non-anhedonic group; Mann-Whitney). (A) Sucrose preference in the anhedonic group is significantly lower than in the non-anhedonic and control mice throughout the entire experiment. (B) Sucrose intake in the anhedonic group is significantly decreased after the termination of stress (vs. control and non-anhedonic group) and during week 1 of the stress-free period (vs. control group). (C, D) Water consumption and total liquid intake are elevated in the anhedonic animals up to 3 weeks after termination of stress as compared to control, and up to 2 weeks when compared to non-anhedonic mice. Data are expressed as mean ± (SEM).

Figure 4

Anhedonic mice display lasting increases in home cage activity during the dark phase of the day. (A) During the dark phase of the diurnal cycle, mean time of horizontal movement of the anhedonic group (dashed line) is significantly elevated during 2nd-4th weeks of stress, and during weeks 1 and 2 after the termination of the stress procedure, as compared to the non-anhedonic (plain thick line) and control (plain thin line) groups (*p < 0.05 vs. control group and # p < 0.05 vs. non-anhedonic group; Mann-Whitney). This parameter does not change in the non-anhedonic group throughout the entire experiment (p > 0.05 vs. control group). (B) During the light phase of the day, no difference between the groups in home cage activity is observed (p > 0.05). Data are expressed as mean ± (SEM).

Figure 5

Effects of citalopram on sucrose test parameters and body weight in anhedonic and non-anhedonic mice. (A-E) Data are expressed as a percentage of the control and presented as medians ± interquartile intervals. Plain lines indicate non-treated groups, dashed lines indicate citalopram-treated groups. NoT: non-treated group; Cit: citalopram-treated group (*p < 0.05 vs. control group; § p < 0.05 vs. citalopram-treated anhedonic mice).