Behavioral tests assessing neuropsychiatric phenotypes in adolescent mice reveal strain- and sex-specific effects

Abstract

In humans, infancy and adolescence are associated with major changes in synaptic functions and ongoing maturation of neural networks, which underlie the major behavioral changes during these periods. Among adult cases with neuropsychiatric disorders including autism spectrum disorder, schizophrenia, attention deficit hyperactivity, and bipolar disorders, 50% have developed behavioral symptoms and received a diagnosis before 15 years of age. However, most of the behavioral studies in mice modeling neuropsychiatric phenotypes are performed in adult animals, missing valuable phenotypic information related to the effect of synaptic maturation during development. Here, we explored which behavioral experiments assessing neuropsychiatric phenotypes can be performed during a specific window of development in adolescent male and female C57BL/6N, DBA/2, and FVB/N mice that are typically used as background strains for generating genetically-modified mouse models. The three wild-type strains were evaluated across anxiety, social behaviors, and cognitive functions in order to cover the main behavioral impairments that occur in neuropsychiatric disorders. During adolescence, the three strains displayed significant differences under certain behavioral paradigms. In addition, C57BL/6N and FVB/N, but not DBA/2 mice revealed some sex-related differences. Our results provide new insights into discrete behaviors during development and emphasize the crucial importance of the genetic background, sex, and experimental settings in the age-dependent regulation of different behaviors.

Figure 1

LABORAS test for C57BL/6N and FVB/N mice at P36 in which mouse movements were continuously monitored for 24 h. (a) High activity was observed in FVB/N mice as indicated by increased locomotion duration, traveled distance and average speed compared to C57BL/6N mice despite the similar maximum speed. (b) Increased repetitive rearing and climbing behaviors in FVB/N compared to C57BL/6N mice, but similar self-grooming counts. (c) Eating and drinking counts were similar between C57BL/6N and FVB/N mice. d) C57BL/6N and DBA/2 mice could not build nests at two different developmental stages. FVB/N mice were able to build simple nests at both developmental stages. (e) The burrowing test revealed a similar ability to burrow food pellets after 2 h in the three strains. However, the amount of burrowed food pellets was significantly increased in FVB/N mice compared to C57BL/6N and DBA/2 mice. Two-way ANOVA followed by Tukey post hoc test, *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001. A black rectangle indicates a significant difference between sexes within a strain (refer to Supplementary Table 1). Blue and red dots refer to males and females, respectively. Error bars indicate the standard error of the mean (SEM).

Figure 2

Anxiety experiments in C57BL/6N, DBA/2, and FVB/N mice. (a) The open field test revealed no difference in total distance in the new arena between C57BL/6N, DBA/2, and FVB/N mice. (b) No difference in latency, number of visits, or duration in the center of the arena between C57BL/6N, DBA/2, and FVB/N mice. (c) DBA/2 mice exhibited significantly higher latency to the light compartment compared to C57BL/6N and FVB/N mice and significantly lower duration compared to FVB/N in the dark/light compartment test. (d) DBA/2 mice showed significantly less visit ratio (open vs closed arms) compared to C57BL/6N and FVB/N mice, and significantly less duration ratio (open vs closed arms) compared to FVB/N. (e) Hole-board test revealed an increased count of head poke in FVB/N mice compared to C57BL/6N and DBA/2 mice. Two-way ANOVA followed by Tukey post hoc test, *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001. A black rectangle indicates a significant difference between sexes within a strain (refer to Supplementary Table 1). Blue and red dots refer to males and females, respectively. Error bars indicate the standard error of the mean (SEM).

Figure 3

sensory and social behaviors in C57BL/6N, DBA/2, and FVB/N mice. (a) C57BL/6N, DBA/2, and FVB/N mice showed a similar average duration in the cold plate test. (b) In the direct social interaction test, C57BL/6N mice showed a decreased latency to the first proximity with a same-sex littermate mouse compared to DBA/2 and increased proximity counts compared to DBA/2 and FVB/N. The cumulative duration percentage of proximity was similar in the three strains. Two-way ANOVA followed by Tukey post hoc test, *P ≤ 0.05, **P ≤ 0.01. Blue and red dots refer to males and females, respectively. Error bars indicate the standard error of the mean (SEM).

Figure 4

Cognitive function in C57BL/6N, DBA/2, and FVB/N mice. (a) In the puzzle box, FVB/N mice showed significantly higher latency to reach the goal in the first sawdust trial (trial 5) and in all trials of the plug (Trials 8, 9, and 10) compared to C57BL/6N mice. DBA/2 mice showed similar results to C57BL/6N mice in all trials except trial 7 showing a decreased latency. (b) In the fear conditioning test, FVB/N mice showed less duration of freezing in the acquisition and context memory trials compared to C57BL/6N and DBA/2 mice. In the cued memory trial, C57BL/6N mice showed a significant increase in the duration of freezing compared to DBA/2 and FVB/N mice. (c) In the active place avoidance test, FVB/N mice showed an increased number of electrical shocks compared to C57BL/6N and DBA/2 mice in all trials except the pre-training trial 1. FVB/N mice showed a significantly decreased latency to enter the shock area compared to C57BL/6N mice in trials 5–9. (d) C57BL/6N mice showed a significantly decreased number of theoretical electrical shocks and increased latency to enter the shock area compared to DBA/2 and FVB/N mice after 24 h. Two-way ANOVA followed by Tukey post hoc test, ($P ≤ 0.05, $$P ≤ 0.01, $$$P ≤ 0.001 FVB/N vs. C57BL/6N); (#P ≤ 0.05, ##P ≤ 0.01 DBA/2 vs. C57BL/6N); (&P ≤ 0.001 FVB/N vs. DBA/2). For (b) and (d), *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001. A black rectangle indicates a significant difference between sexes within a strain (refer to Supplementary Table 1). Blue and red dots refer to males and females, respectively. Error bars indicate the standard error of the mean (SEM).