Longitudinal home-cage automated assessment of climbing behavior shows sexual dimorphism and aging-related decrease in C57BL/6J healthy mice and allows early detection of motor impairment in the N171-82Q mouse model of Huntington's disease

Abstract

Monitoring the activity of mice within their home cage is proving to be a powerful tool for revealing subtle and early-onset phenotypes in mouse models. Video-tracking, in particular, lends itself to automated machine-learning technologies that have the potential to improve the manual annotations carried out by humans. This type of recording and analysis is particularly powerful in objective phenotyping, monitoring behaviors with no experimenter intervention. Automated home-cage testing allows the recording of non-evoked voluntary behaviors, which do not require any contact with the animal or exposure to specialist equipment. By avoiding stress deriving from handling, this approach, on the one hand, increases the welfare of experimental animals and, on the other hand, increases the reliability of results excluding confounding effects of stress on behavior. In this study, we show that the monitoring of climbing on the wire cage lid of a standard individually ventilated cage (IVC) yields reproducible data reflecting complex phenotypes of individual mouse inbred strains and of a widely used model of neurodegeneration, the N171-82Q mouse model of Huntington's disease (HD). Measurements in the home-cage environment allowed for the collection of comprehensive motor activity data, which revealed sexual dimorphism, daily biphasic changes, and aging-related decrease in healthy C57BL/6J mice. Furthermore, home-cage recording of climbing allowed early detection of motor impairment in the N171-82Q HD mouse model. Integrating cage-floor activity with cage-lid activity (climbing) has the potential to greatly enhance the characterization of mouse strains, detecting early and subtle signs of disease and increasing reproducibility in preclinical studies.

Keywords: Huntington’s disease (HD); automated; digital biomarkers; motor function; neurodegeneration; reproducible; welfare.

Figure 1

Distance travelled by mice during the dark phase varies according to sex and across age during passive home-cage monitoring. (A) Distance travelled (mm) over zeitgeber time in female and male cages of 3-month-old mice during the recording session, binned into 6-min time bins and averaged over a 24-h period. The line represents the mean distance over time across cages of a sex group; the shaded error band represents a 95% confidence interval. Data from individual mice within a cage were summed to produce one time series per cage. The gray shaded areas on the plot represent darkness. (B,C) Same as (A) but for 7-month-old and 12-month-old mice, respectively. (D) Distance travelled over zeitgeber time in female cages of mice of all ages. The line represents the mean distance over time across cages of a sex and age group; the error-shaded area represents 95% confidence interval. (E) Same as (D) but for male cages. (F) Boxplot of mean distance travelled during light phase with cages split by age and by sex. Distance travelled within the light phase was averaged across time per cage and per day. Three data points per cage (3 days of recording) were modelled using a linear mixed-effects model to account for repeated-measures and least-squares means estimated to return adjusted p values of levels of factor combinations. (G) Same as (D) but for the dark phase. ns p > 0.05, **p < 0.05, ****p < 0.0001.

Figure 2

Time spent climbing by mice during dark phase varies according to sex and across age during passive home-cage monitoring. (A) Time spent climbing over zeitgeber time in female and male cages of 3-month-old mice during recording session, binned into 6-min time bins and averaged over a 24-h period. Line represents mean time spent climbing over time across cages of a sex group; the shaded error band represents a 95% confidence interval. Data from individual mice within a cage were summed to produce one time series per cage. The gray shaded areas on the plot represent darkness. (B,C) Same as (A) but for 7-month-old and 12-month-old mice, respectively. (D) The time spent climbing over zeitgeber time in female cages of mice of all ages. The line represents the mean distance over time across cages of a sex and age group; the error-shaded area represents a 95% confidence interval. (E) Same as (D) but for male cages. (F) Boxplot of mean time spent climbing moved during light phase with cages split by age and by sex. Time spent climbing within the light phase was averaged across time per cage and per day. Three data points per cage (3 days of recording) were modelled using a linear mixed-effects model to account for repeated-measures and least-squares means estimated to return adjusted p values of levels of factor combinations. (G) Same as (D) but for the dark phase. ns p > 0.05, ****p < 0.0001.

Figure 3

Distance travelled and climbing activity varies according to genotype in female mice and across age at the conclusion of the dark phase, but not at beginning of the dark phase. (A) Distance travelled over zeitgeber time in female-only cages of 8-week-old mice, split according to genotype, during the recording session, binned into 6-min time bins and averaged over a 24-h period. The line represents the mean distance over time across cages of a genotype group, the shaded error band represents a 95% confidence interval. Data from individual mice within a cage was summed to produce one time-series per cage. The gray shaded areas on the plot represent darkness. (B,C) Same as (A) but for 13-week-old and 15–16-week-old mice, respectively. (D) Boxplot of mean distance travelled during first 30 min of darkness within female-only cages split by age and genotype. Distance travelled was averaged across the first 30 min of darkness per cage and per day. Three data points per cage (3 days of recording) were modelled using a linear mixed-effects model to account for repeated-measures and least-squares means estimated to return adjusted p values of levels of factor combinations. (E) Same as (D) but for last 30 min of darkness. (F) Time spent climbing over zeitgeber time in female-only cages of 8-week-old mice, split according to genotype, during the recording session, binned into 6-min time bins and averaged over a 24-h period. The line represents the mean time spent climbing over time across cages of a genotype group, the shaded error band represents a 95% confidence interval. Data from individual mice within a cage was summed to produce one time-series per cage. The gray shaded areas on the plot represent darkness. (G,H) Same as (F) but for 13-week-old and 15–16-week-old mice, respectively. (I) Boxplot of mean time spent climbing during first 30 min of darkness within female-only cages split by age and genotype. Time spent climbing was averaged across the first 30 min of darkness per cage and per day. Three data points per cage (3 days of recording) were modelled using a linear mixed-effects model to account for repeated-measures and least-squares means estimated to return adjusted p values of levels of factor combinations. (J) Same as (I) but for last 30 min of darkness. ns p > 0.05, **p < 0.01, ****p < 0.0001.

Figure 4

Distance travelled and climbing activity varies according to genotype in male mice and across age at conclusion of dark phase, but not at beginning of dark phase. (A) Distance travelled over zeitgeber time in male-only cages of 8-week-old mice, split according to genotype, during the recording session, binned into 6-min time bins and averaged over a 24-h period. Line represents mean distance over time across cages of a genotype group, the shaded error band represents a 95% confidence interval. Data from individual mice within a cage was summed to produce one time-series per cage. The gray shaded areas on the plot represent darkness. (B,C) Same as (A) but for 13-week-old and 15–16-week-old mice, respectively. (D) Boxplot of mean distance travelled during first 30 min of darkness within male-only cages split by age and genotype. Distance travelled was averaged across the first 30 min of darkness per cage and per day. Three data points per cage (3 days of recording) were modelled using a linear mixed-effects model to account for repeated-measures and least-squares means estimated to return adjusted p values of levels of factor combinations. (E) Same as (D) but for last 30 min of darkness. (F) Time spent climbing over zeitgeber time in male-only cages of 8-week-old mice, split according to genotype, during the recording session, binned into 6-min time bins and averaged over a 24-h period. The line represents the mean time spent climbing over time across cages of a genotype group, the shaded error band represents a 95% confidence interval. Data from individual mice within a cage was summed to produce one time-series per cage. The gray shaded areas on the plot represent darkness. (G,H) Same as (F) but for 13-week-old and 15–16-week-old mice, respectively. (I) Boxplot of mean time spent climbing during first 30 min of darkness within male-only cages split by age and genotype. Time spent climbing was averaged across the first 30 min of darkness per cage and per day. Three data points per cage (3 days of recording) were modelled using a linear mixed-effects model to account for repeated-measures and least-squares means estimated to return adjusted p values of levels of factor combinations. (J) Same as (I) but for last 30 min of darkness. ns p > 0.05, *p < 0.05, ***p < 0.001, ****p < 0.0001.