Clicker Training Mice for Improved Compliance in the Catwalk Test

Abstract

The CatWalk test relies on the run of mice across the platform to measure a constant speed with low variation. Mice usually require a stimulus to walk to the end of the catwalk. However, such stimuli are usually aversive and can impair welfare. Positive reinforcement training of laboratory animals is a thriving tool for refinement and contributes to meeting the demands instituted by Directive 2010/63/EU. We have already demonstrated the positive effects of clicker training. In this study, we trained male and female mice to complete the CatWalk protocol while assessing the effects of training on their well-being (Open Filed and Elevated Plus Maze). In the CatWalk test, we observed that clicker training improved the running speed of the mice. In addition, clicker training reduced the number of runs required by mice, which was more pronounced in males. Clicker training lowered anxiety-like behaviors in our mice, especially in females, where a significant difference was observed between trained and untrained ones. Based on our findings, we hypothesize that clicker training is an effective tool to motivate mice and increase performance on the CatWalk test without potentially impairing their welfare (e.g., by puffing them).

Keywords: 3Rs; CatWalk test; clicker training; refinement; welfare.

Figure 3

CatWalk XT performance of trained (gray) and untrained (white) C57BL/6J male and female mice. (a) Scheme of the CatWalk XT platform. (b) Running speed (cm/s). (c) Number of runs per group (#). We used two-way ANOVA followed by the Tukey HSD test for statistical analysis and multiple comparisons, n = 12. Each point represents an individual. Bars indicate the means ± SD. The exact p-value is provided when significant differences are given.

Figure 4

Behavioral activity of trained (gray) and untrained (white) C57BL/6J male and female mice in the open field (OF) test. (a) Schematic representation of the OF experiment. (b) Time the mice spent in the center (s). (c) Distance traveled in the center (cm). (d) Distance traveled in the center/distance traveled in total. Two-way ANOVA followed by the Tukey-HSD test was used for statistical analysis, n = 12 (untrained females n = 11). Each point represents an individual. Bars indicate the means ± SD. The exact p-value is provided when significant differences or trends are given.

Figure 5

Behavioral activity of trained (gray) and untrained (white) C57BL/6J male and female mice in the Elevated Plus Maze (EPM) test. (a) Schematic representation of the EPM experiment. (b) Time spent in open arms (s). (c) Distance traveled in the EPM (cm). (d) Running speed (cm/s). We used Two-way ANOVA followed by the Tukey HSD test for statistical analysis, n = 12 (trained males in 5b n = 11). Each point represents an individual. Bars indicate the means ± SD. The exact p-value is provided when significant differences are given. Two-way ANOVA analysis for open arm duration (b) indicated an effect of training, although the Tukey HSD test did not show differences.

Figure 1

Graphical illustration of the clicker training protocol.

Figure 2

Mouse performing the Catwalk test. The footprint of each paw is visualized in green color.