Effects of Various Cleaning Agents on the Performance of Mice in Behavioral Assays of Anxiety

Abstract

Cleaning behavioral equipment between rodent subjects is important to prevent disease transmission and reduce odor cues from previous subjects. However, the reporting regarding the cleansing procedures used during such experiments is sporadic and often incomplete. In addition, some investigators are reluctant to clean devices between subjects because they are concerned that animals will react negatively to the smell of the cleansing agents. We hypothesized that mice tested on an elevated plus maze (EPM) soiled with excretions from conspecifics would test as being more stressed than mice tested on the same apparatus that was cleaned between animals. We tested the performance of C57BL/6J mice on an EPM sanitized with 3 common cleaning agents-isopropyl alcohol, chlorine dioxide, and bleach-and on an EPM soiled with rodent urine, feces, and presumably pheromones. We further tested the potentially aversive nature of the cleansing agents by using the classic light:dark box and a 2-choice light:dark box. Our data indicate that cleaning the EPM compared with leaving it soiled did not affect performance in male or female C57 mice, nor did cleaning agent choice. In addition, test subjects did not react to the presence of the cleaning agents when incorporated into the classic light:dark test. However, in the 2-choice light:dark test, mice given the option to avoid an area containing a cleaning agent showed aversion to all 3 agents, when all other conditions were equal. Given the lack of an observable effect of cleaning on EPM performance, we recommend cleaning of the EPM device between C57 mice to minimize the potential spread of disease.

Figure 1.

Mean total distance (in meters) traveled in the EPM across sex and cleaning agent during 5-min trials. Initially each group contained 20 mice, but data from one mouse were lost. Error bars, SEM.

Figure 2.

Mean ratio of entries into the open arm divided by the number of entries into the closed arm in the EPM across sex and cleaning agent. Initially each group contained 20 mice, but data from one mouse were lost. Error bars, SEM.

Figure 3.

Mean ratio of time spent in the open arm divided by the time spent in the closed arm in the EPM across sex and cleaning agent. Initially each group contained 20 mice, but data from one mouse were lost. Error bars, SEM.

Figure 4.

Mean ratio of time spent in the light box divided by the time spent in the dark box during the 2-min trial. The dark box contained one of the cleaning agents. Each group contained 8 mice; error bars, SEM.

Figure 5.

In the 2-choice procedure, 2 dark boxes were attached to a central light box. In the control condition for each agent, water was placed in both dark boxes to establish a side preference in a 2-min trial. Data for the control condition for each agent represent the time spent in the nonpreferred dark box divided by the time spent in the preferred dark box (that is, the value will be less than 1). On the next trial for each mouse, the cleaning agent was placed in the preferred dark box and water in the less-preferred dark box, and a 2-min trial followed. Data for each agent are time spent in the dark box with water divided by the time in the dark box with the cleanser; therefore, values greater than 1 indicate the mouse preferred the dark box with the water more than the dark box with the cleanser. Each group contained 8 mice; error bars, SEM; *, significant (P < 0.05) difference between control (water) and odor trials.