Automated video analysis system reveals distinct diurnal behaviors in C57BL/6 and C3H/HeN mice

Abstract

Advances in rodent behavior dissection using automated video recording and analysis allows detailed phenotyping. This study compared and contrasted 15 diurnal behaviors recorded continuously using an automated behavioral analysis system for a period of 14 days under a 14/10 light/dark cycle in single housed C3H/HeN (C3H) or C57BL/6 (C57) male mice. Diurnal behaviors, recorded with minimal experimental interference and analyzed using phenotypic array and temporal distribution analysis showed bimodal and unimodal profiles in the C57 and C3H mice, respectively. Phenotypic array analysis revealed distinct behavioral rhythms in Activity-Like Behaviors (i.e. walk, hang, jump, come down) (ALB), Exploration-Like Behaviors (i.e. dig, groom, rear up, sniff, stretch) (ELB), Ingestion-Like Behaviors (i.e. drink, eat) (ILB) and Resting-Like Behaviors (i.e. awake, remain low, rest, twitch) (RLB) of C3H and C57 mice. Temporal distribution analysis demonstrated that strain and time of day affects the magnitude and distribution of the spontaneous homecage behaviors. Wheel running activity, water and food measurements correlated with timing of homecage behaviors. Subcutaneous (3 mg/kg, sc) or oral (0.02 mg/ml, oral) melatonin treatments in C57 mice did not modify either the total 24 h magnitude or temporal distribution of homecage behaviors when compared with vehicle treatments. We conclude that C3H and C57 mice show different spontaneous activity and behavioral rhythms specifically during the night period which are not modulated by melatonin.

Fig. 1. Phenotypic Arrays of C3H and C57 Mice Homecage Behaviors over a 24 hour Period

Mean values for behavior magnitudes in the C3H (A) and C57 (B) mice were scaled and plotted over the 24 h period as function of color (0 to 4). The fold increase is indicated by the increase in the color red intensity from baseline represented by 1. The gray bar represents the light period (14h) while the dark bar represents the dark period (10h).

Fig. 2. Effects of C3H and C57 Mice Strain on the Temporal Distribution of Homecage Behaviors

The ordinates represent the temporal duration of each behavior over 24 h period expressed as seconds (s) per hour for behaviors included in the following classes: ALB (A–D), ELB (E–I), ILB (J–K) and RLB (L–O). The abscissae represent Zeitgeiber time (ZT) with ZT0 indicating Lights on. Shaded area on the graph indicates the dark period (10 h). The total 24 h duration of each behavior in each strain was compared using two-way ANOVA. The main effect of strain on total behavioral magnitude is indicated by a p value at the upper right corner of each graph. ns.: non-significant. * p<0.05 when compared the behavior magnitude in C3H with C57 mice at each ZT (Bonferoni post-test).

Fig. 3. Effects of Mice Strain on Wheel Running Activity, Ingestion and Weight Parameters

A. The ordinate represents C3H and C57 mice wheel running activity expressed as counts per minute and plotted as a function of Zeitgeiber time (ZT) (ZT0: Light onset). Shaded area indicates the dark period. The C3H and C57 mouse behavioral profiles were compared using two-way ANOVA (p < 0.01). * p<0.05 when comparing running wheel activity in C3H with C57 mice at each ZT (Bonferoni post-test). B. Columns represent total ingestion of water and food by C3H and C57 mice over a week-long period during recording of basal homecage behaviors. **p< 0.001, ****p < 0.0001 when compared with C3H (Student t-test). C. Body weight determined during basal spontaneous behaviors recording in the C3H and C57 mice, and on the last day of the seven day treatment with vehicle (VEH) (0.1 ethanol in water, oral) or melatonin (0.02 mg/ml in vehicle, oral) in C57 mice. (D) Columns represent total ingestion of water and food by C57 mice over a week period during recording of homecage behaviors of C57 mice treated with either oral vehicle or melatonin.

Fig. 4. Effects of Oral Melatonin Treatment on the Temporal Distribution of C57 Mice Behaviors in the Homecage

The ordinates represent the temporal profile of each behavior over a 24 h period expressed as seconds (s) per hour for behaviors included in the following classes: ALB (A–D), ELB (E–I), ILB (J–K) and RLB (L–O). The abscissae represent Zeitgeiber time (ZT) with ZT0 indicating Light on. Shaded area on the graph indicates the dark period (10 hrs.). C57 mice were treated with VEH (0.1% ethanol in water, oral) or melatonin (0.02 mg/ml in vehicle, oral). The effect of treatment on the total behavioral magnitude was compared using two-way ANOVA. ns.: non-significant. Shaded area on the graph indicates the dark period (10 h) (ZT0: Lights on). Data represents behavioral values for the last day of treatment (1 week). The profile for each day of treatment was identical.