Circadian Activity Monitoring
Objective: Continuous monitoring of animal locomotor activity in 5-minute bins to analyze circadian behavioral rhythms and determine free-running periods
Protocol Steps
Animal acquisition and housing setup
Acquire male and female C57BL/6 mice aged 3-5 months weighing 25-30g from Charles River Laboratories. Breed AQP4 KO mice in University vivarium and backcross to C57BL/6 for 20+ generations. House mice in groups with equal numbers of males and females per experimental group.
Note: Minimum of five mice per group; maintain humidity at 30-70%
View evidence from paper
“Male and female C57BL/6 mice (aged 3–5 months, weight between 25 g and 30 g) were acquired from Charles River Laboratories”
Establish lighting conditions
House mice either in a 12:12 light/dark cycle or under constant light conditions. For constant light experiments, house animals two per cage to reduce risk of hypothermia.
Note: Constant light housing requires two mice per cage for thermoregulation
View evidence from paper
“Mice were group-housed either in a 12:12 light/dark cycle or under constant light with ad libitum access to food and water”
Initiate continuous activity monitoring
Place mice in the Comprehensive Lab Animal Monitoring System (Columbus Instruments) and begin continuous monitoring of locomotor activity.
Note: Activity is recorded in 5-minute bins
View evidence from paper
“Activity was monitored continuously in 5 min bins via the Comprehensive Lab Animal Monitoring System (Columbus Instruments)”
Collect activity data
Collect continuous activity data for at least 10 days to obtain sufficient data for circadian analysis and free-running period determination.
Note: Data collected in 5-minute bins
View evidence from paper
“Free-running period of each cage was determined using at least 10 days of activity”
Analyze circadian behavior with ActogramJ
Use ActogramJ software to complete circadian behavioral analysis of the collected activity data.
View evidence from paper
“Circadian behavioral analysis was completed with ActogramJ”
Determine free-running period
Apply chi-square (χ²) periodogram analysis to at least 10 days of activity data to confirm behavioral rhythmicity, estimate experimental times, and determine the free-running period of each cage in conjunction with activity onset.
Note: χ² periodogram used to confirm rhythmicity and estimate experimental timing
View evidence from paper
“Free-running period of each cage was determined using at least 10 days of activity and a χ 2 periodogram, which was used to confirm behavioral rhythmicity and estimate experimental times in conjunction with activity onset of the cage”