Comprehensive lab animal monitoring system clams

At week 5 of treatment, basal metabolic parameters of mice were measured using the Comprehensive Lab Animal Monitoring System (CLAMS, Columbus Instruments, Columbus, OH). The mice were adapted to the metabolic cage for 24 h, and food intake, oxygen consumption (VO₂), and carbon dioxide release (VCO₂) were measured over the next 48 h. The number of times the infrared beams were broken along the X- or Y-axis was used to measure the dynamic activity of mice. The respiratory exchange rate (RER) was calculated using classical Lusk theory and equations RER = VCO₂/VO₂.

Indirect open circuit calorimeter was performed using the Comprehensive Lab Animal Monitoring System (CLAMS; Columbus Instruments, Columbus, OH). In each experiment, thirty two mice were weighed before testing. Each mouse was placed in its own chamber without bedding and allowed to acclimate to its surroundings; mice were not pre-acclimated to the CLAMS cages prior to the experiment. All experiments began at 9–10 am and continued for up to 48 hrs. Phase 1 of the study assessed metabolic preference during a non-feeding state; food was removed from the cages (10AM–4PM). Phase 2 evaluated metabolic preference during a feeding condition; food in excess was provided at 4 PM. The system measured oxygen consumption, carbon dioxide production and calculated the Respiratory Exchange Ratio (RER = VCO2/VO2) for each animal. Energy expenditure was also estimated using the formulas:
$CaloricValue\left(CV\right)=3.815+\left(1.232*RER\right)EnergyExpenditure\left(EE\right)=CV*VO2$
Activity was determined using an array of infrared beams (2.5 cm inter-beam distance) surround each cage. Total activity was monitored continuously and any movement that produced a beam break was counted and summed.

Metabolic parameters were measured by indirect calorimetry. Mice were single housed in open-circuit OxyMax chambers as part of the Comprehensive Lab Animal Monitoring System (CLAMS) (Columbus Instruments) and analyzed using OxyMax software 2.4.2. For exercise tests, using previously described methods^{37 (link)}, mice were acclimated and then subjected to an endurance or graded maximal exercise test^{37 (link)}. In brief, mice were placed on the treadmill at a 0° incline, and the shock grid was activated. The treadmill speed (meters), duration (minutes), and grade (degrees) were then increased until exhaustion as follows: 0 m/min, 3 min, 0°; 6 m/min, 2 min, 0°; 9 m/min, 2 min, 5°; 12 m/min, 2 min, 10°; 15 m/min, 2 min, 15°; 18, 21, 23, 24 m/min, 1 min, 15°; and +1 m/min, each minute thereafter. Exhaustion (endpoint for treadmill cessation) was defined as the point at which mice maintained continuous contact with the shock grid for 5 s. VO_2max was determined by the peak oxygen consumption reached during the test when the respiratory exchange ratio (RER) was greater than 1.0. The maximum running speed was defined as the treadmill speed at which VO_2max was achieved^{37 (link)}.

Mice were placed within the Columbus Instruments Comprehensive Lab Animal Monitoring System (CLAMS, Columbus Instruments, Columbus, OH). The system features 12 cages placed within an enclosed environment-controlled cabinet. Each cage provides real-time monitoring of horizontal and vertical activity, wheel activity, feeding and drinking, oxygen consumption, and CO₂ production every 13 min. These measurements were recorded over a 3-day period. In the first 48 h, the animals were kept at room temperature, followed by a period in which the temperature of the environment-controlled cabinet descended to 4°C (∼2 h) and then held at 4°C for the remaining duration of the study. CLAMS data were placed into the CalR application (22 (link)) where the data were compiled into hourly measurements for further analyses.