Circadian Rhythm Monitoring in Tissue Explants

Within 1.5 h before lights out, tissue explants were prepared as previously described (25 (link)). In the first experiment (Figure S1 in Supplementary Material), SCN, arcuate complex, pituitary, liver, lung, aorta, spleen, and white adipose tissue were collected from each mouse and cultured as previously described (16 (link)). In the subsequent experiments in intact and ovariectomized mice, only liver explants were cultured from female mice. Bioluminescence was measured with the LumiCycle in 10-min intervals (Actimetrics Inc., Evanston, IL, USA). The data were detrended (by subtracting the 24 h moving average) and smoothed (0.5 h adjacent average) using LumiCycle software. Then ClockLab analysis software was used to determine the phase (peak of bioluminescence occurring between 12 h and 36 h in culture) of PER2:LUC expression.