Figure 2. Complete loss of circadian rhythmicity of Bmal1 brain knockout mice in constant darkness and constant light.
(A, B) Representative actograms of daily wheel-running activity of Cre, Fx/Fx, Het, and BKO mice. Activity records were double plotted, with each day being represented beneath and also to the right of the preceding day. Horizontal black and white bars at the top of each actogram represent lights off and on, respectively. Mice were housed in LD, released into DD for 4 weeks, returned to LD for 2 weeks, released into LL for 4 weeks, and then returned to LD. GLM ANOVA and Tukey–Kramer multiple comparison post-tests were used to compare genotypes tested. BKOs showed no significant periodicity in DD or LL. (C–E) Period (C), amplitude of circadian rhythm (D), and activity levels (E) in Cre mice (n = 28 for DD, 12 for LD, 12 for LL), Fx/Fx mice (n = 31 for DD, 18 for LD, 18 for LL), Het mice (n = 31 for DD, 13 for LD, 15 for LL), and BKO mice (n = 31 for DD, 16 for LD, 21 for LL). Bar graphs show mean ± SEM. (C) Free-running period was determined using χ2 periodogram for days 1–28 of DD or LL. A significant effect of genotype on period was found for both DD [F3,120 = 164,319, p < 0.0001] and LL [F3,65 = 20,429, p < 0.0001]. In DD, Cre mice were found to have a slightly longer period than the other two control groups (*p ≤ 0.05). In LL, Het mice were found to have a shorter period than the other two control groups (*p ≤ 0.05). Nevertheless, all three control groups showed a free-running period in both DD and LL similar to that reported for WT mice. (D) DD and LL amplitude of circadian rhythm represented by FFT in the circadian range. A significant effect of genotype on amplitude was found for both DD [F3,120 = 150, p < 0.0001] and LL [F3,65 = 23.72, p < 0.0001], with BKO mice having significantly lower amplitude (*p ≤ 0.05) in both DD and LL. (E) Total daily DD, LD, and LL activity levels, in wheel revolutions per 24 hr. BKO activity levels were found to be significantly higher than those of controls in DD [F3,119 = 9.00, p < 0.0001] (*p ≤ 0.05) and LL [F3,65 = 5.84, p = 0.0014] (*p ≤ 0.05) but not in LD.