Figure 1.

Weak correlation between locomotor activities and PER2::LUC bioluminescence rhythms highlights Bmal1-ELuc as a faithful reporter of intrinsic rhythms. A, Double plots of locomotor activities under constant darkness (top) and of bioluminescent reporter activities of cultured SCNs from the same animals (middle) and photos of corresponding slices (bottom) are shown. The Bmal1-ELuc reporter activity in culture appears to continue the locomotor rhythm (left), whereas PER2::LUC activity in culture deviates from the rhythmic pattern of the locomotor activity when the animal was alive (right). The double plot in culture represents normalized bioluminescence rhythms. The green lines are the guides marking the onsets of significant activity. The light red and light blue in the double plot denote the anterior and posterior SCN explants, respectively (bottom). B, The period of circadian locomotor activity under constant darkness (23.7 ± 0.1 h; n = 8) is statistically indistinguishable from that of Bmal1-ELuc rhythms in both anterior (23.5 ± 0.3 h; n = 39) and mid to posterior SCNs (23.9 ± 0.4 h; n = 43) (top); whereas the periods of locomotor (23.7 ± 0.1 h; n = 4) and PER2::LUC activities from both anterior (24.1 ± 0.2 h; n = 19) and mid to posterior (24.7 ± 0.1 h; n = 29) SCN cultures are significantly different (**p < 0.01; ***p < 0.001) (bottom). There was no significant difference in behavioral rhythm between wild-type and Bmal1-ELuc transgenic strains (p = 0.36; wild type, n = 6) or between wild-type and with PER2::LUC knock-in strains (p = 0.45). Statistical significance was determined by Welch's t test (see Materials and Methods). C, The peak-to-trough amplitude of Bmal1-ELuc bioluminescence is smaller in comparison with its respective baseline bioluminescence (top) than PER2::LUC (bottom). Note the difference in scales. Error bars indicate SEM. D, The SBR is higher in the posterior than the anterior SCN in both reporters, but Bmal1-ELuc is consistently low and ∼65% of PER2::LUC.