Figure 4

Recognition of circadian phase difference between two groups of mice prevents the drawing of false conclusions. Light gray: fully-fed group; dark gray: calorie-restricted group. Two groups of C₃H mice (with differing breast cancer incidence) compared at single but different clock hours, first at near-weekly intervals (1, 2 and 3) and then at about 4- and again about 7-hour intervals (4 and 5) on the same day. The first 3 samplings at weekly intervals were made at earlier and earlier clock hours on two groups whose circadians were in antiphase, since one was fed a calorie-restricted diet in the morning, while the other group was fed ad libitum and fed mostly during the nightly dark span. To validate this assumption, the final two samplings at about 4- and then at about 7-hour intervals on the same day showed, as anticipated, the predicted reversal of the inter-group difference. (A progressive lowering of count associated with repeated blood letting had been demonstrated separately.) The time of day of sampling was the same for the two groups compared, but it differed from comparison to comparison in Figs. 1 2,3 (circled 1, 2 and 3); this fact confounded the results, as documented by repeating sampling at different clock-hours on the same day (circled 4 and 5). This circumstance accounts for the different results in Figs. 1,2,3: 24-h synchronized rhythms were compared on the same lighting but on different feeding regimens, as we realized and then documented the dominant synchronizing role of feeding time (overcoming the effect of lighting) on a diet restricted in carbohydrates and fat by 50% [86].