Figure 1.
Each experiment is represented by a rectangle proportional to the duration of the experiment. Video-recordings are indicated by a camera symbol and days on which recordings occurred are numbered. Feeding times are indicated in italics; for the 18 h interval feeding experiment there were three alternating sets of feeding times. For the first type, the first feeding event is when the video recording begins at ZT 8 and thus anticipation of this feeding event is not possible so we indicated that feeding event in parenthesis.
Figure 2.
12 h interval feeding schedule.
(A) Normalized median +/− IQR high activity data on day 14 of 2X feeding. The normalization is equivalent to dividing the number of frames during which high intensity activity (walking, hanging, jumping, or rearing) is observed per h by the number of frames the high intensity activity is observed during the duration of the 24 h video, yielding a fraction of high activity per hourly bin. Each ZT time represents the total number of s of high activity in the h preceding it (eg. The data point at ZT 9 comprises the amount of high activity that occurred from ZT 8:00-ZT 8:59. (B) The amount of normalized high activity in the 2 h preceding each feeding on day 14. (C) Normalized median +/− IQR high activity data on day 42 of 2X feeding. (D) The amount of normalized high activity in the 2 h preceding each feeding on day 42. Statistical significance was determined using the Mann-Whitney Test with asterisks denoting * = p<0.05, ** = p<0.01, *** = p<0.001. n = 8 for both 2X AL and 2X CR at all time points. Yellow arrows represent feeding times. Note: arrows are offset from feeding times to denote that the data point at the feeding time represents data from before the feeding event. For example, for a feeding at ZT21 the data point at ZT21 represents data from ZT20 to ZT21 and does not include time when food was present, as food was delivered at ZT21. Gray box indicates the 11h dark period. (E) Individual data from 2X CR mouse #1 on day 42, (F) for mouse #4, (G) mouse #5, and (H) mouse #8. Mean 2X AL high activity is sown as a blue line and gray bars indicate the amount of high activity occurring per one h bin for the CR mouse. Dark line indicates the 11 h dark period and arrows indicate feeding times.
Figure 3.
8 h interval feeding schedule.
(A) Normalized median +/− IQR high activity data on day 7 of 3X feeding. (B) The amount of normalized high activity in the 2 h preceding each feeding event on day 7. (C) Normalized high activity data on day 35 of 3X feeding. (D) The amount of normalized high activity in the 2h preceding each feeding event on day 42. For 3X AL data n-6 on day 7 and n = 8 on day 35; for 3X CR, n = 8 on day 7 and 35. (E–H) Individual mouse data for 8 h interval feeding. The s of high intensity activity in each h for individual CR mice on days 35 3X CR feeding for (E) mouse #2, (F) mouse #3, (G) mouse #6, and (H) mouse #8. Black arrows represent feeding times. Weighted black line on the x-axis indicates the 11h dark cycle.
Figure 4.
Four h interval feeding schedule.
(A) Normalized median +/− IQR high activity data on day 21 of 6X feeding. (B) Normalized high activity data on day 32. (C) Normalized high activity data on days 42–43 CR feeding. (C) Normalized high activity data on days 42–43 of 6X feeding. (D–F) The fraction of high activity occurring 1h prior to each meal time for days (D) 21, (E) 32, and (F) 42–43 of 6X feeding. Yellow arrows with a slash through them indicate false feeding events where a small plastic pellet was delivered instead of food. Dashed arrows indicate 4h intervals and do not represent feeding (or false-feeding) events. n = 12–14 mice at for each group at each time point presented. (G–J) Individual mouse data for 4 h interval feeding. The s of high intensity activity in each h for (G) mouse # 5, (H) mouse #8, (I) mouse #9, and (J) mouse #11 on day 32 of 6X CR. Black arrows represent feeding times. Weighted black line on the x-axis indicates the 11 h dark cycle.
Figure 5.
Body temperature and activity measurements during 4h interval feeding.
(A) Mean body temperature (left y-axis and top of the panel) and normalized median high activity data (right y-axis and bottom of the panel panel) plotted against ZT time for day -3 (CR and AL groups are listed separately but no dietary manipulation has occurred at this point). (B) Temperature and activity after 14 days of 4 h interval feeding. (C) Temperature and activity after 33 days of 4 h interval feeding. Normalized high activity for 1h preceding each feeding event on day 14 (D) and day 33 (E). Change in temperature 1 h preceding each feeding event on day 14 (F) and Day 33 (G). Fraction of high activity plotted against change in temperature 1h preceding daytime (H–J) and nighttime (L–N) feeding events for days -3, 14 and 33. Least-square regression lines and r2 values are shown on the top left of each panel. Error bars correspond to SEM for temperature values, and to IQ ranges for activity values. Statistics were performed using t-test for temperature values and Mann-Whitney for behavior values. * denotes p<0.05, ** denotes p<0.01. n = 7 for AL and CR. n = 7 for both AL and CR at all time points. Yellow arrows indicate feeding times.
Figure 6.
18 h interval feeding schedule.
(A) Normalized high activity median +/− IQR profiles during the 48 h recording during days 25–27 for 18 h interval feeding. (B) Normalized high activity profiles for days 37–39. (C) Normalized high activity profiles on days 45–47. Yellow arrows correspond to feeding events. (D) Fraction of high activity during the h preceding feeding events for days 25–27, 37−39, and 45–47. (E–J) Individual high activity mouse data with mean AL values indicated by a blue line for (E) mouse #2 on days 25–27, (F) mouse #3 on days 25–27, (G) mouse #1 on days 37–39, (H) mouse #6 on days 37–39, (I) mouse #3 on days 45–47, and (J) mouse #6 on days 45–47. All statistics were performed using Mann-Whitney * denotes p<0.05, ** denotes p<0.01. n = 6 for AL and n = 6 for CR at all time points.
Figure 7.
Short interval feeding schedule.
(A) Fraction of median +/− IQR normalized high activity for AL and CR mice on day 14 of LS CR, corresponding to the 21st day of 60% CR. (Day -7 is the first day of CR feeding, which occurred as a single feeding event at ZT 8, then the mice were switched to being fed 6 small meals at 30 min intervals beginning at ZT 4.5). (B) Fraction of high activity for AL and CR mice on day 17 (24 total days of 60% CR). (C) Days 21–22 of LS interval feeding (day 28–29 of CR). The yellow arrows indicate the 6 feeding times, whereas the 6 black dashed arrows indicate the expected time of feedings, which were omitted, on day 22 of LS feeding schedule. (D) Fraction of high activity in the two h preceding the first meal delivery for AL and CR mice on days 0, 14, 17, 21, and 22 of LS interval feeding. (E–F) Group median +/− IQR data for high activity shown in one min bins for (E) day 14 and (F) day 17. Min 1140 corresponds to ZT 3 and min 1140 corresponds to ZT 8. Blue line indicated median AL data. (G–I) Individual data for 30 min scheduled CR feeding; (G) s of high activity for CR mouse 901 from ZT3-8 on day 14 of LS interval feeding, (H) s of high activity for CR mouse 903 from ZT3-8 on day 17 of LS interval, and (I) s of high activity for CR mouse 905 from ZT3–8 on day 14 of LS interval feeding. n = 6 for both LS AL and LS CR at all time points.