Feeding monitoring and meal pattern analysis.

The microstructure analysis of feeding behavior was conducted using the rat BioDAQ episodic food intake monitor (BioDAQ, Research Diets, New Brunswick, NJ), as detailed in our previous rat studies (22). DIO rats were placed in the BioDAQ system after 10 wk on the WD, and age-matched rats fed the ND were placed in the BioDAQ system as controls (Fig. 1A). DIO and control animals were acclimated for 2 wk to single housing in a cage containing an enrichment device (red polycarbonate tube) and to learn how to access the food from a front feeding hopper. Then basal 24-h food intake and meal pattern of WD rats were assessed, as well as at 6 wk after switching to the HPD (Fig. 1A). Water was provided ad libitum from a regular water bottle. Food intake and feeding behavior were monitored continuously second by second by the BioDAQ system, which weighs (±0.01 g) the hopper containing the food and algorithmically detects “not eating” as weight stable and “eating” as weight unstable. Feeding bouts (changes in stable weight before and after a bout) are recorded with a start time, duration, and amount consumed. Meals consisted of one or more bouts separated by an intermeal interval. The intermeal interval and minimum meal amount are user-definable. We defined the intermeal interval of 15 min and minimum meal amount of 0.1 g (22). Thus, food intake was considered as one meal when the feeding bouts occurred within 15 min of the previous feeding, and their sum was equal to or greater than 0.1 g. Bouts of feeding longer than 15 min apart were considered a new meal. Meal structures included the number of meals (meal frequency), meal size, meal duration, intermeal interval (time difference between the end of one meal and the initiation of the next meal), and total time spent eating. These parameters were calculated by the software provided by the manufacturer (BioDAQ Monitoring Software 2.3.02). The satiety ratio was determined as the intermeal interval divided by the meal size. Since rats are nocturnal animals, food intake and meal structures were analyzed separately in dark and light phases in addition to daily feeding measurements.