Exercise training device and protocols

When constructing the exercise device, we took advantage of the flies’ natural negative geotaxis behavior to induce upward walking [26]. All exercise group flies started exercise from when they were 2 days old, and underwent a 5 week-long exercise program; we had observed that nearly all HFD group flies totally lost their negative geotaxis behavior by the age of 5 weeks. Vials, with the diet housing 25 flies each, were loaded horizontally into a steel tube that was rotated about its horizontal axis by an electric motor, with a gear regulating its shaft speed. NF/E group and HFD/E group severally had 24 vials training. Thus, with the accompanying rotating steel tube, each vial was rotated along its long axis, which made the flies climb. Most flies continued to respond by climbing throughout the exercise period. The few that failed to climb were actively walking at the inner wall of the vial [17, 27]. Flies were exercised in vials with a 2.8-cm inner diameter, rotated at 0.18 rev/s, 0.20 rev/s, 0.18 rev/s, 0.16 rev/s, and 0.14 rev/s. Flies were exercised for one and a half hours. Flies were exercised for 5 days, followed by a 2-day rest period, for a five-week period.