Figure 4.

Zebrafish larval locomotor activity assay. WT or HS-Hcrt transgenic fish are mated, embryos are collected, and individual larvae are placed in each of 80 wells of a 96-well plate on the fourth day of development. The plate is placed in a box that is continuously illuminated by infrared lights and is illuminated by white lights from 9:00 A.M. to 11:00 P.M. The larvae are monitored by an infrared camera, and the locomotor activity of each larva is recorded by a computer. Sample 40 s activity traces for a single larva during the day and at night are shown. Each movement of the larva is recorded as an upward deflection of the trace. When that deflection reaches a threshold (green), it is recorded as movement by the software. Zebrafish larvae move in short bursts and are mostly inactive at night. The small white deflections at night represent background noise. If necessary, larvae are genotyped by PCR after the experiment to identify HS-Hcrt larvae.