2.3. Sleep disturbance

Sleep was disturbed by frequently arousing the rats during the light-phase for 12 h/day (0600–1800, as rats are nocturnal) on four random days/week to avoid circadian and sleep-pattern adaptations. This was achieved by replacing an old object with a new stimulatory object in their cage when the rat appeared drowsy. Stimulatory objects included plastic toys and tubes of different sizes, tube lids, marbles, and nesting material. When necessary, additional arousal was achieved by disturbing the bedding or introducing objects containing sucrose food reward pellets that could be sensed but not accessed by the animal. Cages were also exchanged twice/day (1200 and 1400) for fresh ones to provide additional stimulation. This method avoids the confounding impact of acute stress associated with current sleep restriction/disturbance methods and better reflects “real-world” poor sleep.