2.11. Chronic restraint stress model

The chronic restraint stress (CRS) model was established following the procedures as previously described⁴⁰. A total of 84 mice were randomly assigned to 7 groups: sham control, CRS model with vehicle control, XE991 (1 mg/kg, s.c.), fluoxetine (20 mg/kg, i.g.) and three dosages of D01 (10, 20, and 40 mg/kg, i.g.) with each group consisting of 12 mice.

Compounds in a volume of 10 mL/kg were administered once a day for 21 days before the initiation of restraint stress. Mice under chronic stress were restrained for 6 h per day in the tunnel of a 50 mL conical transparent plastic centrifuge tube with multiple ostioles in tube walls for ventilation. During the restraint period, the mice were kept in individual cages with no access to food or water.