iDREADDs activation
Each behavioural condition was run both after an infusion of clozapine and after an infusion of saline, which served as a within-subject control. Clozapine was infused on two separate occasions per condition, reflecting the potential for greater within-subject variability. The infusion order was counterbalanced between the two cohorts (Figure 1). There was always an added infusion-free test day between the two clozapine infusions for Conditions 1-4, i.e., apart from the Dark condition.
Animals were first habituated to the infusion procedure, using saline, prior to the commencement of behavioural testing with clozapine. On infusion days, the animals were taken to a separate room in pairs and lightly anaesthetized with an isoflurane-oxygen mixture (5% induction, 1.5-2% maintenance). Double infusion injectors (33-gauge, PlasticsOne, Virginia, USA) were inserted into the guide cannula and either 1µl of sterile saline or 1µl of clozapine (1mg/ml) was infused over 1 minute using an infusion pump (11 Plus, Harvard Apparatus, UK). The injector was held in place for a further minute and the dummy cannula replaced. The infusions lasted no more than 4 minutes per animal, and the animal was returned to its home cage. Animals rested for 15-20 minutes prior to behavioural testing.