Dexamethasone (dex)

In Cohort 4 mice were treated with dexamethasone (0.1 mg/kg ip, Oradexon, Organon, Oss, Hollandia) during the early morning (at the time of the beginning of the dark phase or active period of the animals, with the highest ACTH-corticosterone level) [53 (link)]. Trunk blood was collected 6 h after ip injections in ice-cooled Eppendorf tubes. Serum was separated by centrifugation (3000 rpm for 10 min at 4 °C) and stored at −20 °C until assayed. The ACTH and corticosterone concentrations were determined by radioimmunoassay in duplicates as described earlier [54 (link)].

Each mouse was sedated with an intraperitoneal injection of chlorprothixene (Sigma-Aldrich, UK), at a dose of 0.5 mg/kg, to aid stress-free induction and reduce overall isoflurane concentration during the experimental procedure. After 10–15 min, the animal was transferred to the stereotaxic apparatus and positioned onto the incisor adaptor, facing the experimenter. Surgical anaesthesia was induced with 3–4% isoflurane in O₂, and maintained with 1–1.5% isoflurane in O₂ (Harvard Apparatus, UK) through a customized nose cone that provided minimal obstruction to visual stimulus presentation and stable recordings. Dexamethasone (2 mg/kg) (Organon, UK) and atropine (0.3 mg/kg, 20% in distilled water) (Animalcare, UK) were administered subcutaneously to improve breathing and reduce secretions respectively. Body temperature was monitored and maintained at 37±0.5°C by using a homeothermic heating device (Harvard Apparatus).