Medetomidine

The IOP was measured after sedating the animals with inhalation anesthesia using 2% isoflurane in oxygen (ISOFLO Isoflurane 100% p/p, Zoetis SL, Alcobendas, Madrid, Spain). OHT was induced under general anesthesia using an intraperitoneal injection of a mixture of ketamine (75 mg/kg; Anesketin^®, Dechra Veterinary Products SLU, Barcelona, Spain) and medetomidine (0.26 mg/kg; Medetor^®, Virbac España S.A., Barcelona, Spain). To prevent corneal desiccation and infection after the laser procedure, tobramycin (Tobrex^®; Alcon, Barcelona, Spain) ointment was used. After OHT induction, all the appropriate procedures were adopted to mitigate the pain and discomfort of the animals. The animals were finally sacrificed after the intraperitoneal injection of an overdose of pentobarbital (Dolethal Vetoquinol^®, Especialidades Veterinarias, Alcobendas, Madrid, Spain).

All experiments were performed in adult (3–4 months old) wild-type BDNF^Val/Val and BDNF^Met/Met littermate mice, generated by Chen Z-Y et al. [31 (link)]. Only male mice were used to avoid the potential impact of hormones involved in the ovarian cycle in adipose tissue present in female mice. All experiments were approved by the National Ministry of Health-University of Milan Committee and of DGSA (12/2015 and 349/2015). Surgical procedures were performed in mice anesthetized with ketamine chlorhydrate (75 mg/kg; Intervet, Segrate, Milan, Italy) and medetomidine (1 mg/kg; Virbac, Milan, Italy). Mice were housed under standard conditions (20–22 °C, 12 h light/dark cycle, light on at 7 a.m.) with water and food ad libitum. All efforts were made to minimize animal distress and to reduce the numbers of animals used in this study.