Sealsafe plus gm500

Twenty-four hours after the last extinction session, 2 groups of mice of each housing condition (2 social and 2 isolated groups) were exposed for 22 h to Environmental Enrichment (EE groups).
In the Exp. 1 (social condition), EE consisted in a rat two-level housing cage (each cage: 35.6 × 48.5 × 21.8 cm, Optirat Gen II, Animal Care Systems) where 3 or 4 mice from the same social home cage were housed with various objects (toys with different materials, shapes and colours, i.e. plastic balls and ladders, wood bricks), shelters and tunnels (Fig. 1A). In the Exp. 2 (isolated condition), EE consisted in a novel housing cage (12 × 17.5 × 35.5 cm, Sealsafe Plus GM 500, Tecniplast), where mice from isolation condition were single housed with various objects (toys with different materials, shapes and colours, i.e. plastic ball and ladder, wood brick) and tunnel (Fig. 1B). In both experiments, control NoEE groups were kept in their home cages. In the Exp. 1, home cage bias was controlled counterbalancing the subdivision of littermates in the two environmental manipulations, with half of the mice of each home cage assigned to EE and the other half assigned to NoEE group.

Female BALB/c mice (N = 212) were purchased from Charles River (strain code 028). The mice used in these experiments were 6–10 weeks of age and weighed 15–22 g. All mice received irradiated food (PicoLab Rodent Diet 5053, LabDiet) and water ad libitum. All mice were housed in groups (n = 5) in sanitized solid-bottom individually ventilated cages (Sealsafe Plus GM500, Techniplast) on sterile paper bedding (Alpha-dri, Shepherd Specialty Paper) and were provided with environmental enrichment including a plastic Mouse Igloo with saucer wheel (BioServ) and a single Nestlet square (Ancare). Animals were maintained under a 12:12 hour light/dark cycle, with 10–15 air changes per hour, relative humidity of 30–70%, and room temperature between 68–79ºF (20–26.1ºC). From entry into the facility, all mice were housed under Animal Biosafety Level 2 (ABSL-2) conditions, where they acclimated to the facility for at least 5 days. Animals were identified using cage cards and tail marks with a permanent marker. Prior to the start of the study, all animals were monitored twice daily during routine animal health rounds.