Chronic Restraint Stress
Objective: To expose mice to chronic physical restraint stress for 21 days to induce stress-related behavioral and physiological changes
Protocol Steps
Animal housing and acclimation
Male mice aged 60-90 days were group housed in individually ventilated cages in a pathogen-free room with controlled temperature and light cycles
Note: Light cycle: 12 h cycles, lights on at 0700 h; water and standard regular chow provided ad libitum
View evidence from paper
“Littermates were group housed (Green Line IVC Sealsafe PLUS mouse, Tecniplast, Varese, Italy) in a temperature-specific (22C±1°C) and light-specific (12 h cycles, lights on at 0700 h) pathogen-free room with water and standard regular chow ad libitum”
Baseline behavioral testing
Animals underwent behavioral phenotyping prior to chronic restraint stress exposure. Tests performed in order from least to most invasive: rotarod, elevated plus maze, marble burying test, open field test, sucrose preference test, novelty suppressed feeding, and forced swim test
Note: Investigators were not blinded to group assignment; test order reversed after chronic restraint stress for bell-shaped stress exposure
View evidence from paper
“Behavioral phenotyping was performed between 0900 h and 1600 h. Animals were given 30 min of habituation to the behavioral testing room. Tests were performed from the least to the most invasive”
Chronic restraint stress exposure
After baseline behavioral testing, animals were placed in a horizontal resting position inside a well-ventilated 50 ml falcon tube for restraint stress
Note: Restraint tubes are well-ventilated with 12 holes of 0.5 mm diameter; restraint performed daily at 1000 h
View evidence from paper
“After baseline behavioral testing, animals were submitted to restraint stress for 21 days. Every day, mice were placed in a horizontal resting position inside a well-ventilated (12 holes, 0.5 mm diameter) 50 ml falcon tube at 1000 h and after 4-6 h they were unrestrained”