A Multiple Integrated Social Stress Model for Psychiatric Disorders in Female C57BL/6J Mice

This research aims to understand how chronic social stress leads to psychiatric symptoms in female mice by establishing a multi-integrated social stress model. Most currently available models for psychiatric disorders have been established using male animals. This protocol introduces a new mouse model designed to study these disorders in female mice. This study has developed a reliable animal model that allows us to investigate the neurophysiological mechanisms underlying psychiatric disorders in female mice.

[Narrator] To begin, place a 10 centimeter long, 26 millimeter internal diameter acrylic tube in each cage of female subject mice and winner candidates to allow them to acclimate and learn to pass through it over three days. To train mice for tube competition, gently grasp the tail of each mouse and place it on the table. Then, hold the mouse by the tail and place it at one end of the tube. Release the tail once the mouse enters the tube and allow it to pass through voluntarily. Now, place the mouse at the opposite end of the tube and repeat the same procedure. After repeating the entire process once more, return the mouse to its home cage. For winner screening, prepare a 30 centimeter long, 26 millimeter internal diameter acrylic tube and draw a visible middle line on the tube. Hold both winner candidate mice by their tails and place them at opposite ends of the tube. Allow them to enter and meet at the midpoint, then simultaneously release them and start the timer. Identify the winner as the mouse that pushes the other out of the tube and award it one point. Return both mice to their home cage. Rank the mice based on their total scores. For CD1 aggressor screening, place a C57 male mouse directly into the home cage of a CD1 male for three minutes. Record the latency to the first attack, the number of attacks, and the duration of aggressive behaviors within the three-minute period as indicators of aggressiveness, and repeat the screening procedure for two additional days. For the social competition failure, place a multiple integrated social stress or MISS mouse and a winner mouse at opposite ends of the tube, so that they walk forward and meet at the midpoint. The MISS mouse will be pushed out of the tube by the winner. Switch the positions of the two mice and repeat the interaction. Then, return both the MISS and winner mice to their home cages. For the modified vicarious social defeat stress modeling, place a female MISS mouse inside a CD1 aggressor's cage, confined beneath a perforated acrylic cover to allow vicarious observation of aggressive interactions. Introduce a male C57 intruder mouse into the cage to be physically attacked by the CD1 mouse for 10 minutes. After 10 minutes, remove the male intruder and allow the female MISS mouse and additional five minutes of sensory exposure to the CD1 mouse. After five minutes, replace the CD1 mouse with a novel male C57 intruder mouse, and repeat the entire procedure three times. Then, return the MISS mouse to its home cage. Next, to model inescapable overcrowding stress, place an opaque sleeve inside a standard cage containing bedding. Introduce 12 to 15 female mice, either all MISS mice or mixed with other females into the sleeve for 30 minutes daily. For social isolation, house MISS mice individually when they are not undergoing social competition failure, vicarious defeat stress, or overcrowding stress for the remainder of the 10-day modeling period. The sucrose preference test, which evaluates anhedonia by measuring preference for a sweet solution over water showed that sucrose preference was significantly lower in MISS-exposed mice compared to naive controls. Total liquid intake in the sucrose preference test, including both water and sucrose, did not differ significantly between MISS and naive mice, indicating that the reduced sucrose preference was not due to altered fluid consumption. The tail suspension test, which assesses behavioral despair by measuring passive immobility, revealed that MISS-exposed mice had significantly longer immobility times than naive mice. Additionally, the open field test used to evaluate anxiety by measuring the time spent in the center of an open arena showed that MISS-exposed mice spent significantly less time in the central zone compared to naive mice. The elevated plus maze, which measures anxiety through avoidance of open arms, showed that MISS-exposed mice spent significantly less time in the open arms than naive controls.