Behavior Testing

At P60, we conducted behavioral tests on each group of rats.

Social behavior was tested in a rectangular box divided into three chambers (40×34×24 cm) with side chambers connected to the middle chamber by a corridor (10×10×15 cm). Rats were tested in the device in the dark, and their activities were recorded by infrared cameras. The test was divided into three phases:³⁹ the adaptation stage, the first stage and the second stage. During the adaptation stage, the rats were allowed to freely explore in the chambers for 5 min. In the first stage (Figure 2A), the social preference test phase, a cage containing an unfamiliar rat of the same sex and age was placed on one side of the device as a social stimulus, and an identical empty cage was placed on the other side. The test rat was placed in the middle chamber and allowed to freely explore the entire device for 10 min. During the second stage (Figure 2D), another unfamiliar rat of the same age and sex was placed in an empty cage as a new social stimulus. The test rat was placed in the middle chamber and allowed to freely explore for 10 min. After each test, the entire apparatus was cleaned using 75% alcohol. The amount of time spent sniffing the wire cage was analyzed by a highly trained observer blinded to the genotype as closely social interaction time.

Effect of early swimming intervention on the behavior of Shank3 knockout rats.

Experimental rats were placed inside a rectangular box (40×34×24 cm) for the self-grooming test.³⁶ Infrared cameras were used to record the self-grooming behaviors of rats in the dark. The rats were placed in the box and allowed to adapt for 10 min and then recorded for 10 min. The self-grooming behavior paradigms included rats licking their body and hair, wiping their face with their front paws, and scratching their torso (Figure 2G).