The detailed description of the experimental method is as follows:
In learning and memory experiments in zebrafish, we employ a specific experimental setup to assess their response to novelty objects. First, each zebrafish is transferred to a transparent test pond with a size of 25 20 15 cm (L, W, H). Such a size is enough for zebrafish to swim freely in the pool without feeling crowded.
The new object used in the experiment was a blue plastic cube with dimensions of 3 3 1 cm (length, width and height). This object was chosen to determine the zebrafish's response to novelty. The size, shape, and color of the object are carefully designed to ensure that it attracts the attention of zebrafish but does not cause them to panic or overreact.
The water temperature in the test cell is maintained at 25 1 degrees Celsius and the pH is controlled at 72-7.6, the hardness is 440-61.0 mg CaCO3 L to simulate the living conditions of zebrafish in their natural environment. The water depth is set at 10 cm, which is enough for zebrafish to swim freely and for them to observe and record their behavior.
Before the start of the experiment, the zebrafish underwent a 5-minute acclimatization period in the test pond to reduce stress due to environmental changes. After that, the new object is placed in the corner of the fish tank. For the next 8 minutes, the zebrafish were allowed to explore freely, recording and analyzing their behavioral trajectories using Shanghai Xinsoft FishTrack animal behavior analysis software.
In order to more accurately analyze zebrafish's interest in new objects, the experimental pool was virtually divided into two parts: a new object zone and an objectless zone. By analyzing the total distance (in centimeters) and the time (in seconds) that zebrafish have swam through these two areas, researchers can judge how interested the zebrafish are in the new object.