Experimental method of mouse dark avoidance device
Experimental preparation
Familiarize the environment: Two days before the experiment, familiarize the mice with the laboratory environment, freely eat and drink water.
Grouping design: A stratified random block design is used to ensure that the average weight and gender ratio of animals in each group are similar. The experimental mice were divided into three groups based on different training methods: single adaptation single shock (MOS), single adaptation time limited training (MOL), and multiple adaptation time limited training (MUL). Each group was divided into two subgroups based on different doses: control (C) and model (M), with a total of six groups (n=9 in each group).
Instrument setup: The dark avoidance device is divided into two chambers, bright and dark, with a 3cm diameter circular hole in between. The bottom of both chambers is covered with copper grids. The copper grid at the bottom of the darkroom can be powered on, and the voltage can be adjusted through buttons, usually 40V, 50Hz.
training method
MOS method: The training is completed within one day. First, the mice are placed in a darkroom to adapt to the environment for 3 minutes. During the training, the mice are placed in a bright room with their backs facing the entrance of the cave. Once they enter the dark room and receive an electric shock, they are removed.
MOL method: Training is completed within one day, with mice adapting to the environment for 3 minutes and training for 3 minutes.
MUL method: Place mice in a dark avoidance device for 3 consecutive days at the same time to adapt for 3 minutes. After the adaptation is completed on the third day, start training and remove them after 3 minutes. When adapting to the environment and training, mice should freely shuttle between the open and dark rooms. Those with a latent period greater than 3 minutes should be discarded.
test method
Recording indicators: After 24 hours of training, place the mice back into the darkroom and record the time required for their first entry into the darkroom, i.e. the incubation period (STL), and the number of times they enter the darkroom within 5 minutes, i.e. the number of errors (ETs). If the mouse does not enter the darkroom within 5 minutes, STL is recorded as 300 seconds and ETs are recorded as 0.
Precautions
experimental environment
Environmental cleaning: Before and after the experiment, pay attention to cleaning the excrement of animals that click on the fence, as excrement can affect electrical conductivity. Wipe and air dry with alcohol to remove the previous animal odor as much as possible, and the equipment should have a ventilation system.
Reduce interference: During the experiment, it is necessary to maintain consistency in all experimental procedures, such as shielding noise, reducing odor stimuli, adjusting lighting, etc. The experimental environment should be quiet and dim, and animals should be grasped as lightly as possible to reduce stimulation to the animals. When closing the door between the two boxes, be gentle and do not create too much external noise that affects animal behavior.
Experimental operation
Animal adaptation: 3-7 days before the start of the experiment, grasp and gently stroke the animals every day to allow them to adapt to being grasped by the experimenters and avoid the problem of animals being in a stress state and unreliable experimental results caused by the experimenters' grasping during the experiment.
Electric shock parameters: rat current is 0.65-1.8mA (recommended 0.8mA), or voltage is 65-70V; mouse current is 0.15-0.6mA (recommended 0.25mA), or voltage is 30-36V, frequency is 5-15Hz current.
Equipment maintenance: The copper grid at the bottom of the experimental instrument is prone to rusting, causing an increase in resistance. Before each experiment, it should be sanded smooth with sandpaper.