new object recognition
Summary
New object recognition assesses the memory capacity of the tested animal by the length of time the animal spends exploring familiar old and new objects, which is a simple experimental method for judging the memory capacity of experimental mice and rats.
Operation method
New object recognition
Principle
Put several objects in an open field box and let the mice familiarize themselves with them for a while, then replace one of the objects with a new one. Observe whether the lab mouse is more interested in the new object. If not, the experimental mouse is considered to have a possible memory deficit.
Materials and Instruments
Experimental rats; the experimental field box was 25cm×25cm×40cm without a roof;
Two Lego blocks of 8cm height and 3.2cm width;
The bottom area of the sand-filled box was 25cm
2
Two cell culture flasks filled with sand;
TM-Vision Behavioral Experiment System
Move
Purchased mice were first acclimatized in a rearing cage for 5-7 days. Three days before training and testing, the mice were petted for 1 min per day to avoid high stimulation.
Day 1 Acclimatization Phase
24h before training or testing, the mice are placed in the test room to acclimatize to the test environment.
The mice were placed in an open field box for acclimatization, with their heads facing the experimenter's side, and were allowed to explore freely for 5 min. At the end of the experiment, the mice were taken out and put back into their original cages, and the empty boxes were cleaned with 75% alcohol to remove any residual odors, waiting for or accelerating the evaporation of the alcohol before acclimatizing the next mice.
Day 2 Training phase
Two identical objects (Legos or cell culture flasks filled with sand) were fixed 5 cm from the walls of each box.
The mice were then placed in the empty box with their heads facing the opposite direction of the two identical objects and allowed to move freely in the box.
Immediately after placement, the video equipment was turned on, and the experimenter left the test room immediately to record the mice's contact with the two objects, including the number of times the nose or mouth touched the objects and the time spent exploring within a range of less than 2 cm from the objects.
If the total exploration time reached 20s or the experiment time reached 10min, the experiment was stopped and the time spent on this process was recorded. The mice were removed from the cages at the end of the experiment and immediately returned to their original cages. Allow the mice to rest for 1h before conducting the test again (the mice remain in the test room during this time).
The test can also be performed 24h later, which can be adjusted according to the needs of the experiment. Clean the empty box and objects with 75% alcohol to remove any residual odors and dry them before proceeding to the next mouse.
Day 3 Test Phase
The standard time for each mouse was less than 10 min, and after 24 h, one of the same objects was replaced with another object similar to the training phase.
The position of the new object (left or right) was randomized across mice and across test groups. Mice were timed for the exploration time and number of explorations on each of the two objects, and if the total exploration time reached 20s or the experiment time reached 10 min, the experiment was stopped and the time spent on this process was recorded.
The behavior of the mice during the test phase was tracked and analyzed separately for the time spent exploring the new object, T1, and the time spent exploring the old object, T2, to calculate the cognitive index, CI, which was calculated using the formula CI=T1/(T1+T2).
The cognition index (RI) of the mice can also be calculated.RI(%)=number of times new objects were explored/(number of times new objects were explored+number of times old objects were explored)×100%. Higher cognition index indicates better memory ability of mice.
Caveat
The mice were removed at the end of the experiment, and the empty box and objects were cleaned with 75% alcohol to remove residual odors and dried before proceeding to the next mouse.Object detection was timed when the mice sniffed the object or when they wanted to touch the object while observing it (when the distance between the nose and the object was less than 2 cm).The front paws on the object, sniffing the object, licking the object, etc. were considered as detecting the object.Biting, posing, or climbing on an object without moving are not considered detecting a new object and do not qualify as exploration and are not counted as exploration time.
Common Problems
Different box backgrounds have different effects on the results of NOR test, the animals are less stimulated by stress in the black environment, and can maintain a higher level of concentration on the object, and thus will also maintain a greater interest in the object.
Therefore, when carrying out the new object recognition test, it is important to keep the laboratory environment as dim as possible, without bright light stimulation, and the mice's ability to recognize new objects is significantly improved in a relatively dim environment.
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