Mouse-T maze behavioral protocol
General Notes:
The mice we use are C57BL/6 mice they typically weight 25-40g and are socially housed untill they are implanted. Importantly mice are not little rats they are different species with different behaviors. What these means in terms of behavioral training is that they are not as social nor as intelligent as the rats, and require more habituation and more careful handling procedures. They do not generally "warm up" to the experimenter and become comfortable with handling. Keep this in mind while handling them.
Behavioral Training:
In general behavioral training follows a standard formula with there being some differences from lab to lab. But in general the procedure is habituate the animal to the training apparatus, the trainer, the reward, and the manipulation(s).
This is done in stages starting with habituation to the trainer and reward followed by the apparatus and lastly the manipulations that are done. Importantly animals should always be well habituated to objects or persons they are encountering before behavior measures are started. Unless of course experiment design entails some manipulation should not be familiar.
Feeding Animals: Because the mice are kept socially housed to they are given viruses and implanted with lenses. The food must be distributed across the floor of the maze to prevent one mouse from hogging all the food. For each mouse 1.5-2.0g of chow should be given which is about 1/3 of full size pellet. Take pellets and break them into 1.5-2.0g sizes and scatter them throughout the cage.
Rat smell: Rats can and do act as predators to mice in the wild, and the smell of rats can introduce anxiety in mice, therefore it is important to make sure your lab coat and gloves have not touched any rats.
Not moving: Sometimes mice can be hesitant to run, if a mouse is placed in a maze and doesn't after the block is lifted move after 90 secs remove the mouse and try again in a few minutes.
Arm entry: Deciding what the criterion is for arm entry is important in our lab we do tail tip that is the mouses tip of his tail has passed the edge of the stem on either arm, then mouse counts as having entered that arm.
Important: The maze floor must be wiped down with cholorohexidene to remove any odor cues between the sample and choice trials as well as between each individual animal.
Handling Notes:
When pick up the mouse one of three ways is recommended.
1. The first is to grab them by the base of the tail, and them support them in your open hand.
2. Is to cup your hands together then pick them up out of the cage be sure to keep your hands open as if the mouse feels he couldn't escape he will likely bite. (this is the method that should be used as this will need to be done to place the miniscope on their head in the future).
3. Use a plastic bottle that has been cut open and scoop them into it.
Habituation:
1. mice should be placed on food restriction
2. A small amount of the sweetened condensed milk mixture 1:1 with water should be placed in their cage.
Take two weigh boats on small and one large tape the bottom of the small weigh boat to the large weigh boat and fill the small weigh boat with the condensed milk mixture.
3. Place your hands in the cage without picking up the mice for 5 minutes
4. Then gently pick up each mouse with a cupped hand and hold them for about 30 seconds each. Do this 3 times for each mouse
5. All of this is to be repeated for the first 4 days.
Maze placement 1:
1. Prior to rewarded alternation testing, the animals must be habituated to the T-maze. This is best done by raising the block (on an enclosed maze), filling the food wells and putting an entire home cage group of animals (previously food restricted) in for about 3 min. Replenish the reward if necessary. Do this four times (or more if they are slow to consume the reward) with gaps between exposures of at least 10 min. Do this for 3 days.
Maze placement 2:
1. Next allow individual animals to run from the start arm with one goal arm blocked by its door (or block for an elevated maze). Equal numbers of left and right runs are given for a total of 10 runs or 5 left and 5 right. Do this for 3 days.
Make sure the animal readily consumes the reward and runs to each arm with no issues before moving on.
Task:
Now actual trials of DNMTP can begin there is a total of 10 sample-choice trial pairs, which arm is the correct one for each trail is determined pseudo-randomly. Set up for a sample trial by baiting the sample and choice arms with reward, with access to the correct choice arm denied by the choice point block which is pushed just into the non-sample arm.
Place the animal in the start area, the first few centimeters of the stem which is blocked off by the second block. Stand centrally behind this area so your body presence does not bias the arm choice of the animal, ten seconds after placement remove the block and allow the animal to run to the sample arm and consume all of the reward. When the animal has consumed all the reward, return it to the start arm and re block the stem. Remove the animal placing them in an empty cage, a 20 second delay is then imposed and the maze is cleaned. During this time remove the choice point block
Return the animal to the stem after the 20 second delay. Ten seconds after stem placement remove both the start arm block, and allow the animal to freely choose the arm. If they choose correctly allow them to consume all the reward before returning them to the stem of the maze. If they choose incorrectly give them time to see the food well is empty and remove them after the equivalent time if that arm had been the correct choice.
Place them back into the empty cage and this time a 45 second delay is imposed, the maze is cleaned and the sample-choice trail procedure begins again. The 45 second delay is the intertrial delay and the 20 second delay is the intra-trial delay
Criterion for task is two days at 80% that is 8/10 trials trials correct.
Retrieval Practice Task:
Forthcoming!
References:
Deacon, R. M., & Rawlins, J. N. (January 01, 2006). T-maze alternation in the rodent. Nature Protocols, 1, 1, 7-12.