Mouse training is key to success with experiments in awake mice head-fixed in the Mobile HomeCage. Below we provide some guidelines for the experimental timeline, duration and content of the training sessions.
Prepare mice in advance according to the surgery protocols.
We do not recommend training mice during the first two weeks after surgery (for imaging) or during the first post-surgery week (for electrophysiological recordings). For ephys, acclimation may start 3 days after the first surgery.
Delaying the start of training is necessary for the following reasons:
- the cement helmet must solidify to prevent the mouse from losing it (to ensure cement’s good grip to the skull, remove all periosteum from the exposed bone area during surgery);
- for easy handling and training, the mouse must fully recover from surgery;
- the cranial window requires 3-4 weeks to regain transparency (do not discard windows with bleeding vessels permaturely as they may become transparent over time).
Timeline for imaging experiments
Timeline for electrophysiological recordings
Wear a lab coat and use protective gloves. For best protection from biting use two pairs of gloves, cotton and latex. On the training days, avoid using perfume or other cosmetic substances that generate strong odor. Avoid smelly foods.
It is equally important to prepare yourself mentally: leave your worries and anxieties outside the lab as not to pass them to your mice. Project calm, resolve and the sense of being in control.
Overview of the mouse training
For your convenience, we have broken the entire process of preparation for imaging or recording in awake mice into several components. They appear as acclimation, handling and training blocks in the above diagrams. If you need to break the process or stretch it over a longer period of time, we recommend inserting the breaks between the blocks rather than interrupting the aclimation, handling or training procedures.
To reduce stress, house mice in group cages (at least two animals per cage). Weigh each mouse daily. Record the weight, and exclude the animals with more than 20% weight loss from experiments.
Acclimation (2-3 days)
On the first day, start by acclimating the mouse to the experimental room by leaving the cage in the room for approximately 60 minutes.
On the next day, place the cage in the experimental room next to the Mobile HomeCage device connected to the running air source.
On the following day, you might want add yet another source of noise, such as a radio or a voice recording, which simulates the noise that may occur during the experiment (such as when researchers talk to each other). This step is optional.
Throughout the entire preparation process (acclimation-handling-training), remember to introduce one variable at a time and allow breaks for memory consolidation!
Handling (2-3 days)
Handle mice for at least two days or until they are no longer afraid and don’t try to escape when you hold them on an open palm. Most standard handling techniques as well as accessories (such as e.g. a tube) can be used during handling in preparation for head fixation. However, we also recommend some special procedures. They include holding the mouse by the head plate to restrict head movement, and gently pulling the mouse by the head plate as to force it to walk in a certain direction along your forearm or your palm. These procedures prepare the mouse for the loss of control over head movement. Mice don’t like it, but they get over the initial shock fairly quickly. Visit this page to recap the standard and special handling techniques that can be used to prepare mice for head fixation in the Mobile HomeCage.
Training (2-3 days)
If mice have been handled well, they can be trained in two-three days. Start by placing the mouse into the air-lifted cage and let the mouse explore the cage. After several minutes of exploration, grab the mouse by the head plate and restirct its head movement. Gently pull the mouse by the head plate forcing it to move in a certain direction. Repeat this procedure several times during the first day of training, but keep the training sessions relatively short: 5-10 minutes are sufficient.
On the second day of training, pull the mouse by the head plate all the way into the clamp and quickly fasten the screws. Adjust the air flow making sure that the mouse does not wobble or that its hind legs don’t wrap around its body. Repeat the procedure several times during the day, but keep the training sessions short (5-10 minutes). Short but frequent training sessions produce better results than long training sessions. Mice need time to consolidate new knowledge and skills.
Adjusting the air flow
The right air flow is crucial to minimizing the stress under head fixation. Use this easy trick to check whether the air flow is optimal: slide a piece of office paper between the floating cage and the air dispenser. One sheet of office paper should pass unobstructed, but two sheets or a folded sheet of paper should not pass through. If they do, reduce the air flow.
Two days of training are sufficient for most well-handled mice, but the length of training should be adjusted based on each mouse’s individual performance.
Watch the video below for the overview of acclimation-handling-training procedure. In the video, we also show an alternative way of head-fixing the mouse in the Mobile HomeCage by wrapping it in a soft cloth and immobilizing the limbs. While we do not recommend using the wrapping procedure under normal circumstances, it may be useful if your experimental design does not allow much time for training:
If you are using the Levelt clamp (magnetic head-fixation), watch this video for head-fixation tips: