Intranasal Administration of CNS Therapeutics to Awake Mice

The overall goal of this procedure is to target drugs to the central nervous system using intranasal delivery in mice without anesthesia. This is accomplished by first acclimating the mouse to handling. The second step is to use a modified scruff grip to immobilize the animal's head.

Next droplets of drug are intranasally administered with a pipetter. In the final step, the mouse is returned to its cage with a reward. The main advantages of this technique are that intranasal administration targets drugs to the central nervous system without systemic exposure, and that awake intranasal delivery of drugs allows for chronic treatment in mice without anesthesia.

This method can be used to help answer key questions in the development of drugs for treatment of neurologic disease. Generally, individuals new to this method will struggle because it is difficult to hold the mouth securely to prevent movement of the head. Visual demonstration of this method is critical as the placement of the fingers for the proper grip is key and difficult to explain via text and photographs Two to four weeks prior to intranasal dosing, acclimate the mice to handling by performing the following steps over two to three days each confirming that the mouse exhibits no signs of stress before moving on to the next step.

For the first acclamation step, place the mouse in the palm of the hand for a period of two to three minutes. No more than one foot above the cage. Top next, place the mouse in the palm of the hand for three minutes and pet the animal gently from the head to the tail, allowing the animal to move about freely.

Once the animal tolerates petting, place the mouse in the palm of the hand for three minutes and then use the thumb and pointer finger to lightly pinch together the skin on the back of the animal's neck. For the next acclimation step, lift the mouse by the scruff of the neck for 30 seconds. Then let the animal rest on the cage top for another 30 seconds before repeating the hold again.

Now practice the intranasal grip by first using the thumb and index finger of the dominant hand to hold the mouse by the tail. Then use the middle ring and pinky fingers to lightly pin the lower portion of the animal's body to the cage. Let the mouse grab onto the bars lightly stretching the animal out To better grab the scruff.

Using the thumb and middle finger on the non-dominant hand, proceed with pinching the mouse firmly at the scruff behind the neck and shoulders pulling up on the loose skin of the upper back to make a tent across the shoulder blades. Then quickly place the index finger of the non-dominant hand on the mouse's head as close to the nose as possible, and pull back on the skin. At the same time, slide the thumb and middle fingers toward the head for optimal positioning.

Ideally, the mouse should not be able to move its head vertically or horizontally. During the next session, perform the intranasal grip as just demonstrated. This time inverting the animal so its ventral side faces the ceiling and its neck is parallel to the floor.

Once the mouse is comfortable with the inverted intranasal grip, practice briefly placing a micro pipette with an empty tip close to each nostril. Finally, administer six microliters of saline, first to only one nare, and then in the next session to both using the intranasal drug delivery procedure demonstrated more fully in the next section before beginning the intranasal drug delivery. First, arrange the cages and dosing instruments for easy reach.

Then remove the first mouse from its cage and use the just demonstrated intranasal grip to pick up the animal. Next, using the dominant hand load the micro pipette with six microliters of drug. Place the tip of the filled pipette near the mouse's left nostril at a 45 degree angle.

Then slowly eject the drug to form a small droplet with about half of the volume in the tip, place the droplet close enough to the mouse's nostril that the mouse can inhale it immediately after the mouse inhales the drop, eject the rest of the drug in the pipette tip to form another small droplet for the mouse to inhale through the same nostril about two to three seconds later, hold the mouse in this position for 15 seconds, and then administer the drug to the right nostril in the same fashion. Now, place the mouse back on top of the cage for two minutes, and then deliver the drug again to both nostrils as just demonstrated exactly two minutes after the administration to the first nostril to dose one or two mice at once. This sample schedule can be followed after the mouse has received the full dose of drug.

Return the animal to its cage. After each acclimation or intranasal dosing session, mice should be given a treat such as a cheerio as positive reinforcement to reduce stress, always restore the nests to their original states before returning the animals to their cages, change gloves between cages when dosing group housed animals so as not to transfer scent from cage colony to cage colony, which would induce aggressive cage mate behavior. Once mastered, one person can intranasally administer drug to 15 to 20 mice in one hour While attempting this procedure with more than one mouse at a time.

It's important to take precautions to prevent the mice from accidentally switching cages. After watching this video, you should have a good understanding of how to use intranasal delivery in mice without anesthesia. To target drugs to the central nervous system.

Remember to always take precautions when working with mice such as wearing an N95 mask and gloves to prevent exposure to allergens.