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A Guide to Safe Mouse Handling | Manual on Mouse Transport Protocols and Troubleshooting

[Guide to Selecting Administration Routes] How do you choose the appropriate administration route for mice? Read this article and say goodbye to indecision.

In mouse model studies, the choice of administration route can significantly influence the accuracy and reliability of experimental outcomes. Different routes of administration not only affect drug bioavailability and metabolic pathways but may also exert distinct effects on the animals’ behavior and physiological status.
The routes of administration in experimental animals are: Oral administration, injectable administration, intranasal administration, transdermal administration, intrathecal administration, intracerebral administration, rectal administration, intra-articular administration and various other routes of administration.

Route of administration

| Oral administration

① Oral administration

This is typically accomplished by dissolving a liquid or solid in water and administering it via a feeder. Alternatively, solid particles can be directly mixed into the feed. This method is suitable for most compounds and formulations and is straightforward to implement. It is generally used for the prevention and control of animal diseases or for establishing animal models of human diseases related to diet. 。

② Gavage administration

Using an instrument (gavage needle), the drug is administered directly into the animal’s stomach.

Route of drug circulation:

Esophagus →Stomach → Small intestine → Capillaries of the small intestine (absorption into the bloodstream) → Superior mesenteric vein → Hepatic portal vein → Liver → Hepatic veins → Inferior vena cava → Right atrium → Right ventricle: absorption into the systemic circulation.

Fixing the mouse:

Use the left thumb and index finger to grasp the mouse’s ear and the skin on its neck, and with the ring finger and little finger of the left hand, gently pinch the skin along its back and the base of its tail, aligning its head, neck, and body in a straight line. If the initial grasp is improper, release the mouse and recapture it carefully to avoid compromising subsequent procedures.

Needle insertion:

Holding a syringe in the right hand ( Use a 1 mL syringe fitted with a gavage needle: insert the needle at one corner of the mouse’s mouth, gently press the base of the tongue, and advance it slightly forward. Once the needle enters the esophagus, you will feel a slight “empty” sensation. Continue advancing slowly; typically, when the needle has been inserted 3–4 cm into the mouse—about three-quarters of the way along the gavage tube—you can begin the injection.

Note: If you encounter resistance or the animal struggles vigorously, it indicates that the needle has been inserted in the wrong location; immediately stop advancing the needle or withdraw it, and attempt again once the animal has calmed down.

| Injectable administration

① Intraperitoneal injection

An injection method in which the drug is administered outside the serosa of the gastrointestinal tract and within the peritoneum.

01 Secure the rats and mice in a straight line, with the mouse’s head positioned lower than its tail, to facilitate cephalic organ relocation.

02 Disinfect the needle insertion site on the abdomen of mice and rats with an alcohol swab, moving against the direction of hair growth.

03 Hold the syringe, insert it into the abdomen of the mouse or rat, and slowly administer… 45° Insert the needle into the abdominal cavity from either side, with an insertion depth of less than 1cm The needle insertion is felt as a “loss of resistance,” and aspiration yields no urine or blood.

04 Slowly inject the drug solution, ensuring that the entire dose is delivered into the peritoneal cavity of both mice and rats.

Note to confirm the needle insertion site in mice and rats:

At the intersection of the midline and the penultimate pair of nipples, on either the left or right side—typically, the right side is chosen for needle insertion in mice, as the right abdomen contains the small intestine and is less likely to puncture solid organs.

Route of drug circulation:

Superior mesenteric vein → Hepatic portal vein → Liver → Hepatic vein → Inferior vena cava → Right atrium → Absorption into the bloodstream occurs in the right ventricle;

Following administration, the compound is not absorbed via the gastrointestinal tract; therefore, intraperitoneal administration does not involve first-pass metabolism in the gut, but hepatic first-pass metabolism still occurs, resulting in lower drug loss compared with gavage.

② Subcutaneous injection

Secure the mouse or rat by grasping the head, neck, and dorsal skin with the flexed thumb and index finger of the left hand, while using the palm and the back of the hand to immobilize the animal.

Disinfect the left thumb with an alcohol swab, and pinch the area with the index finger.

With the needle held in the right hand, insert it parallel to the mouse’s body into the skin of the head and neck. Gently move the needle from side to side; if you feel resistance, the needle has entered the subcutaneous tissue. Aspirate to confirm the absence of blood, then slowly inject the drug solution into the subcutaneous space. You may palpate a slight elevation at the injection site between the thumb and index finger.

Note: After needle removal, gently press the puncture site to prevent leakage of the medication and enhance its absorption.

③ Tail vein injection

Prepare a syringe for drawing up the drug solution and set it aside. Place the mouse or rat in the restraint device on its side, with the tail extended through the small opening at the rear of the restraint cylinder.

Straighten the mouse’s tail, locate a vein on one side, and use… 75% Repeated wiping of the veins in mice and rats with alcohol swabs resulted in marked venous dilation.

When inserting the needle, use the index finger and thumb of the left hand to hold the mouse or rat’s tail, allowing the tail to curve downward past the thumb; the injection site is just above the bend.

During injection, the medication normally travels along the vein and flows in smoothly.

Upon needle removal, immediately apply a dry cotton ball to the puncture site for several seconds until bleeding has ceased. During compression, remove the mouse or rat from the restraint device and elevate the tail to promote hemostasis.

Note: When immobilizing rats and mice, be sure to position the fixation clamp so that its small opening aligns with the animal’s nostrils; failure to do so may result in asphyxiation and death.

During injection, position the needle as parallel to the vein as possible, advancing it slowly with the bevel facing upward. The injection should proceed without significant resistance, and the vessel should not become distended or swollen; if either occurs, it indicates that the medication has not been administered into the vessel.

④ Intracerebral administration:

Intracerebral administration is a stereotactic injection. During stereotactic brain injections, a stereotactic apparatus is used to immobilize the head, and external cranial landmarks are employed to localize intracranial nuclei, enabling targeted stimulation, lesioning, or drug administration without exposing the brain. This technique can be applied to establish animal models of Parkinson’s disease, epilepsy, intracranial tumors, and other conditions.

| Nasal administration:

During administration to put The drug is slowly dripped into the mouse’s nostril, allowing the mouse to inhale it through the nasal cavity (note that the solution will disappear into the nostril). Each dose of the solution should be sterile and in a small volume, repeated multiple times until the required total volume is achieved. Nasal drops are commonly used as a route of administration when establishing animal models of asthma and pulmonary arterial hypertension, among others.

| Transdermal administration:

It is non-invasive, delivering drugs into the body through the skin, offering exceptional convenience and a low rejection rate. Transdermal medications are absorbed into the circulation slowly, thereby avoiding the liver’s first-pass effect. , and can also enhance bioavailability.

| Intrathecal administration:

Similar to a minor surgical procedure, this involves injecting medication into the subarachnoid space of the spinal canal and is primarily used for adeno‑associated viruses. (AAV) Injection and recovery from spinal cord injury.

| Rectal administration:

The drug is administered into the rectum via the anus; rectal administration allows absorption through the intestinal mucosa, bypassing hepatic metabolism and entering the systemic circulation directly, thereby enhancing the drug’s bioavailability. Rectal administration typically involves larger doses and is commonly used in animal models of colitis.

| Intra-articular administration:

High-concentration therapeutic agents can be directly delivered into the joint space, typically for the treatment of various musculoskeletal conditions, such as osteoarthritis and rheumatoid arthritis.

Select according to the experimental objective.

Drug studies can generally be categorized into pharmacodynamic studies, pharmacokinetic studies, toxicity studies, and toxicokinetic studies, among others.

| Pharmacological Efficacy Study

When designing pharmacodynamic studies, it is essential to closely align with the intended route of administration in clinical practice and to take into account the drug’s expected therapeutic indications.

For example, intravenous and intraperitoneal injections, owing to their rapid absorption, are particularly well suited for managing emergency situations.

| Pharmacokinetic Experiment

When assessing the bioavailability of a drug, the choice of administration route is critical. If the study focuses on the bioavailability of an orally administered drug, intragastric gavage should be used; if the focus is on the bioavailability of an intraperitoneal formulation, intraperitoneal administration should be employed directly.

| Toxicity Test

In toxicity studies, particularly acute toxicity assays, the choice of administration route significantly influences both the extent of drug exposure and the rate of absorption, potentially leading to substantial variability in observed toxicological outcomes. Consequently, when conducting toxicity assessments, it is essential to carefully account for the potential differences arising from different routes of administration.

When selecting a route of administration, it is essential to comprehensively consider the drug’s solubility, stability, and toxicity. In some cases, the preferred route of administration dictated by the experimental objective may be constrained by the drug’s properties—for example, if the drug is poorly soluble in injection solutions or if oral administration is likely to irritate the gastrointestinal tract—requiring flexible adjustments.

Selecting and implementing an appropriate route of administration based on the drug’s properties, the experimental objectives, the animal’s health status, and the feasibility of the procedure is a critical step in ensuring the accuracy and reproducibility of experimental results.

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