This article describes a method for targeted drug delivery to the cerebrospinal fluid (CSF) in the cisterna magna of anesthetized mice. The technique allows for selective labeling of surface brain cells using a fluorescent marker gene.
Begin with an anesthetized mouse expressing a fluorescent marker gene, and secured in a stereotactic frame.
The head is tilted downward for easier access to the cisterna magna, the cerebrospinal fluid or CSF-filled space between the cerebellum and the brainstem.
Apply eye ointment, shave the neck, and sterilize.
Make a midline incision to expose the membrane covering the cisterna magna.
Retract the tissues with a separator to access the cisterna magna.
Take a microsyringe filled with an inducer drug and fitted with a bent needle.
Insert the needle and puncture the membrane to reach the cisterna magna.
Inject the drug slowly for uniform dispersion within the CSF.
The drug enters the brain surface cells and triggers the expression of the marker gene, producing red fluorescent proteins.
CSF flow prevents drug diffusion into deeper brain regions.
Fluorescence imaging confirms selective labeling of surface cells, demonstrating a targeted drug delivery to CSF-exposed regions.
Anesthetize the animal in the isoflurane chamber.
Then adjust the mouse head holder so that the mouthpiece is approximately at a 30 degree angle from the surface of the surgical table and fix the animal's head onto the holder. To improve accessibility to the cisterna magna, position the animal's body at approximately 30 degrees from the surface of the table with the head tilted downward which will establish an angle of 120 degrees with the rest of the body and extend the back of the neck to facilitate access to the cisterna magna. Once the animal is properly positioned, apply ophthalmic ointment, shave the back of its neck, and sanitize the area with alcohol wipes and Betadine.
Use surgical scissors to make a midline incision starting at the level of the occipital bone and extending posteriorly. Gently separate the superficial connective tissue and neck muscles by pulling sideways from the midline with fine tip tweezers which will expose the dural membrane overlying the cisterna magna. Position a small surgical separator to enable visualization of the cisterna magna throughout the procedure.
Identify the caudal end of the occipital bone and insert the previously bent needle immediately underneath. Once the dura has been perforated, allow the bent tip of the needle to penetrate underneath the surface by gently pulling the syringe upward and parallel to the animal's body which will ensure better stability. Inject the compound slowly to avoid interference with cerebrospinal fluid's natural flow.
After the injection, let the needle rest inside for one minute, then carefully remove it with the help of fine tip forceps.
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