This article describes a method for delivering recombinant adeno-associated virus (AAV) vectors into the bloodstream of anesthetized rats via the lateral tail vein. The procedure facilitates the introduction of genetic material into neurons for neurotherapeutic applications.
Begin with an anesthetized rat positioned on its side with its tail exposed.
Locate the lateral tail vein, identify an injection site along its length, and disinfect the area.
Apply slight pressure above the injection site to dilate the vein.
Take a syringe containing the recombinant adeno-associated virus or AAV vector carrying the target gene.
Align the needle with the vein and pierce the tail skin.
Release the pressure on the vein to restore normal blood flow.
Insert the needle into the tail vein and slowly inject the virus.
After injection, withdraw the needle, apply pressure to stop any bleeding, and monitor the rat's recovery.
The lateral tail vein delivers the virus into the bloodstream, where it circulates through the heart and enters the brain, reaching the neurons.
Inside the neuron, the virus enters the nucleus, releases its genetic material, and facilitates the production of target proteins for neurotherapeutic applications.
Properly anesthetize the rat using the anesthesia system. Ensure an adequate plane of anesthesia by the absence of a reaction to a toe pinch. Then, place the rat in the work area on its side and maintain anesthesia using the nose cone. With the rat lying on its side, one lateral tail vein should be facing up.
The injection site is two thirds down the length of the tail. Wipe the injection site with the alcohol preparation pad. Firmly hold the tail slightly above the injection area with one finger directly over the lateral tail vein. This will dilate the vein. With the bevel facing up, align the needle with the visible tail vein in the same direction.
Pierce the skin over the tail vein while taking great care to avoid the other hand holding the tail and pressing on the tail vein. Release pressure from the tail vein to allow for normal blood flow. Then move the needle into the vein. To ensure that the needle has punctured the tail vein, pull back slightly on the plunger. If the needle is in the vein, blood will flow into the needle.
After repositioning the needle, if required, stabilize the needle by holding the end of the syringe against the tail. Slowly inject the vector into the tail at approximately 20 microliters per second.
In the case of a poor injection, the animal will only be partially transduced and the results will probably not be useful. However, with practice, good injections are very reproducible and reliable.
If the vector has been administered, remove the needle from the tail, and press a gauze pad against the injection site to help stem the bleeding for 30 to 60 seconds. Turn off the isoflurane anesthesia and oxygen. Monitor the rat until it regains consciousness and then return it to its cage. Dispose of materials that may be contaminated with AAV into an appropriate biohazard container, and clean the workstation with 70% ethanol.
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