This article details a method for inducing HIV infection in the rat brain using chimeric viruses. The procedure involves precise surgical techniques to ensure accurate virus delivery to target brain cells.
Begin with an anesthetized rat, disinfect its shaved head, and secure it in a stereotaxic apparatus.
Make a midline incision to expose the skull.
Mark two positions on the skull and drill small holes.
Attach a syringe with chimeric viruses containing ecotropic HIV RNA within a lentiviral vector to the stereotaxic apparatus.
Insert the needle into the hole, targeting the brain cortex cells, including microglia, the brain's immune cells.
Slowly inject the viruses for their even distribution in the cortex.
Post-injection, suture the incision, disinfect the area, and allow the rat to recover.
The injected virus interacts with the brain cell receptors, fuses, and releases HIV RNA into the cytoplasm.
The RNA is reverse-transcribed into DNA and integrates into the cell's DNA.
The infected cell expresses the viral components, producing new RNA and proteins.
These assemble and release as new viruses, which increase the viral load in the brain, establishing an active HIV infection.
For EcoHIV EGFP injection, after confirming a lack of response to pedal reflex, shave the hair from the brain region, and sterilize the exposed skin 2 times with 70% ethanol, and a chlorhexidine-based scrub. Secure the rat in a prone position in a stereotaxic apparatus, and make a 5 to 6-centimeter incision through the skin along the scalp midline.
Mark one drilling position at 0.8 millimeters lateral and one 1.2 millimeters rostral to bregma, and drill a 0.4-millimeter diameter hole at each skull position. Load 1.04 times 10 to the sixth transduction units per milliliter of titered EcoHIV Lentivirus solution into a 10-microliter injection syringe, and secure the syringe to the stereotaxic apparatus.
Move the needle close to the surface of one drilling hole and insert the needle 2.5 millimeters into the hole. Infuse 1 microliter of virus solution at rate of 0.2 microliters of virus per minute. When all of the virus has been injected, leave the needle inside the injection area for five minutes before slowly retracting the needle until it is outside of the rat skull.
Close the skin incision with a 4-0 silk thread suture, and sterilize the solution with 70% ethanol. Then, place the rat in a recovery chamber with a heating pad with monitoring until recumbency.
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