All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. EcoHIV-EGFP [Ecotropic HIV Enhanced Green Fluorescent Protein] virus stereotaxic surgeries
<ol>
	<li>Anesthetize rats using 3% sevoflurane. When the rats are not responsive to noxious stimuli and their reflexes are absent, proceed to step 1.2.</li>
	<li>Shave the hairs from the brain region and sterilize the skin twice with 70% ethanol and a chlorhexidine-based scrub. Secure the rat in a prone position in the stereotaxic apparatus.</li>
	<li>Make an incision (5-6 cm) through the skin along the scalp midline.</li>
	<li>Mark two drilling positions at 0.8 mm lateral, 1.2 mm rostral to bregma. Drill a hole (diameter 0.4 mm) in each skull position.</li>
	<li>Fill titrated EcoHIV lentivirus solution (1.04 &#215; 106 TU/mL) in a 10 &#181;L injection syringe. Secure the syringe to the stereotaxic apparatus.</li>
	<li>Move down the needle close to the surface of the drilling hole. Measure and move 2.5 mm in depth.</li>
	<li>Infuse 1 &#181;L of virus solution at a rate of 0.2 &#181;L/min. Keep the needle inside of the injection area for 5 min. Slowly move the needle up until it is outside of the rat skull.</li>
	<li>Suture the skin with a 4-0 silk thread.</li>
	<li>Sterilize the incision with 70% ethanol once. Subcutaneously inject butorphanol (Dorolex, 0.1 mg/kg body weight).</li>
	<li>Transfer the rat to a recovery chamber with a heating pad until it wakes up.</li>
</ol>

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Antibiotic-Antimycotic solution</td>
			<td>Cellgro</td>
			<td>30004CI</td>
			<td>100X</td>
		</tr>
		<tr>
			<td>Cover glass</td>
			<td>VWR</td>
			<td>637-137</td>
			<td></td>
		</tr>
		<tr>
			<td>Drill</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Ethicon Vicryl Plus Antibacterial, 4-0 Polyglactin 910 Suture, 27in. FS-2</td>
			<td>Med Vet International</td>
			<td>VCP422H</td>
			<td></td>
		</tr>
		<tr>
			<td>Hamilton syringe</td>
			<td>Hamilton</td>
			<td>1701</td>
			<td></td>
		</tr>
		<tr>
			<td>Iris Forceps</td>
			<td>World Precision Instruments</td>
			<td>15914</td>
			<td></td>
		</tr>
		<tr>
			<td>Iris Scissors</td>
			<td>World Precision Instruments</td>
			<td>500216</td>
			<td></td>
		</tr>
		<tr>
			<td>Stereotaxic apparatus</td>
			<td>Kopf Instruments</td>
			<td>Model 900</td>
			<td></td>
		</tr>
		<tr>
			<td>Vannas Scissors</td>
			<td>World Precision Instruments</td>
			<td>500086</td>
			<td></td>
		</tr>
	</tbody>
</table>

establishing an active hiv infection in a rat model

Establishing an Active HIV Infection in a Rat Model

Source: Li, H., McLaurin, et al. <a href='https://app.jove.com/v/62137'>A Rat Model of EcoHIV Brain Infection.</a> J. Vis. Exp. (2021).
This video demonstrates the procedure of injecting chimeric HIV viruses into the cortex of a rat's brain, where the virus RNA integrates into the host genome, replicates, and spreads to establish an active HIV infection in the experimental model.

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. ...

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.

establishing-an-active-hiv-infection-in-a-rat-model

Research

JoVE Encyclopedia of Experiments

Neuroscience

Neuropathology

Infectious Diseases

829 Views. 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 viru...

Establishing an Active HIV Infection in a Rat Model

In This Article

Abstract

Protocol

Disclosures

Materials

Tags

Name	Company	Catalog Number	Comments
Antibiotic-Antimycotic solution	Cellgro	30004CI	100X
Cover glass	VWR	637-137
Drill
Ethicon Vicryl Plus Antibacterial, 4-0 Polyglactin 910 Suture, 27in. FS-2	Med Vet International	VCP422H
Hamilton syringe	Hamilton	1701
Iris Forceps	World Precision Instruments	15914
Iris Scissors	World Precision Instruments	500216
Stereotaxic apparatus	Kopf Instruments	Model 900
Vannas Scissors	World Precision Instruments	500086