inducing a facial nerve injury in a rat model

Inducing a Facial Nerve Injury in a Rat Model

After confirming a lack of response to toe pinch, apply ointment to the eyes of the rat, and shave the surgical area. Establish a method for rat identification, and place a roll of gauze under the neck.
Disinfect the exposed skin with three alternating chlorhexidine and 70% ethanol scrubs, and place the rat under a stereomicroscope. Manipulate the ipsilateral ear in an anterior-posterior direction to determine the natural folding of the postauricular skin. Use a number 15 blade to make a two to three-millimeter incision in the postauricular crease.
The planning and the placement of the incision are the most critical steps for ensuring reliable identification of the facial nerve while minimizing the wound size.
Bluntly dissect through the immediate subcutaneous fascia, and place a micro-Weitlaner retractor to enhance the tissue exposure. Identify the anterior digastric muscle as it travels in an inferior to superior direction towards its insertion along the skull base. Spread gently through the muscle belly along the muscle insertion point to reveal the tendon of the anterior digastric belly.
The tendon will appear as a filmy white process emanating from the muscle with a solid insertion onto the skull base. After identification of the anterior digastric muscle and its tendon, adjust the Weitlaner retractor to further retract the muscle belly.
The exposed region is the three-dimensional space in which the main trunk of the facial nerve lies. Dissect along the nerve in an inferior direction, distally from the exit of the stylomastoid foramen. For a simple transaction, grasp the immediate epineurium overlying the nerve with fine-toothed forceps, and use sharp micro-scissors to cleanly transect the nerve at the desired point with a single cut.
After the experimental injury has been delivered, irrigate the wound with sterile saline, and dry the tissue with sterile gauze, before closing the skin incision according to institutional guidelines.

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All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. Anesthesia and preparation
<ol>
	<li>Place the animal in the anesthesia chamber and induce general anesthesia via 1.8% isoflurane and 0.9 L/min oxygen.
	<ol>
		<li>Confirm an adequate plane of anesthesia via an assessment of spontaneous breathing and an evaluation of consciousness by assessing the animal&#39;s grimace response to a toe pinch.</li>
	</ol>
	</li>
	<li>Apply eye lubricant bilaterally to guard against corneal irritation or dryness.</li>
	<li>Shave the operative site(s) with a razor or automatic clipper.
	<ol>
		<li>Establish a method for rat identification at this time, either via an ear tag or tail label/marking.</li>
	</ol>
	</li>
	<li>Administer a subcutaneous injection of 0.05 mg/kg buprenorphine along the animal&#39;s back for prophylaxis against postoperative pain.</li>
</ol>
2. Surgical approach and injury patterns
<ol>
	<li>Transfer the animal to the operating table and continue the gas flow via a nosecone. Ensure that a warming pad is positioned underneath the animal and the sterile field to maintain its body temperature.</li>
	<li>Place sterilized gauze (rolled up and fastened with tape) on the rat&#39;s neck to use as a neck roll; this will enhance the surgical field&#39;s exposure. Note that the animal&#39;s appropriate positioning is paramount for efficient nerve identification and dissection.</li>
	<li>Prepare the animal&#39;s facial skin for the procedure. To ensure disinfection, scrub the surgical site three times with chlorhexidine or an iodine-based solution alternating with 70% ethanol.</li>
	<li>Plan and mark the surgical incision if desired. Manipulate the ipsilateral ear in an anterior-posterior direction to determine the natural folding of the postauricular skin.</li>
	<li>Fashion a 4-5 mm incision in the postauricular crease using sharp iris scissors or a number 15 blade. This can be expanded later in the procedure as necessary.</li>
	<li>Bluntly dissect through the immediate subcutaneous fascia and place a micro-Weitlaner retractor to enhance exposure. Note that there may be small caliber blood vessels in this area; these are best avoided by retracting superiorly or inferiorly via the Weitlaner retractor.</li>
	<li>Identify the anterior digastric muscle as it travels in an inferior-to-superior direction toward its insertion along the skull base.
	<ol>
		<li>Spread gently through the muscle belly along its insertion point to reveal the tendon of the anterior digastric belly. Note that the tendon appears as a filmy white process emanating from the muscle with a solid insertion onto the skull base.</li>
	</ol>
	</li>
	<li>After identification of the anterior digastric muscle and its tendon, adjust the Weitlaner retractor to further retract the muscle belly. Note that the subsequently exposed region is the three-dimensional space where the main trunk of the facial nerve lies. 
	NOTE: This region is bounded superiorly and medially by the skull base, laterally by the anterior digastric muscle, posteromedially by the ear canal, and inferiorly by the structures of the neck, including the superficial temporal artery.</li>
	<li>After adequate exposure, identify the main trunk of the facial nerve as it travels inferiorly from underneath the tendon of the digastric muscle, where it exits the stylomastoid foramen from the skull base. Note that the nerve appears as a pearly white cord encased in the animal&#39;s parotid-masseteric fascia. Practice caution when further exposing the nerve for the following reasons.
	<ol>
		<li>Avoid aggressive dissection, or perpendicular spreads, to guard against stretch-mediated neuropraxia injury.</li>
		<li>Avoid aggressive posteriorly and medially directed dissection to guard against violating the thin tissues overlying the ear canal, as this could introduce middle ear flora into the surgical field.</li>
		<li>Avoid damaging the superficial temporal artery through broad medially and inferiorly directed dissection. Note that an injury will be identified by brisk, pulsatile bleeding.
		<ol>
			<li>If the artery is injured, apply prompt pressure with a cotton-tipped applicator or sterile gauze via forceps. Hemostatic agents or liquid fibrin sealant can be placed in near proximity. Keep in mind that the animal may require a subcutaneous injection of 0.9% sterile saline for fluid stabilization.</li>
		</ol>
		</li>
	</ol>
	</li>
	<li>Trace the main trunk distally by dissecting along the nerve in an inferior direction, distally from the exit of the stylomastoid foramen.
	<ol>
		<li>Extend the original incision to expose the nerve and its branches fully. Avoid disrupting the parotid gland, as this could result in postoperative sialocele.</li>
	</ol>
	</li>
	<li>Induce the desired injury patterns as follows.
	<ol>
		<li>For a crush injury, use smooth-surfaced jeweler&#39;s forceps to firmly grasp the nerve and compress it9. Apply constant and reproducible pressure to the nerve for 30 seconds to ensure an appropriate crush injury.</li>
		<li>For a simple transection, grasp the fascia overlying the nerve, or the immediate epineurium, with fine-toothed forceps, and use sharp microscissors to cleanly transect the nerve at the desired point with a single cut. Take care to avoid excess traction on the nerve with the forceps.</li>
		<li>For a nerve gap model, create the desired nerve gap using a similar method to the simple transection injury. Use the sterilized shaft of a cotton-tipped applicator cut to the desired nerve gap length-intraoperatively to ensure similarity of injury pattern between animals.</li>
	</ol>
	</li>
</ol>
3. Wound closure
<ol>
	<li>Irrigate the wound with sterile saline and dry it with sterile gauze.</li>
	<li>Approximate the skin edges in a simple, subcuticular fashion with absorbable sutures, or use skin glue or wound clips, which are also acceptable for wound closure. Place a buried stitch by taking a deep-to-superficial bite of one skin edge and then a subsequent superficial-to-deep bite of the opposite skin edge.</li>
</ol>

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>1.8% isoflurane</td>
			<td>VetOne</td>
			<td>13985-030-40</td>
			<td></td>
		</tr>
		<tr>
			<td>11-0 nylon microsutures</td>
			<td>AROSuture</td>
			<td>TK-117038</td>
			<td></td>
		</tr>
		<tr>
			<td>4-0 monocryl suture</td>
			<td>VWR</td>
			<td>75982-084</td>
			<td></td>
		</tr>
		<tr>
			<td>Buprenorphine SR</td>
			<td>ZooPharm</td>
			<td>MIF 900-006</td>
			<td></td>
		</tr>
		<tr>
			<td>Carprofen</td>
			<td>Sigma-Aldrich</td>
			<td>MFCD00079028</td>
			<td></td>
		</tr>
		<tr>
			<td>Chlorhexidine</td>
			<td>VWR</td>
			<td>IC19135805</td>
			<td></td>
		</tr>
		<tr>
			<td>Jeweler forceps</td>
			<td>VWR</td>
			<td>21909-458</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro Weitlaner retractor</td>
			<td>VWR</td>
			<td>82030-146</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro-scissors</td>
			<td>VWR</td>
			<td>100492-348</td>
			<td></td>
		</tr>
		<tr>
			<td>Mini tenotomy scissors</td>
			<td>VWR</td>
			<td>89023-522</td>
			<td></td>
		</tr>
		<tr>
			<td>Number 15 scalpel blade</td>
			<td>VWR</td>
			<td>102097-834</td>
			<td></td>
		</tr>
		<tr>
			<td>Operating microscope</td>
			<td>Leica</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Petrolatum eye gel</td>
			<td>Pharmaderm</td>
			<td>B002LUWBEK</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile water</td>
			<td>VWR</td>
			<td>89125-834</td>
			<td></td>
		</tr>
		<tr>
			<td>Tissue adhesive</td>
			<td>Vetbond, 3M</td>
			<td>NC9259532</td>
			<td></td>
		</tr>
		<tr>
			<td>Water conductor pad</td>
			<td>Aqua Relief System</td>
			<td>ARS2000B</td>
			<td></td>
		</tr>
	</tbody>
</table>

Source: Ali, S. A., et al. <a href="https://app.jove.com/v/59224">Facial Nerve Surgery in the Rat Model to Study Axonal Inhibition and Regeneration.</a> J. Vis. Exp. (2020)
This video demonstrates a surgical protocol on a rat model, detailing steps from anesthesia preparation to facial nerve injury induction. This protocol is useful for studying nerve regeneration and surgical techniques in medical research.

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 and shave the surgical area.
Place a gauze roll under the rat&#39;s neck to enhance exposure to the surgical field.
Disinfect the site with antiseptic.
Position the rat under a stereomicroscope.
Make a small incision behind the ear and dissect the superficial connective tissue.
Use a retractor to access the underlying tissues.
Identify the anterior digastric muscle, a type of jaw muscle. Spread this muscle to expose its tendon.
Adjust the retractor to reveal the facial nerve&#39;s trunk.
Dissect the nerve inferiorly to ensure clear accessibility for inducing injury.
Grasp the epineurium, a thin tissue layer overlying the nerve, and cut the target nerve at the desired site to create a transection injury.
Then, rinse the wound with saline, dry it, and suture the incision.
The rat model with a facial nerve injury is now ready for further studies.

19 ビュー. ラットモデルにおける顔面神経損傷の誘発

ビデオ: ラットモデルにおける顔面神経損傷の誘発 - 実験

Facial Nerve Axotomy in Mice: A Model to Study Motoneuron Response to Injury

The goal of this surgical protocol is to expose the facial nerve, which innervates the facial musculature, at its exit from the stylomastoid foramen and either cut or crush it to induce peripheral nerve injury. Advantages of this surgery are its simplicity, high reproducibility, and the lack of effect on vital functions or mobility from the subsequent facial paralysis, thus resulting in a relatively mild surgical outcome compared to other nerve injury models. A major advantage of using a cranial nerve injury model is that the motoneurons reside in a relatively homogenous population in the facial motor nucleus in the pons, simplifying the study of the motoneuron cell bodies. Because of the symmetrical nature of facial nerve innervation and the lack of crosstalk between the facial motor nuclei, the operation can be performed unilaterally with the unaxotomized side serving as a paired internal control. A variety of analyses can be performed postoperatively to assess the physiologic response, details of which are beyond the scope of this article. For example, recovery of muscle function can serve as a behavioral marker for reinnervation, or the motoneurons can be quantified to measure cell survival. Additionally, the motoneurons can be accurately captured using laser microdissection for molecular analysis. Because the facial nerve axotomy is minimally invasive and well tolerated, it can be utilized on a wide variety of genetically modified mice. Also, this surgery model can be used to analyze the effectiveness of peripheral nerve injury treatments. Facial nerve injury provides a means for investigating not only motoneurons, but also the responses of the central and peripheral glial microenvironment, immune system, and target musculature. The facial nerve injury model is a widely accepted peripheral nerve injury model that serves as a powerful tool for studying nerve injury and regeneration.

We present a surgical protocol detailing how to perform a cut or crush axotomy on the facial nerve in the mouse. The facial nerve axotomy can be employed to study the physiological response to nerve injury and test therapeutic techniques.

マウスにおける顔面神経軸索切断：傷害に運動ニューロン応答を研究するためのモデル

The goal of this surgical protocol is to expose the facial nerve, which innervates the facial musculature, at its exit from the stylomastoid foramen and ...

JoVE Journal

医学

Functional and Physiological Methods of Evaluating Median Nerve Regeneration in the Rat

The main goal of this investigation is to show how to create and repair different types of median nerve (MN) lesions in the rat. Moreover, different methods of simulating postoperative physiotherapy are presented. Multiple standardized strategies are used to assess motor and sensory recovery using an MN model of peripheral nerve lesion and repair, thus permitting easy comparison of the results. Several options are included for providing a postoperative physiotherapy-like environment to rats that have undergone MN injuries. Finally, the paper provides a method to evaluate the recovery of the MN using several noninvasive tests (i.e., grasping test, pin prick test, ladder rung walking test, rope climbing test, and walking track analysis), and physiological measurements (infrared thermography, electroneuromyography, flexion strength evaluation, and flexor carpi radialis muscle weight determination). Hence, this model seems particularly appropriate to replicate a clinical scenario, facilitating extrapolation of results to the human species.
Although the sciatic nerve is the most studied nerve in peripheral nerve research, analysis of the rat MN presents various advantages. For example, there is a reduced incidence of joint contractures and automutilation of the affected limb in MN lesion studies. Furthermore, the MN is not covered by muscle masses, making its dissection easier than that of the sciatic nerve. In addition, MN recovery is observed sooner, because the MN is shorter than the sciatic nerve. Also, the MN has a parallel path to the ulnar nerve in the arm. Hence, the ulnar nerve can be easily used as the nerve graft for repairing MN injuries. Finally, the MN in rats is located in the forelimb, akin to the human upper limb; in humans, the upper limb is the site of most peripheral nerve lesions.

Presented is a protocol to produce different types of median nerve (MN) lesions and repair in the rat. Additionally, the protocol shows how to evaluate the functional recovery of the nerve using several noninvasive behavioral tests and physiological measurements.

ラットの神経再生の中央値を評価する機能と生理学的方法

The main goal of this investigation is to show how to create and repair different types of median nerve (MN) lesions in the rat. Moreover, different ...

The Superficial Inferior Epigastric Artery Axial Flap to Study Ischemic Preconditioning Effects in a Rat Model

Fasciocutaneous flaps (FCF) have become the gold standard for complex defect reconstruction in plastic and reconstructive surgery. This muscle-sparing technique allows transferring vascularized tissues to cover any large defect. FCF can be used as pedicled flaps or as free flaps; however, in the literature, failure rates for pedicled FCF and free FCF are above 5%, leaving room for improvement for these techniques and further knowledge expansion in this area. Ischemic preconditioning (I.P.) has been widely studied, but the mechanisms and the optimization of the I.P. regimen are yet to be determined. This phenomenon is indeed poorly explored in plastic and reconstructive surgery. Here, a surgical model is presented to study the I.P. regimen in a rat axial fasciocutaneous flap model, describing how to safely and reliably assess the effects of I.P. on flap survival. This article describes the complete surgical procedure, including suggestions to improve the reliability of this model. The objective is to provide researchers with a reproducible and reliable model to test various ischemic preconditioning regimens and assess their effects on flap survivability.

This protocol describes harvesting, suturing, and monitoring fasciocutaneous flaps in rats that allow for good visualization and manipulation of blood flow through the superficial inferior epigastric vessels by means of clamping and ligating the femoral vessels. This is critical for studies involving ischemic preconditioning.

ラットモデルにおける虚血性プレコンディショニング効果を研究するための表在性下上腹部動脈軸フラップ

Fasciocutaneous flaps (FCF) have become the gold standard for complex defect reconstruction in plastic and reconstructive surgery. This muscle-sparing ...

Inducing a Facial Nerve Injury in a Rat Model

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Inducing a Facial Nerve Injury in a Rat Model