Name: murine model of controlled cortical impact for the induction of traumatic brain injury
Uploaded: 2019-08-16T14:45:00
Description: Watch this Scientific Journal Video about Murine Model of Controlled Cortical Impact for the Induction of Traumatic Brain Injury at JoVE.com

This work was supported by National Institutes of Health Grant GM117341 and The American College of Surgeons C. James Carrico Research Fellowship to S.J.S.

All procedures were approved by the Northwestern University Institutional Animal Care and Use Committee. C57BL/6 mice were purchased from the Jackson Laboratory and group housed at a barrier facility at the Center for Comparative Medicine at Northwestern University (Chicago, IL). All animals were housed in 12/12 h light/dark cycle with free access to food and water.
1. Induce anesthesia
<ol>
	<li>Anesthetize the mouse with ketamine (125 mg/kg) and xylazine (10 mg/kg) injected intraperitoneal...

<ol>
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There are several steps that are critical for applying a reliable and consistent injury. First, the mouse must reach a deep plane of surgical anesthesia ensuring no movement during the performance of the craniectomy. While numerous anesthetic regimens may be used to induce general anesthesia in rodents, anesthetics that induce respiratory depression such as inhalational anesthetics may result in respiratory arrest when combined with a severe TBI. This protocol utilizes ketamine (125 mg/kg) and xylazine (10 mg/kg) injecte...

The Centers for Disease Control and Injury Prevention estimate that approximately 2 million Americans sustain a traumatic brain injury (TBI) every year1,2. In fact, TBI contributes to over 30% of all injury related deaths in the United States with healthcare costs nearing $80 billion annually and almost $4 million per person per year surviving a severe TBI3,4,5. The impact of TBI is highlighted by the significant long-term neurocognitive and neuropsychiatric complications suffered by its survivors with the insidious onset of behavioral, cognitive, and motor impairments termed Chronic Traumatic Encephalopathy (CTE)6,7,8,9,10. Even subclinical concussive events&#8212;those impacts that do not result in clinical symptoms&#8212;can lead to long-term neurologic dysfunction11,12.
Animal models for the study of TBI have been employed since the late 1800&#8217;s13. In the 1980s, a pneumatic impactor for the purpose of modeling TBI was developed. This method is now referred to as controlled cortical impact (CCI)14. The control and reproducibility of CCI led researchers to adapt the model for use in rodents15. Our laboratory uses this model to induce TBI via a commercially available impactor and electronic actuating device16,17. This model is capable of producing a wide range of clinically applicable TBI states depending on the biomechanical parameters used. Histologic evaluation of TBI brains after a severe injury induced in our laboratory demonstrates significant ipsilateral cortical and hippocampal loss as well as contralateral edema and distortion. Additionally, CCI produces a consistent impairment in motor and cognitive function as measured by behavioral assays18. Limitations to CCI include the need for craniotomy and the expense of acquiring the impactor and actuating device.
Several additional models of TBI exist and are well established in the literature including the lateral fluid percussion model, weight drop model, and blast injury model19,20,21. While each of these models have their own distinct advantages their main drawbacks are mixed injury, high mortality and lack of standardization, respectively22. Furthermore, none of these models offer the accuracy, precision, and reproducibility of CCI. By adjusting the biomechanical parameters input into the actuating device, the CCI model allows the investigator precise control over size of the injury, depth of the injury, and kinetic energy applied to the brain. This gives investigators the ability to apply the entire spectrum of TBI to specific areas of the brain. It also permits the greatest reproducibility from experiment to experiment.

<table>
	<tbody>
		<tr>
			<td>AnaSed Injection Xylazine Sterile Solution</td>
			<td>LLOYD, Inc.</td>
			<td>5939911020</td>
		</tr>
		<tr>
			<td>Buprenorphine SR Lab 0.5mg/mL</td>
			<td>Zoopharm-Wildlife Pharmaceuticals USA</td>
			<td>BSRLAB0.5-182012</td>
		</tr>
		<tr>
			<td>High Speed Rotary Micromotor KiT0</td>
			<td>Foredom Electric Company</td>
			<td>K.1070</td>
		</tr>
		<tr>
			<td>Imapact one for Stereotaxix CCI</td>
			<td>Leica Biosystems Nussloch GmbH</td>
			<td>39463920</td>
		</tr>
		<tr>
			<td>Ketathesia Ketamine HCl Injection USP</td>
			<td>Henry Schein, Inc</td>
			<td>56344</td>
		</tr>
		<tr>
			<td>Mouse Specific Stereotaxic Base</td>
			<td>Leica Biosystems Nussloch GmbH</td>
			<td>39462980</td>
		</tr>
		<tr>
			<td>Trephines for Micro Drill</td>
			<td>Fine Science Tools, Inc</td>
			<td>18004-50</td>
		</tr>
	</tbody>
</table>

murine model of controlled cortical impact for the induction of traumatic brain injury

The Centers for Disease Control and Injury Prevention estimate that almost 2 million people sustain a traumatic brain injury (TBI) every year in the United States. In fact, TBI is a contributing factor to over a third of all injury-related mortality. Nonetheless, the cellular and molecular mechanisms underlying the pathophysiology of TBI are poorly understood. Thus, preclinical models of TBI capable of replicating the injury mechanisms pertinent to TBI in human patients are a critical research need. The controlled cortical impact (CCI) model of TBI utilizes a mechanical device to directly impact the exposed cortex. While no model can full recapitulate the disparate injury patterns and heterogeneous nature of TBI in human patients, CCI is capable of inducing a wide range of clinically applicable TBI. Furthermore, CCI is easily standardized allowing investigators to compare results across experiments as well as across investigative groups. The following protocol is a detailed description of applying a severe CCI with a commercially available impacting device in a murine model of TBI.

The impactor mounts directly on the stereotaxic frame allowing for as much as 10 &#181;m resolution for control of the point of impact, depth and penetration. The electromagnetic forces employed can impart impact velocities ranging 1.5&#8211;6 m/s. This allows for unparalleled precision and reproducibility over the entire range of clinically relevant TBI. Investigators can run pilot experiments changing the injury parameters such as impactor tip size, impact velocity, and impact depth to determine the parameters that bes...

Watch this Scientific Journal Video about Murine Model of Controlled Cortical Impact for the Induction of Traumatic Brain Injury at JoVE.com

Murine Model of Controlled Cortical Impact for the Induction of Traumatic Brain Injury

Here we describe a protocol for the induction of murine traumatic brain injury via an open-head controlled cortical impact.

The Centers for Disease Control and Injury Prevention estimate that almost 2 million people sustain a traumatic brain injury (TBI) every year in the ...

Controlled cortical impact is easy to standardize across subjects and experiments and allows for the application of the entire spectrum of traumatic brain injury to precisely defined regions of the brain. Demonstrating this procedure is Mecca Islam, a research technician in my laboratory. This technique remains the most consistent and reproducible form of inducing traumatic brain injury in rodents.Before beginning the procedure, attach the impacting device to the stereotaxic operating frame. Se the actuating device with the desired biomechanical parameters for the appropriate experimental velocity and well time. Don new personal protective equipment and sterile gloves and confirm a lack of response to toe pinch in the anesthetized experimental mouse.Use clippers to shave the fur for the surgical site and apply ointment to the animal's eyes. Then place the mouse into the operating theater and prep the shaved skin with three sequential iodine based and alcohol surgical scrubs. To induce a controlled cortical traumatic brain injury, first use a scalpel to make a one centimeter incision along the midline of the scalp to expose the skull.Retract the scalp from the operative site and identify the sagittal and coronal sutures on the exposed skull. To perform a craniectomy, equip a microdrill with a five millimeter trephine drill bit to activate the drill at maximum speed and apply the bit perpendicular to the skull two millimeters to the left of the sagittal suture and two millimeters rostral to the coronal suture with gentle even pressure. A slight give will be felt when the drill penetrates the skull.Use forceps and a small gauge hypodermic needle to remove the bone flap, fully exposing the underlying dura mater and rotate the impacter tip into the operative field. Secure the bilateral temporal bones between the miniature ear bars of the stereotaxic frame and lock the incisors within the incisor clamp to create a stable three point hold on the mouse head. Lower the tip until it makes contact with the exposed dura mater.The contact sensor of the instrument will make an audible tone to alert that contact has been made. Retract the impacting tip from the zero point and lower the impacter position on the stereotaxic frame to set the desired impact depth. Then active the impacter on the actuating device before rotating the device back out of the field to allow the animal to be removed from the frame.Immediately after the injury, apply direct pressure from a sterile cotton tipped applicator to the skull and injured cortical surface to control any bleeding and use a fresh applicator to dry the skull. Close the scalp over the craniectomy according to standard techniques and administer postoperative analgesia. Then place the animal in the lateral decubitus recovery position in a clean pre-warmed cage with monitoring until full recumbency and measure the body weight every three days throughout the course of the experiment.Delivery of the injury as demonstrated results in a subdural intraparenchymal and subarachnoid hemorrhage. Histologic evaluation of traumatic brain injury brain tissue sections after severe injury reveals a significant ipsilateral cortical and hippocampal loss as well as contralateral edema and distortion. MR imaging of severely injured brains also demonstrates progressive tissue loss and replacement by cerebra-spinal fluid.Controlled cortical impact induces a reliable and consistent injury that is capable of producing a wide range of clinically applicable brain injuries. Our controlled cortical impact method generates a murine model of severe traumatic brain injury for behavioral and imaging analysis and produces tissue for molecular and cellular analysis.

Research

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