The overall goal of this procedure is to induce a cervical unilateral contusion model using the infinite horizon impactor. This is accomplished by first constructing a custom designed frame and clamp. Next, the cervical spine is mounted onto the clamp.
The animal is then positioned in the impactor in preparation for spinal cord injury. The final step of the procedure is to injure the cervical spinal cord. This procedure produces a consistent injury model with sustained and sufficient functional deficits.
Ultimately, the reproducibility and low morbidity of this injury model reduces the number of animals used for spinal cord injury research. Demonstrating the procedure will be j Lee, a research associate from my laboratory, who has really championed the development of this model over the past few years In order to deliver consistent and reproducible biomechanical forces to the spinal cord. This procedure uses the infinite horizon or IH spinal cord injury, impactor, a clamp and a frame for holding.
The animal was custom designed and built to accommodate the IH impactor. After confirming that the animal has reached a surgical plane of anesthesia, place the animal in a stereotaxic frame and carefully dissect through the dorsal musculature to expose the spine. Once the C four to C six lamina are exposed, Macon an incision underneath the transverse processes on both sides of the spine in order to fit the clamp under the transverse processes of C four to C six with a fine tipped Friedman Pearson ur.
Carefully remove the left C five lamina to visualize the dura and the spinal cord to facilitate clamp insertion. Slide a 1.5 centimeter diameter rod under the arms to prop the animal up and raise the spine, Mount the jaw of the clamp onto lateral transverse processes of C four to C six, and then tighten the screws. Insert the clamp into the metal holders on the two middle rods of the frame that are secured at a 22 and a half degree angle.
Raise the scissor jack until the animal lays flat. Ensure that the clamp is horizontal by placing a small cylinder level on top of the clamp. And by tightening the screws, it is important that all the screws are tight and the setup is rigid without any movement.
Lower and aim the impactor tip using the vertical adjustment knob and the two horizontal adjustment knobs on the IH impactor until the center of the impactor tip is hovering above the apex of the C six spinous process. Once the impactor tip is centered, turn the Y axis adjustment knob 1.4 millimeters to move the tip laterally to the left side. Use the x-axis horizontal adjustment knob to move the tip to the center of C five.
Next, lower the tip until it is just above the dura and the impactor tip should be aimed at the lateral half of the gray matter. Turn the vertical adjustment knob to turns to raise the tip four millimeters above the dura. Ensure that the impact area is dry by using a cotton swab on the IH impactor software.
Set the desired force to 150 kilodons on the program and select start experiment to trigger the impact. This animation illustrates on a cross section of the cervical spinal cord how a unilateral injury is produced at a force setting of 150 kilodons and an angle of 22 and a half degrees. The force versus time graph is shown as an inset.
These graphs show the recorded time versus displacement and time versus force of a typical 150 kilo D contusion delivered with the IH impactor. The red arrow indicates the time the impactor has reached 20 kilodons and the recording of displacement has been initiated. The recordings show that the actual force achieved was 152 kilodons, and the impact or displacement into the cord was measured to be 1, 287 micrometers.Here.
The results of four behavioral tests showed that the spinal cord injury resulted in statistically significant four limb impairments sustained throughout the experimental period, restricted only to the ipsilateral for limb. In all cases, the impairment was sustained for a full six weeks after injury. A series of spinal cord sections that traverse a typical 150 kilo nine contusion shows that the injury resulted in damage to the gray and white matter on the ipsilateral side.
Both rubrospinal and corticospinal tracts on the injury side are included in the injury area. However, some sparing of axons in these tracks cannot be ruled out. More than 80%of the animals had parenchymal damage contained on the ipsilateral side.
The quantification of additional sections provides a gross estimation of the cumulative spread of white and gray matter sparing, which in this case is 2, 400 micrometers, rostral, and coddle to the epicenter of injury. The ipsilateral side had only 51.8%of the white matter s spared and 39.7%of gray matter remaining compared to the contralateral side. After watching this video, you should have a good understanding of how to produce consistent cervical unilateral spinal cord injuries using the infinite horizon impactor.
This injury model will not only provide a valuable tool for testing potential preclinical therapies for spinal cord injury, but the reproducibility of this injury model will also enable us to reduce the number of animals that are utilized for this type of spinal cord injury research. Another aspect of this model, which we feel is very appealing, is the fact that the morbidity for the animals is minimized. The deficits are limited to one forelimb, and we think that these efforts to minimize morbidity is an important aspect of doing preclinical spinal cord injury research.
