This article outlines a surgical procedure to create superficial spinal cord injuries in anesthetized mice. The method focuses on the activation of neural stem cells for tissue repair following injury.
Take an anesthetized mouse and shave its back.
Secure it in a stereotaxic device, with rolled gauze positioned to support the thoracic spine.
Disinfect the skin, make a vertical midline incision, and retract the skin.
Detach the muscle tendons from the thoracic spine and retract them.
Identify the target vertebra, then sever the intervertebral joints to remove it.
Expose the spinal cord and insert a bent needle on either side of the midline.
The needle insertion creates superficial injuries in the white matter, the outer spinal cord layer containing axon bundles, while leaving the central canal intact.
Remove the needle, then suture the muscles and skin.
Damaged cells release signals that activate tissue-resident immune cells, which in turn activate neural stem cells, or NSCs, surrounding the central canal.
Activated NSCs migrate to the injury site, aiding tissue repair.
Before beginning the procedure, use hair clippers to remove the fur from the dorsum of an anesthetized mouse from the mid-back to the neck and ears. After securing the mouse in a stereotaxic device, place four to five pieces of rolled gauze beneath the abdomen, while lightly pulling on the base of the tail to straighten out the body and spine.
Push the thoracic gauze roll rostrally from the mouse abdomen toward the upper thorax to support the thoracic spine. And use laboratory tape to secure the tail and limbs in a star like orientation. After disinfecting the exposed skin with sequential 70% ethanol and povidone iodine swabs, use a number 10 scalpel blade to make a vertical incision parallel to the longitudinal axis of the animal, from the midpoint of both shoulder blades to the curvature of the thoracic spine.
Retract the skin to expose the soft tissue and the spinal column contour, and identify the lower border of the suprascapular fat pad. Then, use the scalpel to carefully, but forcefully cut along both sides of the vertebral T5 to T8 and 9 bones to detach the back muscle tendons from the spinal column. Reposition retractors so that the teeth of the retractors are inserted into the incision site on either side of the spine, and expand each retractor to adjust the exposure until the spine is sufficiently elevated, without putting too much strain on the retracted muscle layers.
Under a surgical microscope, carefully clean the residual muscle and other soft tissue overlying the spine to expose the vertebral bone, clasping the spinous process of the vertebrae with toothed forceps, and moving the forceps slightly up and down to identify the vertebrae that will be removed. Next, insert one tip of a pair of curved, blunted scissors into either side of the exposed intervertebral foramen, caudal to the vertebral lamina to be excised, and cut the connecting intervertebral joints bilaterally. Then, lift the lamina upwards, and cut off the upper attachment of the lamina to isolate and remove the bone.
When cutting away and removing the vertebral lamina, take care to always angle the scissors upwards, so not as to damage the underlying spinal cord.
With the dorsal midline vein serving as a landmark, insert the 45-degree bent shaft of a 30-gauge needle tip bevel side up, 1 millimeter deep into the dorsolateral surface of the spinal cord, and approximately 0.5 millimeters lateral to either side of the midline. Move the needle about two millimeter from the caudal to rostral direction, so that the entire length of the bevel is inserted into the cord. Then, retrace the path of the entry to remove the needle.
To close the wound, remove the retractor, and use A 6-0 absorbable suture to join the back muscles on either side of the injury along the midline, and and a 4-0 silk suture to close the overlying skin.
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