Understanding the pathophysiology of ischemic stroke is limited due to the absence of high quality tissue data in humans. Since such samples are impossible to acquire, well-controlled animal model studies can serve as a surrogate. The rabbit model used in this protocol provides consistent high-quality data for the interrogation of ischemic stroke.
The technique described here allows for precise control and modulation of hemodynamic factors to assess their impact on ischemic stroke pathophysiology. A well-planned procedure can lead to high-quality results, particularly by employing strategies to mitigate vasospasm in the arteries, through a combination of pharmacologic prophylaxis and effective angiographic techniques. To begin, place the rabbit in a supine position on a fluoroscopy compatible operative table.
Extend the head as it optimizes positioning for subsequent angiographic views. To mitigate vasospasm, place 0.5 inches of transdermal nitroglycerin on the inside of the ear after induction of anesthesia. After removing the fur from both inguinal regions using electric clippers, prepare the skin with scrubs of chlorhexidine and alcohol.
Drape the skin in the usual sterile fashion, and palpate the bilateral femoral arterial pulses. Make a five-centimeter surgical incision with a number 10 blade at the site where lidocaine was injected. Use blunt dissection to expose the neurovascular bundle, and if needed, extend the incision to expose an arterial segment large enough for access.
Upon isolation of the neurovascular bundle, drop several drops of 1%lidocaine on the artery to prevent vasospasm. Separate the artery gently from the vein and the adjacent nerve using forceps. Identify the artery by the characteristic appearance of its muscular wall compared to the thin walls of the vein.
After isolating the artery, pass the right angle forceps under the vessel. Then grasp two vessel loops with the instrument and gently pass them under the artery. Situate one each at the upstream and downstream ends of the exposed vessel.
Subject the artery to gentle traction by pulling the vessel loops, and inspect the vessel for any residual tissue to be removed with gentle dissection, increasing the chances of successful access. After dissecting the vessel and preparing the angiocatheter, drip lidocaine on the vessel again. The artery visibly dilates, increasing the chances of successful access and placement of a sheath using the Seldinger technique.
Apply gentle traction to the downstream vessel loop to engorge the artery by reducing outflow and stabilizing the vessel for the access attempt. Next, slowly advance the needle of the angiocatheter into the middle of the exposed arterial segment. On seeing a flash of blood in the angiocatheter and the hub chamber, advance the catheter into the arterial lumen.
On successfully placing the angiocatheter, advance a cope microwire through the angiocatheter lumen and into the aorta. Then remove the angiocatheter over the wire and replace it with a five French slim hydrophilic sheath. Confirm the return of the arterial blood through the sidearm tubing by opening the three-way valve.
Lock the valve closed, and flush the sheath with 0.9%saline. Secure the sheath hub to the adjacent skin with an additional 3.0 silk suture, and repeat this process for the contralateral femoral artery. Under fluoroscopic visualization, advance a four French glide catheter over a 0.035-inch glide wire inserted through the left femoral sheath.
Positioned the tip of the glide catheter in the proximal left vertebral artery. After removing the wire, flush the catheter with heparinized 0.9%saline. Perform angiography by injecting the left vertebral artery with iodinated contrast under lobe magnification to visualize the entire head and neck.
For the left vertebral injection, inject 50%contrast, diluted in normal saline, with a gentle crescendo from a three CC syringe. Determine the injection amount by checking the reflux down the right vertebral artery and into the right subclavian artery. During this injection, also note the posterior cerebral artery or superior cerebral artery, one of which will be the target to occlude with the microcatheter.
Prepare a 2.4 French flow-directed microcatheter with a 0.010 inch microwire, and make a C shape on the tip of the microwire. Under roadmap guidance, advance the microcatheter inside a four French glide catheter using the right femoral sheath and over the wire into the right vertebral artery. Advance the microcatheter through the cervical segment of the left vertebral artery.
To pass the sharp turn as it's best from the V2 to V3 segment, advance the microcatheter alone while the microwire is back proximal to its tip. After passing the sharp turn from V2 to V3, the microcatheter often passes easily to the proximal basal or artery. At this point, advance the microwire and select the desired posterior cerebral and superior cerebellar arteries, as microcatheter injections are not advisable due to the fragile nature of the intracranial arteries.
Further advance the microcatheter over the microwire into the target artery and choose a proximal position, as it is typically safest in the posterior to communicate due to its angulation at its origin. A deeper position is feasible in the superior cerebellar artery. Repeat the angiogram by injecting the left vertebral artery catheter with high magnification over the head to confirm occlusion of the target artery.
For optimal imaging, inject full-strength contrast, typically no more than one CC, which is adequate for pacification of all intracranial arteries. Gently remove the microwire from the microcatheter under fluoroscopic visualization to confirm a stable position. Place a stopcock on the hub of the microcatheter and close the stopcock to prevent blood loss from retrograde blood flow.
Remove the left vertebral catheter to make the left femoral access sheath available. During the ensuing occlusion period, acquire intermittent fluoroscopic images to confirm the stable position of the occlusive microcatheter. Remove the occlusive microcatheter after three hours.
And then continue arterial blood pressure measurement and modulation for the additional desired period. Inflate and deflate the balloon catheter in the aorta to monitor the blood pressure variability. In acute procedures with immediate brain harvesting, remove the calvarium in a piecemeal fashion with rongeurs, starting at the occipital ridge and working anteriorly until the brain can be harvested intact.
Place the brain in formalin or flash freeze, depending on the type of tissue analysis desired. In all the animals, the brain was successfully harvested and histopathological analysis was carried out, demonstrating infarction in the right cerebellum. The blood pressure was traced from a Fogarty balloon catheter placed in the infrarenal aorta, and the data from approximately one hour of monitoring demonstrates real-time arterial pressure changes with changes in balloon inflation.
Short-term tracing of the blood pressure demonstrates the pressure changes throughout the cardiac cycle. Additionally, small rapid changes were noted from respiratory variability, which is physiologically normal. An immediate near-doubling of measured blood pressure was noted following the inflation of the balloon.
While attempting this procedure, meticulous blunt dissection and dripping lidocaine to prevent vasospasm during access of the angiocatheter is critical. Also, angiography of the left vertebral artery provides a roadmap for efficient endovascular access to the targeted intracranial artery. Hemodynamics can be modulated in pharmacological ways or by inflating a balloon catheter in the aorta.
The harvested brain specimens can be analyzed with a variety of techniques, such as histopathology or spatial transcriptomics.
