The Rabbit Blood-shunt Model for the Study of Acute and Late Sequelae of Subarachnoid Hemorrhage: Technical Aspects

The overall goal of this procedure is to establish an examiner independent intracranial pressure, or ICP controlled blood shunt, acute subarachnoid hemorrhage, or SAH model in the rabbit. This is accomplished by first cannulating the subclavian artery. In the second step, digital subtraction angiography is performed, and then neuromonitoring probes and a spinal access needle are placed in the final step.

The spinal access needle and the subclavian artery are connected to a shunt to induce examiner, independent ICP controlled acute SAH, ultimately the ICP cerebral blood flow, or CBF and cardiorespiratory response of the animal to the acute SAH can be monitored. This method may help answer key questions in the neurovascular field, such as how early brain injury and delayed cerebral vessel specimen contributes to un farrow outcome and mortality. Following aneurysmal subic hemorrhage Begin by placing the sedated rabbit in a supine position with both front and hind legs fixed to the operating surface.

Install a three channel electrocardiogram as such. Place three subcutaneous electrodes in a triangular form on the ventral side of the rabbit. Next, disinfect the skin with povidone iodine or similar over the right pictoral muscle around the middle third of the clavicle, cover the animal with sterile sheets.

After infiltrating the pectoral muscle with local anesthetics, make a lateral paramous skin incision and expose the pectoral muscle. Split the muscle fiber superiorly and inferiorly by blunt dissection until the subclavian artery can be visualized. Then under the microscope, dissect the subclavian artery using two straight micro forceps and then ligate it distally, and place a ligature around the exposed proximal end using four O polyfilament sutures and a curved micro forceps.

Next, use a curved micro scissor to incise the wall of the subclavian artery and cannulate the artery retrograde. Then secure the catheter by double knot ligature towards the proximal ligature, and additionally, secure the catheter by double knot ligature towards the distal ligature in order to prevent arterial twisting or bending of the proximal parts of the artery, and to avoid slippage or massive bleeding. Then collect blood samples for the arterial blood gas analysis status, and continuously monitor the standard cardiovascular and respiratory parameters.

Place an external sizing device over the angles of the mandible to calibrate the angiogram and to allow an exact comparison of the post hoc measurements of the baseline and follow up vessel diameter. Then perform retrograde intra arterial bolus injection of non ionic IOP IOL through the cannulated artery, and immediately flush the cannula with saline to prevent occlusion of the ladder. Following baseline digital subtraction angiography, reposition the rabbit to the prone position, taking care not to manipulate or relocate the intra arterial catheter.

Connect the arterial blood pressure monitoring and start to continuously monitor the standard cardiovascular and respiratory parameters, including electrocardiogram, arterial blood pressure, peripheral oxygen saturation, expiratory carbon dioxide, and core temperature. Transfer the data via the analog output interface to an analog digital converter data logger and store. Place the head in a holder at a 30 degree angle oriented down, and then disinfect the skin over the head and neck with povidone iodine three times for one minute each.

After covering the new surgical area with sterile sheets, make a midline skin incision. Insert a small surgical retractor to make three round osteotomies according to outer skull landmarks and calibrate the neuromonitoring probes. Place the intercranial pressure and the inter parenchymal laser doppler blood flow neuromonitoring tips and seal the bur holes with bone wax to keep the cranium fluid tight and to allow for a closed cranium condition in order to keep the steady state of I-C-P-C-P-P and MAP.

Next, insert a 22 gauge 40 millimeter pediatric spinal access needle transcutaneous into the cisterna magna. Then slide the needle down along the bony external occipital protuberance until a gap is detected. Confirm the correct positioning of the needle by observing the spontaneous dripping of cerebral spinal fluid with the rabbit’s head tilted down at a 20 to 30 degree angle for a few minutes.

Now, use a blood filled pressure monitoring tube to connect the spinal access needle in the cisterna magna to the three-way stop cock attached to the previously catheterized subclavian artery. Then open the shunt connection between the subclavian artery and the CI sternum magna to induce subarachnoid hemorrhage. After opening the shunt intracranial pressure starts to increase to values close to the diastolic blood pressure.

At that time, the flow in the shunt stops if the intercranial plateau is maintained for more than 10 seconds, or if the pressure decreases spontaneously, close the shunt after the intercranial pressure reaches its peak, keep the spinal access needle in place until the pressure returns to a steady state close to baseline values. Finally, remove the cerebral blood flow fine needle and intracranial pressure probes. Plug the bur holes with bone wax, then use neomycin sulfate to perform a rigorous wound irrigation and suture the skin.

First, suture the skin over the bur holes. Then after repositioning of the rabbit in supine position and removal of the subclavian artery catheter, suture the pectoral muscle and the skin. The rabid blood shunt model of SAH described in this report produces early brain injury in the hippocampus basal cortex, and cerebral vasculature as early as 24 hours after injury, inducing a characteristic blood distribution.

In addition, this model triggers moderate to severe degrees of delayed cerebral vasospasm of day three. After SAH induction, the mortality rate of the procedure is approximately 20%due to respiratory arrest or severe bradycardia at the time of acute SAH during which the animals are under complete anesthesia. From a technical point of view, this blood shunt model allows for both examiner independent and controllable SAH induction.

While attempting this procedure, it’s important to remember to firmly secure this sub clavon artery, take great car and repositioning the animal, and be very gentle and patient during Cy Sternum acupuncture.