The study utilizes castrated male Yorkshire swine (weighing 45–70 kg). Swine are all subject to an acclimatization period under the care of licensed veterinary staff for at least 48 hours prior to proceduralization. During to this time, animals have free access to food and water until the night before surgery when they were fasted overnight.
The animal protocol begins with animal preparation and instrumentation. This is followed by a baseline period where control values are collected followed by the experimental portion of the protocol, see Figure 1. Then we perform cooling with the ICEPICC device while continuing to monitor the animal 6 hours, followed by 2 hours of monitoring; lastly the animal is either awakened, or sacrificed and histology is sampled (shown in Figure 1).
I. Animal Preparation and Instrumentation:
1. anesthetize the animal with telazol (5mg/kg) and xylazine (2mg/kg) at appropriate doses.
2. transport the animal to the procedure area, with oxygen saturation monitoring and baseline temperature assessment.
3. place the animal under isoflurane targeting 1.0 MAC by facemask.
4. place the animal in sternal recumbency and intubate the animal with a 7.0 endotracheal tube. Transition to generally 10 ccs/kg TV, RR of 12-14 initially but to target a pCO2 of 30-45 and an FiO2 of 40% but adjusted appropriately as needed for the remainder of the cases.
5. make the animal supine and restrain.
6. Place all intravascular catheters using a percutaneous, ultrasound-guided, modified-Seldinger technique. Placement of the pressure catheters/angiographic catheters within the introducer sheaths is performed under direct fluoroscopic guidance. For this study, this consists of at least one 7 Fr. Catheter in a femoral artery through which we place an intra-aortic solid state pressure catheter for blood pressure monitoring, a 5-7 fr intravenous (femoral vein) catheter for IVF and medications as needed. A 7 Fr. Brachial access was obtained for intra-aortic contrast injection for CT perfusion studies. Due to anatomical considerations, the right brachial artery was preferentially cannulated over the left (Edwards et al., 2021).
7. Perform a lower abdominal laparotomy for cystostomy (place a foley catheter into the bladder) to facilitate bladder drainage.
8. Animals enrolled in the non-survival arm had a burr hole drilled and placed, those enrolled in the survival arm did not. Here, once animals are in sternal recumbency, a burr hole is drilled to allow placement of an intracranial temperature probe, laser doppler flowmetry probe, and an ICP probe; this was done in a standard fashion under CT guidance as previously described by our group
- Drill 2 burr holes with the Dremmel ~ 2 cm anterior to the posterior ridge, 1 cm to either side of midline. A third burr hole is made on the right anterior portion of the skull.
- Incise the dura underlying each burr hole with a scalpel. Place the Fogarty balloon into the anterior burr hole, ~ 1 cm deep, ensuring the balloon portion is within the skull. Place a 5Fr pressure catheter through the right posterior burr hole. Seal all burr holes around their respective probes/catheters with bone wax to ensure that there is no leakage of blood or CSF through the burr holes.
- Perform an additional helical CT scan to ensure proper placement of the intracranial instrumentation. Ensure that data is being recorded properly.
9. Perform a TIMEOUT. Confirm all line placements, confirm all sheaths work (drawback and flush), confirm fluids are ready, that the timer is ready and reset, that data is being transduced through LabChart through appropriately labeled channels and saved. Confirm ventilatory settings.
10. Confirm placement of all temperature probes and that they are functioning correctly: rectal, intracranial, middle ear, nasal.
10. Confirm fluoroscopically that all catheters and devices are appropriately positioned.
1. Heparinize the animal with 10k units of heparin
II. Begin Baseline normalization period: (60 min)
1. Once everything is confirmed then start the one hour baseline time period and collect temperature data throughout.
2. Perform baseline CT perfusion. This is a pre-determined protocol. Briefly, it is a 60 second scan that captures the same slice every second. It records the transit of contrast. As such, a power injector is utilized (2.5ml/sec for 7.5ml total) to inject contrast through the angiographic catheter in the proximal aorta. Ensure that contrast is observed transiting through the brain during the scan.
Throughout baseline and observation periods use the following guidelines:
- Treat glucose < 65 with D50
- for pH < 7.2 give on ampule of bicarbonate
- treat pCO2 as necessary with MV changes
In this phase we apply the ICEPICC intranasal cooler system and continuously monitor the animals for 4 hours, see figure 1. Throughout this period we monitor the parameters noted above. Subjects that are randomized to the cooling group then had the ICEPICC turned on, while those randomized to the control group will not. This period ends at the end of the 4 hour mark.
III. Post cooling and monitoring
1. Turn off the ICEPPICC device and monitor the animals for 2 more hours.
2. Obtain a repeat CT perfusion and CT of the head and snout for evaluation
3. If the animals are going to be survived, then after 2 hours awake and wean the animal.
4. If the animals are non-survival, then we will euthanize and collect histology, and place them in formalin. At a later date these will be collected and taken to a pathologist.