The animal protocol begins with animal preparation and instrumentation, followed by the experimental protocol. The experimental protocol consists of the following phases: baseline, injury, post-injury shock, intervention, and resuscitation.
I. Animal Preparation and Instrumentation:
1. Perform sedation with telazol (5mg/kg) and xylazine (2mg/kg) as an intramuscular (IM) injection.
2. Place oxygen saturation/heart rate probe to monitor animal while sedated. Take temperature for baseline measurement.
3. Transport the animal to the procedure area
4. Utilizing the appropriate face mask, place the animal under isoflurane anesthesia, with appropriate quantitative end-tidal CO2 monitoring
5. Attach electrocardiographic monitor, pulse oximetry monitor, and place temperature probe. Ensure data is being collected and recorded
6. Place the animal in sternal recumbency and perform endotracheal intubation. Ensure proper placement with quantitative CO2 monitor. Secure endotracheal tube in place.
7. Place the animal into dorsal recumbency and secure to the operating table. Ensure monitors are still attached and recording properly. Adjust the isoflurane to achieve an MAC ~1.0. Set ventilator tidal volume to ~8-10cc/kg, respiratory rate of 12-14, PEEP of 5, FiO2 of 40%. Titrate minute ventilation to achieve a pCO2 35-45 mmHg based on blood gas analysis. O2 can be titrated based on pulse oximetry.
8. Place all intravascular catheters using a percutaneous, ultrasound-guided, modified-Seldinger technique. Placement of the pressure catheters/angiogrpahic catheters within the introducer sheaths is performed under direct fluoroscopic guidance.
- Place 8 Fr. 24 cm Arrow Superflex introducer sheath in the right brachial artery. This is to house the angiographic catheter for image acquisition. Steer the angiographic catheter into the proximal aorta under ultrasound guidance. Perform fluoroscopy to confirm adequate position proximal to the takeoff of the bicarotid trunk. It is placed in this location for anatomical reasons [Edwards]
- Place 8 Fr. 10 cm Arrow Superflex introducer sheath in the left brachial artery. This is to house the 5Fr pressure catheter to record arterial pressure above the REBOA catheter.
- Place 9 Fr. 25cm Terumo Introducer sheath into the left internal jugular vein. Guide the tip of the catheter into the right atrium. This is to perform venous blood gases to measure mixed venous blood oxygen saturation
- Place 7 Fr. 10cm introducer sheath into the right femoral artery. Place a 5 Fr. pressure catheter through this introducer sheath into the distal aorta under fluoroscopic guidance. This is to measure the blood pressure below the REBOA catheter.
- Place 10 Fr. 10cm introducer sheath into the left femoral artery. Place the REBOA catheter through the sheath and into zone 1 of the aorta under fluoroscopic guidance. Test the REBOA catheter balloon prior to insertion to ensure that the balloon is intact
- Place 8 Fr. 10cm introducer sheath into the left femoral vein. This will be used for controlled hemorrhage, as well as for drugs and resuscitative fluids/blood product
- Place 7 Fr. 10 cm introducer sheath in the right femoral vein. Place a 5Fr. pressure catheter through this introducer sheath and place into the IVC under fluoroscopic guidance to monitor CVP. This introducer will also be used for drug administration.
9. Perform a small lower midline laparotomy using electrocautery for placement of a cystotomy drainage tube. Once intraperitoneal access is established, purse-string suture a Foley catheter in the superior/anterior portion of the bladder. Inflate the foley balloon. Ensure adequate bladder drainage. Close the subcutaneous tissue with a skin stapler.
10. Ensure all introducers are secured into place. Ensure all catheters are performing adequate hemodynamic monitoring. Give the animal 1L or normal saline with 1 amp of D50 to normalize fluid/glycemic status after fasting.
11. Once the fluid is completely infused, prepare animal for placement in sternal recumbency. Disconnect pressure probes, clamp and disconnect foley from foley collection bag, disconnect the endotracheal tube from the ventilator, and disconnect any other monitors that could get tangled during the position change.
12. Place the animal into sternal recumbency and reconnect ventilator, all monitors, and the foley to the foley connection bag.
13. Check the position of all of the pressure catheters and the angiographic catheter using fluoroscopic guidance. Once adequately postitioned and data collection is confirmed to be working appropriately, attention can be paid to central instrumentation.
14. Remove the skin and soft tissue overlying the areas of the proposed burr holes with electrocautery.
15. Perform a helical CT scan to plan for the cerebral probe placement.
16. 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.
17. 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. Place the LDF probe through the left 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.
18. Perform an additional helical CT scan to ensure proper placement of the intracranial instrumentation. Ensure that data is being recorded properly.
19. After performing intracranial instrumentation, provide systemic heparinization to the animal (10k units).
20. Allow for the collection of baseline data for ~15-30 minutes. This includes:
- baseline hemodynamic parameters
- baseline intracranial parameters
- baseline mixed venous blood gas, arterial blood gas
- baseline chemistry
- baseline troponin
21. 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.
III. Injury (30 min):
22. Prepare the blood collection bag with the appropriate anticoagulant, calculate the rates of controlled hemorrhage, and set up the syringe pump to inflate the Fogarty catheter.
23. Utilizing a standard syringe pump connected to the Fogarty catheter, inflate the Fogarty catheter balloon at a rate of 1ml/min until the ICP reading has a nadir of 20mmHg
24. Utilizing a standard peristaltic pump connected to the left femoral vein catheter, withdraw blood into the blood collection bag. The controlled hemorrhage is a total of 25% hemorrhage with 15% of the hemorrhage occurring in the first 15 minutes. Continue until the completion of hemorrhage.
IV: Post-injury Shock (15 min):
25. Collect full set of labs post injury
26. Obtain post-injury shock CTP
27. TIMEOUT: Ensure all sheaths are drawing back for labs, ensure the proper FiO2/FiCO2/paCO2, empty foley bag, ensure labs and CTP are completed
V. Intervention (30 minutes)
28. Perform the intervention (fREBOA vs. pREBOA vs. no REBOA)
- fREBOA is continuous REBOA without any alteration of the balloon
- pREBOA titrated to systolic pressure goal of 100 +/- 10 mmHg
- no REBOA group with additional 30 minutes of no intervention
29. Perform immediate CTP after intervention
30. Collect blood gases at 15 minutes, full set of labs at 30 minutes post intervention
31. Perform CTP 30 minutes post intervention
32. Start infusion of previously shed blood through the left femoral vein via a pressure bag
33. In the fREBOA and pREBOA groups, deflate the balloon as soon as possible
34. If not meeting systolic blood pressure goal of 100 +/- 10 mmHg while infusing blood, start norepinephrine infusion.
35. Once blood is utilized, start normal saline infusion to maintain blood pressure at systolic goal stated above. If not meeting this blood pressure requirement, utilize norepinephrine infusion to meet blood pressure goal.
36. Once 3x the volume of the controlled hemorrhage is utilized in crystalloid, then carrier fluid can be utilized to deliver norepinephrine. No other fluid boluses were allowed.
37. Obtain CTP at 60, 120, 180 minutes post resuscitation
38. Obtain labs at 15, 30, 60, 120, 180 minute marks
39. Interventions during the resuscitation phase, based on laboratory parameters:
- Glucose < 75: amp of D50
- pH < 7.2 with pCO2 < 40 give 1 amp bicarb
- pH < 7.2 with pCO2 > 40, increase RR by 2
- iCa < 1.3, give calcium carbonate
- K+ < 2.5, give KCl
- K+> 7, give insulin + D50
40. If the animal survives the entirety of the protocol, euthanize the animal according to institutional protocol (e.g. rapid KCl injection under heavy inhalant anesthesia).