A Piglet Model of Neonatal Hypoxic-Ischemic Encephalopathy

The overall goal of the following experiment is to demonstrate a piglet model of perinatal hypoxic ischemic encephalopathy. This model can then be used for the investigation of neonatal Brain injury. First, Piglets are sedated, ventilated, and monitored in a setup comparable to that of a neonatal intensive care unit.

The technique described in the present manuscript uses amplitude integrated EEG to titrate hypoxia. According to the resilience of each individual study, animal hypoxia is induced in a controlled manner, keeping the amplitude integrated, EEG suppressed while ensuring survival. In this four day survival model, animals are euthanized at 72 hours post hypoxia.

Finally, according to the purpose of the experiment, cerebral and other outcomes may be evaluated using magnetic resonance imaging, neuropathology and analysis of body fluids and organs. In this study, we visualize step-by-step the technical procedures necessary to establish a piglet model of neonatal hypoxic ischemic encephalopathy. Nerve developmental outcome is a major concern in all areas of neonatal intensive care.

Good animal models are essential to improving the management of neonatal brain injury. The model you’re about to see is a good approximation to the clinical course of hypoxia isch. For this reason, results translate well to the clinical setting in the model presented here, we control the severity and timing of the hypoxic insult, thus minimizing the biological variation.

Complicating the interpretation of clinical studies, we use Danish land raise piglets at 18 to 24 hours postnatal age. They weigh on average 1000 to 2000 grams, and their physiology is in many ways comparable to that of the newborn infant. This model can be used for investigating treatment effects on magnetic resonance imaging, neuropathology and biomarkers in body fluids such as cerebral spinal fluid, urine, and blood.

It is a technically challenging model to work with. In the following, we demonstrate each step necessary for the completion of the model. The equipment needed For initiating anesthesia is a mask for sevo fluorine administration, peripheral vein catheter syringes with saline, propofol, fentanyl, muscle relaxant, and penicillin.

A sling for opening the mouth, a cotton tipped swap, and veterinary laryngoscope with straight blade Sloane spray cuffed endotracheal tubes, SIIS 3 0 2 0.5 and two oh self-inflating bag syringe for cuff inflation and stethoscope. VO fluorine is administered using a mask and the depth of anesthesia is monitored. According to protocol, a peripheral vein catheter is placed in an ear vein and flushed with salan.

At this point, the animal is sedated, but breathing spontaneously. Intravenous anesthesia is initiated by giving fentanyl 30 micrograms per kilo, followed by propofol, five milligrams per kilo and broon one milligram per kilo immediately prior to intubation. For intubation, the piglet is displaced in the supine position.

The length of the endotracheal tube is measured from the tip of the snout to above the sternal nudge. Typically around 13 centimeters, a cotton swap is used to pull the long epiglottis forward so that it may be lifted. Used in the laryngoscope enabling full view of the adenoid cartilages in vocal cords.

Sine spray is applied locally in the larynx to prevent spasm. The endotracheal tube is forwarded slowly through the vocal cords when advanced, according to the premeasured distance, the endotracheal tube is connected to a self-inflating bag for manual ventilation, correct. Endotracheal tube placement is confirmed by observing symmetrical chest movement, osculation by stethoscope, visual confirmation of mist in the tube and tid CO2 curve as seen in white on the monitor.

Finally, the cuff is inflated to ensure a secure airway. The endotracheal tube is now connected to a ventilator for mechanical ventilation. Finally, Infusion of propofol, five milligrams per kilo per hour, and fentanyl.

10 micrograms per kilo per hour is started. The ventilator is set to a volume controlled mode delivering tidal volumes of 10 milliliters per kilo equipment for monitoring. The piglet is ECG electrodes, pulse oximeter, rectal temperature probe, and equipment.

For central arterial pressure monitoring, the pulse oximeter is placed, followed by insertion of a rectal temperature probe. The chest is shaved prior to electrode placement. The screen displays ECG and heart rate central arterial pressure and oxygen saturation.

The equipment needed for placing umbilical catheters is sterile draping scalpel, micro instruments, umbilical catheters size 3.5 French and five French syringes for blood sampling and flushing suture set for visualization of the umbilical vessels. The cord is cut at skin level. The umbilical vein is seen in the right of the picture, and one of two umbilical arteries is seen to the left.

First, the arterial catheter is inserted to a depth equal to the piglet’s weight in kilos times three plus 10 centimeters. Correct position is verified by drawing back blood. Subsequently, the venous catheter is inserted to a depth of five centimeters.

Catheters are secured by suturing. Using a three art suture, the head is shaved for placement of electrodes. For amplitude integrated EEG monitoring, the parietal electrodes should be placed with at least two to four centimeters between them, the reference electrode in the midline and the ground electrode anywhere on the head.

Impotence testing reveals four correctly placed electrodes with good signal to noise ratio. The bottom of the screen displays real time EEG, while the top screen shows the time compressed amplitude integrated, EEG, the piglet is now allowed to rest for 60 minutes prior to induction of hypoxia. Hypoxia is initiated by switching to a ventilator, delivering a mixture of nitrogen and 4%oxygen.

It takes an average two minutes to achieve a flat trace on the amplitude integrated EEG corresponding to significant hypoxia. Leukemia in the brain for 45 minutes. The fraction of inspired oxygen is regulated.

To maintain a flat EEG amplitude below seven microvolts during hypoxia, the piglet initially compensates by increasing the heart rate. As hypoxia continues, the piglet starts to decompensate and the mean material pressure drops. At this point, its critical to adjust fraction of inspired oxygen to ensure survival.

After 45 minutes, the fraction of inspired oxygen is increased to 21%resulting in increased brain activity. Following hypoxia, the piglet is extubated and allowed to wake up for the next four days. The piglet is cared for by an experienced animal technician who feeds the pig and scores it neurologically on a daily basis.

At seven two hours, the pig is sedated and ventilated. Again, genetic resonance imaging and spectroscopy are powerful tools for examining the brain. Magnetic resonance imaging is complimented by neuropathologic examination.

The brain is carefully dissected in each area, investigated by an experienced neuropathologist. Having watched this video, you should have a good understanding of how to establish a piglet model of neonatal hypoxic ischemic encephalopathy. With this model, pertinent questions pertaining to diagnosis and therapy of neonatal brain injury can be investigated.

Ultimately optimizing the management of a affected Infants.