Instrumentation of Near-term Fetal Sheep for Multivariate Chronic Non-anesthetized Recordings

The overall goal of this procedure is to describe the instrumentation of the OV fetus for long-term physiologic monitoring in response to any physiologic perturbation, this research model allows vascular access and electrocardiographic and other bioelectric monitoring of the non anesthetized fetus in utero. This gives us insight into the changes that are occurred during a physiologic perturbation such as vascular occlusion or an injection of a pro-inflammatory mediator. This model is clinically relevant to understanding fetal development in healthy and disease states.

All animal procedures presented here have been approved by the Ethics Committee for the use of animals at the University of Montreal and followed the guidelines set by the Canadian Council for Animal Care. This is a non Survival experiment. The anesthetic protocol described Here has minimal effects on the inflammatory response.

The addition of a small dose of propophol reduces laryngeal reflexes, thus facilitating intubation. A multi-parameter physiologic monitor is used throughout the anesthetic period. All the maternal physiologic parameters measured can be seen here.

The electrocardiogram, direct arterial blood pressure, airway pressure, pulse oximetry, capnography, expired anesthetic level temperature, as well as arterial blood gas analysis connecting the maternal monitor to a Cambridge electronic design data acquisition device. Using a custom made interface cable allows the synchronized display of maternal and fetal physiologic data. The abdomen of the U is prepared using standardized aseptic techniques.

The U is tilted to the right slightly to allow for the exit of the inserted vascular catheters and ECG cable prior to and throughout surgery. The depth of anesthesia is evaluated using a number of neurologic reflexes such as eye position, palp reflex, jaw tone, and The response to surgical stimulation. A midline Incision is made using a scalp blade.

The techniques of hemostasis used in this study were either ligation of the vessels or by simple compression. The median subcutaneous abdominal vein is ligated. The subcutaneous fat is sharply dissected until the linear elbow is clearly identified.

The linear is carefully incised at the umbilicus. A finger is then inserted through this small incision to ensure there is no adhesions or vital structures immediately below the incision. The handle of the nason brown forceps can be used to protect the cutting edge of the scalpel blade during the incision, thus protecting the underlying anatomical structures.

Initially, the uterus is palpated to identify the two horns of the uterus and to count the number of fetuses present. Triplets are possible and the probability of successfully completing the treatment is lower. Therefore, we do not recommend using triplets.

The largest fetus is selected using the interoral distance returned to the abdomen to allow for the insertion of the vascular catheters and ECG cable sponge forceps are inserted into the left aspect of the abdominal cavity along the abdominal wall where they're pushed up against the left flank. A small incision is made over these forceps. The vascular catheters and ECG cable are then passed through this incision and out of the abdominal incision.

It is important to verify that you have all the vascular catheters and ECG cable before starting the rest of the surgery. The head of the fetus is exteriorized and an area of the uterus, which is free of cot. Lead ons is identified along the larger curvature.

A small incision is first made using a scalpel blade. This incision can then be extended over the head of the fetus using scissors to minimize the risk of injury carefully leaving the amnion intact. The amniotic fluid is sampled using a blunt ended catheter before the amnion is opened to minimize the contamination with blood.

This sample can be used to detect the presence of inflammatory changes. The head is removed from the fetal membranes and immediately placed within a surgical glove. The glove is filled with warm water to help maintain norm hermia Babcock forceps are placed on the fetal membranes, uterus, and skin to help reduce loss of amniotic fluid and damage to the placent tone from surgical manipulation.

The surgical field is isolated using sterile drapes. The first step of the fetal instrumentation is the insertion of an arterial catheter. An incision is made along the length of the medial aspect of the humerus.

At the midpoint of the humerus, the brachial artery is bluntly dissected from within between the muscle bellies. A loop of absorbable suture material is then passed underneath the artery. The loop is then cut and the two pieces of suture material are pulled to each end of the artery.

The distal end of the artery is ligated. A loop is made around the proximal end and is used to place traction on the artery, making it easier to create a small incision in the artery using RS scissors. A micro medical grade vinyl catheter is inserted into the artery using forceps.

An alternative to forceps is the use of a vessel dilator. The the point of a vessel dilator device can be inserted into the incision to aid with the insertion of the catheter. Graduated markings on the catheter will help ensure an adequate length of catheter is inserted.

A vascular clamp is placed on the catheter in between the proximal ligature and the insertion site to prevent leakage of blood. The proximal ligature is then secured around the vessel and the catheter. The vascular clamp can now be removed.

A second ligature is placed around the proximal aspect. The free ends of the distal ligature are secured around the catheter. Throughout this process, an assistant is continually aspirating and flushing the catheter to ensure its patency and that the ligatures are not too tight around the catheter.

It is important to minimize the volume of saline injected to avoid fluid overloading the fetus. A fetal blood gas analysis may now be performed to assess the health status of the fetus. A venous catheter can be placed using exactly the same technique.

The cephalic vein can be localized to the proximal aspect of the incision. The vein is stabilized using two ligatures around the vein. The proximal ligature is not tied until the catheter is inserted into the vein.

Again, using our scissors, a small hole is made in the vessel. The catheter is inserted and secured both proximally and distally to the insertion point. The exact location of the placement of these vascular catheters will depend upon the needs of the research model and the target organs being investigated.

Both the venous and arterial catheters exit the same incision site. The skin incision is then closed using a single layer simple continuous suture pattern. The catheters are attached to the anti brachial at various points to allow the leg to be flexed.

In a normal position, the catheters are attached to the shoulder and then in between the scape. A second arterial catheter is placed in the contralateral side as a backup arterial blood pressure and arterial sampling can be achieved during the postoperative period. Four electrocardiographic cables are attached using a 16 gauge needle, the needle is passed subcutaneously.

Notice the denuded section of wire, which is eventually positioned subcutaneously, and the wire is inserted through the bevel of the needle. The needle is removed leaving the wire subcutaneously. Two square knots are used to secure the wire.

Following this cyanoacrylate glue is also applied to each of the knots. A cable is placed on each shoulder joint manubrium and xiphoid process. A fenestrated catheter is attached to the sternum to allow for the measurement of the intrauterine pressure and repeated sampling of the amniotic fluid prior to returning the fetus to the uterus.

30 micrograms of clenbuterol. A beta two agonist is administered slowly intravenously to provide uterine relaxation. Climp erol also reduces the potential abdominal discomfort due to the manipulation of the uterus.

The four legs are carefully positioned back into the uterus or the vascular catheters and ECG cable are passed to the dorsal aspect of the neck where they're a suture to the skin throughout the surgery. It is important to bathe the fetus with warm saline to prevent the drying of exposed tissues and to keep the fetus warm. The surgical glove is removed at the last moment and the head replaced back into The uterus.

The closure of the uterus now Begins. Firstly, the Babcock clamps are removed from the uterine incision. A simple continuous suture pattern is used to close the fetal membranes with a four oh size synthetic multifilament absorbable suture material.

Each of the catheters and ECG cable are incorporated into the closure separately. This permits a tighter closure, thus preventing any leaks. A double layer closure of the serum muscular tissue is performed using a Cushing suture pattern with a O size synthetic multifilament absorbable suture material.

This inverting suture pattern thus helps to minimize adhesion formation and leakage. Again, the catheters and ECG cable are incorporated into the closure separately. Once the uterus is closed, saline is infused into the uterus via the amniotic catheter in order to replace the amniotic fluid lost.

During the surgery. The uterine closure is then covered with the greater omentum. A one poly dione suture material is used to close the linear elbow.

A simple continuous pattern is used. It is important to ensure that the abdominal rectal sheath is incorporated in each pass of the needle as seen. Here, the surgeon has elected to close the linear elbow in two stages.

A simple continuous pattern is also used to close the subcutaneous tissues. At this stage, the assistant surgeon will place a purse string suture around the vascular catheters and ECG cable as they exit through the left flank. Skin staples are used for the skin closure, which will accelerate the closure time.

The surgical wounds are cleaned and a non occlusive bandage is placed over the sites. A stock net Bandage is placed over the torso of the U.Within one hour Post-surgery, the U is up walking around and starting to eat again in a calm, inquisitive manner. Buprenorphine and analgesic Therapy is given during this time throughout the surgical period.

The maternal physiologic parameters were recorded using a multi-parameter physiologic monitor. The maternal ECG and airway pressure were synchronized with the fetal physiologic data for a customized cable linking to the Cambridge electronic design monitor. Fetal and maternal heart rate variabilities can be derived from these data.

An example of the physiologic data captured is shown here. Paired maternal and fetal arterial blood gas samples were taken at predetermined time points during the surgical and postoperative periods to ensure their health status. Ventilator settings, fluid therapy and dobutamine perfusion rate were all adjusted based upon these results to ensure adequate ventilation and perfusion of both the U and the fetus, oxygen nasal therapy was instituted.

Once the U was extubated. Fluid therapy was continued. If any acid base or electrolyte disturbance were detected, we hypothesized the reason for the transient mild metabolic acidosis observed in the fetus was due to the reduction in uterine perfusion associated with use of gentle anesthesia.

For further information regarding results, Please refer to the written article. In summary, this model describes the basic Surgical implantation of two arterial catheters, one venous catheter, the placement of ECG cables and an amniotic catheter. These will allow for the monitoring of the electrocardiogram, arterial blood pressure sampling of both arterial and venous blood, and monitoring of the intrauterine hydrostatic pressure.

We wish to acknowledge the following individuals for their valued assistance with this intensive research project. We also like to thank the Canadian Institutes of Health Research. Lafon do research to Quebec on Santi and the Twell Perinatal Research Foundation for their financial Support of this project.