2025年2月28日
Mechanisms of sudden unexpected death in epilepsy (SUDEP) are poorly understood and challenging to translate from current models. Transgenic rabbits may offer insights into these mechanisms. We describe a method for long-term, continuous electroencephalography and electrocardiography recordings in transgenic rabbit kits to evaluate serious events that may lead to death.
[Instructor] To begin, position the anesthetized kit supine on an infrared heating pad controlled via a rectal thermometer. Place the nose and mouth in a custom 3D printed face mask connected via a swivel connector to a non-rebreathing Jackson-Reese circuit with a 0.5 liter bag. Monitor the depth of anesthesia using a pulse oximeter on either the ear or paw. Place adhesive surgical towels on either side of the kit, and cover it with a large surgical drape. Cut an appropriately sized hole in the drape to expose the abdomen and chest. Open the implant onto the surgical field and place non-absorbable anchor sutures into each of the implant anchor holes, leaving a five to six centimeter tail on each suture. Place the implant in a bowl of warm sterile saline until ready for placement. Using a scalpel, make a three centimeter incision through the skin along the linea alba. And carefully incise through the muscle to open the peritoneal cavity. Then place the implant into the cranial portion of the abdominal cavity, and position it to the left of the incision. Use a trocar to tunnel the positive electrocardiogram, or ECG, and EEG wire out of the peritoneal cavity and skin approximately three to four centimeters to the left of the incision. Tunnel the negative ECG wire two centimeters to the right of the incision, allowing the implant to sit comfortably within the cavity. Now, secure the implant with anchor sutures to the ventral wall of the peritoneal cavity, ensuring that no bowel is entrapped. And close the abdominal wall with an absorbable suture in an interrupted pattern. Then close the skin incision with a non-absorbable suture in an interrupted pattern. Tunnel the negative ECG leads subcutaneously to the right upper chest at the level of the first rib. Bluntly dissect a subcutaneous pocket to loosely coil approximately 10 centimeters of wire. Cut any excess wire and create a loop after tying the exposed end to the insulated wire with a non-absorbable suture. Secure the loop to the muscle with two non-absorbable sutures. Then tunnel the positive ECG lead to the left lower rib, and repeat the demonstrated steps to secure it to the muscle and close the incision. Wrap the exposed EEG wires with sterile aluminum foil. Let a non-sterile assistant remove the sterile drape and leg tie. Turn the kit into the prone position, while ensuring the face mask remains securely in place by rotating the swivel connector between the face mask and circuit. Adjust the pulse oximeter and Doppler monitor as needed to maintain continuous anesthetic monitoring. Then position a Doppler probe over the heart. Place a sterile adhesive towel under the left side, while the assistant holds the aluminum foil packet containing the wires. Gently remove the wires from the aluminum packet and place them on the sterile field, and complete the draping process with the sterile towel. Make a three centimeter incision along the midline of the scalp to expose the skull. Use a trocar to subcutaneously tunnel the EEG leads from the left side to the skull. Then clean and scrape the periosteum from the exposed parietal bones using a scalpel. Insert a handheld drill into a sterile ultrasound cover, and guide a one millimeter drill burr into the drill. Next, drill bilateral burr holes into the parietal bones, approximately 0.5 centimeters anterior to lambda and 0.5 centimeters lateral to the sagittal suture. Use fine forceps to place a screw into the burr hole. Then use a screwdriver to insert the screw approximately halfway into the skull. Now, bluntly dissect a subcutaneous pocket along the back of the neck to loosely coil approximately 10 centimeters of wire. Cut any excess wire and strip the insulation from the tip, stretching the wire. To create a loop at the end of the exposed wire, tie a knot leaving a small loop. Place the loop over the screw and tighten the screw to the skull, ensuring the wire makes contact with the screw. Position the ground wire on the left and the recording wire on the right. Now, secure the screws and wires to the skull using dental acrylic. Finally, inject bupivacaine subcutaneously at each incision site for pain relief. And cover each incision with a small amount of skin glue. Once all wires are in place, assess the telemetry signals using analysis software to verify signal fidelity. Expect EEG signals to appear at low amplitude while the kit is sedated. Implant surgery was successfully performed on 15 rabbit kits, with 12 surviving to the experimental endpoint. Telemetry recordings immediately after returning the kits to home cages showed low amplitude EEG signals, which improved as the kits recovered from anesthesia. ECG signal morphology changed slightly in the first few days post-surgery due to scarring and wire stabilization. But overall, signal quality remained robust over time. The implanted telemetry system allowed for the recording of multiple bio potentials, including electroencephalography, temperature, acceleration, electrocardiography, and signal quality.
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本研究利用转基因兔仔探索癫痫猝死(SUDEP)的机制。文中描述了一种用于长期连续脑电图(EEG)和心电图(ECG)记录的方法,以评估可能导致死亡的严重事件。
在新生转基因兔仔中同步进行脑电图(EEG)和心电图(ECG)遥测,可实现对与SUDEP机制相关的神经和心脏电生理活动进行高保真、长期的监测。该方法通过在一个比啮齿类模型更接近人类生理系统的模型中捕捉与疾病相关的心律失常和癫痫发作事件,弥补了临床前模型中的关键空白。该技术有助于提高对癫痫及心脏功能障碍相关离子通道病中靶点验证的预测可信度,并支持对作用机制的风险评估。
这种具备遥测功能的兔模型在癫痫和心律失常研究中架起了早期发现与临床前验证之间的桥梁,支持先导化合物的筛选及机制研究。