Development of a Neonatal Rat Model for Brachial Plexus Birth Injury

This protocol describes the development of a neonatal rat model for brachial plexus birth injury. Brachial plexus birth injury is a nerve injury that occurs during a difficult delivery. While many infants recover spontaneously, up to 30%developed long-term sequelae.

Current treatments range from physical therapy to microsurgical nerve repair, but outcomes remain viable. A reliable animal model is therefore needed to study the underlying mechanisms and long-term consequences of brachial plexus birth injury. All animal procedures were performed with approval of the Institutional Animal Care and Use Committee.

A multiparous pregnant Sprague Dawley rat at gestational day 14 is housed under standard laboratory conditions. During late gestation, a researcher visited the dam daily and performed olfactory conditioning by placing cotton swabs with ethanol and Betadine in the cage. Additional bedding material was provided to allow the dam to create a bigger nest.

All surgical instruments were sterilized by autoclaving. A surgical bed was prepared from polystyrene. Surgeries are performed on five day old rat pups.

Anesthesia is induced by 2%isoflurane in oxygen, and confirmed by the absence of a toe-pinch reflex. The rat pup is placed on a pre-warmed heating pad. Each pup is marked with a unique number.

Extended release buprenorphine is administered for analgesia and anesthesia is maintained at 1.5 to 2%isoflurane. throughout the procedure. The surgical field is disinfected with alternating scrubs of Betadine and ethanol before the pup is positioned in the polystyrene mold.

The head is secured to the nose cone with tape, and the lower body is stabilized using wrap to minimize movement. Create a midline cervical incision for approximately five millimeters in length, extending from the base of the skull towards the C 71 level. Create the incision using a number 11 blade.

Superficial vessels are cauterized. Dissect through the trapezius and the underlying rhomboid and levator scapulae muscles using cauterization. Be careful to not cauterize the skin edges to avoid thermal injury.

Identify and isolate the brachial plexus via blunt dissection. Cauterize the identified nerve of the brachial plexus. Confirm a successful injury by observing upper extremity muscle twitches.

Place an absorbable gelatin sponge between the proximal and distal segments of the brachial plexus to prevent nerve regeneration. Close the skin using 4-0, 7-0 suture in a continuous pattern. Apply a tissue adhesive to prevent wound dehiscence.

Place the rat pup on a heating pad with nesting material from the rat dam's cage. Monitor the respiratory rate and movements of the rat pup until it returns to normal respiratory patterns. To prevent maternal rejection, rub the rat pup gently with the home cage nesting material before placing it back to the dam.

Postoperatively, the effectiveness of the surgical technique can be evaluated by the absence of movement in the effectively limb. Motor function was assessed daily during the first two weeks postoperatively and weekly thereafter. Briefly, absence of limb movement was scored as zero, partial limb movement was scored as one, and normal symmetric movement was graded as two.

All included rat pups exhibited a pan-plexal injury and scored zero on the movement scale. Upon sacrifice, the degree of muscle atrophy, limb shortening, and the presence of contractors can be evaluated. To conclude this protocol provides a reproducible neonatal rat model to investigate brachial plexus birth injury.