Optimizing Neuromuscular Junction Formation for Enhanced Signal Transduction in the Regenerative Peripheral Nerve Interface.

Abstract

RPNI surgery amplifies motor intent from upstream peripheral nerves, enabling myoelectric prosthetic devices to better capture signals, thereby improving the precision of prosthetic limb control. This study aims to clarify the mechanisms underlying RPNI motor function via neuromuscular junction (NMJ) evaluation, and provide a novel method of enhancing reinnervation to improve signal amplification. Adult wild-type C57BL/6J mice underwent RPNI surgery utilizing free extensor digitorum longus (EDL) muscle grafts transferred to the thigh and neurotized with the common peroneal nerve (CPN) in three ways: A) RPNI: CPN neurotization; B) Double fascicular RPNI: CPN separated into two fascicles then neurotization; and C) Denervated RPNI: no CPN neurotization. In the control groups, the EDL muscles remained in situ with: D) Intact CPN innervation (healthy EDL); and E) CPN cut and repaired (C&R). The endpoint assessments were conducted at 8 weeks and 12 weeks postoperatively with electrodiagnostic tests (mV), target muscle weight measurement, whole-mount immunofluorescence (IF) staining, and western blot analysis. The results showed that RPNI demonstrates adequate NMJ innervation for effective neural signal amplification. Additionally, the double fascicular RPNI exhibited greater compound muscle action potential (CMAP), larger target muscle weight, more reinnervated NMJs, and less motor endplate fragmentation compared to the RPNI. The degree of NMJ reinnervation and the morphology of the motor endplate within the RPNI reveal information about the innervation. Dividing the neural source enhances NMJ reinnervation within the RPNI, with the added potential of improving signal amplification.