Temporal Neuromuscular Adaptations and Proteomic Signatures Following Botulinum Neurotoxin A Injection in Spastic...

Cerebral palsy (CP) is predominantly characterized by spasticity. While botulinum neurotoxin type A (BoNT-A) effectively reduces spasticity, its transient efficacy represents a major clinical limitation. Elucidating the mechanisms that limit its duration of action may inform strategies to prolong therapeutic benefit.

A spastic CP rat model was established via carotid artery ligation and hypoxia in 7-day-old Wistar rats. On postnatal Day 21, BoNT-A (5 U/kg) was administered to the gastrocnemius muscle. Behavioral and molecular biological assessments were performed at 4 and 12 weeks post-injection.

At 4 weeks after injection, the motor performance improved, spasticity decreased (p < 0.05), neuromuscular junction density increased, and neurotrophic factors IGF-1, GAP 43, and S100 increased (all p < 0.05). However, these functional and molecular changes diminished by 12 weeks. Further proteomic analysis revealed a shift in pathway enrichment, from protein synthesis and vesicular transport at 4 weeks to metabolic regulation at 12 weeks. Among the differentially expressed proteins, Sar1b and Rtn1, whose expression patterns paralleled with nerve sprouting, may be key regulatory factors.

These findings indicate that BoNT-A facilitates NMJ recovery through temporal regulation of energy metabolism, protein synthesis, and vesicular transport. Rtn1 and Sar1b may represent candidate molecular targets for extending the therapeutic effects of BoNT-A, although further functional validation is required.