Evaluation of Recombinant Botulinum Neurotoxin Type A1 Efficacy in Peripheral Inflammatory Pain in Mice

Abstract

Well-established efficacy of botulinum neurotoxin type A (BoNT/A) in aesthetic dermatology and neuromuscular hyperactivity disorders relies on canonical interruption of acetylcholine neurotransmission at the neuromuscular junction at the site of the injection. The mechanisms and the site of activity of BoNT/A in pain, on the other hand, remain elusive. Here, we explored analgesic activity of recombinant BoNT/A1 (rBoNT/A1; IPN10260) in a mouse model of inflammatory pain to investigate the potential role of peripheral sensory afferents in this activity. After confirming analgesic efficacy of rBoNT/A1 on CFA-induced mechanical hypersensitivity in C57Bl6J mice, we used GCaMP6s to perform in vivo calcium imaging in the ipsilateral dorsal root ganglion (DRG) neurons in rBoNT/A1 vs. vehicle-treated mice at baseline and following administration of a range of mechanical and thermal stimuli. Additionally, immunohisochemical studies were performed to detect cleaved SNAP25 in the skin, DRGs and the spinal cord. Injection of CFA resulted in reduced mechanical sensitivity threshold and increased calcium fluctuations in the DRG neurons. While rBoNT/A1 reduced mechanical hypersensitivity, calcium fluctuations in the DRG of rBoNT/A1- and vehicle-treated animals were similar. Cleaved SNAP25 was largely absent in the skin and the DRG but present in the lumbar spinal cord of rBoNT/A1-treated animals. Taken together, rBoNT/A1 ameliorates mechanical hypersensitivity related to inflammation, while the signal transmission from the peripheral sensory afferents to the DRG remained unchanged. This strengthens the possibility that spinal, rather than peripheral, mechanisms play a role in the mediation of analgesic efficacy of BoNT/A in inflammatory pain.

Intramuscular Botulinum Neurotoxin Serotypes E and A Elicit Distinct Effects on SNAP25 Protein Fragments, Muscular Histology, Spread and Neuronal Transport: An Integrated Histology-Based Study in the Rat

“…In the lumbar spinal cord, and as expected from previous work [20], no c-SNAP25 staining was observed in vehicle-treated animals. However, the presence of c-SNAP25 was noted with both toxins mainly in the ipsilateral part of the ventral horn close to the large motoneurons, indicating neuronal transport (Figure 7G-I,N-P).…”

“…Although this must be experimentally confirmed, we can postulate that the lower ability of BoNT/E to undergo such a phenomenon could be due to a potential targeting of other types of cargo less involved in neuronal transport. Many works support the importance of neuronal transport in the antinociceptive activity of BoNT/A [20,45,46]. As such behavior is limited in BoNT/E, it might restraint its therapeutic use for such conditions, maybe explaining the paucity of published data on the subject.…”

“…The immunohistochemical staining method was performed using a standard avidinbiotin-peroxidase procedure as previously described [20]. After a heat-induced epitope retrieval step, endogenous peroxidases were blocked by 10 min incubation in a 3% H 2 O 2 solution (Sigma-Aldrich, #H1009).…”

Intraoperative abobotulinumtoxinA alleviates pain after surgery and improves general wellness in a translational animal model

“…Peripheral injections of BoNT-A do not alter normal nociceptive thresholds or acute nociceptive pain in both humans and animals [31][32][33] . In a recent study, we evaluated in vivo calcium imaging in the ipsilateral DRG neurons in mice exhibiting CFA-mediated inflammatory pain, that were treated with recombinant BoNT-A or vehicle 34 . While BoNT-A injections reduced mechanical allodynia (measured with von Frey filaments) in comparison www.nature.com/scientificreports/ to vehicle treatment, calcium fluctuations of BoNT-A and vehicle-treated animals in the DRG were similar 34 .…”

“…In a recent study, we evaluated in vivo calcium imaging in the ipsilateral DRG neurons in mice exhibiting CFA-mediated inflammatory pain, that were treated with recombinant BoNT-A or vehicle 34 . While BoNT-A injections reduced mechanical allodynia (measured with von Frey filaments) in comparison www.nature.com/scientificreports/ to vehicle treatment, calcium fluctuations of BoNT-A and vehicle-treated animals in the DRG were similar 34 . This suggests that peripherally-administered BoNT-A does not change nociception-related signal transmission from the peripheral sensory afferents to the DRG.…”