Structural and functional interactions between transient receptor potential vanilloid subfamily 1 and botulinum neurotoxin serotype A

Abstract

Background: Botulinum neurotoxins are produced by Clostridium botulinumbacteria. There are eight serologically distinct botulinum neurotoxin isoforms (serotypes A.H). Currently, botulinum neurotoxin serotype A (BoNT.A) is commonly used for the treatment ofmany disorders, such as hyperactive musculoskeletal disorders, dystonia, and pain. However, the effectiveness of BoNT.A for pain alleviation and the mechanisms thatmediate the analgesic effects of BoNT.A remain unclear. To define the antinociceptive mechanisms by which BoNT/A functions, the interactions between BoNT.A and the transient receptor potential vanilloid subfamily 1 (TRPV1) were investigated using immunofluorescence, co-immunoprecipitation, and western blot analysis in primary mouse embryonic dorsal root ganglion neuronal cultures. Results: 1) Three-week-old cultured dorsal root ganglion neurons highly expressed transient TRPV1, synaptic vesicle 2A (SV2A) and synaptosomal-associated protein 25 (SNAP-25). SV2A and SNAP-25 are the binding receptor and target protein, respectively, of BoNT.A. 2) TRPV1 colocalized with both BoNT.A and cleaved SNAP-25 when BoNT.A was added to dorsal root ganglia neuronal cultures. 3) After 24 hours of BoNT.A treatment (1 nmol.l), both TRPV1 and BoNT.A positive bands were detected in western blots of immunoprecipitated pellets. 4) Blocking TRPV1 with a specific antibody decreased the cleavage of SNAP-25 by BoNT.A. Conclusion: BoNT/A interacts with TRPV1 both structurally and functionally in cultured mouse embryonic dorsal root ganglion neurons. These results suggest that an alternative mechanism is used by BoNT.A to mediate pain relief.