Capsaicin Protects Mouse Neuromuscular Junctions From The Paralytic Effects Of Botulinum Neurotoxin A

Abstract

Botulinum neurotoxins (BoNTs) are the most toxic naturally occurring proteins. Botulinum serotype A (BoNT/A) selectively cleaves SNAP-25 and inhibits acetylcholine release from motor nerve endings resulting in life threatening paralysis of skeletal muscles. Here we report that capsaicin (8-methyl-N-vanillyl-6-nonenamide), an irritant active principle of chili peppers, protects against the paralytic effects of BoNT/A in mouse. In Triangularis sterini nerve muscle preparations, capsaicin pretreatment in vitro, significantly reduced fluorescent labeled BoNT/A uptake mediated by depolarization with 40 mM KCl or neural stimulations. In vivo injection of capsaicin (3 μl of a 1 mM stock solution) either coinjected or injected 4 or 8 hours before BoNT/A injection protected the mice from the inhibitory effects of BoNT/A measured by toe spread reflex. In controls the toe spreading reflex was inhibited within 24 hrs of poisoning with BoNT/A. Also wortmannin, a PIP5K inhibitor, injected prior to BoNT/A exposure, protected the mice from the paralytic effect of BoNT/A. In vitro muscle tension measurements demonstrate that capsaicin pretreatment partially protected the functions of the mouse Extensor digitorum longus nerve muscle preparations. Also pretreatment of cultured cholinergic neuroblastoma (N2a) cells with 10 μM capsaicin for 10 minutes prior to exposure of 10 pM BoNT/A significantly preserved labeling of synaptic vesicle pools by FM1-43. Our data collectively demonstrate that capsaicin offers protection from the paralyzing effect of BoNT/A by significantly reducing the uptake of the toxin in mouse neuromuscular junction probably by interfering with endocytosis of the toxin.Supported by HDTRA1-07-C-0031 CBT BAA.