Type A1 but Not Type A2 Botulinum Toxin Decreases the Grip Strength of the Contralateral Foreleg Through Axonal Transport From the Toxin-Treated Foreleg of Rats

Torii, Yasushi; Akaike, Norio; Harakawa, Tetsuhiro; Kato, Keiko; Sugimoto, Nakaba; Goto, Yoshitaka; Nakahira, Shinji; Kohda, Tomoko; Kozaki, Shunji; Kaji, Ryuji; Ginnaga, Akihiro

doi:10.1254/jphs.11121fp

Cited by 43 publications

(40 citation statements)

References 28 publications

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“…The symptoms associated with intoxication are different for BoNT/A1 and BoNT/A2 in mice, where intravenous injection of BoNT/A1 caused ruffled fur, labored breathing, descending paralysis, and spasticity before death (52), while intravenous injection of BoNT/A2 caused paralysis of the front legs, hind legs, and whole body, followed by death (24). Further, BoNT/A1 diffuses more readily than BoNT/A2 to the contralateral side due to retrograde movement (27,29,53); whether retrograde trafficking of BoNT plays a role in the symptoms elicited by the toxins remains unknown. The symptoms of other BoNT subtypes appear to be related to their similarity.…”

Section: Discussionmentioning

confidence: 99%

“…While BoNT/A1 and BoNT/A2 cleave SNAP25 with similar kinetics (22)(23)(24) and possess similar potencies in the mouse model of botulism (BoNT/A1 and BoNT/A2 had specific activities of 1.25 ϫ 10 8 and 1.27 ϫ 10 8 50% lethal doses [LD 50 ] [in U/mg], respectively, in mice) (25,26), BoNT/A1 and BoNT/A2 elicit different paralytic symptoms in mice (24). For example, BoNT/A1 appears to be more potent than BoNT/A2 for grip strength on the contralateral side (27), and BoNT/A1 shows more contralateral spread than BoNT/A2, which is prevented by colchicine treatment in a rat model (26,27). In addition, BoNT/A2 is more potent than BoNT/A1 in cultured primary neurons (24,28).…”

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Entry of Botulinum Neurotoxin Subtypes A1 and A2 into Neurons

et al. 2017

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Botulinum neurotoxins (BoNTs) are the most toxic proteins for humans but also are common therapies for neurological diseases. BoNTs are dichain toxins, comprising an N-terminal catalytic domain (LC) disulfide bond linked to a C-terminal heavy chain (HC) which includes a translocation domain (H N ) and a receptor binding domain (H C ). Recently, the BoNT serotype A (BoNT/A) subtypes A1 and A2 were reported to possess similar potencies but different rates of cellular intoxication and pathology in a mouse model of botulism. The current study measured H C A1 and H C A2 entry into rat primary neurons and cultured Neuro2A cells. We found that there were two sequential steps during the association of BoNT/A with neurons. The initial step was ganglioside dependent, while the subsequent step involved association with synaptic vesicles. H C A1 and H C A2 entered the same population of synaptic vesicles and entered cells at similar rates. The primary difference was that H C A2 had a higher degree of receptor occupancy for cells and neurons than HcA1. Thus, H C A2 and H C A1 share receptors and entry pathway but differ in their affinity for receptor. The initial interaction of H C A1 and H C A2 with neurons may contribute to the unique pathologies of BoNT/A1 and BoNT/A2 in mouse models.KEYWORDS botulinum toxin, gangliosides, synaptic vesicles, synaptic vesicle protein 2, TIRF microscopy, clostridial neurotoxins, Clostridium botulinum, toxins B otulinum neurotoxins (BoNTs) are AB exotoxins secreted by several species of the genus Clostridium. BoNTs are single-chain 150-kDa proteins cleaved by either bacterial or host proteases to a dichain comprising a 50-kDa light chain (LC) and a 100-kDa heavy chain (HC) linked by an interchain disulfide bond. The HC includes a translocation domain (H N ) and receptor binding domain (H C ) (1). The H C includes an N-terminal subdomain (H CN ) of limited known function and a C-terminal subdomain (H CC ) that confers neuron specificity by binding dual neuron-specific receptors. There are seven BoNT serotypes (A to G) (2). BoNT serotype A (BoNT/A) binds a ganglioside and synaptic vesicle glycoprotein 2 (SV2) (3-6), which allows rapid toxin entry via synaptic vesicles. Acidification of the synaptic vesicle lumen triggers H N to form a channel to facilitate LC translocation into the cytosol. Intracellular LC cleaves a SNARE (soluble N-ethylmaleimide-sensitive factor [NSF] attachment protein receptor) protein (7-12). SNARE protein cleavage inhibits exocytosis of cholinergic synaptic vesicles at neuromuscular junctions. BoNT LCs are long-lived proteases, sustaining paralysis in humans for several months depending on the serotype (2), which led to the licensing of BoNT/A and BoNT/B for human therapies (13,14).BoNT serotypes include subtypes neutralized by serotype-specific antisera (15-17). Informatics has classified eight BoNT/A subtypes (A1 to A8), which vary by ϳ10 to 15% in amino acid identity (15,16,(18)(19)(20)(21). While BoNT/A1 and BoNT/A2 cleave SNAP25 with similar kinetics (22-2...

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

Entry of Botulinum Neurotoxin Subtypes A1 and A2 into Neurons

et al. 2017

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“…BoNT/As were prepared employing a previously reported method (16) with minor modifications (17). Briefly, Clostridium botulinum type A strains 62A and Chiba-H, which belong to subtypes A1 and A2, respectively, were cultured in a PYG medium, containing 2% peptone, 0.5% yeast extract, 0.5% glucose, and 0.025% sodium thioglycolate, at 30°C for 3 days.…”

Section: Methodsmentioning

confidence: 99%

Spinal Central Effects of Peripherally Applied Botulinum Neurotoxin A in Comparison between Its Subtypes A1 and A2

et al. 2014

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Because of its unique ability to exert long-lasting synaptic transmission blockade, botulinum neurotoxin A (BoNT/A) is used to treat a wide variety of disorders involving peripheral nerve terminal hyperexcitability. However, it has been a matter of debate whether this toxin has central or peripheral sites of action. We employed a rat model in which BoNT/A1 or BoNT/A2 was unilaterally injected into the gastrocnemius muscle. On time-course measurements of compound muscle action potential (CMAP) amplitudes after injection of BoNT/A1 or BoNT/A2 at doses ranging from 1.7 to 13.6 U, CMAP amplitude for the ipsilateral hind leg was markedly decreased on the first day, and this muscle flaccidity persisted up to the 14th day. Of note, both BoNT/A1 and BoNT/A2 administrations also resulted in decreased CMAP amplitudes for the contralateral leg in a dose-dependent manner ranging from 1.7 to 13.6 U, and this muscle flaccidity increased until the fourth day and then slowly recovered. Immunohistochemical results revealed that BoNT/A-cleaved synaptosomal-associated protein of 25 kDa (SNAP-25) appeared in the bilateral ventral and dorsal horns 4 days after injection of BoNT/A1 (10 U) or BoNT/A2 (10 U), although there seemed to be a wider spread of BoNT/A-cleaved SNAP-25 associated with BoNT/A1 than BoNT/A2 in the contralateral spinal cord. This suggests that the catalytically active BoNT/A1 and BoNT/A2 were axonally transported via peripheral motor and sensory nerves to the spinal cord, where they spread through a transcytosis (cell-to-cell trafficking) mechanism. Our results provide evidence for the central effects of intramuscularly administered BoNT/A1 and BoNT/A2 in the spinal cord, and a new insight into the clinical effects of peripheral BoNT/A applications.

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“…It remains unclear how these amino acid differences affect the subtypes' biologic activity, and only a few differences in characteristics have been elucidated. BoNT/A2 has been shown to enter cells faster than BoNT/A1 (20), to more potently inhibit the grip strength in rats after local administration, and to be more potent in the hemidiaphragm assay (21,22). In addition, grip strength analysis after local injection in rats indicated that BoNT/A2 diffusion to the contralateral leg was reduced nearly 3-fold compared to A1 diffusion and no axonal transport was observed (21).…”

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Characterization of Botulinum Neurotoxin A Subtypes 1 Through 5 by Investigation of Activities in Mice, in Neuronal Cell Cultures, and In Vitro

et al. 2013

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Botulinum neurotoxins (BoNTs) are synthesized by Clostridium botulinum and exist as seven immunologically distinct serotypes designated A through G. For most serotypes, several subtypes have now been described based on nominal differences in the amino acid sequences. BoNT/A1 is the most well-characterized subtype of the BoNT/A serotype, and many of its properties, including its potency, its prevalence as a food poison, and its utility as a pharmaceutical, have been thoroughly studied. In contrast, much remains unknown of the other BoNT/A subtypes. In this study, BoNT/A subtype 1 (BoNT/A1) to BoNT/A5 were characterized utilizing a mouse bioassay, an in vitro cleavage assay, and several neuronal cell-based assays. The data indicate that BoNT/A1 to -5 have distinct in vitro and in vivo toxicological properties and that, unlike those for BoNT/A1, the neuronal and mouse results for BoNT/A2 to -5 do not correlate with their enzymatic activity. These results indicate that BoNT/A1 to -5 have distinct characteristics, which are of importance for a greater understanding of botulism and for pharmaceutical applications.

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Type A1 but Not Type A2 Botulinum Toxin Decreases the Grip Strength of the Contralateral Foreleg Through Axonal Transport From the Toxin-Treated Foreleg of Rats

Cited by 43 publications

References 28 publications

Entry of Botulinum Neurotoxin Subtypes A1 and A2 into Neurons

Entry of Botulinum Neurotoxin Subtypes A1 and A2 into Neurons

Spinal Central Effects of Peripherally Applied Botulinum Neurotoxin A in Comparison between Its Subtypes A1 and A2

Characterization of Botulinum Neurotoxin A Subtypes 1 Through 5 by Investigation of Activities in Mice, in Neuronal Cell Cultures, and In Vitro

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