Action Of Botulinum Neurotoxin Research Articles

Mechanism of action of botulinum neurotoxin in hyperhidrosis

Mechanism of action of botulinum neurotoxin in hyperhidrosis

Re: Kanai A, et al. mechanisms of action of botulinum neurotoxins, β3‐adrenergic receptor agonists, and PDE5 inhibitors in modulating detrusor function in overactive bladders: ICI‐RS 2011. Neurourol urodyn 2012;31:300–308

Re: Kanai A, et al. mechanisms of action of botulinum neurotoxins, β3‐adrenergic receptor agonists, and PDE5 inhibitors in modulating detrusor function in overactive bladders: ICI‐RS 2011. Neurourol urodyn 2012;31:300–308

Mechanisms of action of botulinum neurotoxins, β3‐adrenergic receptor agonists, and PDE5 inhibitors in modulating detrusor function in overactive bladders: ICI‐RS 2011

Botulinum neurotoxins type A (BoNT/A), β(3)-adrenergic receptor agonists, and phosphodiesterase type 5 (PDE5) inhibitors are promising agents that mitigate lower urinary tract symptoms by attenuating the sensory system. However, whether they act directly on afferent nerves or indirectly through the other cell types is unclear. Spinal cord transected female mice were used as a model for neurogenic bladder overactivity. In vivo methods utilized decerebrate mouse cystometry. In vitro approaches included optical mapping of Ca(2+) transient, single unit afferent nerve recordings and tension measurements from bladder sheets and wall cross-sections. Immunohistochemistry was used to measure the expression of β(3)-adrenergic receptors on dorsal root ganglion (DRG) neurons. Our unique approaches revealed the direct effects of BoNT/A in inhibiting neuropeptide release and firing rates in afferents following bladder injections. β(3)-adrenergic receptor agonists are demonstrated to directly inhibit afferent nerve firing independent of the relaxing effects on bladder smooth muscle. Moreover, data suggest the expression of these receptors on DRG neurons that send projections to the bladder. The mechanism of action of PDE5 inhibitors on bladder overactivity is discussed. The questions raised during the plenary session of the 2011 International Consultation on Incontinence-Research Society meeting regarding the benefits of BoNT/A, β(3)-adrenergic receptor agonist and PDE5 inhibitor treatments of overactive bladder are addressed. Our findings suggest that the abovementioned agents, in low enough concentrations, can directly inhibit afferent excitability without decreasing detrusor contractility. Accordingly, they have considerable potential for treating the sensory component of lower urinary tract dysfunctions.

Botulinum Neurotoxin for Pain Management: Insights from Animal Models

The action of botulinum neurotoxins (BoNTs) at the neuromuscular junction has been extensively investigated and knowledge gained in this field laid the foundation for the use of BoNTs in human pathologies characterized by excessive muscle contractions. Although much more is known about the action of BoNTs on the peripheral system, growing evidence has demonstrated several effects also at the central level. Pain conditions, with special regard to neuropathic and intractable pain, are some of the pathological states that have been recently treated with BoNTs with beneficial effects. The knowledge of the action and potentiality of BoNTs utilization against pain, with emphasis for its possible use in modulation and alleviation of chronic pain, still represents an outstanding challenge for experimental research. This review highlights recent findings on the effects of BoNTs in animal pain models.

Use of botulinum toxin in the neurology clinic

Botulinum neurotoxin (BoNT) is an effective treatment for conditions associated with overactivity of glandular, smooth or skeletal muscle, and this toxin can also ameliorate certain painful conditions. Electromyography, endoscopy and imaging techniques such as ultrasonography and fluoroscopy have been used to increase the accuracy of BoNT injections. This Review assesses the mechanisms of action of BoNT, and examines the use of BoNT injections in numerous neurological conditions, including dystonia, spasticity, headaches and other painful disorders, hemifacial spasm, essential tremor, motor tics, hyperhidrosis, and sialorrhea and drooling. Important practical aspects, such as the reconstitution of BoNT, dosing, and methods of administration, are also addressed.

International consensus statement for the use of botulinum toxin treatment in adults and children with neurological impairments – introduction

Botulinum neurotoxin (BoNT) is most commonly used to reduce focal over-activity in skeletal muscle, although newer indications such as management of drooling, pain and tremor are emerging. Treatment of spasticity incorporating BoNT is usually part of an integrated multidisciplinary rehabilitation programme. Prior to initiating this therapy, specific functional limitations, goals and expected outcomes of treatment should be discussed with the patient/carers. Muscle selection and the order/priority of treatment should be agreed. Treatment goals may involve increasing active or passive function or the avoidance of secondary complications or impairment progression. This paper describes the basic science mechanisms of the action of BoNT and subsequent nerve recovery and introduces a supplement comprising the best available evidence and expert opinion from international panels on questions of assessment, indications, BoNT regimen, adjunctive therapy, expected outcomes and recommended monitoring. Speciality areas reviewed include Paediatric Lower Limb Hypertonicity, Paediatric Upper Limb Hypertonicity, Adult Lower Limb Hypertonicity, Adult Upper Limb Hypertonicity, Cervical Dystonia, Drooling and Pain and Niche Indications. There is good quality scientific evidence to support the efficacy of BoNT to reduce muscle over-activity in the limbs secondary to central nervous system disorders in adults and children, to address primary or secondary cervical dystonia, to reduce saliva flow and to treat some pain syndromes. There is emergent evidence for the efficacy of BoNT to reduce focal tremor, to treat other types of pain including neuropathic pain and also to improve function following treatment of focal muscle over-activity.

Rapid product analysis and increased sensitivity for quantitative determinations of botulinum neurotoxin proteolytic activity

The ultimate molecular action of botulinum neurotoxin (BoNT) is a Zn-dependent endoproteolytic activity on one of the three SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins. There are seven serotypes (A–G) of BoNT having distinct cleavage sites on the SNARE substrates. The proteolytic activity is located on the N-terminal light chain (Lc) domain and is used extensively as the primary target toward therapeutic development against botulism. Here we describe an improved method using ultra-performance liquid chromatography (UPLC) whereby quantitative data were obtained in 1/10th the time using 1/20th the sample and solvent volumes compared with a widely used high-performance liquid chromatography (HPLC) method. We also synthesized a VAMP (vesicle-associated membrane protein)-based peptide containing an intact V1 motif that was efficiently used as a substrate by BoNT/D Lc. Although serotype C1 cleaves the serotype A substrate at a bond separated by only one residue, we were able to distinguish the two reactions by UPLC. The new method can accurately quantify as low as 7pmol of the peptide substrates for BoNT serotypes A, B, C1, and D. We also report here that the catalytic efficiency of serotype A can be stimulated 35-fold by the addition of Triton X-100 to the reaction mixture. Combining the use of Triton X-100 with the newly introduced UPLC method, we were able to accurately detect very low levels of proteolytic activity in a very short time. Sensitivity of the assay and accuracy and rapidity of product analysis should greatly augment efforts in therapeutic development.

A Targeted Therapeutic Rescues Botulinum‐A Poisoned Neurons

Botulinum neurotoxin (BoNT) is the most poisonous poison known to mankind for which currently no antidote is available to rescue poisoned nerves. An effective medical countermeasure strategy will be required and would include its efficient delivery specifically to presynaptic nerve terminals. Here we present the report of rescue of botulinum poisoned nerve cells by Mastoparan‐7, a peptide constituent of bee venom that was delivered through a drug delivery vehicle (DDV) constructed from the non‐toxic fragment of botulinum neurotoxin itself. We found that mastoparan‐7 that was delivered into spinal cord cells rescued BoNT/A poisoned cells by restoring over 40% neurotransmitter release. The rescue of the cell poisoning does not seem to occur from inhibition of the endopeptidase activity of BoNT/A against its well known substrate, SNAP‐25, mechanistically involved in the exocytosis process. Rather, Mastoparan‐7 induces a physiological host response apparently unrelated to SNAP‐25 but linked to the phospholipase signal transduction pathway. Our results open new avenues to examine mechanism of exocytosis process, and also to examine alternative cellular mechanism of botulinum neurotoxin action. Successful demonstration of BoNT‐based DDV opens door for its utility for drug delivery against many neuronal and neuron‐muscular disorders.This work was supported JSTO‐CBD/DTRA #3.10003_05_WR_B

Is botulinum toxin useful in treating headache? No

The discrepancy between the widespread use of botulinum neurotoxin (BoNT) in managing headache and the supporting clinical evidence is unprecedented. No substance seems to have inspired more physicians and patients to undertake spirited treatment attempts. Tremendous treatment success in small, uncontrolled clinical trials has been repeatedly reported, but no substance that has been studied to an equal extent has so utterly failed to provide proof of effect in controlled clinical trials. Nevertheless, even though most randomized, controlled clinical trials have not met their defined primary outcome criterion, BoNT is still considered a promising treatment alternative for primary headache disorders. Experimental approaches to the pathophysiologic impact of BoNT on the perception of pain have been equally unsuccessful. Although most studies have been unable to find a direct antinociceptive effect in humans, some researchers continue to seek specific injection sites or injection techniques that may promise more successful results. Others look for a positive effect by narrowing the indications for BoNT to more homogenous symptoms or special patient subgroups. The results of randomized, controlled studies involving a total of 3552 patients indicate that BoNT injection is probably ineffective for patients with migraine and chronic tension-type headache regardless of injection site, dosage, or injection regimen, and there is insufficient evidence to draw a conclusion about its effectiveness for the treatment of chronic daily headache or subforms. The lack of direct experimental or clinical trial evidence that BoNT has a direct antinociceptive effect in humans must be addressed before more trials are conducted, involving even more patients. Additional pathophysiologically oriented research is also needed to unravel the mechanisms of action of BoNT in human pain perception or, alternatively, to bring it all down to the placebo effect.

A Targeted Therapeutic Rescues Botulinum Toxin-A Poisoned Neurons

AbstractBotulinum neurotoxin (BoNT), a Category A biothreat agent, is the most potent poison known to mankind. Currently no antidote is available to rescue poisoned synapses. BoNT acts specifically by blocking neurotransmission primarily at peripheral nerve-muscle junctions causing severe flaccid muscle paralysis, which is fatal if proper medical care is not provided. The neurotoxin acts by specifically entering the presynaptic nerve endings where it interferes with the biochemical machinery involved in the process of neurotransmitter release, i.e., neuroexocytosis. Most serotypes of BoNT are known to remain active for weeks to months after entering the nerves, but BoNT/A is the most potent and long lasting in causing muscle paralysis. An effective medical countermeasure strategy requires developing a drug that could rescue poisoned neuromuscular synapses, and would include its efficient delivery specifically to presynaptic nerve terminals. Here we report rescuing of botulinum poisoned nerve cells by Mastoparan-7 (Mas-7), a peptide constituent of bee venom, that was delivered through a drug delivery vehicle (DDV) constructed from the non-toxic fragment of botulinum neurotoxin itself. We found that Mas-7 that was delivered into BoNT/A intoxicated cultured mouse spinal cord cells restored over 40% of stimulated neurotransmitter release. The rescue of the cell poisoning did not occur from inhibition of the endopeptidase activity of BoNT/A against its well known substrate, SNAP-25 that is mechanistically involved in the exocytosis process. Rather, Mas-7 induced a physiological host response apparently unrelated to SNAP-25, but linked to the phospholipase signal transduction pathway. In addition to providing the first effective antidote against botulism, our results open new avenues to study the mechanism of exocytosis, and also to examine an alternative cellular mechanism of botulinum neurotoxin action. An effective BoNT-based DDV can also be utilized for drug delivery against many neuronal and neuromuscular disorders.

Multiple events triggered by the use of botulinum neurotoxins at the vertebrate neuromuscular junction

The three main cellular components of the neuromuscular junction (NMJ) are affected by a single local injection of botulinum neurotoxin (BoNT). Motor nerve terminals are known to synthesize and release acetylcholine (ACh) spontaneously, or in response to nerve stimuli. The main action of BoNTs is to reduce the number of ACh quanta released, without affecting transmitter synthesis and storage. According to the serotype used, quantal ACh release remains synchronized (BoNT/A, /E, and /C), or is desynchronized (BoNT/B, /D, /F/ and /G). These effects are related to the synaptic proteins cleaved by BoNTs. The reduction in quantal ACh release prevents nerve-evoked synaptic responses to reach the threshold for action potential generation in muscle fibres and blocks nerve- evoked muscle contractions. The duration of the neuromuscular block depends on the BoNT serotype, the dose injected, and the animal species considered. Muscle inactivity in turn induces an important atrophy of muscle fibres, with no evident damage to NMJ components, and triggers an overgrowth of intramuscular axons, nerve terminals, and perisynaptic Schwann cells. The events leading to the maturation of the newly formed sprouts, the development of the first nerve–muscle contacts, the differentiation of new endplates, and the elimination of superfluous synapses after BoNTs injection is a remarkable demonstration of synaptic plasticity at the mature NMJ.

Proteomic changes in rat thyroarytenoid muscle induced by botulinum neurotoxin injection

Botulinum neurotoxin (BoNT) injection into the thyroarytenoid (TA) muscle is a commonly performed medical intervention for adductor spasmodic dysphonia. The mechanism of action of BoNT at the neuromuscular junction is well understood, however, aside from reports focused on myosin heavy chain isoform abundance, there is a paucity of data addressing the effects of therapeutic BoNT injection on the TA muscle proteome. In this study, 12 adult Sprague Dawley rats underwent unilateral TA muscle BoNT serotype A injection followed by tissue harvest at 72 h, 7 days, 14 days, and 56 days postinjection. Three additional rats were reserved as controls. Proteomic analysis was performed using 2-D SDS-PAGE followed by MALDI-MS. Vocal fold movement was significantly reduced by 72 h, with complete return of function by 56 days. Twenty-five protein spots demonstrated significant protein abundance changes following BoNT injection, and were associated with alterations in energy metabolism, muscle contractile function, cellular stress response, transcription, translation, and cell proliferation. A number of protein abundance changes persisted beyond the return of gross physiologic TA function. These findings represent the first report of BoNT-induced changes in any skeletal muscle proteome, and reinforce the utility of applying proteomic tools to the study of system-wide biological processes in normal and perturbed TA muscle function.

Evaluation of toosendanin as a botulinum neurotoxin antagonist

Toosendanin (TSN), a tetranortriterpenoid compound from the plant Melia toosendan, has been reported to protect rats and non-human primates from the lethal actions of botulinum neurotoxin A (BoNT/A). TSN was tested in isolated mouse phrenic nerve-hemidiaphragm preparations against BoNT serotypes A, B and E. When added prior to toxin, TSN was found to delay the onset of muscle paralysis with all 3 BoNT serotypes. TSN had little or no effect on BoNT-induced muscle paralysis when applied 30 min after BoNT. It is suggested that the antagonist activity of TSN may result from inhibition of BoNT translocation.

Manipulation of Signal Transduction by Botulinum Neurotoxins and their Derivatives

Botulinum neurotoxins produced by various Clostridium sp are the most potent acute lethal toxins known, and yet they have found increasing use in the clinical treatment of diseases or conditions involving neuromuscular or autonomic neuronal transmission. By a process involving proteolytic cleavage of one or more SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins, botulinum toxins inhibit the release of acetylcholine from peripheral cholinergic nerve terminals. Cleavage of SNARE proteins is both specific and effective, and leads to blockade of secretory vesicle fusion that is maintained for many weeks or months. Though inhibition of release of transmitter is currently the most clinically-relevant endpoint, SNARE proteolysis can also inhibit presentation of receptors, channels and other surface-membrane located materials. The implications for signal transduction are only just becoming apparent, but are an important facet of the mechanism of action of botulinum neurotoxins. Additionally, the ubiquitous distribution of the SNARE proteins ensures that proteolysis-dependent inhibition of transmitter release and receptor/channel presentation is theoretically not confined to neuronal targets. This review will consider the potential of botulinum neurotoxins as inhibitors of intercellular communication, and highlight the general concept of deriving novel therapeutic molecules from the neurotoxins. Keywords: Clostridium, botulinum toxin, SNARE, exocytosis, neurotoxin

Future aspects of botulinum neurotoxins

The future of botulinum neurotoxin (BoNT) development is expected to proceed along two lines: the development of novel indications and the development of novel products. New indications will likely be based on the neuromuscular mode of action of BoNTs, as well as action on primary sensory fibers and other neuronal types. Novel BoNT products may be designed for increased specificity or enhanced duration. As new products enter the market, it will be important for each to demonstrate efficacy and safety. Unfortunately, the future of BoNTs will also likely include attempts to obtain and distribute unlicensed and illegal BoNT products that may pose substantial risks to patients.

Action of botulinum neurotoxins in the central nervous system: Antiepileptic effects

Botulinum neurotoxins (BoNTs) are metalloproteases which act on nerve terminals and cause a long-lasting inhibition of neurotransmitter release. BoNTs act by cleaving core proteins of the neurotransmitter release machinery, namely the SNARE (soluble NSF-attachment receptors) proteins. The action of BoNTs in the peripheral nervous system (PNS) has been extensively documented, and knowledge gained in this field laid the foundations for the use of BoNTs in human disorders characterized by hyperfunction of peripheral nerve terminals. Much less is known about the action of BoNTs on the central nervous system (CNS). In vitro studies have demonstrated that BoNTs can affect the release of several neurotransmitters from central neurons. Recent studies have provided the first characterization of the effects of BoNT/E on CNS neurons in vivo. It has been shown that BoNT/E injected into the rat hippocampus inhibits glutamate release and blocks spike activity of pyramidal neurons. Intrahippocampal injection of BoNT/E resulted in significant inhibition of seizure activity in experimental models of epilepsy, suggesting a potential therapeutic use of BoNTs in the CNS.

Botulinum a toxin for the treatment of benign prostatic hyperplasia/lower urinary tract symptoms

Botulinum neurotoxin (BoNT) has been called the most poisonous poison and a potential bioterrorism weapon, and yet modern medicine has been able to harvest the elegant and specific activity of this toxin to treat a variety of medical conditions. BoNT application recently has been extended to prostate disorders, and this article reviews the literature on the mechanisms of action and clinical efficacy of BoNT treatment in the prostate. BoNT has demonstrated promising preliminary results for male lower urinary tract symptoms, and translational research suggests novel mechanism of action of BoNT in the prostate. It is important to remember that the application of BoNT in the prostate is not approved by the regulatory agencies and caution should be applied until larger randomized clinical studies are completed.

The potential and promise of using botulinum toxin in the prostate gland

Botulinum neurotoxin (BoNT) has been called the 'most poisonous poison' and a potential bioterrorist weapon. However, it is possible to use the elegant and specific activity of this toxin to treat various medical conditions. Recently, the application of BoNT was extended to prostate disorders, and we review published reports on the mechanisms of action and clinical efficacy of BoNT in the prostate. The information was gathered from Medline, abstracts from recent urological meetings, and from personal experience. BoNT has shown promising preliminary results for male lower urinary tract symptoms, and translational research suggests a novel mechanism of action of BoNT in the prostate. Importantly, the application of BoNT in the prostate is not approved by the regulatory agencies and caution should be used until larger randomized clinical studies are completed.

Beyond BOTOX: advantages and limitations of individual botulinum neurotoxins

In the past decade, botulinum toxin has been widely used for treatment of neuromuscular disorders, as well as for cosmetic purposes. This review provides some insights into the mechanisms of action of botulinum neurotoxins; it also provides a condensed comparative guide to the seven botulinum toxins that target the synaptic release machinery to cause synaptic blockade of various durations. Botulinum neurotoxins are the most potent toxins designed by nature. They are produced by Clostridium bacteria to cause long-lasting paralysis and death. Botulinum neurotoxin type A is the toxin currently widely employed in clinical neurology and in cosmetic clinics. The seven botulinum neurotoxins all act by targeting the same vesicle fusion machinery, and yet they cause paralysis of different durations. The distinct behaviors of botulinum neurotoxins are discussed, and advantages and limitations for their future exploitation are highlighted.—Valérie Biousse

Ethyl acetate extract from black tea prevents neuromuscular blockade by botulinum neurotoxin type A in vitro

Botulinum neurotoxin produced by Clostridium botulinum is the strongest neurotoxin and causes botulism in mammals. The current study aimed to find an inactivator for botulinum neurotoxin in black, oolong, roasted, and green teas. The ability of the four teas to inactivate the neuromuscular blocking action of botulinum neurotoxin was determined. Water extracts from black, oolong, and roasted teas protected against the toxicity of botulinum neurotoxin type A in mouse phrenic nerve–diaphragm preparations. The order of potency of the water extracts was black tea > oolong tea > roasted tea > green tea (no effect). The effects of several organic solvent extracts of black tea water extract were examined, and the order of potency was ethyl acetate extract > butanol extract = remaining extract > chloroform extract (no effect). Ethyl acetate extracts from oolong, roasted, and green tea water extracts also exhibited a stronger protecting effect than chloroform, butanol, and remaining extracts from these teas, but they had weaker protective effect than ethyl acetate extract from black tea water extract. These protective effects occurred only when each extract was pre-mixed with the toxin before the assay, and they were not modified by mixing each extract with bovine serum albumin (BSA) before adding the toxin. These results indicate that ethyl acetate extract from black tea is the best source for searching for tea-derived inactivating substance(s) of botulinum neurotoxin.