Acetylcholine Receptor Density Research Articles

Neuroprotective effect of long-term resistance physical exercise against memory damage elicited by a lipopolysaccharide-induced neuroinflammation model in malerats.

Resistance exercise training (RET) is considered an excellent tool for preventing diseases with an inflammatory background. Its neuroprotective, antioxidant, and anti-inflammatory properties are responsible for positively modulating cholinergic and oxidative systems, promoting neurogenesis, and improving memory. However, the mechanisms behind these actions are largely unknown. In order to investigate the pathways related to these effects of exercise, we conducted a 12-week long-term exercise training protocol and used lipopolysaccharide (LPS) to induce damage to the cortex and hippocampus of male Wistar rats. The cholinergic system, oxidative stress, and histochemical parameters were analyzed in the cerebral cortex and hippocampus, and memory tests were also performed. It was observed that LPS: (1) caused memory loss in the novel object recognition (NOR) test; (2) increased the activity of acetylcholinesterase (AChE) and Iba1 protein density; (3) reduced the protein density of brain-derived neurotrophic factor (BDNF) and muscarinic acetylcholine receptor M1 (CHRM1); (4) elevated the levels of lipid peroxidation (TBARS) and reactive species (RS); and (5) caused inflammatory damage to the dentate gyrus. RET, on the other hand, was able to prevent all alterations induced by LPS, as well as increase per se the protein density of the alpha-7 nicotinic acetylcholine receptor (nAChRα7) and Nestin, and the levels of protein thiols (T-SH). Overall, our study elucidates some mechanisms that support resistance physical exercise as a valuable approach against LPS-induced neuroinflammation and memory loss.

The M 1 -muscarinic acetylcholine receptor subtype may play a role in learning and memory performance in the hippocampus of neonatal monosodium glutamate-obese rats.

Here, we investigate the effects of obesity induced by monosodium glutamate (MSG) on cognitive impairment and whether this model induces any alteration in the affinity, density, and subtypes of muscarinic acetylcholine receptors (mAChRs) in rat hippocampus. Healthy rats were used as controls, and MSG-obese rats were selected via the Lee index > 0.300. The effects of MSG-induced obesity on hippocampal spatial learning and memory processes were evaluated by using the working memory versions of the Morris' water maze task and the evaluation of mAChRs by binding assay and their subtypes by immunoprecipitation assays. [ 3 H]Quinuclidinyl benzilate specific binding analysis showed that the equilibrium dissociation constant (K D ) did not differ between control and MSG, indicating that affinity is not affected by obesity induced by MSG. The maximum number of binding sites (B max ) obtained in MSG subjects was lower than that obtained from control rats, indicating a decrease in the expression of total mAChRs. Immunoprecipitation assays reveal a decrease in the expression of M 1 subtype of MSG when compared with control rats (M 2 to M 5 subtypes did not differ between control and MSG). We also observed that MSG promotes a disruption of the spatial working memory which was accompanied by a decrease in the M 1 mAChR subtype in rat hippocampus, thus suggesting deleterious long-term effects besides the obesity. In conclusion, these findings provide new insights into how obesity can influence spatial learning and memory that is hippocampal-dependent. The data suggest that the M 1 mAChR subtype protein expression is a potential therapeutic target.

Swimming exercise and clove oil can improve memory by molecular responses modification and reduce dark cells in rat model of Alzheimer's disease

Alzheimer's disease (AD) is marked by reduced acetylcholine receptor (AChR) density and an increase in nucleotide oligomerization domain (NOD)-like receptors NLR family, pyrin domain containing 1 (NLRP1). We examined the effect of swimming and consumption of clove supplements on memory, dark cells, and α7nAChR and NLRP1 mRNA and protein expression in the hippocampus of the rat model of AD. Forty-eight rats were divided into six groups: sham (sh), healthy-control (HC), Alzheimer (-control (AC), -training (AT), -training-supplement (ATS), and -supplement (AS)). Alzheimer was induced by injection of amyloid β1–42 (Aβ1–42). Swimming exercise protocol (30 min) and gavaging clove supplement (0.1 mg/kg) were administered daily for three weeks. The results indicated that in response to AD, α7 nicotinic acetylcholine receptor (α7nAChR) mRNA and protein rate (p = 0.001) and memory (p = 0.003) were significantly decreased. In contrast, NLRP1 mRNA and protein rate (p = 0.001) and dark cells (p = 0.001) were significantly increased. This is while exercise and clove supplementation improved Alzheimer-induced changes in α7nAChR, NLRP1, memory, and dark cells (p < 0/05). The present study indicated that exercising and consuming clove supplementation could improve memory by increasing α7nAChR and decreasing NLRP1 and dark cells.

Mapping the Nicotinic Acetylcholine Receptor Nanocluster Topography at the Cell Membrane with STED and STORM Nanoscopies.

The cell-surface topography and density of nicotinic acetylcholine receptors (nAChRs) play a key functional role in the synapse. Here we employ in parallel two labeling and two super-resolution microscopy strategies to characterize the distribution of this receptor at the plasma membrane of the mammalian clonal cell line CHO-K1/A5. Cells were interrogated with two targeted techniques (confocal microscopy and stimulated emission depletion (STED) nanoscopy) and single-molecule nanoscopy (stochastic optical reconstruction microscopy, STORM) using the same fluorophore, Alexa Fluor 647, tagged onto either α-bungarotoxin (BTX) or the monoclonal antibody mAb35. Analysis of the topography of nanometer-sized aggregates (“nanoclusters”) was carried out using STORMGraph, a quantitative clustering analysis for single-molecule localization microscopy based on graph theory and community detection, and ASTRICS, an inter-cluster similarity algorithm based on computational geometry. Antibody-induced crosslinking of receptors resulted in nanoclusters with a larger number of receptor molecules and higher densities than those observed in BTX-labeled samples. STORM and STED provided complementary information, STED rendering a direct map of the mesoscale nAChR distribution at distances ~10-times larger than the nanocluster centroid distances measured in STORM samples. By applying photon threshold filtering analysis, we show that it is also possible to detect the mesoscale organization in STORM images.

Exercise-induced gene expression changes in skeletal muscle of old mice

Exercise is believed to be beneficial for skeletal muscle functions across all ages. Regimented exercise is often prescribed as an effective treatment/prophylaxis for age-related loss of muscle mass and function, known as sarcopenia, and plays an important role in the maintenance of mobility and functional independence in the elderly. However, response to exercise declines with aging, resulting in limited gain of muscle strength and endurance. These changes likely reflect age-dependent alterations in transcriptional response underlying the muscular adaptation to exercise. The exact changes in gene expression accompanying exercise, however, are largely unknown, and elucidating them is of a great clinical interest for understanding and optimizing the exercise-based therapies for sarcopenia.In order to characterize the exercise-induced transcriptomic changes in aged muscle, a paired-end RNA sequencing was performed on rRNA-depleted total RNA extracted from the gastrocnemius muscles of 24 months-old mice after 8 weeks of regimented exercise (exercise group) or no formal exercise program (sedentary group).Differential gene expression analysis of aged skeletal muscle revealed upregulations in the group of genes involved in neurotransmission and neuroexcitation, as well as equally notable absence of anabolic gene upregulations in the exercise group. In particular, genes encoding the transporters and receptor components of glutaminergic transmission were significantly upregulated in exercised muscles, as exemplified by Gria 1, Gria 2 and Grin2c encoding glutamate receptor 1, 2 and 2C respectively, Grin1 and Grin2b encoding N-methyl-d-aspartate receptors (NMDARs), Nptx1 responsible for glutaminergic receptor clustering, and Slc1a2 and Slc17a7 regulating synaptic uptake of glutamate. These changes were accompanied by an increase in the post-synaptic density of NMDARs and acetylcholine receptors (AChRs), as well as their innervation at neuromuscular junctions (NMJs).These results suggest that neural responses predominate the adaptive response of aged skeletal muscle to exercise, and indicate a possibility that glutaminergic transmission at NMJs may be present and responsible for synaptic protection and neural remodeling accompanying the exercise-induced functional enhancement in aged skeletal muscle. In addition, the absence of upregulations in the anabolic pathways highlights them as the area of potential pharmacological targeting for optimizing exercise-led sarcopenia therapy.

GABAB receptors blockage modulates somatic and aversive manifestations induced by nicotine withdrawal

There is substantial evidence that GABAB agonist, baclofen, prevents somatic and motivational responses induced by nicotine withdrawal and may target drug cue vulnerabilities in humans. In this context, we explored different aspects associated with the possible mechanisms whereby the GABAB receptors might influence nicotine withdrawal. Male mice received nicotine (2.5 mg/kg, s.c.) 4 times daily, for 7 consecutive days. Nicotine-treated mice received the nicotinic acetylcholine receptor antagonist, mecamylamine (MEC, 2 or 3.5 mg/kg, s.c.), to precipitate the withdrawal state. A second group of dependent mice received 2-hydroxysaclofen (GABAB receptor antagonist, 1 mg/kg, s.c.) before MEC-precipitated abstinence. Somatic signs of nicotine withdrawal were measured for 30 min. Anxiogenic-like response associated to nicotine withdrawal was assessed by the elevated plus maze test. The dysphoric/aversive effect induced by nicotine withdrawal was evaluated using conditioned place aversion paradigm. Dopamine, serotonin and its metabolites concentrations were determined by HPLC in the striatum, cortex and hippocampus. Finally, α4β2 nicotinic acetylcholine receptor density was determined in several brain regions using autoradiography assays. The results showed that MEC-precipitated nicotine withdrawal induced somatic manifestations, anxiogenic-like response and dysphoric/aversive effect, and 2-hydroxysaclofen potentiated these behavioral responses. Additionally, 2-hydroxysaclofen was able to change striatal dopamine levels and α4β2 nicotinic acetylcholine receptor density, both altered by MEC-precipitated nicotine withdrawal. These findings provide important contributions to elucidate neurobiological mechanisms implicated in nicotine withdrawal. We suggest that GABAB receptor activity is necessary to control alterations induced by nicotine withdrawal, which supports the idea of targeting GABAB receptors to treat tobacco addiction in humans.

Control of Neuronal Excitability by Cell Surface Receptor Density and Phosphoinositide Metabolism.

Phosphoinositides are members of a family of minor phospholipids that make up about 1% of all lipids in most cell types. Despite their low abundance they have been found to be essential regulators of neuronal activities such as action potential firing, release and re-uptake of neurotransmitters, and interaction of cytoskeletal proteins with the plasma membrane. Activation of several different neurotransmitter receptors can deplete phosphoinositide levels by more than 90% in seconds, thereby profoundly altering neuronal behavior; however, despite the physiological importance of this mechanism we still lack a profound quantitative understanding of the connection between phosphoinositide metabolism and neuronal activity. Here, we present a model that describes phosphoinositide metabolism and phosphoinositide-dependent action potential firing in sympathetic neurons. The model allows for a simulation of activation of muscarinic acetylcholine receptors and its effects on phosphoinositide levels and their regulation of action potential firing in these neurons. In this paper, we describe the characteristics of the model, its calibration to experimental data, and use the model to analyze how alterations of surface density of muscarinic acetylcholine receptors or altered activity levels of a key enzyme of phosphoinositide metabolism influence action potential firing of sympathetic neurons. In conclusion, the model provides a comprehensive framework describing the connection between muscarinic acetylcholine signaling, phosphoinositide metabolism, and action potential firing in sympathetic neurons which can be used to study the role of these signaling systems in health and disease.

Binding properties to nicotinic acetylcholine receptors can explain differential toxicity of neonicotinoid insecticides in Chironomidae

Neonicotinoids are neuroactive insecticides commonly detected in freshwater ecosystems. Recent studies have indicated that these compounds are markedly toxic to Chironomidae, a widespread family of ecologically important aquatic insects. However, despite their sensitivity, the pharmacological mechanisms driving neonicotinoid toxicity have yet to be characterized in these insect species. Here, we used a combination of saturation and competition binding studies to characterize neonicotinoid binding properties to nicotinic acetylcholine receptors (nAChR) in two different Chironomidae (Chironomus riparius and Chironomus dilutus) at two different life stages (larval and adult). Using radiolabeled imidacloprid ([3H]-IMI), we characterized and compared receptor density (Bmax), imidacloprid binding affinity (KD), and receptor binding affinity (Ki) to three different neonicotinoid competitors (imidacloprid, clothianidin, and thiamethoxam). We then compared receptor density and binding affinity parameters derived for Chironomidae to data previously generated for other dipterans and agricultural pests. We found that there were limited differences in neonicotinoid binding between C. riparius and C. dilutus, with both organisms demonstrating high affinities for imidacloprid (KD = 0.22−0.87 nM) and high receptor densities (Bmax = 0.92–6.53 pmol/mg). However, there were significant differences between life-stages, with larvae expressing higher densities of nicotinic acetylcholine receptors and higher imidacloprid affinities than adults. Moreover, there were compound-specific differences in receptor affinity, with larval stages displaying relative affinities (Ki) that generally correlated with acute neonicotinoid toxicity (e.g. clothianidin ≥ imidacloprid >>> thiamethoxam). Finally, compared to other dipterans and agricultural pests, Chironomidae display very high densities of high affinity nicotinic acetylcholine receptors, which likely contribute to their sensitivity. Results indicated that receptor-level differences in neonicotinoid binding may be responsible for ecotoxicological differences amongst distinct insect species, and they likely play a role in life stage-, and compound-level toxicity differences previously observed for Chironomidae. Overall, this study highlights the value of understanding the toxicological mechanisms of action of neonicotinoids in sensitive, non-target aquatic insects, to better predict adverse effects associated with unintentional neonicotinoid exposure.

Influence of different energy patterns on efficacy of radial shock wave therapy

BackgroundOne of the mechanisms of the efficacy of extracorporeal shock wave therapy (ESWT) for impaired muscle coordination of limbs is the destruction of Acetylcholine receptors (AChRs) at neuromuscular junction. The highly increased density of AChRs can change the acoustic impedance, and this change of the acoustic impedance can be the reason that radial shock wave (rESW) destroy AChRs selectively. However, the relationship between applied energy of rESWs and the therapeutic efficacy remains unclear, although some studies compared the clinical efficacy of rESWT between high- and low-energy applications. This study aimed to compare the CMAP change among different energy and pulses of rESW application. MethodsMale Sprauger–Dawley rats were used. A device that generates radial shock waves pneumatically, was used to apply the following six patterns of radial extracorporeal shockwaves in different energy flux densities and pulses to the right calf of each rat: 1, 8000 pulses at 0.045 mJ/mm2; 2, 4000 pulses at 0.09 mJ/mm2; 3, 2000 pulses at 0.18 mJ/mm2; 4, 4000 pulses at 0.045 mJ/mm2; 5, 2000 pulses at 0.09 mJ/mm2; 6, 1000 pulses at 0.18 mJ/mm2. Left calf muscles were considered controls. ResultsThere was a significant reduction in CMAP amplitude between control and rESW-exposed muscles in the group applied 4000 pulses with EFD at 0.09 mJ/mm2 and the group applied 2000 pulses with EFD at 0.18 mJ/mm2. However, the group applied 8000 pulses with EFD at 0.045 mJ/mm2 and all groups which was exposed to total 180 mJ rESW application did not show a significantly decreased CMAP amplitude compared with the untreated side. ConclusionsTotal energy and energy flux density correlate with a decrease in CMAP amplitude by rESW application. These findings could be availed by clinicians in actual clinical setting for the proper application of rESW.

The disassembly of the neuromuscular synapse in high-fat diet-induced obese male mice

ObjectiveA sustained high fat diet in mice mimics many features of human obesity. We used male and female Non-Swiss albino mice to investigate the impact of short and long-term high-fat diet-(HFD)-induced obesity on the peripheral neuromuscular junction (NMJ) and whether obesity-related synaptic structural alterations were reversible after switching obese mice from HFD to a standard fat diet (SD). MethodsHFD-induced obese and age-matched control mice fed SD were used. We carried out in vivo time lapse imaging to monitor changes of synapses over time, quantitative fluorescence imaging to study the regulation of acetylcholine receptor number and density at neuromuscular junctions, and high resolution confocal microscope to study structural alterations in both the pre- and postsynaptic apparatus. ResultsTime-lapse imaging in vivo over a 9 month period revealed that NMJs of HFD obese male mice display a variety of obesity-related structural alterations, including the disappearance of large synaptic areas, significant reduction in the density/number of nicotinic acetylcholine receptor (AChRs), abnormal distribution of AChRs, high turnover rate of AChRs, retraction of axons from lost postsynaptic sites, and partially denervated synapses. The severity of these synaptic alterations is associated with the duration of obesity. However, no substantial alterations were observed at NMJs of age-matched HFD obese female mice or male mice fed with a standard or low fat diet. Intriguingly, when obese male mice were switched from HFD to a standard diet, receptor density and the abnormal pattern of AChR distribution were completely reversed to normal, whereas lost synaptic structures were not restored. ConclusionsThese results show that the obese male mice are more vulnerable than female mice to the impacts of long-term HFD on the NMJ damage and provide evidence that diet restriction can partially reverse obesity-related synaptic changes.

Increased thalamic volume and decreased thalamo-precuneus functional connectivity are associated with smoking relapse

The thalamus, with the highest density of nicotinic acetylcholine receptor (nAChR) in the brain, plays a central role in thalamo-cortical circuits that are implicated in nicotine addiction. However, little is known about whether the thalamo-cortical circuits are potentially predictive of smoking relapse. In the current study, a total of 125 participants (84 treatment-seeking male smokers and 41 age-matched male nonsmokers) were recruited. Structural and functional magnetic resonance images (MRI) were acquired from all participants. After a 12-week smoking cessation treatment with varenicline, the smokers were then divided into relapsers (n = 54) and nonrelapsers (n = 30). Then, we compared thalamic volume and seed-based thalamo-cortical resting state functional connectivity (rsFC) prior to the cessation treatment among relapsers, nonrelapsers and nonsmokers to investigate the associations between thalamic structure/function and smoking relapse. Increased thalamic volume was detected in smokers relative to nonsmokers, and in relapsers relative to nonrelapsers, especially on the left side. Moreover, decreased left thalamo-precuneus rsFC was detected in relapsers relative to nonrelapsers. Additionally, a logistic regression analysis showed that the thalamic volume and thalamo-precuneus rsFC predicted smoking relapse with an accuracy of 75.7%. These novel findings indicate that increased thalamic volume and decreased thalamo-precuneus rsFC are associated with smoking relapse, and these thalamic measures may be used to predict treatment efficacy of nicotine addiction and serve as a potential biomarker for personalized medicine.

Loss of Protein Kinase Csnk2b/CK2β at Neuromuscular Junctions Affects Morphology and Dynamics of Aggregated Nicotinic Acetylcholine Receptors, Neuromuscular Transmission, and Synaptic Gene Expression.

The protein kinase Csnk2/CK2 is important for cell proliferation, differentiation, and survival. Previously, we showed that CK2 binds distinctive proteins at neuromuscular junctions (NMJs) of mice and phosphorylates some of them. CK2 likely stabilizes clustered nicotinic acetylcholine receptors (AChRs). In the absence of the β-subunit of CK2 in skeletal muscle fibers, mice develop an age-dependent decrease of grip strength accompanied by NMJ fragmentation and impairments of neuromuscular transmission. However, the precise role of CK2β regarding the clustering of AChRs and downstream signaling at NMJs is unknown. Here, we compared conditional CK2β-deficient mice with controls and found in the mutants (1) a lower decrement of endplate potentials after repetitive stimulation and decrements of nerve-evoked compound muscle action potentials decayed more rapidly after synaptic transmission was partially blocked, (2) that their muscle weakness was partially rescued by administration of an acetylcholine esterase inhibitor, (3) fragmented NMJs and impaired AChR clustering was detected in muscles and cultured muscle cells, (4) enlarged myonuclei, (5) impaired synaptic gene expression, and (6) a high turnover rate of their AChR clusters in vivo. Altogether, our data demonstrate a role for CK2 at the NMJ by maintaining a high density of AChRs and ensuring physiological synaptic gene expression.

α7 nicotinic ACh receptors are necessary for memory recovery and neuroprotection promoted by attention training in amyloid-β-infused mice.

Attention training reverses the neurodegeneration and memory loss promoted by infusion of amyloid-β (Aβ) peptide in rats and increases the density of α7 nicotinic ACh receptors (α7nAChRs) in brain areas related to memory. Hence, we aimed to assess the role of α7nAChRs in the memory recovery promoted by attention training. C57Bl/6 mice were chronically infused with Aβ, Aβ plus the α7 antagonist methyllycaconitine (MLA), or MLA alone. Control animals were infused with vehicle. Animals were subjected weekly to the active avoidance shuttle box for 4weeks (attention training). The brain and serum were collected for biochemical and histological analysis. Aβ caused cognitive impairment, which was reversed by the weekly training, whereas Aβ+MLA also promoted memory loss but with no reversal with weekly training. MLA alone also promoted memory loss but with only partial reversal with the training. Animals infused with Aβ alone showed senile plaques in hippocampus, no change in BDNF levels in cortex, hippocampus, and serum, but increased AChE activity in cortex and hippocampus. Co-treatment with MLA increased AChE activity and senile plaque deposition in hippocampus as well as reducing BDNF in hippocampus and serum, suggesting a lack of α7nAChR function leads to a loss of neuroprotection mechanisms. The α7nAChR has a determinant role in memory recovery and brain resilience in the presence of neurodegeneration promoted by Aβ peptide. These data support further studies concerning these receptors as pharmacological targets for future therapies.

Nicotinic Acetylcholine Receptor Density in the "Higher-Order" Thalamus Projecting to the Prefrontal Cortex in Humans: a PET Study.

The parcellation of the thalamus into different nuclei involved in different corticothalamocortical loops reflects its functional diversity. The connections between the mediodorsal nucleus and the prefrontal cortex play a major role in cognition, particularly in the rapid processing of behaviorally relevant information. The thalamus is the brain region with the highest density in α4β2 nicotinic acetylcholine receptors, the main human nicotinic acetylcholine receptor subtype. The aim of this study was to investigate the possible role of the nicotinic cholinergic system in the thalamo-cortical loops measuring receptor density in different subregions of the thalamus, based on their cortical connectivity. We studied α4β2 nicotinic acetylcholine receptors using positron emission tomography and [18F]Fluoro-A-85380, a radiotracer specific for this receptor subtype, in 36 non-smoking male subjects, including 12 healthy controls and 24 patients with epilepsy. [18F]Fluoro-A-85380 ratio index of binding potential was compared by a repeated measures general linear model, including the thalamic subregions and the brain hemisphere as within-subject factor and clinical groups as between-subject factor. The "prefrontal" thalamus, the subregion including the mediodorsal nucleus, had a significantly higher nicotinic acetylcholine receptor density than all other thalamic subregions. These findings were confirmed when analyzing solely the 12 healthy controls. This particular neurochemical organization of the thalamus supports a major role of the cholinergic system in the loops between the thalamus and the prefrontal cortex. The highest nicotinic acetylcholine receptor density in the « higher-order thalamus » could partly explain the beneficial effect of acute nicotine on attentional and executive functions and possibly the pathophysiology of some neuropsychiatric disorders.

Investigational RNAi Therapeutic Targeting C5 Is Efficacious in Pre-clinical Models of Myasthenia Gravis

Complement-mediated damage to the neuromuscular junction (NMJ) is a key mechanism of pathology in myasthenia gravis (MG), and therapeutics inhibiting complement have shown evidence of efficacy in the treatment of MG. In this study, we describe the development of a subcutaneously administered N-acetylgalactosamine (GalNAc)-conjugated small interfering RNA (siRNA) targeting the C5 component of complement that silences C5 expression in the liver (ALN-CC5). Treatment of wild-type rodents with ALN-CC5 resulted in robust and durable suppression of liver C5 expression. Dose-dependent serum C5 suppression was observed in non-human primates, with a lowering of serum C5 of up to 97.5% and the concomitant inhibition of serum complement activity. C5 silencing was efficacious in ameliorating disease symptoms in two standard rat models of MG, demonstrating the key role of circulating C5 in pathology at the NMJ. Improvement in disease activity scores and NMJ pathology was observed at intermediate levels of complement activity inhibition, suggesting that complete ablation of complement activity may not be required for efficacy in MG. The pre-clinical studies of ALN-CC5 and efficacy of C5 silencing in rat models of MG support further clinical development of ALN-CC5 as a potential therapeutic for the treatment of MG and other complement-mediated disorders.

Salbutamol modifies the neuromuscular junction in a mouse model of ColQ myasthenic syndrome.

The β-adrenergic agonists salbutamol and ephedrine have proven to be effective as therapies for human disorders of the neuromuscular junction, in particular many subsets of congenital myasthenic syndromes. However, the mechanisms underlying this clinical benefit are unknown and improved understanding of the effect of adrenergic signalling on the neuromuscular junction is essential to facilitate the development of more targeted therapies. Here, we investigated the effect of salbutamol treatment on the neuromuscular junction in the ColQ deficient mouse, a model of end-plate acetylcholinesterase deficiency. ColQ−/− mice received 7 weeks of daily salbutamol injection, and the effect on muscle strength and neuromuscular junction morphology was analysed. We show that salbutamol leads to a gradual improvement in muscle strength in ColQ−/− mice. In addition, the neuromuscular junctions of salbutamol treated mice showed significant improvements in several postsynaptic morphological defects, including increased synaptic area, acetylcholine receptor area and density, and extent of postjunctional folds. These changes occurred without alterations in skeletal muscle fibre size or type. These findings suggest that β-adrenergic agonists lead to functional benefit in the ColQ−/− mouse and to long-term structural changes at the neuromuscular junction. These effects are primarily at the postsynaptic membrane and may lead to enhanced neuromuscular transmission.

Molecular and clinical relationship between live-attenuated Japanese encephalitis vaccination and childhood onset myasthenia gravis.

ObjectiveThe incidence of childhood onset myasthenia gravis (CMG) in China is higher than that in other countries; however, the reasons for this are unclear.MethodsWe investigated the clinical and immunological profiles of CMG, and assessed the potential precipitating factors. For the mouse studies, the possible implication of vaccination in the pathogenesis was explored.ResultsIn our retrospective study, 51.22% of the 4,219 cases of myasthenia gravis (MG) were of the childhood onset type. The cohort study uncovered that the pathophysiology of CMG was mediated by immune deviation, rather than through gene mutations or virus infections. The administration of the live‐attenuated Japanese encephalitis vaccine (LA‐JEV), but not the inactivated vaccine or other vaccines, in mice induced serum acetylcholine receptor (AChR) antibody production, reduced the AChR density at the endplates, and decreased both muscle strength and response to repetitive nerve stimulation. We found a peptide (containing 7 amino acids) of LA‐JEV similar to the AChR‐α subunit, and immunization with a synthesized protein containing this peptide reproduced the MG‐like phenotype in mice.InterpretationOur results describe the immunological profile of CMG. Immunization with LA‐JEV induced an autoimmune reaction against the AChR through molecular mimicry. These findings might explain the higher occurrence rate of CMG in China, where children are routinely vaccinated with LA‐JEV, compared with that in countries, where this vaccination is not as common. Efforts should be made to optimize immunization strategies and reduce the risk for developing autoimmune disorders among children. Ann Neurol 2018;84:386–400

Autoregulation of Acetylcholine Release and Micro-Pharmacodynamic Mechanisms at Neuromuscular Junction: Selective Acetylcholinesterase Inhibitors for Therapy of Myasthenic Syndromes.

Neuromuscular junctions (NMJs) are directly involved into such indispensable to life processes as respiration and locomotion. However, motor nerve forms only one synaptic contact at each muscle fiber. This unique configuration requires specific properties and constrains to be effective. The very high density of acetylcholine receptors (AChRs) of muscle type in synaptic cleft and an excess of acetylcholine (ACh) released under physiological conditions make this synapse extremely reliable. Nevertheless, under pathological conditions such as myasthenia gravis and congenital myasthenic syndromes, the safety factor can be markedly reduced. Drugs used for short-term symptomatic therapy of these pathological states, cause partial inhibition of cholinesterases (ChEs). These enzymes catalyze the hydrolysis of ACh, thus terminate its action on AChRs. Extension of the lifetime of ACh molecules compensates muscular AChRs abnormalities and, consequently, rescues muscle contractions. In this mini review, we will first outline the functional organization of the NMJ, and then, consider the concept of the safety factor and how it may be changed. This will be followed by a look at autoregulation of ACh release that influences the safety factor of NMJs. Finally, we will consider the morphological features of NMJs as a putative reserve to increase effectiveness of pathological muscle weakness therapy by ChEs inhibitors due to opportunity to use micro-pharmacodynamic mechanisms.

Brain PET Imaging of α7-nAChR with [18F]ASEM: Reproducibility, Occupancy, Receptor Density, and Changes in Schizophrenia.

BackgroundThe α7 nicotinic acetylcholine receptor increasingly has been implicated in normal brain physiology, as well as in neuropsychiatric disorders. The highly cortical distribution of α7 nicotinic acetylcholine receptor suggests a role in cognition.MethodsWe expanded the first-in-human PET imaging of α7 nicotinic acetylcholine receptor with [18F]ASEM from 5 to 21 healthy nonsmoking volunteers and added a feasibility study in 6 male patients with schizophrenia. Study aims included: (1) confirmation of test-retest reproducibility of [18F]ASEM binding, (2) demonstration of specificity by competition with DMXB-A, an α7 nicotinic acetylcholine receptor partial agonist, (3) estimation of [18F]ASEM binding potentials and α7 nicotinic acetylcholine receptor density in vivo in humans, and (4) demonstrating the feasibility of studying α7 nicotinic acetylcholine receptor as a target for schizophrenia.ResultsTest-retest PET confirmed reproducibility (>90%) (variability ≤7%) of [18F]ASEM volume of distribution (VT) estimates in healthy volunteers. Repeated sessions of PET in 5 healthy subjects included baseline and effect of inhibition after oral administration of 150 mg DMXB-A. From reduction of binding potentials, we estimated the dose-dependent occupancy of α7 nicotinic acetylcholine receptor by DMXB-A at 17% to 49% for plasma concentrations at 60 to 200 nM DMXB-A. In agreement with evidence postmortem, α7 nicotinic acetylcholine receptor density averaged 0.67 to 0.82 nM and inhibitor affinity constant averaged 170 to 385 nM. Median VT in a feasibility study of 6 patients with schizophrenia was lower than in healthy volunteers in cingulate cortex, frontal cortex, and hippocampus (P = 0.02, corrected for multiple comparions, Mann–Whitney test).ConclusionsThe current results confirm the reproducibility of [18F]ASEM VT estimates and the specificity of the tracer for α7 nicotinic acetylcholine receptor. Preliminary findings from our feasibility study of [18F]ASEM binding in patients with schizophrenia are suggestive and provide guidance for future studies with more subjects.

Extracorporeal shock wave treatment can selectively destroy end plates in neuromuscular junctions.

This study assesses the effect of radial extracorporeal shock wave (rESW) exposure on neuromuscular transmission and neuromuscular junction (NMJ) morphology. We applied 2,000 rESWs at 0.18 mJ/mm2 and a frequency of 15 Hz to the right calf of male rats, measured the compound muscle action potential (CMAP), and examined NMJ morphology using electron microscopy. Left calf muscles were used as controls. rESW exposure significantly reduced CMAP amplitude without delayed latency in exposed muscles compared with controls. All rESW-exposed muscles exhibited NMJs with irregular end plates. Mean interjunctional fold interval was significantly increased compared with controls. However, axon terminals and muscle fibers surrounding NMJs with irregular end plates were unchanged. This localized destruction of end plates may be caused by differences in acoustic impedance induced by the density of acetylcholine receptors. These results provide a possible mechanism for the effectiveness of rESW treatment for spasticity and dystonia. Muscle Nerve 57: 466-472, 2018.