scholarly journals Bioengineered model of the human motor unit with physiologically functional neuromuscular junctions

2020 ◽  
Author(s):  
Rowan P. Rimington ◽  
Jacob W. Fleming ◽  
Andrew J. Capel ◽  
Patrick C. Wheeler ◽  
Mark P. Lewis
Keyword(s):  
Motor Neuron ◽  
Motor Unit ◽  
Neuromuscular Junctions ◽  
Motor Nerve ◽  
Motor Units ◽  
Model Organisms ◽  
Synaptic Membrane ◽  
Type I ◽  
Dependent Manner ◽  
Human Motor

AbstractInvestigations of the human neuromuscular junction (NMJ) have predominately utilised experimental animals and model organisms. Consequently, there remains a paucity of data regarding the development of the human NMJ and a lack of systems that enable temporal investigation of the motor unit. This work addresses this need, providing the methodologies to bioengineer 3D models of the human motor unit. Separate maturation of primary human skeletal muscle and iPSC derived motor neurons seeks to accurately represent neuromuscular development via controlled addition of motor axons following primary myogenesis. Spheroid cultures of motor neuron progenitors augmented the transcription of OLIG2, ISLET1 and SMI32 motor neuron mRNAs ∼400, ∼150 and ∼200-fold respectively. Axon projections of adhered motor neuron spheroids exceeded 1000μm in monolayer, with transcription of SMI32 and VACHT mRNAs further enhanced in a concentration dependent manner within optimised 3D type I collagen extracellular matrices. Bioengineered skeletal muscles produce functional forces, demonstrate increased acetylcholine receptor (AChR) clustering, and transcription of MUSK and LRP4 mRNAs indicating enhanced organisation of the post-synaptic membrane. Dosed integration of motor neuron spheroids outlined the motor pool required to functionally innervate muscle tissues in 3D, generating physiologically functional human NMJs that evidence pre- and post-synaptic membrane and motor nerve terminal co-localisation. Spontaneous firing was significantly elevated in 3D motor units, confirmed to be driven by the motor nerve via antagonistic inhibition of the AChR. Finally, functional analyses outlined decreased time to peak twitch and half relaxation times, indicating enhanced physiology of excitation contraction coupling of NMJs within innervated motor units.

scholarly journals Bioengineered model of the human motor unit with physiologically functional neuromuscular junctions

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Rowan P. Rimington ◽  
Jacob W. Fleming ◽  
Andrew J. Capel ◽  
Patrick C. Wheeler ◽  
Mark P. Lewis
Keyword(s):  
Neuromuscular Junction ◽  
Motor Neuron ◽  
Motor Unit ◽  
Motor Nerve ◽  
Motor Units ◽  
Extracellular Matrices ◽  
Synaptic Membrane ◽  
Type I ◽  
Dependent Manner ◽  
Human Motor

AbstractInvestigations of the human neuromuscular junction (NMJ) have predominately utilised experimental animals, model organisms, or monolayer cell cultures that fail to represent the physiological complexity of the synapse. Consequently, there remains a paucity of data regarding the development of the human NMJ and a lack of systems that enable investigation of the motor unit. This work addresses this need, providing the methodologies to bioengineer 3D models of the human motor unit. Spheroid culture of iPSC derived motor neuron progenitors augmented the transcription of OLIG2, ISLET1 and SMI32 motor neuron mRNAs ~ 400, ~ 150 and ~ 200-fold respectively compared to monolayer equivalents. Axon projections of adhered spheroids exceeded 1000 μm in monolayer, with transcription of SMI32 and VACHT mRNAs further enhanced by addition to 3D extracellular matrices in a type I collagen concentration dependent manner. Bioengineered skeletal muscles produced functional tetanic and twitch profiles, demonstrated increased acetylcholine receptor (AChR) clustering and transcription of MUSK and LRP4 mRNAs, indicating enhanced organisation of the post-synaptic membrane. The number of motor neuron spheroids, or motor pool, required to functionally innervate 3D muscle tissues was then determined, generating functional human NMJs that evidence pre- and post-synaptic membrane and motor nerve axon co-localisation. Spontaneous firing was significantly elevated in 3D motor units, confirmed to be driven by the motor nerve via antagonistic inhibition of the AChR. Functional analysis outlined decreased time to peak twitch and half relaxation times, indicating enhanced physiology of excitation contraction coupling in innervated motor units. Our findings provide the methods to maximise the maturity of both iPSC motor neurons and primary human skeletal muscle, utilising cell type specific extracellular matrices and developmental timelines to bioengineer the human motor unit for the study of neuromuscular junction physiology.

scholarly journals Human motor units in microfluidic devices are impaired by FUS mutations and improved by HDAC6 inhibition

2020 ◽  
Author(s):  
Katarina Stoklund Dittlau ◽  
Emily N. Krasnow ◽  
Laura Fumagalli ◽  
Tijs Vandoorne ◽  
Pieter Baatsen ◽  
...  
Keyword(s):  
Motor Neuron ◽  
Neurite Outgrowth ◽  
Motor Neurons ◽  
Neuromuscular Junctions ◽  
Induced Pluripotent Stem Cell ◽  
Microfluidic Devices ◽  
Motor Units ◽  
Human Motor ◽  
Induced Pluripotent

AbstractNeuromuscular junctions (NMJs) ensure proper communication between motor neurons and muscle through the release of neurotransmitters. In motor neuron disorders, such as amyotrophic lateral sclerosis (ALS), NMJs degenerate resulting in muscle atrophy, paralysis and respiratory failure. The aim of this study was to establish a versatile and reproducible in vitro model of a human motor unit to study the effect of ALS-causing mutations. Therefore, we generated a co-culture of human induced pluripotent stem cell-derived motor neurons and human primary mesoangioblast-derived myotubes in microfluidic devices. A chemotactic and volumetric gradient facilitated the growth of motor neuron neurites through microgrooves resulting in the interaction with myotubes and the formation of NMJs. We observed that ALS-causing FUS mutations resulted in a reduced neurite outgrowth and in a decreased NMJ number. Interestingly, the selective HDAC6 inhibitor, Tubastatin A, improved the neurite outgrowth and the NMJ morphology of FUS-ALS co-cultures, further prompting HDAC6 inhibition as a potential therapeutic strategy for ALS.

scholarly journals Activity-Driven Synaptic and Axonal Degeneration in Canine Motor Neuron Disease

2004 ◽  
Vol 92 (2) ◽  
pp. 1175-1181 ◽  
Author(s):  
Dario I. Carrasco ◽  
Mark M. Rich ◽  
Qingbo Wang ◽  
Timothy C. Cope ◽  
Martin J. Pinter
Keyword(s):  
Motor Neuron ◽  
Motor Neuron Disease ◽  
Motor Unit ◽  
Neuromuscular Junctions ◽  
Muscular Atrophy ◽  
Motor Axon ◽  
Medial Gastrocnemius ◽  
Neuron Activity ◽  
Functional Changes ◽  
Unit Function

The role of neuronal activity in the pathogenesis of neurodegenerative disease is largely unknown. In this study, we examined the effects of increasing motor neuron activity on the pathogenesis of a canine version of inherited motor neuron disease (hereditary canine spinal muscular atrophy). Activity of motor neurons innervating the ankle extensor muscle medial gastrocnemius (MG) was increased by denervating close synergist muscles. In affected animals, 4 wk of synergist denervation accelerated loss of motor-unit function relative to control muscles and decreased motor axon conduction velocities. Slowing of axon conduction was greatest in the most distal portions of motor axons. Morphological analysis of neuromuscular junctions (NMJs) showed that these functional changes were associated with increased loss of intact innervation and with the appearance of significant motor axon and motor terminal sprouting. These effects were not observed in the MG muscles of age-matched, normal animals with synergist denervation for 5 wk. The results indicate that motor neuron action potential activity is a major contributing factor to the loss of motor-unit function and degeneration in inherited canine motor neuron disease.

Physiological responses of rat plantaris motor units to overload induced by surgical removal of its synergists

1988 ◽  
Vol 60 (6) ◽  
pp. 2138-2151 ◽  
Author(s):  
A. E. Olha ◽  
B. J. Jasmin ◽  
R. N. Michel ◽  
P. F. Gardiner
Keyword(s):  
Motor Unit ◽  
Fatigue Resistance ◽  
Motor Units ◽  
High Threshold ◽  
Surgical Removal ◽  
Type I ◽  
Myofibrillar Atpase ◽  
Force Output ◽  
Cross Sectional ◽  
Single Motor Units

1. Rat plantaris muscles were subjected to chronic overload by the surgical removal of the soleus and most of the gastrocnemius muscles. Twelve to 16 wk later whole muscle and motor unit (ventral root dissection technique) contractile properties as well as histochemistry were determined. 2. Motor units were categorized as fast, fatigable (FF), fast, intermediate fatigue-resistant (FI), fast, fatigue-resistant (FR), and slow (S) based on contractile characteristics. Muscle fibers were identified as type I and type II according to myofibrillar ATPase staining. 3. Whole muscles demonstrated increases in wet weight, tetanic force, proportion of type I fibers, and mean cross-sectional areas of both type I and II fibers, as a result of chronic overload. 4. Tetanic tension increased by the same relative magnitude in all motor units whereas twitch tension remained unchanged. A significant change in the proportions of the motor unit types occurred in overloaded muscles, such that the latter contained higher proportions of FF and S units, and lower proportions of FI and FR units, than normal muscles. 5. The fatigue profile of a composite constructed from a summation of motor unit responses revealed that the overloaded plantaris displayed fatigue resistance similar to that of the normal plantaris for a given absolute force output. 6. Glycogen-depleted fibers of hypertrophied single motor units demonstrated uniform myofibrillar ATPase and SDH staining characteristics suggesting that metabolic adaptations among fibers of the same unit were similar after 12-16 wk of overload. 7. The finding that overload caused a uniform increase in the tetanic strength of all motor units, whereas alterations in fatigue resistance varied in degree and direction among unit types, demonstrate that these two properties are not controlled in parallel in this model. The smallest units maintain or even increase their fatigue resistance during the hypertrophic process, whereas high threshold units actually decrease in fatigue resistance.

scholarly journals Assessing Rat Forelimb and Hindlimb Motor Unit Connectivity as Objective and Robust Biomarkers of Spinal Motor Neuron Function

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Markus E. Harrigan ◽  
Angela R. Filous ◽  
Andrew P. Tosolini ◽  
Renee Morris ◽  
Jan M. Schwab ◽  
...  
Keyword(s):  
Motor Neuron ◽  
Motor Unit ◽  
Compound Muscle Action Potential ◽  
Motor Units ◽  
Longitudinal Assessment ◽  
Muscle Contractility ◽  
Cross Sectional ◽  
Muscle Action Potential ◽  
Experimental Findings ◽  
Neuron Function

Abstract Sensitive and objective biomarkers of neuronal injury, degeneration, and regeneration can help facilitate translation of experimental findings into clinical testing. Whereas measures of upper motor neuron connectivity have been readily established, functional assessments of lower motor neuron (LMN) innervation of forelimb muscles are lacking. Compound muscle action potential (CMAP) and motor unit (MU) number estimation (MUNE) are well-established methods that allow longitudinal MU integrity monitoring in patients. In analogy we refined CMAP and MUNE methods for assessing spinal MU input in the rat forelimb and hindlimb. Repeated CMAP and MUNE recordings are robust (coefficients of variability: 4.5–11.3%), and MUNE measurements from forelimb wrist flexor muscles (415 ± 8 [SEM]) align with back-traced anatomical LMN counts (336 ± 16 [SEM]). For disease validation, cross-sectional blinded electrophysiological and muscle contractility measurements were obtained in a cohort of G93A SOD1 mutant overexpressing rats and compared with controls. Longitudinal assessment of mutant animals demonstrated progressive motor unit decline in the hindlimb to a greater extent than the forelimb. Hindlimb CMAP and MUNE demonstrated strong correlations with plantarflexion muscle contractility. Cross-species assessment of upper/fore- limb and lower/hind- limb motor units using objective electrophysiological CMAP and MUNE values as biomarkers will guide and improve bi-directional translation.

scholarly journals Model-Based Analysis of Muscle Strength and EMG-Force Relation with respect to Different Patterns of Motor Unit Loss

2021 ◽  
Vol 2021 ◽  
pp. 1-9
Author(s):  
Chengjun Huang ◽  
Maoqi Chen ◽  
Yingchun Zhang ◽  
Sheng Li ◽  
Ping Zhou
Keyword(s):  
Muscle Strength ◽  
Motor Neuron ◽  
Motor Unit ◽  
Muscle Force ◽  
Motor Units ◽  
Force Variability ◽  
Valuable Insight ◽  
Model Based ◽  
Force Relation ◽  
Pool Model

This study presents a model-based sensitivity analysis of the strength of voluntary muscle contraction with respect to different patterns of motor unit loss. A motor unit pool model was implemented including simulation of a motor neuron pool, muscle force, and surface electromyogram (EMG) signals. Three different patterns of motor unit loss were simulated, including (1) motor unit loss restricted to the largest ones, (2) motor unit loss restricted to the smallest ones, and (3) motor unit loss without size restriction. The model outputs including muscle force amplitude, variability, and the resultant EMG-force relation were quantified under two different motor neuron firing strategies. It was found that motor unit loss restricted to the largest ones had the most dominant impact on muscle strength and significantly changed the EMG-force relation, while loss restricted to the smallest motor units had a pronounced effect on force variability. These findings provide valuable insight toward our understanding of the neurophysiological mechanisms underlying experimental observations of muscle strength, force control, and EMG-force relation in both normal and pathological conditions.

The motor unit

2016 ◽  
Author(s):  
David Burke ◽  
James Howells
Keyword(s):  
Motor Unit ◽  
Neuromuscular Junctions ◽  
Discharge Rate ◽  
Motor Units ◽  
Motor Unit Action Potential ◽  
Muscle Fibres ◽  
Reflex Gain ◽  
Active Motor ◽  
Motor Unit Action ◽  
Fast Twitch

The motor unit represent the final output of the motor system. Each consists of a motoneuron, its axon, neuromuscular junctions, and muscle fibres innervated by that axon. The discharge of a motor unit can be followed by recording its electromyographic signature, the motor unit action potential. Motoneurons are not passive responders to the excitatory and inhibitory influences on them from descending and segmental sources. Their properties can change, e.g. due to descending monoaminergic pathways, which can alter their responses to other inputs (changing ‘reflex gain’). Contraction strength depends on the number of active motor units, their discharge rate, and whether the innervated muscle fibres are slow-twitch producing low force, but resistant to fatigue, fast-twitch producing more force, but susceptible to fatigue, or intermediate fast-twitch fatigue-resistant. These properties are imposed by the parent motoneurons, and the innervated muscle fibres have different histochemical profiles (oxidative, glycolytic, or oxidative-glycolytic, respectively).

Effects of Aging on Motor-Unit Control Properties

1999 ◽  
Vol 82 (5) ◽  
pp. 2081-2091 ◽  
Author(s):  
Zeynep Erim ◽  
M. Faisal Beg ◽  
David T. Burke ◽  
Carlo J. de Luca
Keyword(s):  
Firing Rate ◽  
Motor Unit ◽  
Motor Units ◽  
Elderly Subjects ◽  
Compensatory Mechanism ◽  
Type I ◽  
Firing Behavior ◽  
Firing Rates ◽  
Age Related ◽  
Young Subjects

It was hypothesized that the age-related alterations in the morphological properties of a motor unit would be accompanied by modifications in the control aspects of the motor unit, as either an adaptive or compensatory mechanism to preserve smooth force production. In specific, the objective of the study was to investigate the age-related alterations in the concurrent firing behavior of multiple motor units in the first dorsal interosseous (FDI) muscle in isometric contractions at 20 and 50% of the subject's voluntary contraction level. Analysis of the data collected from 10 young (24–37 yr of age) and 10 elderly (65–88 yr of age) subjects led to three novel observations regarding the firing behavior of aged motor units. 1) Among elderly subjects, there is a decrease in the common fluctuations that are observed among the firing rates of motor units in the young. 2) The relationship observed between the firing rate and recruitment threshold of young subjects is disturbed in the elderly. Although in young subjects, at any point in a given submaximal contraction, earlier recruited motor units have higher firing rates than later-recruited units; in aged subjects this dependency of firing rate on recruitment rank is compromised. 3) The progressive decrease observed in the firing rates of concurrently active motor units in constant-force contractions in the young is not seen in the aged. In addition to these original findings, this study provided support for earlier reports of 1) decreased average firing rates probably reflecting the slowing of the muscle, 2) a shift in recruitment thresholds toward lower force levels in line with the shift toward type I fibers, and 3) multiphasic action potential shapes indicative of the reinnervation process that takes place during aging. Taken as a whole, these findings indicate significant age-related modifications in the control properties of human motor units.

scholarly journals Hypoexcitability precedes denervation in the large fast-contracting motor units in two unrelated mouse models of ALS

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
María de Lourdes Martínez-Silva ◽  
Rebecca D Imhoff-Manuel ◽  
Aarti Sharma ◽  
CJ Heckman ◽  
Neil A Shneider ◽  
...  
Keyword(s):  
Motor Unit ◽  
Mouse Models ◽  
Neuromuscular Junctions ◽  
Motor Units ◽  
Loss Of Function ◽  
Disease Markers ◽  
Physiological Type ◽  
Lateral Sclerosis ◽  
Motoneuron Degeneration

Hyperexcitability has been suggested to contribute to motoneuron degeneration in amyotrophic lateral sclerosis (ALS). If this is so, and given that the physiological type of a motor unit determines the relative susceptibility of its motoneuron in ALS, then one would expect the most vulnerable motoneurons to display the strongest hyperexcitability prior to their degeneration, whereas the less vulnerable should display a moderate hyperexcitability, if any. We tested this hypothesis in vivo in two unrelated ALS mouse models by correlating the electrical properties of motoneurons with their physiological types, identified based on their motor unit contractile properties. We found that, far from being hyperexcitable, the most vulnerable motoneurons become unable to fire repetitively despite the fact that their neuromuscular junctions were still functional. Disease markers confirm that this loss of function is an early sign of degeneration. Our results indicate that intrinsic hyperexcitability is unlikely to be the cause of motoneuron degeneration.

scholarly journals Reduced Endplate Currents Underlie Motor Unit Dysfunction in Canine Motor Neuron Disease

2002 ◽  
Vol 88 (6) ◽  
pp. 3293-3304 ◽  
Author(s):  
Mark M. Rich ◽  
Robert. F. Waldeck ◽  
Linda C. Cork ◽  
Rita J. Balice-Gordon ◽  
Robert E. W. Fyffe ◽  
...  
Keyword(s):  
Motor Neuron ◽  
Motor Neuron Disease ◽  
Motor Unit ◽  
Conduction Velocity ◽  
Motor Neurons ◽  
Muscle Fiber ◽  
Muscular Atrophy ◽  
Motor Units ◽  
Postnatal Maturation ◽  
Axonal Conduction

Hereditary canine spinal muscular atrophy (HCSMA) is an autosomal dominant degenerative disorder of motor neurons. In homozygous animals, motor units produce decreased force output and fail during repetitive activity. Previous studies suggest that decreased efficacy of neuromuscular transmission underlies these abnormalities. To examine this, we recorded muscle fiber endplate currents (EPCs) and found reduced amplitudes and increased failures during nerve stimulation in homozygotes compared with wild-type controls. Comparison of EPC amplitudes with muscle fiber current thresholds indicate that many EPCs from homozygotes fall below threshold for activating muscle fibers but can be raised above threshold following potentiation. To determine whether axonal abnormalities might play a role in causing motor unit dysfunction, we examined the postnatal maturation of axonal conduction velocity in relation to the appearance of tetanic failure. We also examined intracellularly labeled motor neurons for evidence of axonal neurofilament accumulations, which are found in many instances of motor neuron disease including HCSMA. Despite the appearance of tetanic failure between 90 and 120 days, average motor axon conduction velocity increased with age in homozygotes and achieved adult levels. Normal correlations between motor neuron properties (including conduction velocity) and motor unit properties were also observed. Labeled proximal motor axons of several motor neurons that supplied failing motor units exhibited little or no evidence of axonal swellings. We conclude that decreased release of transmitter from motor terminals underlies motor unit dysfunction in HCSMA and that the mechanisms determining the maturation of axonal conduction velocity and the pattern of correlation between motor neuron and motor unit properties do not contribute to the appearance or evolution of motor unit dysfunction.

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