scholarly journals The engulfment receptor Draper organizes the postsynaptic spectrin cytoskeleton into corrals containing synaptic proteins and promotes synaptic renewal

2019 ◽  
Author(s):  
Simon Wang ◽  
Mannan Wang ◽  
Hae-yoon Kim ◽  
Nicole Yoo ◽  
Matias Raski ◽  
...  
Keyword(s):  
Neuromuscular Junction ◽  
Motor Neurons ◽  
Postsynaptic Membrane ◽  
Synaptic Proteins ◽  
Muscle Membrane ◽  
Lateral Mobility ◽  
Present Evidence ◽  
Spectrin Cytoskeleton ◽  
Symmetric Patterns

SummaryThe spectrin cytoskeleton is required for development of the Drosophila neuromuscular junction (NMJ) but its role is unclear. Here we show that the muscle spectrin lattice functions to corral membrane-associated synaptic proteins and limit their lateral mobility. Drosophila adducin, Hts, is required for integrity of the spectrin cytoskeleton and disruption of Hts function results in failure of the corrals. The spectrin cytoskeleton is itself patterned at the muscle membrane by the engulfment receptor Draper (Drpr) through regulation of Hts. We find patches of membrane where the spectrin cytoskeleton is organized into bilaterally symmetric patterns, which coincide with a field of Drpr-dependent structures similar to phagocytic pseudopods. The bilaterally symmetric patterns are likely created by folds of the muscle membrane in the pseudopods. We present evidence that the folds trap nascent boutons of motor neurons, leading to boutons with a bilaterally symmetric organization of the postsynaptic membrane. Drpr thus acts as a sensor of synaptic damage that promotes synaptogenesis.

scholarly journals Agrin regulates CLASP2-mediated capture of microtubules at the neuromuscular junction synaptic membrane

2012 ◽  
Vol 198 (3) ◽  
pp. 421-437 ◽  
Author(s):  
Nadine Schmidt ◽  
Sreya Basu ◽  
Stefan Sladecek ◽  
Sabrina Gatti ◽  
Jeffrey van Haren ◽  
...  
Keyword(s):  
Neuromuscular Junction ◽  
Acetylcholine Receptor ◽  
Three Dimensional ◽  
Postsynaptic Membrane ◽  
Dimensional Structure ◽  
Muscle Membrane ◽  
Synaptic Membrane ◽  
Dense Network ◽  
Phosphatidylinositol 3 Kinase ◽  
Neuronal Regulation

Agrin is the major factor mediating the neuronal regulation of postsynaptic structures at the vertebrate neuromuscular junction, but the details of how it orchestrates this unique three-dimensional structure remain unknown. Here, we show that agrin induces the formation of the dense network of microtubules in the subsynaptic cytoplasm and that this, in turn, regulates acetylcholine receptor insertion into the postsynaptic membrane. Agrin acted in part by locally activating phosphatidylinositol 3-kinase and inactivating GSK3β, which led to the local capturing of dynamic microtubules at agrin-induced acetylcholine receptor (AChR) clusters, mediated to a large extent by the microtubule plus-end tracking proteins CLASP2 and CLIP-170. Indeed, in the absence of CLASP2, microtubule plus ends at the subsynaptic muscle membrane, the density of synaptic AChRs, the size of AChR clusters, and the numbers of subsynaptic muscle nuclei with their selective gene expression programs were all reduced. Thus, the cascade linking agrin to CLASP2-mediated microtubule capturing at the synaptic membrane is essential for the maintenance of a normal neuromuscular phenotype.

Nerve-Derived Trophic Factors and DNA Elements Controlling Expression of Genes Encoding Synaptic Proteins in Skeletal Muscle Fibers

1998 ◽  
Vol 23 (4) ◽  
pp. 366-376 ◽  
Author(s):  
Bernard J. Jasmin ◽  
Anthony O. Gramolini ◽  
Feisal A. Adatia ◽  
Lindsay Angus ◽  
Céline Boudreau-Larivière ◽  
...  
Keyword(s):  
Neuromuscular Junction ◽  
Transcriptional Activation ◽  
Recombinant Dna ◽  
Signal Transduction Pathway ◽  
Regulatory Elements ◽  
Postsynaptic Membrane ◽  
Synaptic Proteins ◽  
Trophic Factors ◽  
Recombinant Dna Technology ◽  
Genes Encoding

The neuromuscular junction represents an excellent model system for studying various critical issues in neurobiology at the molecular, cellular, and physiological levels. Our understanding of the basic events underlying synpase formation, maintenance, and plasticity has progressed considerably over the last few years primarily because of the numerous studies that have focused on this synapse and used sophisticated recombinant DNA technology. Recent data indicate that myonuclei located in the vicinity of the postsynaptic membrane are in a differential state of transcription compared to nuclei of the extrasynaptic sarcoplasm. Thus, renewal of postsynaptic membrane proteins appears to occur via a mechanism involving the local transcriptional activation of genes encoding these specialized proteins and extracellular cues originating from motoneurons. Such interaction between presynaptic nerve terminals and the postsynaptic sarcoplasm indicates that the entire signal transduction pathway is compartmentalized at the level of the neuromuscular junction. Expression of these genes appears less coregulated than originally anticipated, indicating that maintenance of the postsynaptic membrane requires the contribution of multiple extracellular signals, which ultimately urge target transcription factors to distinct DNA regulatory elements via various second messenger systems. Key words: neuromuscular junction, synapse, acetylcholinesterase, utrophin, agrin, CGRP, promoter, mRNA

Distribution of acetylcholine receptors in the vicinity of nerve terminals on skeletal muscle of the frog

1972 ◽  
Vol 181 (1065) ◽  
pp. 431-440 ◽  
Keyword(s):  
Skeletal Muscle ◽  
Connective Tissue ◽  
Neuromuscular Junction ◽  
Muscle Fibre ◽  
Nerve Terminal ◽  
Acetylcholine Receptors ◽  
Postsynaptic Membrane ◽  
Muscle Membrane ◽  
Muscle Fibres ◽  
Nerve Terminals

1. The acetylcholine (ACh) sensitivity of muscle fibres at the neuromuscular junction of the frog was investigated in preparations in which the nerve terminals could be clearly seen. 2. ACh released iontophoretically from a micropipette that was precisely positioned at various points along the muscle fibre in the vicinity of the synapse showed that the peak chemosensitivity (up to 1900 mV/nC) is confined to an area of postsynaptic membrane within a few micra of the nerve terminal; a tenfold decline in sensitivity was obtained when the ACh was released only 5 to 10 μm from the terminal’s edge. It is estimated that most of the response obtained when ACh is released within 40 μm from the terminal (the area covered in this study) is due to diffusion to the immediate postsynaptic area. The extrasynaptic chemosensitivity of the muscle membrane was too low to be measured with the present methods. 3. The accuracy with which micropipettes could be positioned in synaptic areas and the clarity of viewing nerve terminals were improved by bathing the tissue in collagenase, which reduced the amount of connective tissue. The distribution of chemosensitivity remained unchanged by such treatment. The ACh response was not detectably altered when nerve terminals were lifted off the muscle, exposing the subsynaptic muscle surface.

scholarly journals Processing of ARIA and release from isolated nerve terminals

1999 ◽  
Vol 354 (1381) ◽  
pp. 411-416 ◽  
Author(s):  
Bomie Han ◽  
Gerald D. Fischbach
Keyword(s):  
Neuromuscular Junction ◽  
Action Potential ◽  
Acetylcholine Receptors ◽  
Molecular Layer ◽  
Postsynaptic Membrane ◽  
High Density ◽  
Small Contribution ◽  
Presynaptic Nerve Terminal ◽  
Voltage Gated

The neuromuscular junction is a specialized synapse in that every action potential in the presynaptic nerve terminal results in an action potential in the postsynaptic membrane, unlike most interneuronal synapses where a single presynaptic input makes only a small contribution to the population postsynaptic response. The postsynaptic membrane at the neuromuscular junction contains a high density of neurotransmitter (acetylcholine) receptors and a high density of voltage–gated Na + channels. Thus, the large acetylcholine activated current occurs at the same site where the threshold for action potential generation is low. Acetylcholine receptor inducing activity (ARIA), a 42 kD protein, that stimulates synthesis of acetylcholine receptors and voltage–gated Na + channels in cultured myotubes, probably plays the same roles at developing and mature motor endplates in vivo . ARIA is synthesized as part of a larger, transmembrane, precursor protein called proARIA. Delivery of ARIA from motor neuron cell bodies in the spinal cord to the target endplates involves several steps, including proteolytic cleavage of proARIA. ARIA is also expressed in the central nervous system and it is abundant in the molecular layer of the cerebellum. In this paper we describe our first experiments on the processing and release of ARIA from subcellular fractions containing synaptosomes from the chick cerebellum as a model system.

Promoting Long‐Term Cultivation of Motor Neurons for 3D Neuromuscular Junction Formation of 3D In Vitro Using Central‐Nervous‐Tissue‐Derived Bioink

2021 ◽  
pp. 2100581
Author(s):  
Jeong Sik Kong ◽  
Xuan Huang ◽  
Yeong‐Jin Choi ◽  
Hee‐Gyeong Yi ◽  
Junsu Kang ◽  
...  
Keyword(s):  
Neuromuscular Junction ◽  
Motor Neurons ◽  
Nervous Tissue ◽  
Central Nervous Tissue ◽  
Junction Formation

scholarly journals A three-dimensional culture model of innervated human skeletal muscle enables studies of the adult neuromuscular junction and disease modeling

2018 ◽  
Author(s):  
Mohsen Afshar Bakooshli ◽  
Ethan S Lippmann ◽  
Ben Mulcahy ◽  
Nisha R Iyer ◽  
Christine T Nguyen ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Neuromuscular Junction ◽  
Motor Neuron ◽  
Motor Neurons ◽  
Human Skeletal Muscle ◽  
Disease Modeling ◽  
De Novo ◽  
Muscle Fibers ◽  
Three Dimensional ◽  
Adult Human

SummaryTwo-dimensional (2D) human skeletal muscle fiber cultures are ill equipped to support the contractile properties of maturing muscle fibers. This limits their application to the study of adult human neuromuscular junction (NMJ) development, a process requiring maturation of muscle fibers in the presence of motor neuron endplates. Here we describe a three-dimensional (3D) co-culture method whereby human muscle progenitors mixed with human pluripotent stem cell-derived motor neurons self-organize to form functional NMJ connections within two weeks. Functional connectivity between motor neuron endplates and muscle fibers is confirmed with calcium transient imaging and electrophysiological recordings. Notably, we only observed epsilon acetylcholine receptor subunit protein upregulation and activity in 3D co-culture. This demonstrates that the 3D co-culture system supports a developmental shift from the embryonic to adult form of the receptor that does not occur in 2D co-culture. Further, 3D co-culture treatments with myasthenia gravis patient sera shows the ease of studying human disease with the system. This work delivers a simple, reproducible, and adaptable method to model and evaluate adult human NMJ de novo development and disease in culture.

scholarly journals Dynein acts to cluster glutamate receptors and traffic the PIP5 kinase, Skittles, to regulate postsynaptic membrane organization at the neuromuscular junction

2021 ◽  
Author(s):  
Amanda L. Neisch ◽  
Thomas Pengo ◽  
Adam W. Avery ◽  
Min-Gang Li ◽  
Thomas S. Hays
Keyword(s):  
Neuromuscular Junctions ◽  
Cytoplasmic Dynein ◽  
Postsynaptic Membrane ◽  
Independent Function ◽  
Protein Levels ◽  
Cluster Organization ◽  
Synaptic Terminals ◽  
Cargo Transport ◽  
Spectrin Cytoskeleton ◽  
Phosphatidylinositol 4

Cytoplasmic dynein is essential in motoneurons for retrograde cargo transport that sustains neuronal connectivity. Little, however, is known about dynein's function on the postsynaptic side of the circuit. Here we report distinct postsynaptic roles for dynein at neuromuscular junctions (NMJs). Intriguingly, we show that dynein punctae accumulate postsynaptically at glutamatergic synaptic terminals. Moreover, Skittles, a phosphatidylinositol 4-phosphate 5-kinase that produces PI(4,5)P2 to organize the spectrin cytoskeleton, also localizes specifically to glutamatergic synaptic terminals. Depletion of postsynaptic dynein disrupts the accumulation of Skittles, PI(4,5)P2 phospholipid, and organization of the spectrin cytoskeleton at the postsynaptic membrane. Coincidental with dynein depletion, we observe an increase in the clusters size of ionotropic glutamate receptor (iGluR), and an increase in the amplitude and frequency of mEJPs. However, PI(4,5)P2 levels do not affect iGluR clustering and dynein does not affect the protein levels of iGluR subunits at the NMJ, suggesting a separate, transport independent function for dynein in iGluR cluster organization. As dynein punctae closely associate with iGluR clusters, we propose that dynein physically tethers iGluR clusters at the postsynaptic membrane to ensure proper synaptic transmission.

Barbiturates depress vagal motor pathway to ferret trachea at ganglia

1982 ◽  
Vol 53 (1) ◽  
pp. 253-257 ◽  
Author(s):  
B. E. Skoogh ◽  
M. J. Holtzman ◽  
J. R. Sheller ◽  
J. A. Nadel
Keyword(s):  
Neuromuscular Junction ◽  
Vagus Nerve ◽  
Muscle Tension ◽  
Postsynaptic Membrane ◽  
Nerve Fibers ◽  
Motor Pathway ◽  
Parasympathetic Ganglia ◽  
Before And After ◽  
Nerve Muscle

To determine which site in the vagal motor pathway to airway smooth muscle is most sensitive to depression by barbiturates, we recorded isometric muscle tension in vitro and stimulated the vagal motor pathway at four different sites before and after exposure to barbiturates. In isolated tracheal rings from ferrets, we stimulated muscarinic receptors in the neuromuscular junction by exogenous acetylcholine, postganglionic nerve fibers by electrical fluid stimulation, and the postsynaptic membrane in ganglia by 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP). We also developed a tracheal nerve-muscle preparation to stimulate preganglionic fibers in the vagus nerve electrically. Activation of ganglia by DMPP or by vagus nerve stimulation was depressed by barbiturates at 10-fold lower concentrations than those depressing the activation of postganglionic nerves or the neuromuscular junction. These findings suggest that the postsynaptic membrane in parasympathetic ganglia is the site in the vagal motor pathway most sensitive to depression by barbiturates.

Synthesis and Transport of Agrin—Like Molecules in Motor Neurons

1990 ◽  
Vol 153 (1) ◽  
pp. 1-10 ◽  
Author(s):  
CATHERINE MAGILL-SOLC ◽  
U. J. MCMAHAN
Keyword(s):  
In Situ Hybridization ◽  
Neuromuscular Junction ◽  
Motor Neurons

Several lies of evidence indicate that agrin, or a protein very similar to it, directs the formation and maintenance of the postsynaptic apparatus at the neuromuscular junction. We discuss the results of studies involving immunohistochemical, biochemical and in situ hybridization techniques that support hypothesis that agrin or agrin—like molecules active at the junction are produced bymotor neurons.

A role for PS integrins in morphological growth and synaptic function at the postembryonic neuromuscular junction of Drosophila

Development ◽  
1999 ◽  
Vol 126 (24) ◽  
pp. 5833-5846 ◽  
Author(s):  
K.J. Beumer ◽  
J. Rohrbough ◽  
A. Prokop ◽  
K. Broadie
Keyword(s):  
Neuromuscular Junction ◽  
Synapse Formation ◽  
Postsynaptic Membrane ◽  
Synaptic Function ◽  
Beta Subunit ◽  
Target Cells ◽  
Type I ◽  
Critical Determinant ◽  
Beta Subunit Gene ◽  
Mutant Phenotypes

A family of three position-specific (PS) integrins are expressed at the Drosophila neuromuscular junction (NMJ): a beta subunit ((betaPS), expressed in both presynaptic and postsynaptic membranes, and two alpha subunits (alphaPS1, alphaPS2), expressed at least in the postsynaptic membrane. PS integrins appear at postembryonic NMJs coincident with the onset of rapid morphological growth and terminal type-specific differentiation, and are restricted to type I synaptic boutons, which mediate fast, excitatory glutamatergic transmission. We show that two distinctive hypomorphic mutant alleles of the beta subunit gene myospheroid (mys(b9) and mys(ts1)), differentially affect betaPS protein expression at the synapse to produce distinctive alterations in NMJ branching, bouton formation, synaptic architecture and the specificity of synapse formation on target cells. The mys(b9) mutation alters betaPS localization to cause a striking reduction in NMJ branching, bouton size/number and the formation of aberrant ‘mini-boutons’, which may represent a developmentally arrested state. The mys(ts1) mutation strongly reduces betaPS expression to cause the opposite phenotype of excessive synaptic sprouting and morphological growth. NMJ function in these mutant conditions is altered in line with the severity of the morphological aberrations. Consistent with these mutant phenotypes, transgenic overexpression of the betaPS protein with a heat-shock construct or tissue-specific GAL4 drivers causes a reduction in synaptic branching and bouton number. We conclude that betaPS integrin at the postembryonic NMJ is a critical determinant of morphological growth and synaptic specificity. These data provide the first genetic evidence for a functional role of integrins at the postembryonic synapse.

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