scholarly journals In Vivo Single-Molecule Tracking at the Drosophila Presynaptic Motor Nerve Terminal

10.3791/56952 ◽  
2018 ◽  
Author(s):  
Adekunle T. Bademosi ◽  
Elsa Lauwers ◽  
Rumelo Amor ◽  
Patrik Verstreken ◽  
Bruno van Swinderen ◽  
...  
Keyword(s):  
Single Molecule ◽  
Nerve Terminal ◽  
Motor Nerve ◽  
Motor Nerve Terminal ◽  
Single Molecule Tracking

scholarly journals In vivo single-molecule imaging of syntaxin1A reveals polyphosphoinositide- and activity-dependent trapping in presynaptic nanoclusters

2016 ◽  
Vol 7 (1) ◽  
Author(s):  
Adekunle T. Bademosi ◽  
Elsa Lauwers ◽  
Pranesh Padmanabhan ◽  
Lorenzo Odierna ◽  
Ye Jin Chai ◽  
...  
Keyword(s):  
Single Molecule ◽  
Neurotransmitter Release ◽  
Motor Nerve ◽  
Living Organism ◽  
Neurosecretory Cells ◽  
Single Particle Tracking ◽  
Snare Complex ◽  
Motor Nerve Terminal ◽  
Activity Dependent

Abstract Syntaxin1A is organized in nanoclusters that are critical for the docking and priming of secretory vesicles from neurosecretory cells. Whether and how these nanoclusters are affected by neurotransmitter release in nerve terminals from a living organism is unknown. Here we imaged photoconvertible syntaxin1A-mEos2 in the motor nerve terminal of Drosophila larvae by single-particle tracking photoactivation localization microscopy. Opto- and thermo-genetic neuronal stimulation increased syntaxin1A-mEos2 mobility, and reduced the size and molecular density of nanoclusters, suggesting an activity-dependent release of syntaxin1A from the confinement of nanoclusters. Syntaxin1A mobility was increased by mutating its polyphosphoinositide-binding site or preventing SNARE complex assembly via co-expression of tetanus toxin light chain. In contrast, syntaxin1A mobility was reduced by preventing SNARE complex disassembly. Our data demonstrate that polyphosphoinositide favours syntaxin1A trapping, and show that SNARE complex disassembly leads to syntaxin1A dissociation from nanoclusters. Lateral diffusion and trapping of syntaxin1A in nanoclusters therefore dynamically regulate neurotransmitter release.

scholarly journals Transport and Secretion of the Wnt3 Ligand by Motor Neuron-like Cells and Developing Motor Neurons

Biomolecules ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 1898
Author(s):  
Cristina Pinto ◽  
Viviana Pérez ◽  
Jessica Mella ◽  
Miguel Albistur ◽  
Teresa Caprile ◽  
...  
Keyword(s):  
Motor Neuron ◽  
Motor Neurons ◽  
Nerve Terminal ◽  
Motor Nerve ◽  
Motor Nerve Terminal ◽  
In Ovo ◽  
Wnt Ligands ◽  
Postsynaptic Differentiation

The vertebrate neuromuscular junction (NMJ) is formed by a presynaptic motor nerve terminal and a postsynaptic muscle specialization. Cumulative evidence reveals that Wnt ligands secreted by the nerve terminal control crucial steps of NMJ synaptogenesis. For instance, the Wnt3 ligand is expressed by motor neurons at the time of NMJ formation and induces postsynaptic differentiation in recently formed muscle fibers. However, the behavior of presynaptic-derived Wnt ligands at the vertebrate NMJ has not been deeply analyzed. Here, we conducted overexpression experiments to study the expression, distribution, secretion, and function of Wnt3 by transfection of the motor neuron-like NSC-34 cell line and by in ovo electroporation of chick motor neurons. Our findings reveal that Wnt3 is transported along motor axons in vivo following a vesicular-like pattern and reaches the NMJ area. In vitro, we found that endogenous Wnt3 expression increases as the differentiation of NSC-34 cells proceeds. Although NSC-34 cells overexpressing Wnt3 do not modify their morphological differentiation towards a neuronal phenotype, they effectively induce acetylcholine receptor clustering on co-cultured myotubes. These findings support the notion that presynaptic Wnt3 is transported and secreted by motor neurons to induce postsynaptic differentiation in nascent NMJs.

scholarly journals Single molecule tracking of Ace1p in Saccharomyces cerevisiae defines a characteristic residence time for non-specific interactions of transcription factors with chromatin

2016 ◽  
Vol 44 (21) ◽  
pp. e160-e160 ◽  
Author(s):  
David A Ball ◽  
Gunjan D Mehta ◽  
Ronit Salomon-Kent ◽  
Davide Mazza ◽  
Tatsuya Morisaki ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Residence Time ◽  
Single Molecule ◽  
Specific Binding ◽  
Underlying Mechanism ◽  
Specific Interactions ◽  
Single Molecule Tracking ◽  
Reliable Performance ◽  
Transient Interactions

Abstract In vivo single molecule tracking has recently developed into a powerful technique for measuring and understanding the transient interactions of transcription factors (TF) with their chromatin response elements. However, this method still lacks a solid foundation for distinguishing between specific and non-specific interactions. To address this issue, we took advantage of the power of molecular genetics of yeast. Yeast TF Ace1p has only five specific sites in the genome and thus serves as a benchmark to distinguish specific from non-specific binding. Here, we show that the estimated residence time of the short-residence molecules is essentially the same for Hht1p, Ace1p and Hsf1p, equaling 0.12–0.32 s. These three DNA-binding proteins are very different in their structure, function and intracellular concentration. This suggests that (i) short-residence molecules are bound to DNA non-specifically, and (ii) that non-specific binding shares common characteristics between vastly different DNA-bound proteins and thus may have a common underlying mechanism. We develop new and robust procedure for evaluation of adverse effects of labeling, and new quantitative analysis procedures that significantly improve residence time measurements by accounting for fluorophore blinking. Our results provide a framework for the reliable performance and analysis of single molecule TF experiments in yeast.

Motor nerve terminal loss from degenerating muscle fibers

Neuron ◽  
1989 ◽  
Vol 3 (6) ◽  
pp. 677-688 ◽  
Author(s):  
Mark Rich ◽  
Jeff W. Lichtman
Keyword(s):  
Nerve Terminal ◽  
Motor Nerve ◽  
Muscle Fibers ◽  
Motor Nerve Terminal

Motor nerve terminal morphologic plasticity induced by small changes in the locomotor activtty of the adult rat

1990 ◽  
Vol 14 ◽  
pp. 227
Author(s):  
J TOMAS ◽  
R FENOLL ◽  
J BATLLE ◽  
M SANTAFE ◽  
V PIERA ◽  
...  
Keyword(s):  
Nerve Terminal ◽  
Motor Nerve ◽  
Motor Nerve Terminal ◽  
Adult Rat

scholarly journals Comparison of in-vitro and in-vivo DNA Hybridization Kinetics using 3D Single-Molecule Tracking Method

2020 ◽  
Vol 118 (3) ◽  
pp. 616a
Author(s):  
Yuan-I Chen ◽  
Yin-Jui Chang ◽  
Trung D. Nguyen ◽  
Cong Liu ◽  
Stephanie Phillion ◽  
...  
Keyword(s):  
Single Molecule ◽  
Dna Hybridization ◽  
Single Molecule Tracking ◽  
Tracking Method

scholarly journals Interaction of 125I-labeled botulinum neurotoxins with nerve terminals. I. Ultrastructural autoradiographic localization and quantitation of distinct membrane acceptors for types A and B on motor nerves.

1986 ◽  
Vol 103 (2) ◽  
pp. 521-534 ◽  
Author(s):  
J D Black ◽  
J O Dolly
Keyword(s):  
Plasma Membrane ◽  
Neuromuscular Junction ◽  
Nerve Terminal ◽  
Motor Nerve ◽  
Type A ◽  
Nerve Trunk ◽  
Metabolic Inhibitors ◽  
Motor Nerve Terminal ◽  
Electron Microscopic Autoradiography

The labeling patterns produced by radioiodinated botulinum neurotoxin (125I-BoNT) types A and B at the vertebrate neuromuscular junction were investigated using electron microscopic autoradiography. The data obtained allow the following conclusions to be made. 125I-BoNT type A, applied in vivo or in vitro to mouse diaphragm or frog cutaneous pectoris muscle, interacts saturably with the motor nerve terminal only; silver grains occur on the plasma membrane, within the synaptic bouton, and in the axoplasm of the nerve trunk, suggesting internalization and retrograde intra-axonal transport of toxin or fragments thereof. 125I-BoNT type B, applied in vitro to the murine neuromuscular junction, interacts likewise with the motor nerve terminal except that a lower proportion of internalized radioactivity is seen. This result is reconcilable with the similar, but not identical, pharmacological action of these toxin types. The saturability of labeling in each case suggested the involvement of acceptors; on preventing the internalization step with metabolic inhibitors, their precise location became apparent. They were found on all unmyelinated areas of the nerve terminal membrane, including the preterminal axon and the synaptic bouton. Although 125I-BoNT type A interacts specifically with developing terminals of newborn rats, the unmyelinated plasma membrane of the nerve trunk is not labeled, indicating that the acceptors are unique components restricted to the nerve terminal area. BoNT types A and B have distinct acceptors on the terminal membrane. Having optimized the conditions for saturation of these binding sites and calibrated the autoradiographic procedure, we found the densities of the acceptors for types A and B to be approximately 150 and 630/micron 2 of membrane, respectively. It is proposed that these membrane acceptors target BoNT to the nerve terminal and mediate its delivery to an intracellular site, thus contributing to the toxin's selective inhibitory action on neurotransmitter release.

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