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Dissecting and Recording from The C. Elegans Neuromuscular Junction

Neurotransmission is the process by which neurons transfer information via chemical signals to their post-synaptic targets, on a rapid time scale. This complex process requires the coordinated activity of many pre- and post-synaptic proteins to ensure appropriate synaptic connectivity, conduction of...

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Autor principal: Richmond, Janet
Formato: Texto
Lenguaje:English
Publicado: MyJove Corporation 2009
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2789031/
https://www.ncbi.nlm.nih.gov/pubmed/19488020
http://dx.doi.org/10.3791/1165
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author Richmond, Janet
author_facet Richmond, Janet
author_sort Richmond, Janet
collection PubMed
description Neurotransmission is the process by which neurons transfer information via chemical signals to their post-synaptic targets, on a rapid time scale. This complex process requires the coordinated activity of many pre- and post-synaptic proteins to ensure appropriate synaptic connectivity, conduction of electrical signals, targeting and priming of secretory vesicles, calcium sensing, vesicle fusion, localization and function of postsynaptic receptors and finally, recycling mechanisms. As neuroscientists it is our goal to elucidate which proteins function in each of these steps and understand their mechanisms of action. Electrophysiological recordings from synapses provide a quantifiable read out of the underlying electrical events that occur during synaptic transmission. By combining this technique with the powerful array of molecular and genetic tools available to manipulate synaptic proteins in C. elegans, we can analyze the resulting functional changes in synaptic transmission. The C. elegans NMJs formed between motor neurons and body wall muscles control locomotion, therefore, mutants with uncoordinated locomotory phenotypes (known as unc s) often perturb synaptic transmission at these synapses (1). Since unc mutants are maintained on a rich supply of a bacterial food source, they remain viable as long as they retain some pharyngeal pumping ability to ingest food. This, together with the fact that C. elegans exist as hermaphrodites, allows them to pass on mutant progeny without the need for elaborate mating behaviors. These attributes, coupled with our recent ability to record from the worms NMJs (2,3,7) make this an excellent model organism in which to address precisely how unc mutants impact neurotransmission. The dissection method involves immobilizing adult worms using a cyanoacrylic glue in order to make an incision in the worm cuticle exposing the NMJs. Since C. elegans adults are only 1 mm in length the dissection is performed with the use of a dissecting microscope and requires excellent hand-eye coordination. NMJ recordings are made by whole-cell voltage clamping individual body wall muscle cells and neurotransmitter release can be evoked using a variety of stimulation protocols including electrical stimulation, light-activated channel-rhodopsin-mediated depolarization (4) and hyperosmotic saline, all of which will be briefly described.
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spelling pubmed-27890312011-07-21 Dissecting and Recording from The C. Elegans Neuromuscular Junction Richmond, Janet J Vis Exp Neuroscience Neurotransmission is the process by which neurons transfer information via chemical signals to their post-synaptic targets, on a rapid time scale. This complex process requires the coordinated activity of many pre- and post-synaptic proteins to ensure appropriate synaptic connectivity, conduction of electrical signals, targeting and priming of secretory vesicles, calcium sensing, vesicle fusion, localization and function of postsynaptic receptors and finally, recycling mechanisms. As neuroscientists it is our goal to elucidate which proteins function in each of these steps and understand their mechanisms of action. Electrophysiological recordings from synapses provide a quantifiable read out of the underlying electrical events that occur during synaptic transmission. By combining this technique with the powerful array of molecular and genetic tools available to manipulate synaptic proteins in C. elegans, we can analyze the resulting functional changes in synaptic transmission. The C. elegans NMJs formed between motor neurons and body wall muscles control locomotion, therefore, mutants with uncoordinated locomotory phenotypes (known as unc s) often perturb synaptic transmission at these synapses (1). Since unc mutants are maintained on a rich supply of a bacterial food source, they remain viable as long as they retain some pharyngeal pumping ability to ingest food. This, together with the fact that C. elegans exist as hermaphrodites, allows them to pass on mutant progeny without the need for elaborate mating behaviors. These attributes, coupled with our recent ability to record from the worms NMJs (2,3,7) make this an excellent model organism in which to address precisely how unc mutants impact neurotransmission. The dissection method involves immobilizing adult worms using a cyanoacrylic glue in order to make an incision in the worm cuticle exposing the NMJs. Since C. elegans adults are only 1 mm in length the dissection is performed with the use of a dissecting microscope and requires excellent hand-eye coordination. NMJ recordings are made by whole-cell voltage clamping individual body wall muscle cells and neurotransmitter release can be evoked using a variety of stimulation protocols including electrical stimulation, light-activated channel-rhodopsin-mediated depolarization (4) and hyperosmotic saline, all of which will be briefly described. MyJove Corporation 2009-02-25 /pmc/articles/PMC2789031/ /pubmed/19488020 http://dx.doi.org/10.3791/1165 Text en Copyright © 2009, Journal of Visualized Experiments http://creativecommons.org/licenses/by/2.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Neuroscience
Richmond, Janet
Dissecting and Recording from The C. Elegans Neuromuscular Junction
title Dissecting and Recording from The C. Elegans Neuromuscular Junction
title_full Dissecting and Recording from The C. Elegans Neuromuscular Junction
title_fullStr Dissecting and Recording from The C. Elegans Neuromuscular Junction
title_full_unstemmed Dissecting and Recording from The C. Elegans Neuromuscular Junction
title_short Dissecting and Recording from The C. Elegans Neuromuscular Junction
title_sort dissecting and recording from the c. elegans neuromuscular junction
topic Neuroscience
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2789031/
https://www.ncbi.nlm.nih.gov/pubmed/19488020
http://dx.doi.org/10.3791/1165
work_keys_str_mv AT richmondjanet dissectingandrecordingfromthecelegansneuromuscularjunction