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Ultrastructural Imaging of Activity-Dependent Synaptic Membrane-Trafficking Events in Cultured Brain Slices
Electron microscopy can resolve synapse ultrastructure with nanometer precision, but the capture of time-resolved, activity-dependent synaptic membrane-trafficking events has remained challenging, particularly in functionally distinct synapses in a tissue context. We present a method that combines o...
Autores principales: | , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Cell Press
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7736621/ https://www.ncbi.nlm.nih.gov/pubmed/32991831 http://dx.doi.org/10.1016/j.neuron.2020.09.004 |
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author | Imig, Cordelia López-Murcia, Francisco José Maus, Lydia García-Plaza, Inés Hojas Mortensen, Lena Sünke Schwark, Manuela Schwarze, Valentin Angibaud, Julie Nägerl, U. Valentin Taschenberger, Holger Brose, Nils Cooper, Benjamin H. |
author_facet | Imig, Cordelia López-Murcia, Francisco José Maus, Lydia García-Plaza, Inés Hojas Mortensen, Lena Sünke Schwark, Manuela Schwarze, Valentin Angibaud, Julie Nägerl, U. Valentin Taschenberger, Holger Brose, Nils Cooper, Benjamin H. |
author_sort | Imig, Cordelia |
collection | PubMed |
description | Electron microscopy can resolve synapse ultrastructure with nanometer precision, but the capture of time-resolved, activity-dependent synaptic membrane-trafficking events has remained challenging, particularly in functionally distinct synapses in a tissue context. We present a method that combines optogenetic stimulation-coupled cryofixation (“flash-and-freeze”) and electron microscopy to visualize membrane trafficking events and synapse-state-specific changes in presynaptic vesicle organization with high spatiotemporal resolution in synapses of cultured mouse brain tissue. With our experimental workflow, electrophysiological and “flash-and-freeze” electron microscopy experiments can be performed under identical conditions in artificial cerebrospinal fluid alone, without the addition of external cryoprotectants, which are otherwise needed to allow adequate tissue preservation upon freezing. Using this approach, we reveal depletion of docked vesicles and resolve compensatory membrane recycling events at individual presynaptic active zones at hippocampal mossy fiber synapses upon sustained stimulation. |
format | Online Article Text |
id | pubmed-7736621 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | Cell Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-77366212020-12-18 Ultrastructural Imaging of Activity-Dependent Synaptic Membrane-Trafficking Events in Cultured Brain Slices Imig, Cordelia López-Murcia, Francisco José Maus, Lydia García-Plaza, Inés Hojas Mortensen, Lena Sünke Schwark, Manuela Schwarze, Valentin Angibaud, Julie Nägerl, U. Valentin Taschenberger, Holger Brose, Nils Cooper, Benjamin H. Neuron NeuroResource Electron microscopy can resolve synapse ultrastructure with nanometer precision, but the capture of time-resolved, activity-dependent synaptic membrane-trafficking events has remained challenging, particularly in functionally distinct synapses in a tissue context. We present a method that combines optogenetic stimulation-coupled cryofixation (“flash-and-freeze”) and electron microscopy to visualize membrane trafficking events and synapse-state-specific changes in presynaptic vesicle organization with high spatiotemporal resolution in synapses of cultured mouse brain tissue. With our experimental workflow, electrophysiological and “flash-and-freeze” electron microscopy experiments can be performed under identical conditions in artificial cerebrospinal fluid alone, without the addition of external cryoprotectants, which are otherwise needed to allow adequate tissue preservation upon freezing. Using this approach, we reveal depletion of docked vesicles and resolve compensatory membrane recycling events at individual presynaptic active zones at hippocampal mossy fiber synapses upon sustained stimulation. Cell Press 2020-12-09 /pmc/articles/PMC7736621/ /pubmed/32991831 http://dx.doi.org/10.1016/j.neuron.2020.09.004 Text en © 2020 The Authors http://creativecommons.org/licenses/by/4.0/ This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | NeuroResource Imig, Cordelia López-Murcia, Francisco José Maus, Lydia García-Plaza, Inés Hojas Mortensen, Lena Sünke Schwark, Manuela Schwarze, Valentin Angibaud, Julie Nägerl, U. Valentin Taschenberger, Holger Brose, Nils Cooper, Benjamin H. Ultrastructural Imaging of Activity-Dependent Synaptic Membrane-Trafficking Events in Cultured Brain Slices |
title | Ultrastructural Imaging of Activity-Dependent Synaptic Membrane-Trafficking Events in Cultured Brain Slices |
title_full | Ultrastructural Imaging of Activity-Dependent Synaptic Membrane-Trafficking Events in Cultured Brain Slices |
title_fullStr | Ultrastructural Imaging of Activity-Dependent Synaptic Membrane-Trafficking Events in Cultured Brain Slices |
title_full_unstemmed | Ultrastructural Imaging of Activity-Dependent Synaptic Membrane-Trafficking Events in Cultured Brain Slices |
title_short | Ultrastructural Imaging of Activity-Dependent Synaptic Membrane-Trafficking Events in Cultured Brain Slices |
title_sort | ultrastructural imaging of activity-dependent synaptic membrane-trafficking events in cultured brain slices |
topic | NeuroResource |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7736621/ https://www.ncbi.nlm.nih.gov/pubmed/32991831 http://dx.doi.org/10.1016/j.neuron.2020.09.004 |
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