A protocol for preparing GFP-labeled neurons previously imaged in vivo and in slice preparations for light and electron microscopic analysis.

Détails

ID Serval
serval:BIB_1B090B527406
Type
Article: article d'un périodique ou d'un magazine.
Collection
Publications
Institution
Titre
A protocol for preparing GFP-labeled neurons previously imaged in vivo and in slice preparations for light and electron microscopic analysis.
Périodique
Nature Protocols
Auteur⸱e⸱s
Knott G.W., Holtmaat A., Trachtenberg J.T., Svoboda K., Welker E.
ISSN
1750-2799[electronic]
Statut éditorial
Publié
Date de publication
2009
Peer-reviewed
Oui
Volume
4
Numéro
8
Pages
1145-1156
Langue
anglais
Notes
Publication types: Journal Article ; Research Support, Non-U.S. Gov't
Publication Status: ppublish
Résumé
In vivo imaging of green fluorescent protein (GFP)-labeled neurons in the intact brain is being used increasingly to study neuronal plasticity. However, interpreting the observed changes as modifications in neuronal connectivity needs information about synapses. We show here that axons and dendrites of GFP-labeled neurons imaged previously in the live mouse or in slice preparations using 2-photon laser microscopy can be analyzed using light and electron microscopy, allowing morphological reconstruction of the synapses both on the imaged neurons, as well as those in the surrounding neuropil. We describe how, over a 2-day period, the imaged tissue is fixed, sliced and immuno-labeled to localize the neurons of interest. Once embedded in epoxy resin, the entire neuron can then be drawn in three dimensions (3D) for detailed morphological analysis using light microscopy. Specific dendrites and axons can be further serially thin sectioned, imaged in the electron microscope (EM) and then the ultrastructure analyzed on the serial images.
Mots-clé
Animals, Axons/metabolism, Axons/ultrastructure, Dendrites/metabolism, Dendrites/ultrastructure, Green Fluorescent Proteins/analysis, Green Fluorescent Proteins/genetics, Imaging, Three-Dimensional/methods, Immunohistochemistry, Mice, Microscopy, Confocal, Microscopy, Electron, Transmission, Microtomy, Neurons/cytology, Neurons/metabolism, Software, Staining and Labeling, Synapses/ultrastructure
Pubmed
Web of science
Création de la notice
28/07/2009 14:54
Dernière modification de la notice
20/08/2019 13:51
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