In Utero Electroporation of Multiaddressable Genome-Integrating Color (MAGIC) Markers to Individualize Cortical Mouse Astrocytes - Archive ouverte HAL Accéder directement au contenu
Article Dans Une Revue Journal of visualized experiments : JoVE Année : 2020

In Utero Electroporation of Multiaddressable Genome-Integrating Color (MAGIC) Markers to Individualize Cortical Mouse Astrocytes

Laura Dumas
  • Fonction : Auteur
Solène Clavreul
  • Fonction : Auteur
Jason Durand
  • Fonction : Auteur
Edwin Hernandez-Garzon
  • Fonction : Auteur
Lamiae Abdeladim
  • Fonction : Auteur
Raphaëlle Barry-Martinet
Alicia Caballero-Megido
  • Fonction : Auteur
Emmanuel Beaurepaire
  • Fonction : Auteur
Jean Livet
  • Fonction : Auteur
Karine Loulier
  • Fonction : Auteur
  • PersonId : 1354980

Résumé

Protoplasmic astrocytes (PrA) located in the mouse cerebral cortex are tightly juxtaposed, forming an apparently continuous three-dimensional matrix at adult stages. Thus far, no immunostaining strategy can single them out and segment their morphology in mature animals and over the course of corticogenesis. Cortical PrA originate from progenitors located in the dorsal pallium and can easily be targeted using in utero electroporation of integrative vectors. A protocol is presented here to label these cells with the multiaddressable genome-integrating color (MAGIC) Markers strategy, which relies on piggyBac/Tol2 transposition and Cre/lox recombination to stochastically express distinct fluorescent proteins (blue, cyan, yellow, and red) addressed to specific subcellular compartments. This multicolor fate mapping strategy enables to mark in situ nearby cortical progenitors with combinations of color markers prior to the start of gliogenesis and to track their descendants, including astrocytes, from embryonic to adult stages at the individual cell level. Semi-sparse labeling achieved by adjusting the concentration of electroporated vectors and color contrasts provided by the Multiaddressable Genome-Integrating Color Markers (MAGIC Markers or MM) enable to individualize astrocytes and single out their territory and complex morphology despite their dense anatomical arrangement. Presented here is a comprehensive experimental workflow including the details of the electroporation procedure, multichannel image stacks acquisition by confocal microscopy, and computer-assisted three-dimensional segmentation that will enable the experimenter to assess individual PrA volume and morphology. In summary, electroporation of MAGIC Markers provides a convenient method to individually label numerous astrocytes and gain access to their anatomical features at different developmental stages. This technique will be useful to analyze cortical astrocyte morphological properties in various mouse models without resorting to complex crosses with transgenic reporter lines.
Fichier principal
Vignette du fichier
Dumas - Jove 2020.pdf (1.2 Mo) Télécharger le fichier
Origine : Fichiers éditeurs autorisés sur une archive ouverte

Dates et versions

hal-04469353 , version 1 (20-02-2024)

Identifiants

Citer

Laura Dumas, Solène Clavreul, Jason Durand, Edwin Hernandez-Garzon, Lamiae Abdeladim, et al.. In Utero Electroporation of Multiaddressable Genome-Integrating Color (MAGIC) Markers to Individualize Cortical Mouse Astrocytes. Journal of visualized experiments : JoVE, 2020, 159, ⟨10.3791/61110⟩. ⟨hal-04469353⟩
4 Consultations
14 Téléchargements

Altmetric

Partager

Gmail Facebook X LinkedIn More