Ex utero electroporation and organotypic slice culture of mouse hippocampal tissue

J Vis Exp. 2015 Mar 4:(97):52550. doi: 10.3791/52550.

Abstract

Mouse genetics offers a powerful tool determining the role of specific genes during development. Analyzing the resulting phenotypes by immunohistochemical and molecular methods provides information of potential target genes and signaling pathways. To further elucidate specific regulatory mechanisms requires a system allowing the manipulation of only a small number of cells of a specific tissue by either overexpression, ablation or re-introduction of specific genes and follow their fate during development. To achieve this ex utero electroporation of hippocampal structures, especially the dentate gyrus, followed by organotypic slice culture provides such a tool. Using this system to generate mosaic deletions allows determining whether the gene of interest regulates cell-autonomously developmental processes like progenitor cell proliferation or neuronal differentiation. Furthermore it facilitates the rescue of phenotypes by re-introducing the deleted gene or its target genes. In contrast to in utero electroporation the ex utero approach improves the rate of successfully targeting deeper layers of the brain like the dentate gyrus. Overall ex utero electroporation and organotypic slice culture provide a potent tool to study regulatory mechanisms in a semi-native environment mirroring endogenous conditions.

Publication types

  • Research Support, Non-U.S. Gov't
  • Video-Audio Media

MeSH terms

  • Animals
  • Dentate Gyrus / cytology
  • Dentate Gyrus / embryology
  • Dentate Gyrus / physiology*
  • Desmoplakins / physiology
  • Down-Regulation
  • Electroporation / methods*
  • Female
  • Gene Expression Regulation, Developmental
  • Mice
  • Organ Culture Techniques / methods*
  • Pregnancy
  • Repressor Proteins / physiology
  • Single-Cell Analysis / methods
  • Tumor Suppressor Proteins / physiology
  • Up-Regulation

Substances

  • Bcl11b protein, mouse
  • Desmoplakins
  • Dsp protein, mouse
  • Repressor Proteins
  • Tumor Suppressor Proteins