Ex utero electroporation and whole hemisphere explants: a simple experimental method for studies of early cortical development

J Vis Exp. 2013 Apr 3;(74):50271. doi: 10.3791/50271.

Abstract

Cortical development involves complex interactions between neurons and non-neuronal elements including precursor cells, blood vessels, meninges and associated extracellular matrix. Because they provide a suitable organotypic environment, cortical slice explants are often used to investigate those interactions that control neuronal differentiation and development. Although beneficial, the slice explant model can suffer from drawbacks including aberrant cellular lamination and migration. Here we report a whole cerebral hemisphere explant system for studies of early cortical development that is easier to prepare than cortical slices and shows consistent organotypic migration and lamination. In this model system, early lamination and migration patterns proceed normally for a period of two days in vitro, including the period of preplate splitting, during which prospective cortical layer six forms. We then developed an ex utero electroporation (EUEP) approach that achieves -80% success in targeting GFP expression to neurons developing in the dorsal medial cortex. The whole hemisphere explant model makes early cortical development accessible for electroporation, pharmacological intervention and live imaging approaches. This method avoids the survival surgery required of in utero electroporation (IUEP) approaches while improving both transfection and areal targeting consistency. This method will facilitate experimental studies of neuronal proliferation, migration and differentiation.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Video-Audio Media

MeSH terms

  • Animals
  • Brain / embryology*
  • DNA / administration & dosage
  • DNA / genetics
  • Electroporation / methods*
  • Female
  • Green Fluorescent Proteins / biosynthesis
  • Green Fluorescent Proteins / genetics
  • Male
  • Mice
  • Plasmids / administration & dosage
  • Plasmids / genetics
  • Pregnancy

Substances

  • Green Fluorescent Proteins
  • DNA