Original Research

Cellular and Molecular Neurobiology

, Volume 31, Issue 5, pp 715-727

First online:

Open Access This content is freely available online to anyone, anywhere at any time.

In Vitro Differentiation of Mouse Embryonic Stem Cells into Neurons of the Dorsal Forebrain

  • Ying JingAffiliated withSchool of Medicine, Zhengzhou UniversityOslo University Hospital, Unit for Cell Signalling
  • , Ondrej MachonAffiliated withOslo University Hospital, Unit for Cell SignallingInstitute of Molecular Genetics, Academy of Sciences
  • , Ales HamplAffiliated withDepartment of Histology and Embryology, Faculty of Medicine, Masaryk UniversityInstitute of Experimental Medicine, Academy of Sciences of the Czech Republic
  • , Petr DvorakAffiliated withDepartment of Biology, Faculty of Medicine, Masaryk University
  • , Ying XingAffiliated withSchool of Medicine, Zhengzhou University
  • , Stefan KraussAffiliated withOslo University Hospital, Unit for Cell Signalling Email author 


Pluripotent embryonic stem cells (ESCs) are able to differentiate into all cell types in the organism including cortical neurons. To follow the dynamic generation of progenitors of the dorsal forebrain in vitro, we generated ESCs from D6-GFP mice in which GFP marks neocortical progenitors and neurons after embryonic day (E) 10.5. We used several cell culture protocols for differentiation of ESCs into progenitors and neurons of the dorsal forebrain. In cell culture, GFP-positive cells were induced under differentiation conditions in quickly formed embryoid bodies (qEBs) after 10–12 day incubation. Activation of Wnt signaling during ESC differentiation further stimulated generation of D6-GFP-positive cortical cells. In contrast, differentiation protocols using normal embryoid bodies (nEBs) yielded only a few D6-GFP-positive cells. Gene expression analysis revealed that multiple components of the canonical Wnt signaling pathway were expressed during the development of embryoid bodies. As shown by immunohistochemistry and quantitative qRT-PCR, D6-GFP-positive cells from qEBs expressed genes that are characteristic for the dorsal forebrain such as Pax6, Dach1, Tbr1, Tbr2, or Sox5. qEBs culture allowed the formation of a D6-GFP positive pseudo-polarized neuroepithelium with the characteristic presence of N-cadherin at the apical pole resembling the structure of the developing neocortex.


Mouse embryonic stem cell Embryonic forebrain Stem cell differentiation Wnt signaling