Developing HiPSC Derived Serum Free Embryoid Bodies for the Interrogation of 3-D Stem Cell Cultures Using Physiologically Relevant Assays

J Vis Exp. 2017 Jul 20:(125):55799. doi: 10.3791/55799.

Abstract

Although a number of in vitro disease models have been developed using hiPSCs, one limitation is that these two-dimensional (2-D) systems may not represent the underlying cytoarchitectural and functional complexity of the affected individuals carrying suspected disease variants. Conventional 2-D models remain incomplete representations of in vivo-like structures and do not adequately capture the complexity of the brain. Thus, there is an emerging need for more 3-D hiPSC-based models that can better recapitulate the cellular interactions and functions seen in an in vivo system. Here we report a protocol to develop a 3-D system from undifferentiated hiPSCs based on the serum free embryoid body (SFEB). This 3-D model mirrors aspects of a developing ventralized neocortex and allows for studies into functions integral to living neural cells and intact tissue such as migration, connectivity, communication, and maturation. Specifically, we demonstrate that the SFEBs using our protocol can be interrogated using physiologically relevant and high-content cell based assays such as calcium imaging, and multi-electrode array (MEA) recordings without cryosectioning. In the case of MEA recordings, we demonstrate that SFEBs increase both spike activity and network-level bursting activity during long-term culturing. This SFEB protocol provides a robust and scalable system for the study of developing network formation in a 3-D model that captures aspects of early cortical development.

Publication types

  • Video-Audio Media

MeSH terms

  • Animals
  • Cell Culture Techniques / methods*
  • Cell Differentiation
  • Coculture Techniques / methods*
  • Embryoid Bodies / physiology*
  • Humans
  • Induced Pluripotent Stem Cells / physiology*
  • Mice
  • Neurons / physiology