Neuronal Differentiation of hPSC-Derived Neural Progenitor Cells

Human pluripotent stem cells (hPSCs), including embryonicstem (ES) and induced pluripotent stem (iPS) cells, can undergoneural induction to generate neural progenitor cells (NPCs). Highly enriched cultures of CNS-type NPCs may be obtained through the use of standard monolayer culture or embryoid body (EB) protocols. Further differentiation of NPCs into neurons and glia provides a physiologically relevant model for neurological disease research, drug discovery and cell therapy validation. Many published protocols for neuronal differentiation use Brewer’s B27 supplement and/or Bottenstein’s N2 supplement with a basal medium of choice. While traditional neuronal basal media support cell survival, they impair neurological activities, including action potential generation and synaptic activity. BrainPhys™ was designed by Dr. Cedric Bardy in Dr. Fred H. Gage’s laboratory to better support in vitro neuronal function. In this technical bulletin, we describe a method (based on Yuan et al.) for neuronal differentiation (Figure1), using BrainPhys™ Neuronal Medium as a basal medium, supplemented with NeuroCult™ SM1,

N2 Supplement-A and other factors. Using this protocol, forebrain-type neurons can be generated from NPCs in 2 – 4 weeks. Further culture in BrainPhys™ Neuronal Medium results in cultures that are phenotypically mature and, after 65 days in culture, synaptically active.