Functional maturation of postmitotic neurons from human embryonic stem cell-derived neuroepithelium

The successful differentiation of neurons from human embryonic stem (hES) cells has offered potentially new therapeutic transplant material to treat a variety of neurological diseases. Although great progress has been made in creating specific neuronal subtypes required to treat different diseases, methods to create safe and effective transplantable neuronal cells have not been carefully studied. In this thesis, I have attempted to understand how the proliferation of hES-derived neuronal progenitors, the most likely stage of cells for transplantation, is controlled. I also assessed the ability of astrocytes and their secreted factors to promote the functional maturation and integration of hES derived neurons into neuronal circuits, an essential property for functional neural transplantation. Specifically, I present evidence that hES-derived neuronal progenitors undergo spontaneous Ca2+ transients that can be blocked by inhibitors of transient receptor potential (Trp) channels, which also inhibit proliferation within the progenitor pool. Furthermore, I have demonstrated that newly born post-mitotic neurons depend on members of the TrpC family for proper neurite extension. I have demonstrated that the temporal pattern of physiologic maturation of hES derived neurons recapitulates that which occurs during human development and that the onset of synaptic activity within these cultures is modified by astrocytes. This work represents the first systematic analysis of hES derived neuronal progenitor proliferation and of the functional maturation of hES-derived neurons. Information gained from this study will provide a foundation for studying the functions of human neurons as well as designing safe functional neural transplants in humans.