The effects of intrinsic and extrinsic factors on neural stem cell populations

Neural stem cells (NSCs) are uncommitted cells of the central nervous system (CNS), defined by their ability to self renew and to generate neurons, astrocytes and oligodendrocytes. These cells contribute to the formation of the CNS during embryonic development and in adults for tissue homeostasis. Currently, the best in vitro assay for NSCs is their ability to clonally form neurospheres. Neurospheres are free-floating spherical structures, composed of NSCs, neural progenitor cells (NPCs) as well as differentiated neural cells. To better understand the formation and regulation of NSCs, the developmental time points at which neurospheres can form from the mouse CNS were determined. Novel culture conditions for chick neurosphere culture were then defined and used to characterise the stages neurospheres can be generated from chick CNS tissue. Second, the role of Sox9 in neurosphere formation was investigated. This is a member of the class E group of HMG box containing SOX transcription factors and studies have shown it to be a vital component of the neuron-glia switch. In recent years, radial glia have been reported to exhibit NSC-like properties. Evidence is provided that SOX9 expression coincides with the timing of neurosphere formation from the mouse CNS. Moreover, gain and loss-of-function experiments indicate Sox9 is both necessary and sufficient for neurosphere formation. In addition, Sox9 was able to significantly increase neurosphere generation from the adult SVZ. The secreted molecule sonic hedgehog (SHH) has been implicated in promoting NSC formation and growth. In support of this, experiments described in this thesis suggest that SHH increases both the number of neurosphere forming cells and the proliferation of cells within the neurosphere in El 1.5 mouse spinal cord cultures. Experiments using neurospheres derived from embryonic chick spinal cord and telencephalon cultures reveal a similar proliferative response to SHH as mouse CNS cultures, suggesting conservation of molecular mechanisms. In addition, evidence is provided that SHH can induce early neurosphere formation, both in the embryonic mouse and chick. Moreover, SHH is shown to significantly increase neurosphere generation from the adult NSC niche, the subventricular zone (SVZ). Finally evidence is provided that Sox9 acts downstream of SHH, establishing a functional link between extrinsic and intrinsic factors that control NSCs.