To understand how the complex architecture of the brain is formed during

development, it is essential to identify the signals that regulate the behaviors of neural progenitor cells. To date, many of the signals that regulate cell proliferation and differentiation of neuronal and glial progenitors of the neocortex are unknown. For my primary thesis work, I investigated the role of the Wnt-β-catenin intercellular signaling pathway in the behaviors of neural progenitors of the mouse neocortex. In this study, we show that the Wnt-β-catenin pathway (WBP) regulates the differentiation of intermediate progenitors (IPs; also known as basal progenitors) into neurons. Upregulation of the WBP by overexpression of Wnt3a secreted agonist in the neocortex induced early differentiation of IPs into neurons and the accumulation of these newly born neurons at the border of the subventricular zone and intermediate zone. Long-term overexpression of Wnt3a led to cortical dysplasia associated with the formation of large neuronal heterotopias. Conversely, downregulation of the WBP with Dkk1, a secreted Wnt antagonist, during mid and late stages of neurogenesis inhibited neuronal production. Lastly, consistent with previous reports, we also show that the WBP also promotes radial glia (RG) expansion. Thus, our findings show differential effects of the WBP on distinct groups of cortical neuronal progenitors: RG expansion and IP differentiation. Moreover, these findings suggest that dysregulation of the WBP can lead to developmental defects similar to human cortical malformation disorders. In a collaborative project, I contributed to address the role of the WBP in oligodendrogenesis. The WBP has previously been shown to inhibit the maturation of oligodendrocyte precursor cells (OPCs). Here, we show that the WBP also inhibits the production of OPCs using both in vitro and in vivo approaches. Our in vivo results show that downregulation of the WBP with a dominant negative form of LEF1 transcription factor or Dkk1 promoted early production of OPCs in the neocortex. Together with previous work, these findings identify multiple roles for the WBP in both neurogenesis and oligodendrogenesis.

Myelin is the insulating membrane concentrically wrapped around some axons, which serves to increase the speed and efficiency of action potential propagation. In the central nervous system (CNS), myelin is formed by oligodendrocytes, which differentiate from self-renewing oligodendrocyte precursor cells (OPCs) throughout life. While factors intrinsic to oligodendrocyte lineage cells control certain features of myelination, extrinsic cues also play an important role in regulating myelination both in development and in response to experience. Thus far, identifying these extrinsic molecular players has proved challenging. Chapter 1 of this thesis describes the known patterns of CNS myelination and provides a framework to understand how neuronal signaling could influence this architecture, using it to evaluate previous work in this field. Chapter 2 presents our experimental work identifying one of these extrinsic neuronal cues for myelination as the neuropeptide dynorphin. We demonstrate that dynorphin, which is released from neurons upon high levels of activity, promotes experience-dependent myelination. Following forced swim stress, an experience that induces striatal dynorphin release, we observe increased striatal OPC differentiation and myelination, which is abolished by deleting dynorphin or blocking its endogenous receptor, kappa opioid receptor (KOR). We find dynorphin also promotes developmental OPC differentiation and myelination, and demonstrate that this effect requires KOR expression specifically on OPCs. We characterize dynorphin-expressing neurons and use genetic sparse-labeling to trace their axonal projections. Surprisingly, we find they are unmyelinated normally and following forced swim stress, in part due to their small size. We propose a new model whereby experience-dependent and developmental myelination is mediated by unmyelinated, neuropeptide-expressing neurons that promote OPC differentiation for the myelination of neighboring axons. Chapter 3 evaluates our findings in the context of the framework presented in Chapter 1 and comments on the significance of our work.

Adult neurogenesis, or the birth of new neurons in the mature brain, is a process that occurs continuously and robustly from neural stem cells located in two discreet regions of the adult mammalian brain, the subventricular zone (SVZ) of the lateral ventricles, and the subgranular zone (SGZ) of the dentate gyrus region of the hippocampus. Though some progress has been made in understanding the factors which regulate the maintenance and fate of adult neural stem cells, our understanding remains limited. Fezf2, a conserved forebrain-specific transcription factor, is expressed during development in both zebrafish and mouse in regions where neural progenitor cells are present, and previous studies point to a role for Fezf2 in embryonic neurogenesis in both systems. Here, we show that fezf2 is expressed in the adult zebrafish forebrain, most notably in radial glial-like cells of the telencephalic ventricular zone, which label with markers of neural stem cells and proliferation. Further analysis using a fezf2-GFP transgenic zebrafish line indicates that these fezf2-expressing cells have neural stem cell-like properties, as they are able to self-renew and can likely give rise to glutamatergic neurons in the adult zebrafish telencephalon. Analysis of too few (Fezf2) homozygous mutant zebrafish crossed to our fezf2-GFP reporter line reveals a previously unreported adult telencephalic phenotype, including an increase in proliferation of fezf2-GFP+ cells, and an increase in adult neurogenesis. No major differences are observed in the mutant telencephalon at early larval stage, suggesting that the phenotype occurs largely at the late-larval to adult stage. Moreover, transplantation of fezf2-GFP+ mutant cells into wiltype hosts reveals a cell-autonomous role for fezf2 in maintaining the non-proliferative state of telencephalic radial glial cells. Taken together, our findings suggest a role for fezf2 in regulating the proliferation and differentiation of neural progenitor cells in the adult vertebrate telencephalon.

Neurogenesis occurs in two places in the postnatal brain: the subventricular zone (SVZ) surrounding the lateral ventricles and the subgranular zone of the dentate gyrus in the hippocampus. Once cell fate is determined in new neurons, they must begin a process of maturation that allows them to functionally integrate into an existing neural network. An essential aspect of their maturation is their morphological development, as neuronal functionality depends on branched signaling processes called dendrites. Wnt signaling has been implicated in several aspects of postnatal neurogenesis and morphological maturation, but which Wnt pathways are involved has been ambiguous. Moreover, these pathways have only been examined in the dentate gyrus and never in vivo. My dissertation project examines the role of Wnt signaling in the morphological maturation of neurons born in the SVZ that eventually become olfactory interneurons. We found that Wnt5a is essential for the morphological maturation of olfactory interneurons in vitro and in vivo. In our experiments Wnt5a signals through a non-canonical Wnt pathway and works in direct opposition to the actions of canonical Wnt signaling on dendrite development.