Browsing by Subject "Neurogenesis"

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    Investigating neurogenesis and cell type specification in the mammalian thalamus.
    (2012-06) Bluske, Krista K.
    The thalamus mediates a variety of important brain functions that are critical for behavior and survival. A key feature that enables the thalamus to perform such diverse functions is its parcellation into anatomically and functionally distinct groups of neurons called nuclei. The purpose of this project was to identify the origin of neuronal diversity within the thalamus by investigating the process of neurogenesis. During neurogenesis, proliferating progenitor cells begin to divide asymmetrically to generate neurons. The central hypothesis of the research presented herein is that thalamic organization requires the appropriate number and types of neurons to be generated and that these critical processes are regulated during neurogenesis. This work has characterized the different types of progenitor cells present during thalamic neurogenesis. We confirmed the existence of a special population of thalamic progenitor cells, intermediate (or basal) progenitor cells, and identified transcription factors that regulate their formation and/or maintenance. We also addressed the origin of distinct subtypes of neurons. The spatial organization of thalamic progenitor cells into two distinct progenitor domains during neurogenesis is thought to drive the formation of different subtypes of thalamic neurons. Signaling molecules have been proposed to induce the formation of distinct progenitor domains in numerous brain areas, including the thalamus. We provided a detailed characterization of components of the Wnt/β-catenin-mediated transcriptional pathway during thalamic neurogenesis. Based on the pattern of signaling activity, we hypothesized that Wnt/β-catenin-mediated transcription has a function in forming the two progenitor domains during thalamic neurogenesis. Using conditional genetic manipulations of β-catenin, we found that β-catenin-mediated transcription is required for the specification of thalamic progenitor domains. Furthermore, we found that the Wnt/β-catenin signaling pathway functions in parallel with the sonic hedgehog (Shh) signaling pathway, which had been previously shown to specify thalamic progenitor identity in an opposing manner, by independently regulating transcriptional networks in thalamic progenitor cells. Collectively, the process of neurogenesis involves the generation of the correct number of neurons by regulating asymmetric progenitor divisions and generation of appropriate neuronal subtypes through the functions of signaling pathways and transcriptional networks. These mechanisms provide a broad map for the generation and positioning of appropriate types of neurons in the correct locations within the thalamus.
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    Progenitor cell maturation and initiation of neurogenesis in the developing vertebrate neural retina.
    (2009-10) Yang, Hyun-Jin
    The mature vertebrate central nervous system is composed of an enormous number of neuronal and glial cells. A relatively small number of progenitor cells generate these cells during a finite period of time of development. Progenitor cells during early stages of central nervous system development divide so that each division produces two progeny that divide again. This `preneurogenic' mode of division is essential for the exponential increase of number of progenitor cells. Later, progenitor cells change their mode of division to `neurogenic', where one or both daughter cells produced by a division withdraw from the mitotic cycle and differentiate. This more mature, neurogenic division is critical for generation of a functional nervous system. The aim of the project described in this thesis was to understand: 1) the molecular differences that dictate the two modes of progenitor cell division, namely preneurogenic and neurogenic, 2) the mechanism that regulates the switch in the mode of division, and 3) the molecular trigger that initiates differentiation. Molecular differences between preneurogenic and neurogenic progenitor cells were identified, and are described in more detail in chapter II. The early, preneurogenic progenitor cells express the transcription factor, Sox2, and a ligand for the Notch receptor, Delta1. The more mature, neurogenic progenitor cells express Sox2 and the bHLH transcription factor, E2A, and do not express Delta1. Perturbation of Notch signaling resulted in conversion of progenitor cells from preneurogenic to neurogenic and in premature neurogenesis. Furthermore, Sonic hedgehog was found to be expressed by a subset of newly differentiating cells. Misexpression of Sonic hedgehog led to premature maturation of preneurogenic progenitor cells and neurogenesis. These results suggest that Notch signaling maintains progenitor cells in the preneurogenic state and that Sonic hedgehog initiates progenitor cell maturation. Certain proneural bHLH transcription factors were found to initiate neurogenesis, and are described in more detail in chapter III. Expression of a number of proneural bHLH factors comes up in a stereotypic temporal sequence prior to the onset of ganglion cell differentiation. Ascl1 and Neurog2 were expressed first, which was followed by expression of Neurod1 and Neurod4. Finally, Atoh7 was expressed, which preceded the appearance of ganglion cells. Individual progenitor cells expressed heterogeneous combinations of proneural genes prior to ganglion cell genesis. Misexpression of Ascl1 or Neurog2 in preneurogenic retina was sufficient to initiate ganglion cell genesis. Misexpression of Neurog2 initiated the stereotypic sequence of proneural gene expression that normally preceded ganglion cell genesis. Ascl1 expression was also sufficient to initiate ganglion cell genesis. However, it functioned by a mechanism distinct from that of Neurog2. These results suggest that ganglion cell genesis may be initiated by two different mechanisms.
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    The role of cellular calcium channels in planarian biology
    (2011-11) Zhang, Dan
    Identification of signaling pathways and therein drugable targets, to manipulate stem cell behavior in vivo is a major focus of regenerative medicine. This dissertation focuses on the role of Ca2+ channels in stem cell differentiation and regeneration in a simple in vivo model, the planarian flatworm. These animals maintain a totipotent population of stem cells that give rise to all the cell types in the worm. Previously, we discovered that the isoquinoline drug praziquantel (PZQ) caused a robust (100%) and complete duplication of the entire anterior-posterior (AP) axis during flatworm regeneration to yield two-headed (bipolar) organisms. My studies mechanistically dissect these observations to show that PZQ subverted regeneration via activation of a specific neuronal voltage-gated Ca2+ channel (VGCC) isoform (Cav1A). Surprisingly, another isoform Cav1B was found to play opposing roles in axis formation to promote tail regeneration, suggesting a delicate interplay between Ca2+ signals critical for nervous system regeneration. Further dissection of the downstream pathway showed that RNAi of Cav1A blocked PZQ-evoked bipolar regeneration, Ca2+ entry and decreases in Wnt levels, the output of Hedgehog signaling. Thus, these data demonstrated that calcium signaling regulated regeneration through modulating Hedgehog signaling, a pathway that has been shown to regulate neuronal stem cell behavior, patterning and growth in diverse development processes. Taken together, these findings add new insights into the mechanisms that govern planarian regeneration. Additionally, my work on intracellular Ca2+ release channels in this system led to the identification of the planarian inositol 1, 4, 5-trisphosphate receptor (IP3R). Studies designed to elucidate the biological significance of this protein by in vivo RNAi knockdown led to the discovery that sexual planarians underwent severe defects of laying eggs in the absence of IP3R, although it failed to produce an obvious phenotype in asexual worms. Thus, these data provided genetic evidence that IP3R plays an important role in regulating reproductive physiology in planarian flatworms. In summary, the data obtained in this thesis have revealed essential roles of Ca2+ signaling in regulating planarian stem cell differentiation and reproductive physiology.
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    Thalamocortical axons regulate superficial layer neurogenesis and cell fate in the embryonic and neonatal sensory cortex
    (2021-10) Monko, Timothy
    The mammalian neocortex is composed of diverse types of neurons that are sequentially generated during embryonic development and are arranged in an orderly manner across layers. In the adult neocortex, the distribution and density of each neuronal type differ across the cortex. In primary sensory cortex—visual, somatosensory, and auditory—neurons that reside in layer 4 are targeted by axons from corresponding principal sensory nuclei of the thalamus. Synaptic activity of these thalamocortical axons are known to shape cellular organization, characteristic morphology, and patterns of gene expression of layer 4 neurons in postnatal sensory cortex. Even though experiments nearly half a century ago found evidence for this thalamocortical axon based extrinsic regulation of the mammalian neocortex, most scientists today attribute the specification of sensory cortex to be driven by intrinsic gene regulatory mechanisms. However, whether these axons influence earlier stages of development, including neurogenesis and neuronal fate specification is poorly understood. We first found that thalamocortical axons arrive in the cortex during the early period of superficial layer neurogenesis. Analysis of mutant mice lacking the majority of thalamocortical axons showed a decreased number of superficial layer neurons in primary visual and somatosensory areas, but not in motor areas, at early postnatal stages. During embryonic development, lack of thalamocortical axons decreased the number and division of radial glia and intermediate progenitor cells in a sensory area-specific manner. Cell fate analysis using pulse-labeling of progenitor cells with a thymidine analog revealed that thalamocortical axons are required for proper specification of superficial layer neurons towards layer 4. Further evidence revealed that this role of thalamocortical axons on cell fate is exerted as early as neonatal stages, demonstrating that these axons are required for the proper number of layer 4 neurons to be generated and specified in primary sensory areas even before mature synaptic input. We found that these area-specific roles are in part played by the thalamus-derived molecule, VGF. In summary, our study provides a mechanism by which afferent input to the neocortex from the thalamus complements the intrinsic program of early cortical development, allowing the sequential generation of diverse neuron types in an area-specific manner.