Adult hippocampal neurogenesis—a unique form of plasticity in the dentate gyrus (DG)—is regulated by experience, and when manipulated can have specific effects on behavior. Different methods have been used over the years to study new neurons’ functional role in the hippocampus, many of which focus on ablating neurogenesis. While ablation methods can test the necessity of adult-born granule cells (abGCs) for behavior, these techniques remove all abGCs from the circuit and thus do not allow one to determine which properties of abGCs are required for behavior. Such information is required to understand the mechanism of their action. Thus, new strategies are needed to determine what properties of young abGCs allow them to distinguish themselves from their mature counterparts and uniquely impact behavior. Recent hypotheses have suggested that the enhanced synaptic plasticity exhibited by 4–6-week-old abGCs allows them to uniquely contribute to hippocampal circuit function, and thus behavior. The primary goal of this thesis was to explore the contribution young abGCs’ heightened synaptic plasticity makes to hippocampal function. This was achieved using a transgenic mouse approach that allowed for the conditional deletion of NR2B from abGCs. Overall, iNR2BNes mice generated the same number of new neurons in adulthood as control mice at baseline. These neurons survived and matured with only a slight reduction in dendritic complexity. However, a potentially important electrophysiological property of these neurons—their enhanced synaptic plasticity—had been eliminated. From an electrophysiological standpoint, iNR2BNes mice resemble mice with ablated neurogenesis, while from all other neurogenic standpoints examined they most closely resemble wild-type mice. Consequently, these mice provided a novel model to test the extent to which young abGCs’ enhanced plasticity contributes to hippocampal-dependent behaviors. The results reveal that eliminating NR2B-containing NMDA receptors from abGCs does not alter baseline anxiety or antidepressant (AD)-like behavior. However, iNR2BNes mice differed from controls in measures of cognitive function. These mice were able to learn in the contextual fear conditioning test, but were impaired in the more difficult contextual fear discrimination test. Mice also exhibited a decreased novelty exploration phenotype that impaired their performance in the novel object recognition test. Together, these results indicate that the NR2B-dependent heightened plasticity exhibited by 4–6-week-old abGCs is necessary for responses to novelty and fine contextual discrimination, but does not contribute to baseline anxiety or emotionality. AD treatment increases levels of adult neurogenesis in the hippocampus, and these newborn neurons have been shown to be necessary for some of the behavioral effects of ADs seen in rodents. In addition, the maturation timeline of adult neurogenesis correlates with the onset of behavioral responses to ADs. ADs also enhance a neurogenesis-dependent form of long-term potentiation (LTP) in the DG evoked by medial perforant path stimulation under intact GABAergic tone called ACSF-LTP. Thus, a potential mechanism by which abGCs may contribute to AD behavioral efficacy is by providing extra plastic units to the DG circuit. This theory was tested by once again using the mouse line in which NR2B can be conditionally deleted from abGCs in the DG. Here, we found that deletion of the NR2B subunit significantly attenuated a neurogenesis-dependent behavioral response to fluoxetine in the novelty suppressed feeding test, and additionally blocked fluoxetine’s ability to enhance young abGCs’ maturation and subsequent integration into the hippocampal network. This suggests that eliminating abGCs’ enhanced plasticity decreases their ability to influence DG output resulting in an AD response that is less robust than seen in control mice. Control experiments revealed the specificity of this effect, as NR2B deletion did not impact the effect of fluoxetine in a neurogenesis-independent behavioral assay (tail suspension test) or in an assay that was insensitive to fluoxetine in this strain of mice (elevated plus maze). Our efforts to isolate the contribution of abGCs’ unique physiology from the neurogenic effects of fluoxetine were not entirely successful as the results presented here also revealed slight group differences in neurogenesis between control mice and mice lacking NR2B in young neurons. Yet, this data still supports the idea that fluoxetine increases the ability of abGCs to participate in DG output by increasing the chance that new neurons will be activated during DG stimulation. This may be achieved either by increasing their overall number, increasing their potential to make synaptic connections, or increasing their ability to strengthen their connections. However, due to the close link between activity and maturation that appears to be enhanced with fluoxetine treatment, a different approach with greater temporal resolution is needed to separate the neurogenic effects of fluoxetine from the physiological contribution abGCs make to hippocampal output. With this in mind, a mouse line in which abGCs could be temporally inhibited was also generated. Cellular and behavioral characterization of mice conditionally expressing hM4Di—a mutated muscarinic acetylcholine receptor that is insensitive to endogenous acetylcholine, but can be activated by the biologically inert, highly bioavailable compound, clozapine N-oxide (CNO)—has begun. Results show that acute CNO treatment in mice expressing this designer receptor exclusively activated by a designer drug (DREADD) in DG granule cells can impair encoding of contextual fear memory. Chronically treating these mice had an anxiogenic effect in the open field test, but otherwise anxiety and emotionality in these mice were comparable to controls. Chronic CNO treatment in mice expressing hM4Di in young abGCs effectively decreased these cells’ dendritic complexity, but did not alter proliferation or early survival. Thus, hM4Di DREADDs represent a novel tool that can be used to modulate activity of neurons in a temporally restricted manner, allowing for both acute and chronic manipulations of hippocampal granule cells. The experiments put forth in this thesis will highlight the importance of abGCs enhanced plasticity. The utility as well as potential pitfalls of the mouse models used here to test theories of abGC function will also be explored. Hopefully this analysis will provide an improved framework in which future experiments can be developed with the aim of uncovering novel insights into the hippocampal circuitry that underlies learning and memory and discovering new strategies for the treatment of neurological and psychiatric disorders.

Brain development is an exquisitely coordinated process of progenitor cell proliferation followed by the migration of progeny to their final location in the developing brain. There are a myriad of points at which this process can be disturbed, and the examination of these perturbations help us further understand basic science, as well as epidemics sweeping through the world around us. Microcephaly, which is defined as a head circumference greater than 2 standard deviations below the mean, can occur through genetic, infectious, vascular, or metabolic etiologies, and the studies herein examine two forms by which microcephaly occurs. First, we investigate the role of the dynein regulatory protein Nde1 in the development of the neocortex, which is the outer region of the forebrain. NDE1 mutations are associated with severe microcephaly, and we find that unlike most microcephaly genes whose products have one role in the cell cycle, Nde1 is required at three discrete points in neuronal progenitors, termed radial glia progenitors (RGPs). We initially find that Nde1 is required to recruit dynein to the nuclear envelope to allow for interkinetic nuclear migration (INM) during G2. Additionally, Nde1 helps to initiate primary cilia resorption at the G1-to-S transition. Finally, there is a necessity for Nde1 at the G2-to-M transition after the completion of INM and prior to nuclear envelope breakdown. These three distinct roles for Nde1 illustrate the breadth of functions that the protein has during RGP proliferation, and help to explain why patients with NDE1 mutations have such severe microcephaly. As this work was ongoing there was a global outbreak of a new pathogen that had previously been dormant throughout Africa and Asia, only to emerge at epidemic proportions in the Western Hemisphere. This pathogen, the Zika Virus (ZIKV), is particularly alarming because of its subclinical course in adults but devastating consequences for fetal development, with the hallmark symptom being microcephaly. Using our organotypic brain slice model system, we demonstrate the ability of a variety of ZIKV isolates to infect and replicate in embryonic brain tissue. All ZIKV isolates that infect the organotypic slices lead to increases in apoptosis, though these increases are particularly pronounced in isolates from the Asian/American lineages. Notably, one isolate from a patient in Nigeria (termed 30656) does not replicate in mouse neuronal tissue, but electroporation of the 30656 ZIKV genome allows for a single cycle replication, suggesting that this isolate is unable to enter RGPs. All infectious isolates are pathogenic in early- and mid- gestation embryonic tissue, but only one isolate infects and replicates in late- gestation embryonic tissue. This was the most recently isolated sample tested, and it demonstrates a predilection for neurons, suggesting that ZIKV may be mutating as it spreads. These results provide foundational insight into the pathogenesis of ZIKV- associated microcephaly, and illustrate how studies of genetic forms of microcephaly can enhance and facilitate our understanding of infectious causes of the disease.

Abstract not available

Thesis (Ph. D.)--University of Hong Kong, 2009. / Includes bibliographical references (p. 200-224). Also available in print.

Thesis (Ph. D.)--University of Hong Kong, 2009. / Includes bibliographical references (p. 200-224). Also available online.

published_or_final_version / Anatomy / Master / Master of Philosophy

MicroRNAs are short RNA molecules composed of 20-22 nucleotides. They highly accurately indicate cell identity and hence they are useful in labeling cells and tacking lineage commitment. However, this requires accurate microRNA profiling of cells in individual developmental stages. Since microRNAs are important negative regulators of eukaryotic gene expression, microRNA profiling allows better understanding of molecular regulatory networks of important cellular events, such as adult neurogenesis. Adult neurogenesis is the process in which neurons, as well as glia, are generated from neural stem cells. It was found to be responsible for brain regeneration, olfactory discrimination, memory formation and learning. Depression was suggested to be related to dysregulation of neurogenesis. Thus, knowledge in cellular and molecular mechanisms of adult neurogenesis will lay solid foundation to develop therapies to regenerate neural cells after injuries or onsets of neurodegenerative diseases and to understand the cognitive ability, memory formation and learning of the brain. In spite of its importance, investigation into the miRNA profiles and functions in neurogenesis is still infant. This project aimed to establish a preliminary microRNA profile on neurogenesis. Although this was not completed, the project could be extended to a large-scale microRNA profiling in neurogenesis. This would enable future workers to track the lineage commitment, the migration, and the distribution of NSCs and their derived cells accurately by in situ hybridization. Also, the future workers may construct a 2D representation of the changes in miRNA profiles and this may lead to discovery of previously unknown molecular and cellular differences among cells of same cell identity. / published_or_final_version / Anatomy / Master / Master of Medical Sciences

Adult neurogenesis is a fundamental feature of mammalian nervous systems. Curiously, neurogenesis in the dentate gyrus is typically regarded as homogenous. This thesis challenges that view, and reports the discovery and characterization of a novel region of the dentate gyrus that consistently lacks basal neurogenesis. We demonstrate that this area, referred to as the neurogenically quiescent zone, represents approximately 1.5% of the total volume of the dentate gyrus, and that its location is invariant among animals. This region contains several critical cell types and molecular factors that are known to be critical to the neurogenic niche, including stem cells. We also present data that attempt to conceptualize the existence of this region in the context of early agerelated declines in neurogenesis. Finally, we demonstrate that, under some behavioural conditions, neurogenesis can be induced in this region, suggesting that, under basal conditions, it may simply lack the presence of pro-neurogenic factors. / xvi, 125 leaves : ill. ; 29 cm. --

No description available.

Thesis (M.A.)--University of North Carolina Wilmington, 2008. / Title from PDF title page (October 20, 2008) Includes bibliographical references (59-71)