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Item Determining function of the IKAP protein in the peripheral nervous system for targeted therapeutic intervention in familial dysautonomia(Montana State University - Bozeman, College of Letters & Science, 2017) Ohlen, Sarah Beth; Chairperson, Graduate Committee: Frances Lefcort; Magdalena L. Russell, Michael J. Brownstein and Frances Lefcort were co-authors of the article, 'BGP-15 prevents the death of neurons in a mouse model of familial dysautonomia' in the journal 'Proceedings of the National Academy of Sciences' which is contained within this thesis.Familial Dysautonomia (FD) is a recessive genetic disorder that leads to devastation of the peripheral nervous system and is the result of incomplete neurodevelopment and progressive neurodegeneration. The disorder is also marked by a continual loss of retinal ganglion cells that leads to blindness. Even with early identification and treatment, the disorder is ultimately fatal. FD is caused by mutation in the IKBKAP gene that leads to cell-type specific loss of the IKAP protein, also known as ELP1. IKAP functions as a part of the six-unit Elongator complex. The role of Elongator is unresolved, although data has accumulated that support Elongator function in tRNA modification and efficient translation of proteins and that its absence leads to cell stress and neurological impairment. We have a mouse model of FD in which mouse Ikbkap is deleted from the peripheral nervous system, and it recapitulates the death of autonomic and TrkA+ sensory neurons observed in FD patients. As we can culture TrkA+ neurons in vitro, while also studying this neuronal population in vivo, we have a system to investigate our goals of (1) determining cellular processes that go awry in absence of Ikap and (2) targeting these cell types and events to prevent their progressive death. We have determined that mitochondrial and cytoskeletal function are disrupted in Ikbkap -/-, TrkA+ neurons and show activation of stress signaling. Interestingly, disrupted mitochondrial function is an emerging hallmark common to most neurodegenerative diseases. We have identified that the compound, BGP-15, is able to restore aspects of mitochondrial function and stress signaling in vitro and can restore neuronal survival of TrkA+ neurons lacking Ikap in vitro and in vivo. BGP-15 also improves actin cytoskeletal function and target innervation. Additionally, we have determined that introduction of the C-terminal half of human IKAP is sufficient to increase neuronal survival in vitro. This smaller protein fragment is compatible with viral delivery to retinal ganglion cells and could be utilized for gene therapy, potentially preventing this neuronal death that lead to blindness. Our goals now are to further explore stress pathways common to many neurodegenerative disorders and optimize rescue strategies in vivo.Item Analysis of the expression and function of chicken protocadherin 1 in neural crest cell migration and peripheral nervous system formation(Montana State University - Bozeman, College of Letters & Science, 2007) Bononi, Judy; Chairperson, Graduate Committee: Roger Bradley.The necessary steps of development from a single cell to a multi-celled functional organism are complex. Many molecules have been identified and their roles characterized in this process. One interesting population of cells includes the highly migratory neural crest cells (NCCs) unique to the vertebrate embryo and existing transiently during early embryonic development. The NCCs migrate along specific pathways at specific timepoints, stop at target locations, differentiate and give rise to a variety of cell types and tissues. Trunk NCCs must choose between two different migratory pathways: the ventral route, giving rise to neurons and glia of the dorsal root ganglia (DRG), sympathetic ganglia (SG), Schwann cells of the ventral root (VR); or the dorsolateral pathway, giving rise to melanocytes. Although many aspects of neural crest migration have been elucidated, cessation of migration and subsequent differentiation at target structures is not clearly defined. One family of molecules involved in various steps of NCC migration is the cell-cell adhesion molecules, the cadherins. To investigate the involvement of cadherins in NCC migration and differentiation during development using the avian model system, a combination of experiments and techniques including a library screen, in situ hybridization, in ovo electroporation, immunohistochemical and immunofluorescence staining as well as live time-lapse confocal imaging were performed. Results from these experiments produced the discovery and isolation of a novel molecule in the family of cadherin adhesion molecules, chicken protocadherin-1 (cPcdh1). Expression analysis showed cPcdh1 expressed in migrating NCCs, the DRG, SG and Schwann cells along the VR. A distinct expression pattern showed cPcdh1 along the periphery of the DRG, where crest cells are in an undifferentiated and mitotically active state. Further testing with deletion constructs and siRNA demonstrated when cPcdh1 function is inhibited, a greater percentage of cells migrate to the SG and VR at the expense of the DRG. Time-lapse confocal imaging showed cPcdh1 cells having an elongated cell shape with contact primarily being formed with neighboring cells along the periphery and longer cell-cell contact than observed in the control. Collectively, the results provide evidence for cPcdh1 involvement in NCC migration arrest and DRG formation.Item Characterization of the neural codebook in an invertebrate sensory system(Montana State University - Bozeman, College of Letters & Science, 2007) Aldworth, Zane Nathan; Chairperson, Graduate Committee: John P. Miller; Tomas Gedeon (co-chair)An outstanding problem in neuroscience is to describe the relationship between various stimulus sources in the environment and how they are represented by patterns of activity in nervous systems, a problem generically referred to as 'neural coding'. Most previous methods developed to address this problem have assumed a linear relationship between environmental stimuli and neural responses, and generally relied on measures of the mean state of the environment preceding neural activity to characterize the stimulus-response transformation. The goal of this thesis is to develop new methods of characterization that extend earlier work, and to demonstrate the utility of these new methods through application to an invertebrate sensory system. All applications of the methods developed in this thesis were carried out in the cercal system of crickets. The cercal system mediates the detection and analysis of low velocity air currents, and is implemented around an internal representation of air current direction that demonstrates the essential features of a continuous neural map. The stimulus feature selectivity, timing precision and coding characteristics of two bilateral pairs of primary sensory interneurons of the cercal system were characterized using three novel techniques. First, estimates of the cells' feature selectivity that take the natural variance in stimulus-response latency (i.e., spike 'jitter') into account were derived. Second, the cells' stimulusresponse relationship was probed for specific non-linear aspects that could constitute 'temporal' encoding. Third, an iterative stimulation paradigm was used to test and refine the predictions of the cercal system's stimulus selectivity. Compared to earlier characterization of this system, these new analytical procedures yield significantly different estimates of the stimulus feature selectivity of these cells. A 'code book' for the stimulus-response characteristics of these cells is presented, with emphasis on demonstrating instances where a cell represents different stimuli with distinct spike 'code-word' patterns.