Doctor of Philosophy, The Ohio State University, 2015, Molecular, Cellular and Developmental Biology

Sight-threatening diseases of the eye are prevalent across the world and result in a progressive loss of visual acuity that often culminates in blindness. These ocular diseases are caused by the degeneration and death of neurons in the retina, the neurosensory tissue of the eye. A promising possible treatment for retinal diseases is to stimulate neuronal regeneration from the Muller glia. Muller glia regenerate the retina in cold blooded vertebrates, but fail to do so mammalian retinas. To achieve retinal regeneration, Muller glia must de-differentiate into Muller glia-derived progenitor cells (MGPCs), proliferate, differentiate into neurons, and functionally integrate into neuronal circuits. Identification of the signaling pathways that influence the reprogramming of Muller glia into MGPCs is key to harnessing the potential of these cells to regenerate the retina. This dissertation focuses on understanding the role Glucocorticoid Receptor (GCR)- and Wnt-signaling pathways, in regulating proliferative, regenerative and neuroprotective properties of Muller glia in the avian retina in vivo. In the first part of this dissertation, we analyze the impact that the GCR- signaling pathway has on the MGPCs in the presence and absence of damage. The primary amino acid sequence of GCR and its expression by Muller glia is highly conserved across vertebrate species, including chickens, mice, guinea pigs, dogs and humans. We find that, in damaged retinas, activation of GCR-signaling suppresses the formation MGPCs via inhibition of MAPK-signaling, and inhibition of GCR-signaling stimulates the formation of proliferating MGPCs. In undamaged retina, FGF2/MAPK-signaling stimulates the formation of MGPCs, and activation of GCR-signaling reduces the number of proliferating MGPCs in FGF2 treated retinas. We also find that inhibition of the GCR-signaling enhances the neuronal differentiation of MGPC-derived cells in damaged retina. The second part of this dissertation describes th (open full item for complete abstract)

Committee: Andy Fischer (Advisor) Subjects: Biology; Cellular Biology; Developmental Biology; Molecular Biology; Neurosciences

PhD, University of Cincinnati, 2014, Medicine: Molecular and Developmental Biology

A large body of studies revealed the extensive conservation of eye development and function between the fly and vertebrates. In the case of the fruit fly (Drosophila melanogaster), compound eye development begins in late larval life in an epithelial sheet known as the eye imaginal disc in which the photoreceptors and corneal lens secreting cone cells are born. This is followed by a less studied stage of eye development, the pupal stage, in which the pigment cells are recruited and patterned to form the distinct hexagonal shape of each eye unit (ommatidia). Finally, midway through pupation, the cells in the eye terminally differentiate in preparation of performing the adult function. This final stage of development is by far the least studied and in the case of the cone cells, almost nothing is known about these cells other than their ability to secrete the corneal lens and underlying pseudocone. In this body of work, we analyzed the putatively conserved roles of the transcription factors prospero and dPax2 during cone cells throughout their development. From these studies, we found that these transcription factors have antagonistic effects on Notch and Ras/MAPK signaling during cone cell specification, creating an asymmetry between the cone cells. Using this information, we further show that the asymmetry created by pros and dPax2 in the cone cells is fundamental during pupation for the shape and positions of the surrounding retinal pigment epithelium that gives each ommatidium its classical hexagonal form. Finally, we show that adult cone cells are radial cells that use pros and dPax2 to control glial functions (physiological and structural support) for the underlying retina. Taken together, these studies provide a foundation for studying the basic principles of cell specification and pattering, as well as develop a new model system for studying glial biology.

Committee: Tiffany Cook Ph.D. (Committee Chair); Zubair M. Ahmed Ph.D. (Committee Member); Elke Buschbeck Ph.D. (Committee Member); Masato Nakafuku M.D. Ph.D. (Committee Member); Steven Potter Ph.D. (Committee Member) Subjects: Developmental Biology

PhD, University of Cincinnati, 2023, Arts and Sciences: Biological Sciences

Arthropods are arguably the most diverse group in regard to eye types that are observed in the animal kingdom. Despite their anatomical and functional differences, numerous evolutionarily conserved parallels have been identified between various eye types. Among arthropods these parallels include shared gross developmental plans, similar cell types and functionally relevant gene regulatory networks. These parallels suggest the existence of a set of common features that are necessary for the formation of a functional eye, regardless of its origin. Most of the studies that have explored these parallels in arthropods have been done in the photoreceptor cells of Drosophila melanogaster compound eyes and to some extent in the compound eyes of Tribolium castaneum. Only a handful of studies in D. melanogaster have specifically explored similarities in the non-neuronal support cells, but evidence has emerged that support cells in D. melanogaster are glia. However, it is unclear if such a function is conserved in other arthropod eyes like the camera eyes. We here turned to the highly specialized principal camera eyes of the sunburst diving beetle Thermonectus marmoratus larvae to investigate whether the support cells of camera type eyes also have glia-typical functions. We probed this question in a multi-faceted manner using in depth tissue specific transcriptomics, physiology, and molecular manipulations. We found that the support cells in the principal camera eyes are enriched in glia-typical genes that are found in D. melanogaster Semper cells (a type of eye glia). Our physiological experiments showed that intervening with the osmotic environment of freshly molted T. marmoratus larvae attenuated the support cell mediated eye elongation and focus. Suggesting the presence of a glia-typical osmoregulatory function with significant functional implications. Lastly, we found a conserved expression of the transcription factor Cut, a marker for D. melanogaster Semper cells (open full item for complete abstract)

Committee: Elke Buschbeck Ph.D. (Committee Chair); Joshua Benoit Ph.D. (Committee Member); Nathan Morehouse Ph.D. (Committee Member); Mark Charlton-Perkins Ph.D. (Committee Member); Tiffany Cook Ph.D. (Committee Member); Daniel Buchholz Ph.D. (Committee Member) Subjects: Developmental Biology

Doctor of Philosophy, Case Western Reserve University, 2023, Neurosciences

Synaptic circuitry is sculpted by glia. We sought to develop the Drosophila antennal lobe as a new model for identifying conserved principles underlying synapse-glia interactions. Antennal lobe organization is highly stereotyped and characterized by individual glomeruli comprised of unique olfactory receptor neuronal (ORN) populations. The antennal lobe interacts extensively with two glial subtypes. Ensheathing glia wrap individual glomeruli, while astrocytes ramify considerably within them. Roles for ensheathing glia and astrocytes in phagocytosis of terminal arbors and synapses in the uninjured antennal lobes are undefined. Thus, we tested whether Draper is required in either glial subtype to regulate ORN terminal arbor size, shape, or presynaptic content in two representative glomeruli: VC1 and VM7. We find that glial Draper limits the size of individual glomeruli and restrains their presynaptic content. Moreover, glial refinement of ORN terminals is apparent in young adults, a period of rapid terminal arbor and synapse growth, indicating that synapse addition and elimination occur simultaneously in this circuit. Surprisingly, Draper plays differential roles in ensheathing glia and astrocytes in VC1 and VM7, arguing for local heterogeneity of neuron-glia interactions. We also tested whether Draper is required for activity-dependent remodeling of VM7. We find that glial Draper is required for the shrinkage of ORN arbors projecting to VM7 that occurs in response to ethyl butyrate (EB) exposure during early adulthood. Finally, we recorded spontaneous activity of projection neurons (PNs) downstream of VC1 and VM7 ORNs in animals with glial subtype-specific Draper loss. Either astrocytic or ensheathing glial knockdown of Draper leads to a dramatic increase in spontaneous activity and a reduction in processing performance of PNs demonstrating functional consequences to Draper loss. Together, these data demonstrate that the phagocytic activities of distinct glial (open full item for complete abstract)

Committee: Heather Broihier Ph.D. (Advisor); Pola Philippidou Ph.D. (Committee Chair); Masashi Tabuchi Ph.D. (Committee Member); Catherine Collins Ph.D. (Committee Member); Helen Miranda Ph.D. (Committee Member); Andrew Pieper M.D., Ph.D. (Committee Member) Subjects: Neurobiology; Neurosciences

Doctor of Philosophy, Northeast Ohio Medical University, 2019, Integrated Pharmaceutical Medicine

Glaucoma, the leading cause of irreversible blindness worldwide, is a group of neurodegenerative disorders characterized by progressive and irreversible retinal ganglion cell (RGC) loss beginning with an optic neuropathy. Due to our lack of knowledge of the complicated etiology and mechanisms of glaucoma progression, no cure is available to prevent blindness. My purpose in this dissertation was to explore early contributors to glaucoma pathology in an effort to develop targeted therapy. Glaucoma is a multifactorial neurodegenerative disease in which various factors have been identified to play a significant part in its pathogenesis. For example, increased intraocular pressure can disrupt ocular blood flow to the eye, resulting in hypoxia. Although hypoxia was identified as a contributing factor for glaucoma, no one has yet identified the hypoxic cell types, hypoxia timeline and extent during glaucoma. In this dissertation, I contributed to glaucoma research by filling this gap in our knowledge to deepen our understanding of early changes after ocular hypertension. First, I determined the timeline of hypoxia in different cell types in the visual pathway using two hypoxia detection methods, an antibody against hypoxia-associated protein adducts, and a reporter protein expressed as a result of HIF1 stabilization. In addition, during hypoxia, autophagy, degradation of damaged organelles, and mitophagy, selective degradation of mitochondria, are induced. Second, I characterized hypoxia, autophagy, and oxidative stress using two models of glaucoma, the DBA/2J mouse model and a model of ocular hypertension. Finally, I used mass spectrometry to explore the half-life of mitochondrial electron transport chain complexes to better understand mitochondrial turnover, and possibly mitophagy, during glaucoma. Through these investigations, I have utilized multiple techniques, such as: Intraocular injection; intraperitoneal injection; tail vein injection; tissue sectioning using mic (open full item for complete abstract)

Committee: Denise Inman Dr. (Advisor); Brett Schofield Dr. (Committee Member); Vahagn Ohanyan Dr. (Committee Member); John Johnson Dr. (Committee Member); Samuel Crish Dr. (Committee Member) Subjects: Neurobiology; Neurosciences

Doctor of Philosophy, The Ohio State University, 2021, Neuroscience Graduate Studies Program

Retinal degeneration is a leading cause of blindness. Progressive death of retinal neurons is irreversible and results in loss of vision. Currently, there are no therapeutic treatments for loss of vision caused by retinal degeneration. However, the ability of various vertebrate species to utilize endogenous cellular sources for neuronal replacement provides promise for retinal regeneration. Muller glia are the primary support cell of the retina, and they possess the capacity to de-differentiate from mature support cells into progenitor cells that give rise to various types of neurons throughout the retina. However, the capacity for Muller glia-mediated neuronal regeneration varies widely across phylogeny. Cold-blooded vertebrates, such as zebrafish, exhibit a robust regenerative capacity where Muller glia reprogram into proliferating progenitor cells that subsequently differentiate into all the various subtypes of retinal neurons to restore a fully functional retina after damage. This capacity is diminished in avian models where only a small proportion of neurons regenerate from Muller glia-derived progenitor cells. This capacity for regeneration is refractory in the mammalian system, with Muller glia failing to reprogram after damage. Understanding the molecular mechanisms that control Muller glia reprogramming and identifying mechanisms that permit reprogramming in the fish and prevent reprogramming in the mammalian system is essential to harnessing the capacity of these cells for potential therapeutic interventions to regenerate neurons and prevent blindness. The primary focus of this dissertation is how neuroinflammation regulates glial response to damage in the retina. The first data chapter focuses on how neuroinflammation impacts neuronal death after excitotoxin-induced retinal degeneration. We describe how microglia, the resident immune cell of the retina, become activated after damage and communicate with astrocytes, via cytokine signaling, to convey (open full item for complete abstract)

Committee: Andrew Fischer (Advisor); Colleen Cebulla (Committee Member); Karl Obrietan (Committee Member); Dana McTigue (Committee Member) Subjects: Neurosciences

MA, Kent State University, 2021, College of Arts and Sciences / Department of Anthropology

Neurodevelopment and brain aging varies between species and across the lifespan of all mammals. The brain undergoes significant changes through fetal development, early life development, and through aging and senescence even in the absence of pathology. Animal models are commonly used to learn more about the development and aging of the human brain however little is known about the development and aging of one of humans' closest evolutionary relatives, chimpanzees. The objective of this study was to determine age and/or species related effects on neuron density (Nv), glia density (Gv), and glia to neuron ratio (G:N) in Brodmann's area 9 (BA9) of the prefrontal cortex (PFC) using direct comparative methods in stereology to compare Nv, Gv, and G:N between age cohorts within-species and between-species in humans and chimpanzees. We found that there were no age-related effects on Nv, Gv, or G:N ratio within species across the lifespan but there were differences between species in Nv and Gv with chimpanzees having increased cell densities of both types. Additionally, there were significant differences between the developmental and young adult human cohorts in Gv with the young adult cohort having decreased Gv as compared to the developmental group. Additional analysis revealed differences in the way that cell densities changed across the lifespan between species indicating that aging has different effects on the PFC of the human brain in comparison to the chimpanzee brain. However, this research sample was limited with few young individuals in the chimpanzee sample restricting our ability to detect early age-related patterns in chimpanzees or between species in the younger cohorts.

Committee: Mary Ann Raghanti (Advisor); Richard Meindl (Committee Member); Anthony Tosi (Committee Member) Subjects: Aging; Neurobiology; Neurosciences; Physical Anthropology

Doctor of Philosophy, Case Western Reserve University, 2021, Genetics

Interrogating how somatic stem cells interpret and transmit the intricate web of extracellular and intracellular signals that regulate cell state is foundational to biology and informs regenerative medicine approaches for treating disease. Oligodendrocyte progenitor cells (OPCs) are stem cells in the developing and adult brain that form oligodendrocytes, which are responsible for myelinating and supporting neuronal axons. Here, we elucidate transcriptional regulators of the oligodendrocyte lineage in both physiologic and pathologic contexts to identify strategies for promoting myelin regeneration in disease. Oligodendrocyte formation follows a multi-step process of OPC differentiation into immature oligodendrocytes, followed by subsequent maturation to myelinating oligodendrocytes. The transcriptional regulators of oligodendrocyte maturation remain unknown. Here, we discovered that the transcription factor Sox6 forms developmental condensates across gene bodies in immature oligodendrocytes to stabilize this intermediate state. Loss of Sox6 prevented condensate gene activation and accelerated maturation of OPCs directly to myelinating oligodendrocytes. This work offers a novel approach to regenerate myelinating oligodendrocytes in disease. This multi-step differentiation process is impaired by low oxygen (hypoxia) as seen in stroke, premature birth, and respiratory distress syndromes. Foundational to the hypoxic response is the accumulation of evolutionarily conserved transcription factors called hypoxia-inducible factors (HIFs). While HIFs are transiently protective, chronic HIF accumulation drives distinct pathological responses in numerous tissues and exerts a powerful influence over cell fate decisions in a multitude of stem cell types, including impairing oligodendrocyte formation from OPCs. In this study, we demonstrate that non-canonical, cell-type-specific targets of HIF1a are sufficient to impair the expression of Sox10, which is required for oligode (open full item for complete abstract)

Committee: Paul Tesar PhD (Advisor); Drew Adams PhD (Committee Chair); Tara DeSilva PhD (Committee Member); Anthony Wynshaw-Boris MD/PhD (Committee Member) Subjects: Genetics; Neurosciences

PHD, Kent State University, 2021, College of Arts and Sciences / School of Biomedical Sciences

Alzheimer's disease (AD) is a progressive neurodegenerative disorder that is characterized by the accumulation of beta-amyloid plaques and neurofibrillary hyperphosphorylated tau tangles. According to the most recent report from the U.S. Centers for Disease Control and Prevention, Alzheimer's disease is the 6th leading cause of death in the United States and the number of cases as increased almost 150% in the last 20 years in the United States. The disease starts as a subtle change in memory that progresses into noticeable cognitive decline, often referred to as mild cognitive impairment, and progresses into severe and pervasive memory loss. One of the hallmarks of Alzheimer's disease is the accumulation of the misfolded protein product of the amyloid precursor protein, beta-amyloid. Beta-amyloid has been long considered and widely supported by genetics and biochemical observation to promote Alzheimer's disease pathology. One of the earliest and long-supported theories, the amyloid hypothesis, posits beta-amyloid accumulation as having a central role in the pathogenesis of Alzheimer's disease as a promoter of tau seeding and inflammatory signaling which causes immune dysfunction. Beta-amyloid therapeutics represent the majority of the clinical trial candidates for Alzheimer's disease therapeutics. Recent clinical failures of anti-beta-amyloid trials, including Aducanumab and Gantenerumab, have raised concerns for the ability to manage beta-amyloid accumulation. The current pipeline of drugs targeting both beta-amyloid and tau are ineffective; therefore warranting alternative therapeutic options. A possible alternative non-pharmacological option for targeting beta-amyloid plaque aggregation is using energy to disrupt large beta-amyloid plaques into smaller fragments that may be cleared by microglia, the innate immune cells of the brain. One manner in which we can generate sufficient energy in a minimal to non-invasive safe manner is to use an alternating magnetic (open full item for complete abstract)

Committee: Min-Ho Kim Ph.D. (Advisor); Fayez Safadi Ph.D. (Advisor); Colleen Novak Ph.D. (Committee Chair); Gary Koski Ph.D. (Committee Member); Songping Huang Ph.D. (Committee Member) Subjects: Biomedical Engineering; Biomedical Research; Nanoscience; Nanotechnology; Neurobiology; Neurology; Neurosciences

Doctor of Philosophy, The Ohio State University, 2021, Neuroscience Graduate Studies Program

Degenerative retinal diseases result in neuronal cell death, which causes permanent vision loss in humans. The response to the loss of retinal neuron across species is diverse, with some species capable of complete and functional retinal regeneration. One source of these regenerated neurons is Muller glia (MG), the primary glial cell of the retina. In zebrafish, MG respond to damage by dedifferentiating into progenitor cells (MGPCs) which proliferate before they undergo neurogenesis. While mice completely lack a regenerative response by MG, chick MG form progenitors with limited neurogenesis, proving to be an effective phenotype for understanding molecular pathways that influence reprogramming and cell fate. The focus of this dissertation was on understanding various cellular factors that regulate MG de-differentiation and proliferation or reactive gliosis. The work outlined in these chapters utilize new advancements in single cell RNA sequencing technology (scRNA-seq) to better understand the glial biology of cellular reprogramming. In recent years, we have been able to quantitatively capture the transcriptome of tens of thousands of individual cells in an individual library. Generating these libraries has allowed us probe for transcriptional changes in MG in response to damage or growth factor treatment. These large libraries have enabled us to model the transition from MG to MGPC, probe for novel factors, and guide hypotheses about the functional implications of these target genes. Chapter one focuses on Matrix metalloproteinases (MMP) in damaged retinas, and their regulation in response to retinal damage, and how these changes feed back onto Muller glia to influence their regenerative potential in the chick. Gelatinases in the retina are temporally regulated in retinal development, cellular migration, and axonal guidance. We found that oligodendrocytes were the primary expressors of gelatinase MMP-2, and that expression did not fluctuate with damage. We did ob (open full item for complete abstract)

Committee: Andy Fischer (Advisor); Dana McTigue (Committee Member); Colleen Cebulla (Committee Member); Karl Obrietan (Committee Member) Subjects: Bioinformatics; Biology; Cellular Biology; Neurobiology; Neurosciences

Doctor of Philosophy, Case Western Reserve University, 2021, Molecular Medicine

In contrast to humans, animals such as zebrafish are able to regenerate their retinas after injury. Retinal cells called Muller glia facilitate regeneration in fish, yet contribute to scar formation in humans. Zebrafish Muller glia respond to injury by dedifferentiating and undergoing an asymmetric cell division, giving rise to a Muller glia and a proliferating progenitor cell. Subsequent progenitor cell migration and differentiation restores proper retina structure and function. The mechanisms governing the migration and functional integration of these progenitor cells, however, are not yet understood. A cell-surface adhesion molecule called Alcama, a novel marker of activated zebrafish Muller glia, contributes to cell migration, ganglion cell axonal guidance, and retinal lamination during development in both fish and mice. We hypothesize that Alcama, expressed in Muller glia following injury, facilitates the migration of progenitor cells and is important for zebrafish retinal regeneration. Here, we describe how Muller glia were isolated from the heterogenic population of retinal cells in Tg(apoe:gfp) fish with fluorescence activated cell sorting (FACS). The GFP-high population of cells represent Muller glia, which only express Alcama after injury, a pattern that is unique to this population. We have previously shown that proliferating progenitor cells migrate from the inner to outer nuclear layer of the retina following injury under normal conditions. To determine Alcama's role in regeneration, we knocked down its expression in the adult retina with in vivo electroporation of antisense morpholinos targeting Alcama. Using EdU lineage tracing, we have observed significant differences between control and Alcama morpholino-treated eyes in the proportion of EdU-positive cells in the inner and outer nuclear layers over time, suggesting that Alcama is playing a role in progenitor cell migration during regeneration. We have also seen a delay in overall regeneration after (open full item for complete abstract)

Committee: Alex Yuan MD, PhD (Advisor); Bela Anand-Apte MBBS, PhD, MBA (Committee Chair); Aleksandra Rachitskaya MD (Committee Member); Tara DeSilva PhD (Committee Member); Takuya Sakaguchi PhD (Committee Member) Subjects: Biomedical Research; Cellular Biology; Molecular Biology; Ophthalmology

PHD, Kent State University, 2020, College of Arts and Sciences / School of Biomedical Sciences

The roles of glia in cognition, pathology, and brain evolution are becoming increasingly clear, although our understanding is far from complete. The goal of this research was to further our understanding of glia through a comparative study of mammalian brains utilizing immunohistochemical and stereological techniques. Glia density and the ratio of glia to neurons increases with overall brain size, likely reflecting an increase in neuronal energy expenditure. This finding led to the assumption that the number of astrocytes, a glia subtype involved in regulating metabolism, will also increase with brain size. To test this assumption, we quantified glia density (Gv), astrocyte density (Av), and astrocyte soma volume (SV) in layer I and white matter in the frontal cortex of 10 mammalian species of varying brain weights. We found that Av, SV, and Av to Gv (Av/Gv) significantly increases in layer I as brain weight increases. Within primates, humans have a significantly greater Av, SV, and Av/Gv in layer I. These results indicate an expansion of layer I astrocytes with increasing brain size. Astrocytes specifically regulate neuronal metabolism through the astrocyte-neuron lactate shuttle via two important transporters, excitatory amino acid transporter two (EAAT2) and glucose transporter one (GLUT1). Since the human brain is metabolically more expensive compared to those of other primate species, we hypothesized that these transporters would have increased in humans compared to three other catarrhine primates. We found that humans have a greater EAAT2 density, GLUT1 vessel volume, and GLUT1 area fraction compared to baboons and chimpanzees, but did not differ from macaques. Therefore, EAAT2 and GLUT1 are not related to the increased energetic demands of the expanded human brain. In addition, aquaporin 4 (AQP4) which plays a role in astrocyte migration and water homeostasis was examined among macaques, baboons, chimpanzees, and humans. We found through a qualitative examina (open full item for complete abstract)

Committee: Mary Ann Raghanti Ph.D. (Advisor); Richard Meindl Ph.D. (Committee Member); Anthony Tosi Ph.D. (Committee Member); Gemma Casadesus Smith Ph.D. (Committee Member); John Gunstad Ph.D. (Committee Member) Subjects: Biomedical Research; Neurobiology; Neurosciences; Physical Anthropology

Doctor of Philosophy, Case Western Reserve University, 2019, Neurosciences

It has been known for over a century that microglia change their phenotype and function in neurodegenerative diseases, but it has remained an open question whether cells actively modulate the disease process. This question was resolved when variants in the gene TREM2, which encodes a microglia-specific receptor, were identified as risk factors for Alzheimer's disease. Our work has focused on understanding TREM2's function in order to gain insight into how microglia contribute to neurodegeneration and normal brain function. When we looked at when and where TREM2 was expressed in the context of Alzheimer's disease, we were surprised to find that cells with the highest levels of TREM2 also expressed markers of peripherally derived immune cells, rather than brain-resident microglia. This finding suggested the provocative possibility that peripheral immune cells play an essential role in Alzheimer's disease pathology. In order to determine the functional role of TREM2 on those cells, we examined how microglia and peripheral immune cell function was altered in Alzheimer's disease mouse models lacking TREM2 expression. We found that, without TREM2, peripherally-derived immune cells were virtually absent from the brain, and resident microglia failed to respond to amyloid pathology. Interestingly, this also prevented astrocytes from responding to amyloid deposition. TREM2 deficient mice exhibited altered pathogenesis and progression of amyloid pathology and plaque-associated neuritic dystrophy. In addition to these disease-related functions, we also identified an important role for TREM2 in normal brain development. Mice lacking TREM2 expression were found to have reduced synapse number. Because microglia play a role in synaptic elimination during circuit refinement, we thought that perhaps overactive synaptic pruning by microglia would be responsible for this synaptic loss. However, we instead found fewer synapses engulfed by TREM2 deficient microglia. Rather, this synapse (open full item for complete abstract)

Committee: Gary Landreth (Advisor); Bruce Lamb (Advisor); Heather Broihier (Committee Chair); Evan Deneris (Committee Member); George Dubyak (Committee Member) Subjects: Neurosciences

Doctor of Philosophy, Case Western Reserve University, 2018, Physiology and Biophysics

Peripheral organ injuries serve as potent stimuli for the immune system, triggering local inflammatory activation that can become increasingly amplified as the injury progresses, this in turn can lead to a severe systemic immune activation, which wreaks havoc over physiologic function. This is exemplified by lung injury, which can induce failure of multiple organ systems, or cause profound disruptions to brain function. When lung injury occurs in perinatal settings it may induce developmental impairments that last into adulthood. Here, using an experimental model of lung injury I investigated the mechanisms responsible for altering brainstem control of cardiorespiratory function in neonatal and adult rats. In neonatal rats, I determined that when lung injury is induced just before a transition period for the neural control of breathing (spanning P11-15), it promotes: i) an increase to the number of apnea directly preceded by a sigh, and ii) depression of viscerosensory synaptic transmission to 2nd-order neurons located in the nucleus tractus solitarii (nTS). This depression occurs through a postsynaptic mechanism that increases the contribution of Ca2+-impermeable (CI) AMPA receptors, and is mediated by the immune response to lung injury; minocycline, an inhibitor of microglia/macrophage activation, prevented the lung injury dependent increase in post-sigh apnea and the CI-AMPAR mediated synaptic depression. In rat-pups that were injured just after the transition period, viscerosensory synaptic transmission was also depressed, but occurred in a CI-AMPAR independent manner that contrastingly was presynaptically mediated. Thus, discrete mechanisms are responsible for these synaptic changes. In adult rats, where lung injury also increased the frequency of post-sigh apnea, I determined a novel immune-to-brain communication (I¿Bc) pathway utilized by the injury, which involves the glial-barrier separating the area postrema (a circumventricular organ) from the imm (open full item for complete abstract)

Committee: Frank Jacono M.D. (Advisor); George Dubyak Ph.D. (Committee Chair); Corey Smith Ph.D. (Committee Member); Thomas Dick Ph.D. (Committee Member); Roberto Galan Ph.D. (Committee Member) Subjects: Biophysics; Neurobiology; Physiology

Doctor of Philosophy, The Ohio State University, 2017, Neuroscience Graduate Studies Program

Eye diseases that result in blindness are often caused by the death of retinal neurons. Muller glia are the major glial support cells in the retina and possess the potential to reprogram into neurogenic progenitor cells. In the fish, Muller glia are able to regenerate a fully functional retina following severe retinal damage. In avian and mammalian retinas, Muller glia retain their regenerative potential but to an insufficient extent to restore lost vision. A better understanding of the mechanisms that govern the reprogramming of Muller glia into neurogenic Muller glia-derived progenitor cells (MGPCs) may allow us to harness these cells in a therapeutically useful context. This dissertation examines the cell signaling mechanisms that regulate Muller glia homeostasis, de-differentiation, proliferation, and neurogenesis. The first chapter focuses on how Hedgehog signaling stimulates Muller glia to reprogram into proliferating progenitors in the chick retina. We show that the Hedgehog-pathway components are up-regulated in damaged retinas when MGPCs are known to form. Furthermore, we find that the Shh-ligand is potentially released by retinal ganglion cells and received by proliferating Muller glia. We report that activation of the Hedgehog pathway increases Muller glia proliferation in damaged and FGF2-stimulated retinas. Consistent with these findings, inhibition of Hedgehog-signaling at the level of the ligand, receptor, and transcription factors attenuate MGPC formation. Activation of Hedgehog signaling in the absence of damage or FGF2-application has no effect on Muller glia. We propose a model wherein retinal damage or FGF2-stimulation renders Muller glia responsive to the mitogenic effects of Hedgehog-signaling. The second chapter examines how Jak/Stat signaling impacts the regenerative capacity of the avian retina. We find that Jak/Stat signaling is rapidly activated in Muller glia in response to retinal damage. We also show that inhibition of the Jak/Sta (open full item for complete abstract)

Committee: Andrew Fischer PhD (Advisor); Dana McTigue PhD (Committee Member); Heithem El-Hodiri PhD (Committee Member); Karl Obreitan PhD (Committee Member) Subjects: Neurosciences

Master of Science, Miami University, 2017, Biology

Our overall objective was to understand the role of activated microglia in the spinal cord plasticity observed following transection of sympathetic axons. Microglia activation was dampened with the antibiotic minocycline (Mino) and microglia proliferation and survival, plasticity of other glial cells as well as neuronal plasticity and long term survival were examined. At one week post injury, microglia exhibited a reduced activation state, with fewer amoeboid microglia and more ramified microglia following Mino treatment, but no effects on microglia proliferation or survival were observed. Mino treatment also resulted in reduced numbers of astrocytes and oligodendrocytes in the spinal cord of injured animals and blunted the typical decrease in choline acetyltransferase (ChAT) expression by the injured neurons. Dampened microglia activation and reduced neurotransmitter plasticity of the injured neurons during the first week post injury may have contributed to the observed long term loss of ChAT+ neurons. However, Mino had no effect on the regrowth of ChAT+ axons into the SCG at 16 weeks post injury, suggesting that target derived factors are more important in facilitating target reinnervation. These results provide evidence of beneficial crosstalk between spinal cord microglia and both glia and neurons following peripheral injury, possibly to promote neuronal survival.

Committee: Lori Isaacson Ph.D (Advisor); Kathleen Killian Ph.D (Committee Member); Dawn Blitz Ph.D (Committee Member) Subjects: Biology; Neurosciences

Doctor of Philosophy, The Ohio State University, 2014, Neuroscience Graduate Studies Program

Spinal cord injury (SCI) is a devastating condition that causes marked and lasting functional impairment. There are currently no treatments available to either prevent tissue destruction, or repair the tissue that is damaged. One of the earliest consequences of a traumatic SCI is blood entering the spinal cord. This sets off a cascade of progressive hemorrhagic necrosis that is highly destructive to the tissue. Therefore intraspinal hemorrhage is critical to SCI outcome, and has profound long-term consequences. In this body of work we developed a model of intraspinal hemorrhage (ISH) that enabled us to better study its role in post-injury processes. With the ISH model we demonstrated the importance of intraspinal hemorrhage not only in instigating tissue pathology, but also in initiating endogenous repair responses such as NG2 cell proliferation and formation of new oligodendrocytes. We detailed the similarities and differences between a purely hemorrhagic injury and a contusion to gain insight into mechanisms of damage and endogenous repair. We used that knowledge to investigate a potential mechanism for blood-mediated signaling within the spinal cord: Protease Activated Receptor 1 (PAR1). PAR1 is a G-protein coupled receptor that is activated by serine proteases, many of which are present within the blood as part of the coagulation cascade. By using both a synthetic activating peptide (TFLLR) and an endogenous agonist (thrombin) in the uninjured spinal cord, we shed light on the effects of two different modes of PAR1 activation. In particular the effects on microglia morphology, axonal pathology, and NG2 cell proliferation were examined. Lastly, as our studies and others provided evidence for detrimental vascular effects of acute PAR1 activation following injury, we investigated the potential of blocking PAR1 immediately after SCI as a neuroprotective treatment strategy. Our results in three different SCI models indicate that PAR1 inhibition shows promise in dec (open full item for complete abstract)

Committee: Dana McTigue PhD (Advisor); Phillip Popovich PhD (Committee Member); Jonathan Godbout PhD (Committee Member); Christine Beattie PhD (Committee Member); Stephanie Di Stasi PhD (Committee Member) Subjects: Neurosciences

Doctor of Philosophy, Case Western Reserve University, 2015, Molecular Medicine

Autism Spectrum Disorder (ASD) is a neurodevelopmental disorder affecting 1 in 68 children within the United States, whose high heritability is complicated by genetic heterogeneity. Studying the functions of genes mutated in syndromic forms of ASD is one method for identifying common pathogenic processes. In these studies, we used a murine model based on germline alterations to PTEN that can occur in patients with PTEN Hamartoma Tumor Syndrome, a cancer predisposition syndrome with high rates of ASD. In the Ptenm3m4 mouse, the localization of the Pten protein is shifted toward cytoplasmic predominance. In Chapter 2, we describe how this mislocalization leads to a range of cellular and behavioral phenotypes. The Ptenm3m4 brain overproduces glial cells, and displays hallmark signs of neuroinflammation. Behaviorally, the mice have increased social motivation and balance problems reminiscent of high functioning ASD. After discovering this connection to human ASD, we hypothesized that the neural transcriptome of the Ptenm3m4 mouse would show alterations that were relevant to those seen in idiopathic ASD. Examining gene expression in this model at 2 and 6 weeks of age revealed a progressive increase in expression differences that fell into categories related to increased neuroinflammation and decreased synaptic transmission – areas disrupted in multiple studies of the ASD brain transcriptome. Additionally, the differentially expressed genes in Ptenm3m4 mice were enriched for ASD susceptibility genes. These findings point to PTEN as an upstream regulator of many pathways altered in idiopathic ASD. Finally, our studies suggest that the Ptenm3m4 mouse is a valid model of high functioning autism – from the original genetic alteration, through a range of transcriptional changes, to a behavioral phenotype that reflects the human disorder in ways previously unmodeled in mice.

Committee: Charis Eng MD,PhD (Advisor); Robert Miller PhD (Committee Chair); Thomas Frazier II/PhD (Committee Member); Bruce Lamb PhD (Committee Member); Jonathan Smith PhD (Committee Member) Subjects: Genetics; Neurosciences

Master of Science, The Ohio State University, 2014, Human Ecology: Human Nutrition

Obesity and major mood disorders (MMD), namely bipolar disorder and major depressive disorder, have drastically increased in recent years in the United States adolescent population. Evidence indicates higher levels of circulating pro-inflammatory cytokines in both obesity and mood disorders. The effect of adipokines released by adipose tissue on the central nervous system (CNS) and inflammation in the progression of mood disorders remains understudied. In our study, we aimed to elucidate mediators between adipose and neural tissue that may link progression of mood disorders and metabolic imbalance. We hypothesized that adipose tissue can produce and secrete signaling molecules such as neuroadipokines or inflammatory cytokines that influence CNS function. We investigated 1) production and secretion of glia maturation factor ß (GMFß) in WT and Aldh1a1-/- adipocytes and 2) inflammatory and anti-inflammatory factors in mood disorder patients' serum and their cumulative inflammatory impact on adipocytes measured by NF-kB, a master transcriptional regulator of inflammation. We found that GMFß is produced and secreted by adipocytes and its expression depends on aldehyde dehydrogenase 1A1. Thus, GMFß is a neuroadipokine that can function in adipose as well as neural tissue. In addition, we found increasing variability of NF-kB and its target cytokines if severity of MMD in patients increases. These data suggest that both suppression and increases in inflammation may influence MMD progression that warrants studies in specific MMD populations.

Committee: Ouliana Ziouzenkova Dr. (Advisor); Barbara Gracious Dr. (Committee Member); Earl Harrison Dr. (Committee Member) Subjects: Nutrition

Master of Science (MS), Wright State University, 2013, Anatomy

Peripheral nerve injuries (PNI) cause alternations in central synapses leading to loss of function. The C-bouton synapses onto a-motoneurons in the ventral horn, and has a role in regulating motor output. Following tibial nerve ligation, the somatic C-bouton coverage is depleted (Alvarez et al., 2011), however, it is unknown what happens following crush type injuries. PNI causes neuroglia activation and proliferation that contribute to synaptic alterations, a response that has not been well-characterized in the ventral horn, where motoneurons are located. Therefore, I hypothesize that glia activation following peripheral nerve injury correlates to the degree of depletion of synaptic coverage of C-boutons. To test, I performed Immunohistochemical analysis of rat spinal cord to characterize C-bouton coverage, and activation of glia following two types of PNI. Our results indicate less C-bouton depletion following tibial nerve crush than ligation injuries. In addition, we found less glia activation following crush than ligation injuries.

Committee: Robert Fyffe Ph.D. (Advisor); Adrian Corbett Ph.D. (Committee Co-Chair); Larry Ream Ph.D. (Committee Co-Chair) Subjects: Neurobiology; Neurosciences