Doctor of Philosophy, The Ohio State University, 2024, Neuroscience Graduate Studies Program

Patients with breast cancer experience debilitating behavioral side effects and exhibit elevated concentrations of circulating inflammatory mediators, even prior to treatment. Inflammation is a proposed mechanism underlying the etiology of cognitive and mood disturbances and thus may be contributing to tumor-induced behavioral side effects. Preclinical rodent models likewise display impairments in learning/memory and anxiety- and depressive-like behaviors. In addition to tumor-induced elevations of circulating inflammatory mediators, tumor-bearing rodent models display neuroinflammation in brain regions important in regulating behavior. However, the cellular and molecular mechanisms by which tumor-induced behavioral side effects occur remain unknown. Estrogen is not only implicated in the etiology of breast cancer, but also in regulating inflammation and behavior; therefore, estrogen signaling may mediate the relationships between tumor-induced neuroinflammation and behavioral side effects. Here, I examine the extent to which 1. estrogen modulates tumor-induced neuroinflammatory and behavioral outcomes and 2. different cells in the brain (i.e., microglia and endothelial cells) contribute to the observed tumor-induced neuroinflammatory phenotype using orthotopic and syngeneic mammary tumors in female mice. In Chapter 2, I demonstrate that ovarian status modulates tumor-induced endocrine and neuroinflammatory outcomes. Most notably, I show that a mammary tumor in ovary-intact mice reduces circulating estradiol and alters estrogen-related signaling in the brain. Further, tumors in ovariectomized mice increase circulating estradiol and modestly exacerbate tumor-induced peripheral and central inflammation. Next, in Chapter 3 I demonstrate that brain-specific estradiol supplementation attenuates tumor-induced fatigue in group-housed mice, but not singly housed mice; the attenuation of tumor-induced fatigue in group-housed mice is not mediated by neuroinflammatory mediato (open full item for complete abstract)

Committee: Leah Pyter (Advisor); Kathryn Lenz (Committee Member); Olga Kokiko-Cochran (Committee Member); Ruth Barrientos (Committee Member) Subjects: Neurobiology; Neurosciences

Doctor of Philosophy, The Ohio State University, 2024, Biomedical Engineering

Glioblastoma (GBM) is an aggressive and incurable primary tumor of the brain with high mortality and abysmal survival rate. Currently, available treatment options include chemotherapy, surgical resection, and radiotherapy, which only increase survival minimally. This underscores the urgent need for in vitro models that accurately recapitulate the complex tumor microenvironment, including the cellular and extracellular matrix (ECM) conditions. This dissertation discusses the development of a 3D in vitro model of glioblastoma to evaluate therapy responses and study the interaction between immune cells and GBM cells. Initially, we fabricated patient-derived tumor constructs with a hyaluronic acid-based hydrogel that can maintain the patient cells in their original genetic state and be amenable to accurate drug testing. Then we fabricated an optimized hydrogel that maintained the microglia cells in their resting state and employed them in co-culture with GBM constructs. The GBM-microglia co-culture was used to evaluate the reciprocal activation between the GBM tumor cells and microglia cells. Then the impact of this reciprocal activation on the altered drug response in the GBM constructs was analyzed. Finally, we explored strategies to improve the immunogenicity of GBM constructs under various culture conditions. Then the cytotoxicity of natural killer cells against GBM constructs in the presence of GBM-activated microglia constructs was analyzed.

Committee: Aleksander Skardal (Advisor); Jessica Winter (Committee Member); Gina Sizemore (Committee Member); Monica Venere (Committee Member) Subjects: Biomedical Engineering

Doctor of Philosophy, The Ohio State University, 2024, Neuroscience Graduate Studies Program

Chronic stress is associated with the development of psychiatric disorders, including anxiety and post-traumatic stress disorder (PTSD). Inflammation is a key component of chronic stress and the development of psychiatric disorders. Repeated social defeat (RSD) is a rodent model of chronic stress that induces neuronal activation, microglial reactivity, and monocyte accumulation. RSD is associated with the development of anxiety-like behavior, social and cognitive deficits, and enhanced fear memory. RSD is clinically relevant as it shares key cellular and behavioral features with PTSD. Microglia play a key role in facilitating monocyte accumulation and anxiety-like behavior following RSD. Additionally, microglia have a primed profile 24d after the cessation of RSD, but how microglia mediate social and cognitive deficits or how microglia communicate with other cells after RSD is unknown. Therefore, single-cell(sc)RNAseq and single-nuclei(sn)RNAseq of the hippocampus was done 14h after RSD (Chapter 2). Here, novel stress-associated microglia were identified and uniquely characterized by cytokine/chemokine, EIF2, and phagocytotic signaling. Microglia depletion with PLX5622 (CSF1R inhibitor) prevented RSD-induced transcriptional changes to endothelia and astrocytes, but had limited, yet regionally specific effects on neurons. Furthermore, RSD-induced social withdrawal and cognitive impairment were microglia dependent. Collectively, these stress-associated microglia influenced transcriptional profiles in the hippocampus linked to social and cognitive deficits. RSD also induces stress-sensitization, where mice have amplified neuronal, immune, and behavioral responses to acute stress 24d later. The mechanisms underlying stress-sensitization are unclear. Thus, the next step of this study was to determine the influence of microglia and neuronal IL-1R1 (nIL-1R1) in stress-sensitization and fear memory after RSD (Chapter 3). Here, RSD enhanced fear memory acutely. Importantly, (open full item for complete abstract)

Committee: John Sheridan (Advisor); Jonathan Godbout (Advisor) Subjects: Immunology; Mental Health; Neurosciences

Doctor of Philosophy, The Ohio State University, 2024, Neuroscience Graduate Studies Program

Pregnancy is a time of dramatic peripheral immune alterations to promote the development and later care for the offspring. However, these changes lend vulnerability to mental illness for some mothers, the most prominent being Postpartum Depression (PPD). In this dissertation, I aimed to investigate the role of the innate brain immune cell, microglia, in the maternal brain and its role in governing maternal mental health and behavior. Microglia decrease during late pregnancy, and these decreases persist postpartum. I first investigated whether microglia decreases are permissive for the onset and display of maternal care. Depleting microglia in a nulliparous maternal sensitization paradigm, I found that loss of microglia was sufficient for the display of caretaking behaviors. This was accompanied by changes in cellular activation across the maternal brain network. After establishing the functional importance of decreased microglia in facilitating maternal behavior, I investigated whether perturbations during pregnancy disrupted the maternal neuroimmune milieu. I also examined if neuroimmune changes were associated with impairments in care. I employed two different models of environmental perturbations I hypothesized would lead to neuroimmune alterations: gestational stress and chronic opioid exposure. Following exposure to chronic stress from gestational days (GD) 7-20, I examined microglia activity at GD21 and at postpartum day 8 (PD8). Immunolabeling of microglia and phagolysosomal marker CD68 increased in the postpartum medial prefrontal cortex (mPFC) in stressed mothers. A Nanostring nCounter panel revealed broad downregulation in stressed mothers of transcripts involved in immune activating and immune dampening effects, indicating dysregulated neuroimmune function at both GD21 and PD8 in the mPFC. Transcript alterations were also found for factors that control excitatory and inhibitory neuronal signaling, as well as components of the extracellular matr (open full item for complete abstract)

Committee: Kathryn Lenz (Advisor); Benedetta Leuner (Advisor); Tamar Gur (Committee Member); Laurence Coutellier (Committee Member); Ruth Barrientos (Committee Member) Subjects: Immunology; Neurosciences

Doctor of Philosophy (PhD), Ohio University, 2024, Biological Sciences (Arts and Sciences)

The neurovascular unit (NVU) is an anatomical and functional feature of the mammalian brain that encompasses the brain microvasculature and is pivotal for maintaining healthy nutrient, waste, and ion exchange between the blood supply and parenchymal tissue. Specializations of the NVU tightly regulate this exchange, thereby restricting the entry/exit of solutes to/from brain tissue and forming a functional blood-brain barrier (BBB). This complex network is composed of tightly interlinked cellular components – including vascular endothelial cells (ECs), pericytes, neurons, astrocytes, and microglia – making the brain vulnerable to pathologies affecting any single NVU element. Disruption in one part can lead to wider dysfunction, exacerbating neurological disease pathogenesis and symptoms. Cells of the NVU interact closely to maintain overall brain health. Pericytes are vital for vascular development and homeostasis, regulating cerebral blood flow and establishing the BBB. Astrocytes are also critical to the development and maintenance of the BBB and allow for vessel-parenchyma crosstalk through perivascular astrocytic end-feet. Microglia serve as regulators of immunogenic function and vascular development and are key players in maintenance of vessel integrity and diameter. Brain arteriovenous malformations (AVMs) present as high-flow vascular anomalies, distinguished by dilated vascular channels that forego a capillary bed, resulting in direct arterial-to-venous connections. This abnormality predisposes patients to spontaneous intracerebral hemorrhages. Currently, therapeutic options are restricted to invasive procedures, including stereotactic radiosurgery, endovascular embolization, and surgical excision. Such limitations underscore the urgent demand for the development of preventive strategies and less invasive treatment approaches. This dissertation reports on the molecular and cellular impacts to the NVU during pathogenesis of brain AVMs. Our lab uses a genetic m (open full item for complete abstract)

Committee: Corinne Nielsen (Advisor); Shiyong Wu (Committee Member); Daewoo Lee (Committee Member); Janet Duerr (Committee Member) Subjects: Biology; Cellular Biology; Molecular Biology; Neurobiology; Neurosciences

PhD, University of Cincinnati, 2023, Medicine: Neuroscience/Medical Science Scholars Interdisciplinary

Microglia are essential components for normal brain functioning at homeostasis and during disease or injury state. Recently it has become clear that TGF-ß signaling is essential for microglial development and homeostatic maintenance. However, it remains unclear which cell types produce TGF-ß1 ligand and how TGF-ß1 is spatially regulated in the brain. Additionally, it is unclear how dyshomeostasis in microglia influences astrocyte and neuronal function. This thesis focuses on addressing these gaps in addition to carefully evaluating the tools used to ensure rigor and integrity of collected data. The first study aims to determine how best to apply microglia targeted transgenic Cre mouse lines for studies of microglial gene function. To accomplish this a thorough and detailed comparison of their properties is needed. We examined four different microglial CreER lines (Cx3cr1CreER(Litt), Cx3cr1CreER(Jung), P2ry12CreER, Tmem119CreER), focusing on (1) recombination specificity; (2) leakiness - degree of tamoxifen-independent recombination in microglia and other cells; (3) efficiency of tamoxifen-induced recombination; (4) extra-neural recombination -the degree of recombination in cells outside the CNS, particularly myelo/monocyte lineages; (5) off-target effects in the context of neonatal brain development. We identify important caveats and strengths for each line to provide guidance for researchers interested in performing conditional gene deletion in microglia. We also provide data emphasizing the potential of these lines for injury models that result in the recruitment of splenic immune cells. We then used the described inducible cre lines to selectively knockout Tgfb1 in different cell types in adult mouse brains to examine which cells produce the ligand for microglial homeostatic maintenance. Our data support that microglia are the primary producers of TGF-ß1 needed for microglial homeostatic maintenance and that astrocyte and neuronal TGF-ß1 ligand are not r (open full item for complete abstract)

Committee: Steven Crone Ph.D. (Committee Chair); Yu (Agnes) Luo Ph.D. (Committee Member); Eric Wohleb Ph.D. (Committee Member); Thomas Thompson Ph.D. (Committee Member); Renu Sah Ph.D. (Committee Member) Subjects: Neurosciences

PhD, University of Cincinnati, 2023, Medicine: Neuroscience/Medical Science Scholars Interdisciplinary

Psychological loss is something that most people will experience in their lifetime. Losing something of value (e.g., interpersonal relationships, financial stability, secure housing, health) can erode well-being and negatively impact quality of life, generating a unique set of depression-like symptoms. While loss is common, it is difficult to track clinically and has received little attention in preclinical studies. Previously, our lab developed a protocol to study this experience of losing a positive stimulus in rats, called enrichment removal (ER). Environmental enrichment is a well-known positive, rewarding experience, and ER generates behavioral phenotypes that are reminiscent of loss symptoms in humans. Given this face validity, my dissertation work seeks to better understand the molecular mechanisms of loss using enrichment removal and a range of molecular and behavioral techniques. We first identified the basolateral amygdala (BLA) as an important region in ER. We then utilized a comprehensive multi-omics approach to identify novel candidate mechanisms in this region, identifying microglia and the extracellular matrix (ECM) as potential targets driving removal phenotypes. Further investigation into these candidates revealed that ER leads to a loss of function in microglia and an accumulation of ECM. The latter effects were especially intriguing, prompting further study of the matrix itself and the parvalbumin interneurons which it typically surrounds in the form of perineuronal nets. Overall, the present molecular results demonstrate that ER increases ECM in the BLA, decreasing overall plasticity in the region and altering parvalbumin phenotypes to promote inhibition. Next, we expanded our behavioral profiling of ER, focusing on behaviors regulated by the BLA. These results revealed an interesting phenotype of impaired salience evaluation, in which the strength of various stressors and the behavioral responses that they elicited were mismatc (open full item for complete abstract)

Committee: Teresa Reyes Ph.D. (Committee Chair); James Herman Ph.D. (Committee Member); Eric Wohleb (Committee Member); Diego Perez-Tilve Ph.D. (Committee Member); Robert McCullumsmith M.D. (Committee Member) Subjects: Neurosciences

Doctor of Philosophy, The Ohio State University, 2023, Neuroscience Graduate Studies Program

Sensorineural hearing loss (SNHL) is a common hearing health disorder caused by damage to cochlear sensory hair cells and neurons, which can occur during normal aging and from acoustic insults throughout the lifespan. Age-related hearing loss (ARHL) describes the form of SNHL that happens in aging patients with progressive loss of hearing perception. Noise-induced hearing loss (NIHL) occurs from noise damage and can happen at any age. Both ARHL and NIHL share similar features in cochlear tissue damages, including the loss of sensory hair cells, auditory neurons, and synapses in the cochlea. Further synaptopathy occurs in the cochlear nucleus (CN), which is where all arriving auditory information from the ear first enters the central nervous system. Acute and chronic cochlear inflammation are associated with worsening outcomes in SNHL. Resident innate immune cells modulate local inflammatory responses and are implicated in both the cochlea and the brain during SNHL. Tissue damage and age-related degeneration increase the activation of both cochlear macrophages and microglia in the brain. These activated immune cells release factors that recruit additional immune cells and alter tissues in ways that can contribute to the progression of pathology. Example signals potentially released by resident immune cells include complement factors, which are proteins also found in circulation that can accumulate in damaged or diseased tissues. This dissertation investigates the activation of innate immune cells, including cochlear macrophages and microglia during ARHL and NIHL, and whether the activation of complement pathways in the cochlea and CN may contribute to SNHL pathophysiology. CBA/CaJ mice were used as the animal model in this research, which were either aged in ambient noise environments to develop ARHL or exposed to various intensities of noise to cause NIHL. Hearing was assessed in all mice and tissues were collected to measure changes in the resident immune cells and (open full item for complete abstract)

Committee: Ruili Xie (Advisor); Eric Bielefeld (Committee Member); Ruth Barrientos (Committee Member); Dana McTigue (Committee Member) Subjects: Neurobiology; Neurosciences

Doctor of Philosophy, The Ohio State University, 2023, Neuroscience Graduate Studies Program

Traumatic brain injury (TBI) is damage to the brain that is caused by an external force. As a leading cause of disabilities worldwide, TBIs can lead to long-term symptoms such as physical, cognitive, and psychological symptoms, including headaches, photosensitivity, seizures, attentional impairment, memory problems, impulsivity, increased anxiety, depression, and substance use disorders. Nearly 40% of TBI patients report two or more psychiatric symptoms after injury. Despite the high proportion of individuals that report psychiatric symptoms after TBI, there are no FDA-approved treatments for such impairments. Additionally, the mechanisms behind the development of psychiatric symptoms are poorly understood, but chronic neuroinflammation is believed to play an important role. Following injury, microglia propagate a chronic neuroinflammatory response that lasts years after the initial event and is linked to increases in psychiatric conditions. This work aimed to better understand the relationship between neuroinflammation and the development of psychiatric symptoms of lack of motivation, impulsivity, and inattention. First, it was examined if lipopolysaccharide (LPS) could model the chronic depressive-like symptoms (lack of motivation) of TBI to establish that long-term neuroinflammation leads to psychiatric symptoms. Motivation was measured with an operant task, the progressive ratio, which works by progressively increasing the effort required to earn a single reinforcer. A 14-day continuous LPS exposure (5 mg/kg/week) subcutaneously did not induce motivational deficits. LPS infused directly into the lateral ventricles (10.5 ug/kg/week) for 14 days caused a significant drop in motivation. However, this decrease in motivation did not mimic the effects of post-TBI depressive-like motivational impairments. Collectively demonstrating that chronic LPS cannot be used as a model for post-TBI depressive-like phenotypes. The next study focused on find (open full item for complete abstract)

Committee: Cole Vonder Haar (Committee Chair); Jonathan Godbout (Committee Member); Olga Kokiko-Cochran (Committee Member); Kathryn Lenz (Committee Member) Subjects: Neurosciences

Doctor of Philosophy, The Ohio State University, 2023, Neuroscience Graduate Studies Program

Traumatic brain injury (TBI) is the leading cause of injury-induced disabilities that decrease quality of life. Neuroinflammation propagated by immune cells like microglia causes or worsens post-TBI maladies and is impacted by environmental factors after injury. Stress is a significant post-TBI factor that profoundly influences inflammation and provides a gateway to dysfunctional neuroimmune responses and behavioral decline following TBI. Mounting preclinical and clinical evidence finds TBI suppresses hypothalamic-pituitary-adrenal (HPA)-axis activity in response to stressors. Even though the HPA-axis is a key mediator of stress-immune bidirectional communication, little is known about the consequences of aberrant stress responses after injury. Sleep disturbances are a clinically relevant post-TBI stressor that could better elucidate the relationship between HPA-axis dysfunction and maladaptive inflammation after TBI. Indeed, acute sleep disruption engages the dysfunctional HPA-axis after lateral fluid percussion (lFPI) with suppressed production of stress hormone, corticosterone (CORT) (Chapter 2). Sleep disruption further exacerbates TBI-induced glial-mediated inflammation and causes lasting anxiety-like behavior. Prevalence of HPA-axis dysfunction in preclinical and clinical TBI populations is better reported, however mechanisms of how TBI alters HPA-axis regulation remain unknown. To determine how TBI influences stress-responsive circuitry and long-term inflammatory and behavioral consequences of sleep disturbances, transient sleep fragmentation (SF) was used as a transient, daily stressor chronically after lFPI (Chapter 3). SF engages the intact HPA-axis of uninjured sham mice, eliciting robust neuronal activity of the first step in the HPA-axis, the paraventricular nucleus (PVN). TBI enhances glucocorticoid receptor (GR) sensitivity in the hippocampus, a major regulator of HPA-axis reactivity that could confer TBI-induced suppression, associated with suppresse (open full item for complete abstract)

Committee: Olga Kokiko-Cochran (Advisor); Leah Pyter (Committee Member); John Sheridan (Committee Member); Jonathan Godbout (Advisor) Subjects: Neurosciences

Doctor of Philosophy, The Ohio State University, 2023, Biomedical Sciences

Every 90 minutes, someone is diagnosed with Amyotrophic lateral sclerosis (ALS), an adult-onset neurodegenerative disease. In ALS, motor neurons that innervate skeletal muscle degenerate, resulting in muscle atrophy and progressive weakness. This leads to difficulty in activities such as walking, swallowing, and breathing and ultimately leads to paralysis and death. As there is currently no treatment for ALS capable of substantially altering the disease course, this disease remains 100% fatal. 90% of ALS cases occur with no family history, while up to 10% of cases are inherited. In inherited (familial) ALS, up to 20% are caused by mutations in superoxide dismutase 1 (SOD1), with multiple mechanisms of cell death being attributed to SOD1-mutation related pathogenesis. Neuroinflammation is a main culprit in accelerating many neurodegenerative diseases, including ALS. However, treatments targeting neuroinflammation alone have shown disappointing results in clinical trials. Microglia, the innate immune cells of the central nervous system are critical mediators of neuroinflammation. Though initially protective in early disease stages, microglia become chronically activated and proinflammatory, and lead to motor neuron death. Previous studies from our lab and others have shown that reduction of mutant SOD1 expression is highly effective in preserving motor function and extending survival of multiple rodent models of ALS. Utilizing adeno-associated virus serotype 9 (AAV9), we developed a gene therapy for ALS that decreases SOD1 expression via short hairpin RNA (shRNA) (AAV9.SOD1.shRNA). Though this approach is extremely effective at targeting and correcting motor neurons and astroctyes, microglia are not efficiently targeted. Thus, microglia-mediated motor neuron toxicity continues. We hypothesized that the most successful therapeutic approach will target as many cell types involved in non-cell autonomous toxicity as possible. The work described in this thesis ev (open full item for complete abstract)

Committee: Kathrin Meyer (Advisor); Stephen Kolb (Committee Member); Jonathan Godbout (Committee Member); Allison Bradbury (Committee Member) Subjects: Biomedical Research

PhD, University of Cincinnati, 2022, Medicine: Neuroscience/Medical Science Scholars Interdisciplinary

Chronic stress reduces glutamatergic synapses in the medial prefrontal cortex (PFC), leading to decreased signaling from pyramidal neurons and behavioral changes. Conversely, the antidepressant effects of low-dose ketamine are linked to increased synaptic density. One explanation for this is through fluctuations in brain-derived neurotrophic factor (BDNF), a regulator of synaptic plasticity that is downregulated by stress and upregulated by ketamine. Historically, this has been attributed to neuronal mechanisms, however recent evidence suggests microglia express BDNF and phagocytose synaptic material after stress exposure. Still, our understanding of neuron-microglia interactions in the contexts of stress and ketamine treatment remains limited. Here, we first characterize the effects of chronic stress on neuron-microglia interactions in the medial PFC of male and female mice. We found that male, but not female, mice displayed behavioral changes after 14 days of CUS. qPCR of sorted PFC microglia revealed that CUS upregulated phagocytosis marker expression and decreased Bdnf levels in males. Confocal imaging showed that CUS reduced synaptic density and increased microglia-mediated neuronal remodeling in males. In subsequent experiments, we found behavioral changes in both sexes after 28 days of CUS, but limited stress effects on phagocytosis markers and microglia-mediated dendritic remodeling. Further, synaptic density remained unaltered in the PFC of females while males showed persisting deficits in spine density and microglial Bdnf expression after 28 days of CUS. These findings indicate chronic stress causes sex-specific and temporally dynamic changes in microglial function. Given that spine density and microglial Bdnf expression were decreased in males at both time points, we next sought to determine the significance of microglial Bdnf to synaptic effects of stress in male mice. To this end, we utilized transgenic mice with microglia-specific depleti (open full item for complete abstract)

Committee: Teresa Reyes Ph.D. (Committee Member); Steve Danzer Ph.D. (Committee Member); Eric Wohleb Ph.D. (Committee Member); Renu Sah Ph.D. (Committee Member); James Herman Ph.D. (Committee Member) Subjects: Neurosciences

MS, University of Cincinnati, 2022, Medicine: Cancer and Cell Biology

Microglia is the primary brain resident macrophage and is an integral component of brain development and functions including neurogenesis, angiogenesis, and synapse formation. The microglia exhibit spatial and temporal diversity in their morphology and functions. The microglial functions depend on the region and stage of development. Despite the advancement in research on microglia, the diversity profile of the microglia has yet been fully understood. Besides microglia, immune cells in the normal brain also include macrophages like the choroid plexus, meningeal, and perivascular macrophages. The role of microglia and different brain resident macrophages in normal brain physiology, neurological diseases, and brain tumors remains to be poorly defined. As the deadliest immune-privileged brain tumor, glioblastomas are largely treatment-resistant, which is in part due to the immunosuppressive tumor microenvironment. Specifically, the tumor-associated macrophages (TAM) act as a pro- and anti-tumorigenic component in GBM and are emerging as a potential target for developing anti-tumor drugs. Here, we discuss the origin, biomarkers, regulatory factors, and physiology of disease-associated microglia/macrophages. A better understanding of disease-associated microglia/macrophages and tumor microenvironment may facilitate the development of effective treatment strategies for neurodegenerative diseases and brain cancers.

Committee: Qing Lu Ph.D. (Committee Member); Ty Troutman Ph.D. (Committee Member); Ziyuan Guo Ph.D. (Committee Member); Theresa Alenghat V.M.D. Ph.D. (Committee Member) Subjects: Cellular Biology

Doctor of Philosophy, The Ohio State University, 2022, Neuroscience Graduate Studies Program

The glucocorticoid receptor (GR), part of the nuclear receptor superfamily of transcription factors, is ubiquitously expressed in all cell types and regulates cellular responses to glucocorticoids (GCs). Stress, a recurring and unavoidable experience throughout life, causes the release of GCs into the bloodstream which exert actions through binding GR. While GRs mainly function to maintain homeostasis and regulate metabolism, they also act in specific cell types to diversely modulate their function. GCs induce structural plasticity in neurons and many other cells throughout the nervous system. While plasticity is essential for adaptation and learning and memory stress-induced plasticity can be maladaptive. Elevated GCs can alter neuron structure and function in the brain leading to cognitive impairment and contribute to the development of neuropsychiatric disorders. GCs also exacerbate neuropathic pain, but less is known about GR mediated structural plasticity in sensory neurons and in the context of pain. Here, we show that GR mediates growth and regeneration in sensory neurons. We found that basal GR expression in dorsal root ganglia (DRG) sensory neurons is 15-fold higher than in neurons in canonical stress-responsive brain regions, making them uniquely sensitive to GCs. In response to stress or applied GCs, adult DRG neurite growth in vitro increases through mechanisms involving GR-dependent gene transcription. We also identify a new set of regeneration associated genes (RAGs) (e.g., Gilz, Cebpa) that are increased by stress. In vivo, acute stress increases peripheral nerve regeneration, providing evidence for a structural correlate of stress-exacerbated pain. Building on these findings, we further investigate sensory neuron GRs in the context of a newly developed clinically relevant model of neuroma. We characterize this new model and find that sural nerve ligation induces allodynia in both male and female mice and develop neuromas with features consi (open full item for complete abstract)

Committee: Phillip Popovich (Advisor) Subjects: Neurosciences

Doctor of Philosophy, The Ohio State University, 2022, Biomedical Sciences

The projects described within address a knowledge gap regarding specific epigenetic regulatory mechanisms underlying the dysfunction of microglia in Alzheimer's disease. AD is the 6th leading cause of death in the US. Of the top 10 causes of death, AD is the only one which currently cannot be prevented or cured. AD brain pathology is characterized by build-up of amyloid-β (Aβ) plaques and neurofibrillary tangles of tau protein. Previous research has established that neuroinflammation also contributes to the synaptic loss, neuronal death, and symptomatic decline of AD patients. Accumulating evidence suggests a critical role for microglia, innate immune phagocytes of the central nervous system, in AD pathogenesis. For instance, microglia are unable to effectively phagocytose and degrade Aβ aggregates, and they instead respond by releasing pro-inflammatory cytokines which are implicated in AD neurodegeneration. How microglia initiate these inflammatory responses without simultaneous clearance of Aβ is unknown. Autophagy is a normal, degradative process that discards non-functional cellular organelles, internalized microbes, protein aggregates, and regulates release of inflammatory proteins. Extensive research in AD established that diseased neurons exhibit dysfunctional autophagy, which contributes to neuronal death. However, autophagy function in other brain cells in AD, such as microglia, remains overlooked. This work uncovered the first-described mechanism underlying dysfunctional autophagy in AD microglia. The MicroRNA (miR) Mirc1/Mir17-92 cluster, known to diminish autophagy effectors, is elevated in AD microglia from human patient samples. Microglia from a mouse model of AD fail to clear Aβ due to the elevation of the Mirc1/Mir17-92 cluster. However, autophagy function and clearance of Aβ can be recovered by inhibiting miR-17 in diseased mouse microglia. Moreover, this work established a nanoparticle-based therapeutic (open full item for complete abstract)

Committee: Amal Amer (Advisor); Amy Lovett-Racke (Committee Chair); Douglas Scharre (Committee Member); Stephanie Seveau (Committee Member); Kathryn Lenz (Committee Member); Ruth Barrientos (Committee Member) Subjects: Immunology; Neurosciences

Doctor of Philosophy, The Ohio State University, 2021, Oral Biology

Inflammation within the central nervous system (CNS) is called “neuroinflammation” and, depending on the context, i.e. magnitude, duration, stimulus type, can cause beneficial or detrimental consequences. The innate immune cells within the CNS that influence the properties of neuroinflammation are the microglia; making microglia targets to understand how CNS immune responses relate to damage or reparative processes. A key molecule that microglia produce is Interleukin-1 (IL-1)—a master cytokine that can initiate all aspects of the inflammatory processes. IL-1 is known to induce both adverse and beneficial effects on CNS processes; however, how these diametrically opposite results arise from a single immune molecule are unknown. To understand how IL-1 signaling can result in protective and destructive outcomes, a double knock-in mouse model which restricts the expression of the cognate receptor for IL-1, Interleukin-1 Receptor Type 1 (IL-1R1), to a specific cell type was used. In this dissertation, multiple models of lasting neuroinflammation were used to dissect how cell type-specific IL-1/IL-1R1 signaling can cause neuroprotection or neuropathologies. Chapters 1 and 2 show that a model of systemic inflammation induced-neuroinflammation initiates a neuroprotective environment within the CNS via the endothelial and microglial IL-1R1. In Chapter 3, chronically expressed IL-1 can induce anti-neurogenic responses that are mediated via endothelial and myeloid IL-1R1, but not the adjacent neuronal IL-1R1. Finally, Chapter 4 investigates how neuronal IL-1R1 influences seizure severity and neuronal activation patterns following induction of epilepsy. These findings highlight the biological mechanisms through which cell type-specific IL-1R1 causes a variety of neuroinflammatory responses.

Committee: John Sheridan (Advisor); Ning Quan (Advisor) Subjects: Immunology; Neurobiology; Neurosciences

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

Neuroinflammation is implicated in the progression and pathology of Alzheimer's disease (AD). While AD presents differently in individual patients, advancing age and presence of the strongest known genetic risk factor, APOE4 (E4) genotype, have been shown to contribute greatly to the increased risk of AD. In addition, females have an increased risk of developing AD at a younger age which is modified by the APOE genotype. APOE protein is predominantly expressed in astrocytes and microglia; E4 glia from both humans and mice have been consistently shown to have a more reactive phenotype compared to APOE3 (E3). Primary microglia (PMG) and astrocytes (PMA) from APOE4 humanized mice have consistently been shown have a higher response to inflammatory stimuli than APOE3 PMG and PMA. However, all studies to date have been performed in mixed sex cultures. Here, we sought to assess the response of sex-specific E3 and E4 PMG and PMA in comparison to traditional mixed sex cultures. Analysis of inflammatory gene expression and secretion of cytokines revealed E4 genotype and female sex, together contribute to a greater inflammatory response in both PMG and PMA. Next, we assessed the contribution of age, sex and APOE genotype on LPS-induced neuroinflammation in humanized targeted replacement E3 and E4 mice. Gene expression and quantification of cytokines in the hippocampus and frontal cortex indicated that a peripheral LPS challenge induces a higher increase in proinflammatory cytokine mRNA expression in aged E4 mice compared to E3 and this effect appears to be sex and region-specific. Taken together, these data demonstrate that multiple factors contribute to susceptibility to neuroinflammation and provide data demonstrating significant roles for age, sex, brain region and APOE genotype in the response to peripheral inflammation.

Committee: Jason Richardson Dr. (Advisor); Ernest Freeman Dr. (Committee Member); John Johnson Dr. (Committee Member); Kim Tieu Dr. (Committee Member); Priya Raman Dr. (Committee Chair) Subjects: 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

Biological aging is a stressor causing several forms of dysfunction at the cellular, tissue, and organismal levels. As inflammatory signaling in the central nervous system is critical for normal physiological and behavioral responses to infection, the dysfunctional neuro-immune communication resulting from normal aging leads to heightened risk of mortality and co-morbidity of depression or dementia after peripheral infection. Moreover, elderly individuals suffering from an infection often experience mood disorders and cognitive dysfunction. Indeed, rodent studies have shown this altered neuroinflammatory state is primarily facilitated by dysfunctional signaling from microglia and astrocytes in the brain. Microglia, the innate immune cells of the central nervous system, develop a pro-inflammatory, “primed” profile with age. Once activated by an inflammatory stimulus, these primed cells produce an exaggerated and prolonged neuroinflammatory response mediated by cytokines and chemokines. To test the hypothesis that this primed phenotype is the result of the accumulation of cellular damage through aging, I pharmacologically forced the turnover of microglia in the brain with a colony-stimulating factor 1 receptor (CSF1R) antagonist. While this approach showed microglia in the aged brain are capable of rapidly and efficiently repopulating after cessation of CSF1R antagonism, indicating these cells are senescent in the classical sense, the repopulated cells still exhibited a pro-inflammatory response to peripheral immune challenge. Moreover, soluble signals from aged brain tissue conferred a primed phenotype to neonatal microglia ex vivo, potentiating their transcriptional response to lipopolysaccharide. Taken together, these data show age-associated priming likely results from microglia-extrinsic signals within the aged brain. Interestingly, forced turnover of microglia exerted beneficial effects on neurons within the aged brain. While microglia showed no effect of forc (open full item for complete abstract)

Committee: Jonathan Godbout (Advisor); Ruth Barrientos (Committee Member); Phillip Popovich (Committee Member); Ning Quan (Committee Member) Subjects: Immunology; Neurosciences

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

Alzheimer's disease (AD) is a chronic neurodegeneration characterized by cognitive deficits and memory loss. It is currently the 6th leading cause of death and is the only one of the top ten that cannot be prevented, slowed, or cured. Though defects in cognition and memory are hallmarks of AD, non-cognitive symptoms occur early in the disease process. Some of the most common symptoms early in AD are visual deficits, which can occur years before noticeable cognitive decline. Accumulations of amyloid-beta (Aβ), one of the neurotoxic proteins associated with AD, are detectable in the retina before they are present in higher order central nervous system structures. However, little is known of the relationship between retinal Aβ and visual dysfunction. The experiments of this dissertation examined the impact of Aβ on retinal ganglion cell (RGC) structure and function and investigated a potential mechanism underlying amyloid-mediated degeneration in the anterior visual system. Using a model of induced retinal amyloidosis, I found that exogenous Aβ induces regional RGC death in the exposed retina. Intriguingly, this effect spreads to other areas of the central nervous system, most notably the opposite, un-injected eye. These findings are reported in Chapter 2. Chapter 3 reports the effect this degeneration has on overall neurophysiological signaling. Here, I found that these intraocular injections of Aβ selectively affects RGC function, sparing upstream neurons in the retina. Finally, I began investigating mechanisms of Aβ-mediated degeneration by performing the same experiments in transgenic mice lacking a major scavenger receptor responsible for glial cell interaction with amyloid plaques, cluster of differentiation 36 (CD36). I found that mice lacking this receptor were resistant to amyloid-induced degeneration. Finally, in chapter 4, I summarize my findings, discuss where they fit within our current understanding of AD, and discuss some future directions of this work (open full item for complete abstract)

Committee: Samuel Crish PhD (Advisor); Christine Dengler-Crish PhD (Committee Member); Denise Inman PhD (Committee Member); Jordan Renna PhD (Committee Member); Michael Lehman PhD (Committee Member) Subjects: Neurosciences