Medical Neuroscience Department Theses and Dissertations

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    The Role of the Lung-Brain Axis in the Ozone-Impaired Amyloid Associated Astrocytic and Vascular Phenotype
    (2024-06) Ahmed, Chandrama; Oblak, Adrian; Block, Michelle; Baucum, A. J.; Bissel, Stephanie J.; Nass, Richard M.
    Air pollution has been associated with an increased risk of Alzheimer’s Disease (AD). Studies show ozone (O3), a major component of urban air pollution, can exacerbate amyloid pathology. However, O3 reacts in its entirety with lung epithelial lining after inhalation, hence does not translocate to brain. Studies have implicated the lung−brain axis in O3 induced central nervous system (CNS) pathology. However, the mechanistic underpinnings of its role in amyloid pathology is obscure. Here, we explored the impact of O3 on the astrocytic and vascular response to amyloid plaque in 5xFAD mice and its link to the O3 lung response. O3 exposure increased GFAP positive astrocyte density correlating with increased plaque burden in the cortex. Focusing on the plaque microenvironment, we found O3 qualitatively altered plaque associated astrocytes, evidenced by both proteomic and transcriptomic changes. Along with loss of protein expression, proteomic changes reflected increased cell-cell interaction in plaque microenvironment. Specifically, we found increased astrocyte-microglia contact selectively in periplaque space from O3 exposure. Transcriptional analysis of periplaque astrocytes revealed an accelerated shift towards disease associated astrocyte (DAA) phenotype. Elevated circulating HMGB1 was previously found from O3 exposure. In this study we demonstrate deleting HMGB1 selectively in peripheral myeloid cells and not in CNS microglia ameliorates the lung immune response to O3 as well as downregulates DAA marker in the CNS, providing a potential link between peripheral HMGB1 and O3 induced astrocytic dysregulation. On examining vascular response to O3 we found increased vascular amyloid accumulation associated with an altered vascular proteomic profile. Our analysis indicates O3 potentially disrupts vascular function such as amyloid clearance. Taken together, our study demonstrates that astrocyte and neurovasculature are contributors to O3 lung-brain axis with important implications towards amyloid pathology progression and identifies peripheral myeloid HMGB1 as its potential modulator. Further studies are required to fully understand the consequences of this impact and its role in amyloid pathology.
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    Modulation of Intralocular Pressure by Tuning Transcriptional Control of Lipid Synthesis
    (2024-06) Wang, Ting; Morral, Nuria; Pattabiraman, Padmanabhan; Corson, Timothy W.; Landreth, Gary E.; Perrin, Benjamin J.
    Glaucoma is an age-related optic neuropathy and is one of the leading causes of irreversible blindness. Primary open-angle glaucoma (POAG) is the predominant subtype of glaucoma. Elevated intraocular pressure (IOP) is a major risk factor for POAG and lowering IOP is the most effective therapeutic strategy. IOP is maintained by the balance of aqueous humor (AH) generation by the ciliary body and drainage by conventional outflow pathway including trabecular meshwork (TM). TM is a highly contractile and mechanosensitive tissue, and its contractility regulated by the actin cytoskeleton and extracellular matrix (ECM) is directly related to IOP regulation. Using multiomics analysis in human TM (HTM) cells, I identified that mechanical stretch caused the activation of sterol regulatory element binding proteins (SREBPs) related-lipid biogenesis pathways. Further, using immunofluorescence, and constitutive activation of each SREBP isoform, I discovered the mechanosensing role of SREBPs in HTM cells and mechanistically deciphered the attributes of SREBPs in regulating the contractile properties of TM. The pharmacological inhibition of SREBPs by fatostatin and molecular inactivation of SREBPs ex vivo and in vivo resulted in significant IOP lowering. Conversely, significantly elevated IOP was observed after using the pharmacological activator of SREBPs by clozapine and constitutive activation of SREBPs ex vivo and in vivo, respectively. As a proof of concept, fatostatin significantly decreased the SREBPs responsive genes and enzymes involved in lipogenic pathways and phospholipids, cholesterol, and triglyceride levels. The increased lipid biogenesis was found after constitutive activation of SREBP isoforms in HTM cells but with slightly different effects between each isoform. Further, I showed that fatostatin mitigated actin polymerization machinery and stabilization, and identified that SREBPs activation is a critical regulator of ECM engagement to the matrix sites. Lastly, I identified that cholesterol levels play an important role in regulating actin polymerization, focal adhesion formation, cell-ECM interactions, and membrane tension in HTM cells. Therefore, we have established the direct connection between cholesterol and TM contractility. Overall, I postulate that lowering de novo lipogenesis in the TM outflow pathway can hold the key to lowering IOP by modifying the TM biomechanics.
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    Contributions of the Presynaptic Protein Bassoon to Tau Pathogenesis and Neurodegeneration
    (2024-05) Patel, Henika Sanjaybhai; Oblak, Adrian; Lasagna-Reeves, Cristian; McKinzie, David; Kim, Jungsu; Webb, Ian; Murray, Melissa
    Neurodegenerative tauopathies, characterized by the aggregation of misfolded tau protein, pose a significant clinical and scientific challenge. A high-molecular-weight (HMW) tau species is known to be involved in spreading tau pathology. However, the nature and composition of this species remain elusive, hindering targeted interventions. There are four main chapters in this dissertation. The first chapter highlights the existing knowledge about tau and its role in neurodegenerative tauopathies and discusses the possible contribution of protein interactors in the pathogenesis of tau pathology. The second chapter investigates the association between pathological hallmarks and functional deficits in the aged PS19 tauopathy model. The findings indicate that a diverse spectrum of pathological tau species may underly different symptoms and that neuroinflammation might contribute to functional deficits independent of tau pathology. In the third chapter, we isolated and characterized the HMW tau species with seeding capabilities from the PS19 brains. Using unbiased quantitative mass spectrometry analysis, we identified Bassoon (BSN), a presynaptic protein, as a crucial interactor of the HMW tau seed. BSN overexpression exacerbated tau-seeding and toxicity both in vitro and in the Drosophila model of tauopathy. Conversely, the downregulation of BSN reduced tau spreading and overall disease pathology in the PS19 mice, indicating the important role of BSN in taumediated pathogenesis. In chapter four, we studied the disease-associated p.Pro3866Ala missense mutation in BSN and further evaluated the mechanisms through which BSN could induce toxicity and neurodegeneration. Using CRISPR-Cas9 technology, we developed a knock-in mouse model harboring the BSN P3866A missense mutation in the endogenous murine Bsn. We observed somatic BSN accumulation suggesting that the P3866A mutation might be enhancing the aggregation propensity of BSN and provide a conducive environment to promote tau aggregation. Furthermore, we observed dysregulation in protein degradation pathways, neuroinflammation, and enhanced synapse elimination by microglia. These findings underscore the pivotal role of BSN in providing a favorable environment for tau aggregation and influencing the properties of the tau seed, thereby contributing to neurodegenerative processes. Overall, our results indicate that targeting BSN could be a potential therapeutic intervention for neurodegenerative diseases.
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    Brain Responses to Sugar: Implications for Alcohol Use Disorder and Obesity
    (2024-05) Alessi, Jonathan P.; Yoder, Karmen K.; Kareken, David A.; Džemidžić, Mario; Considine, Robert V.; Harezlak, Jaroslaw
    Obesity and alcohol use may together account for 640,000 adult deaths each year in the United States. In both cases, overconsumption drives untoward effects. Alcohol use and obesity also both relate to sweet liking, as sugar consumption is consistently linked to weight gain and intense sweet liking has been linked to an inherited risk for alcohol use disorder (AUD). However, the neural underpinnings of these associations are largely unknown. Thus, we used sugar-sweetened water administration during functional magnetic resonance imaging (fMRI) to probe these relationships in two studies. In the first, we tested the relationship between a known AUD risk factor, subjective response to alcohol, and the brain response to both sucrose and monetary reward in 140 young adults. We found a significant positive correlation between the enjoyable component of subjective responses to a standardized intravenous alcohol exposure and activation to high-concentration sucrose (but not monetary reward) in the right dorsal anterior insula and the supplementary motor area, supporting a role for these regions in AUD risk. In the second study, we investigated the neural mechanisms of sweet liking decreases following bariatric surgery, the most effective obesity treatment. Here, we evaluated the change in brain activation to sucrose in 24 women before (BMI 47.0 + 6.9 kg/m2) and 21 women after (BMI 37.6 + 6.5 kg/m2) bariatric surgery and compared the pre- and post-surgical activation patterns to those of 21 normal to overweight (BMI 23.5 + 2.5 kg/m2) control participants. Brain activation did not differ between controls and surgery participants at either time point. However, activation to sucrose in reward, but not sensory, regions decreased significantly after surgery, consistent with reduced drive to consume sweet foods. Together, these studies highlight the utility of quantifying brain responses to sweet taste as a method to understand the mechanisms underlying overconsumptive behavior.
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    The Role of ABI3 in Obesity and Metabolic Regulation
    (2024-04) Smith, Daniel Curtis; Oblak, Adrian; Kim, Jungsu; Flak, Jonathan; Lasagna-Reeves, Cristian; Evans-Molina, Carmella
    Abelson Interactor Protein 3 is an adaptor protein involved in cytoskeletal remodeling. ABI3 is predominantly expressed within mononuclear phagocytotic immune cells within the brain, such as macrophages, peripherally, and microglia. Until recently, little was known about the function of the ABI3 protein, and even less was known regarding its role in disease. Following the identification of a rare mutation within ABI3 that increases the risk of developing Alzheimer’s disease, our laboratory began to investigate the impact of deleting Abi3 in mouse models. While we initially set out to investigate ABI3 in the context of neurodegeneration, we unexpectedly discovered that loss of Abi3 led to obesity in mice. This discovery and the subsequent efforts to uncover the mechanisms by which loss of Abi3 induces obesity are the subject of this dissertation. First, we demonstrate that deletion of Abi3 leads to severe obesity in aged mice. We identified significant Abi3-dependent transcriptomic changes within the hypothalamus, but not adipose tissue, of these mice. These changes occurred within pathways related to immune function, and subsequent immunostaining revealed decreased microglia number and area within the mediobasal hypothalamus of Abi3-/- mice. Next, we performed a longitudinal high-fat diet study to explore the impact of loss of Abi3 on mouse body weight and metabolic regulation during chronic nutrient excess and control conditions. Intriguingly, we found that only female Abi3-/- mice exhibited increased body weight during high-fat diet feeding. Subsequent transcriptomics from the hypothalamus of female Abi3+/+ and Abi3-/- mice from both high-fat and control diet groups revealed cytoskeletal-related changes only in the obese, high-fat diet-fed female Abi3-/- mice. Follow-up immunostaining revealed decreased microglia coverage within the mediobasal hypothalamus of the obese, high-fat diet-fed female Abi3-/- mice. While much remains to be explored regarding the precise role of ABI3 in the setting of energy balance regulation and obesity, our investigations revealed that loss of ABI3 is sufficient to induce obesity and appears to occur through altered microglia function within the hypothalamus. This dissertation represents a critical first step in the investigation of a novel regulator of obesity pathology.
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    Follow Your Heart: Heart Rate Variability Reveals Sex Differences in Autonomic Regulation During Anxiety-Like Behavior and During Alcohol Drinking in Rats
    (2024-02) Frasier, Raizel Michele; Yoder, Karmen; Hopf, Woody; McKinzie, David; Lukkes, Jodi; Conroy, Susan
    Mental health conditions remain a substantial and costly challenge to society. Of note, women have a higher prevalence of anxiety disorders than men, and alcohol misuse in women has risen sharply in recent years. However, critical mechanisms underlying these observed sex differences remain incompletely understood. Measures of cardiac function, including heart rate (HR) and HR variability (HRV), reflect dynamic balance between the two opposing branches of the autonomic nervous system: sympathetic (SNS, “fight or flight”) and parasympathetic (PNS, “rest and digest”). Furthermore, recent evidence strongly suggests these measures are potential biomarkers for pathological states, including mental health conditions. To better understand sex differences in autonomic mechanisms related to pathological anxiety and alcohol misuse, we utilized cardiac telemetry to measure HR and HRV. This allowed observation of real-time autonomic tone in awake, freely behaving Wistar rats of both sexes. At baseline, female rats had greater HR and lower SNS influence than males, which concords with human studies. In both anxiety-like behavior and alcohol drinking studies, we observed that females tend to utilize a higher PNS influence to overcome challenge, whereas males tend to utilize higher SNS. Furthermore, female (but not male) baseline HR and HRV are related to within-task behavior, suggesting that baseline state impacts drinking and anxiety-like behavior in a sex-specific way. Again, these data concord with human research suggesting a similar PNS bias in women and SNS bias in men when responding, especially under challenge. Taken together, these data have contributed new knowledge to sex differences in autonomic engagement, especially for anxiety states and alcohol drinking. Importantly, these findings are likely translationally relevant for the development of novel, and more personalized, therapies.
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    The Influence of Genetic Diversity on Tauopathy
    (2024-02) Acri, Dominic James; Oblak, Adrian L.; Kim, Jungsu; Landreth, Gary E.; Lasagna-Reeves, Cristian A.; Liu, Yunlong
    The presence of misfolded tau proteins is a hallmark of a group of neurodegenerative diseases termed tauopathies. This heterogeneous group of disorders includes Alzheimer’s disease, frontotemporal dementia, chronic traumatic encephalopathy, and dozens of related dementias. One common approach to research these diseases is to use clonally inbred animal strains to investigate druggable pathways that can be translated to human patients. Although clonally inbred lines offer a standard to limit inter-laboratory variability, selecting a single genetic background does not allow researchers to consider the natural population-level genetic diversity seen in humans. Recent studies have demonstrated that introducing genetic diversity into animal models of complex diseases improves translatability to patient populations. Within the context of neurodegeneration, several studies have found that brain pathology in mouse models of amyloid-β accumulation is modified by genetic variance within a subset of wild-derived mouse strains. Several findings have demonstrated that non-C57BL/6J mouse strains alter tau pathology, however, the effect of these wild-derived strains on tau is unknown. To understand how genetically diverse animal models respond to the presence of pathological tau, we generated cohorts of wild-derived mice and fruit flies that express humanized mutant tau. We found that the formation of proteopathic prion-like tau seeds was dependent on the genetic backgrounds of these wild-derived animal models. Interestingly, we showed that multi-omic analysis of PWK/PhJ reveals immune dysregulation not seen in classically inbred models of tau pathology. Furthermore, genetic mapping in a novel panel of wild-derived fruit flies revealed novel genetic modifiers of tau seeds. We demonstrate how tauopathy is influenced by genetic diversity and how these models may be used as a resource to study tau-driven neurodegenerative diseases. Taken together, the data suggest that introducing genetic diversity into animal models of mutant tau expression may increase translation to patients suffering from tauopathy.
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    Increased Excitability of Pyramidal Neurons in the Secondary Motor Cortex Enhances Cocaine-Seeking
    (2023-09) Huang, Donald; Atwood, Brady K.; Baucum, AJ; Hopf, Woody; Ma, Yao-Ying; Sheets, Patrick L.
    Cocaine addiction is a brain disorder characterized by chronic relapse. Although drug-seeking behaviors have been recognized to be associated with relapse, the role of the motor cortex, including the primary (M1) and secondary (M2) motor cortex, which are functionally important mediators of complex behaviors remains unclear in addiction. Here we use a rat cocaine intravenous self-administration (IVSA) model to investigate the intrinsic excitability of pyramidal neurons in the medial prefrontal cortices and motor cortices during withdrawal. Cocaine IVSA-trained rats performed a cocaine-seeking test on withdrawal day (WD) 1 or WD 45. Relative to WD 1 an increase in cocaineseeking was detected on WD 45. Whole-cell patch clamp recordings revealed an increase in intrinsic excitability in pyramidal neurons in Layer 2 of the secondary motor cortex (M2-L2) in cocaine trained rats on WD 45. Using a pharmacological approach, bath application of GABAA receptor agonist, muscimol, dosedependently (0.1 mM, 0.3 mM, and 1.0 mM) decreased the excitability of M2-L2 pyramidal neurons in cocaine IVSA-trained rats on WD 45. Pharmacological inactivation of M2-L2 by bilateral intra-M2 injection of muscimol (324 ng/1.0 μl) attenuated cocaine-seeking on WD 45. A chemogenetic approach was used to validate that M2-L2 pyramidal neurons play a contributing role in the increase in cocaine-seeking, a microinjection of rAAV5-CaMKIIa-hM4di-mCherry was performed to express Gi-DREADD receptors on M2-L2 pyramidal neurons. Activating Gi-DREADD with an intraperitoneal injection of compound 21 on WD 45 attenuated cocaine-seeking. To elucidate the mechanism that contributes to the increased excitability of M2-L2 pyramidal neurons, an analysis of the action potential kinetics revealed that calcium-activated small conductance potassium (SK) channel-mediated medium afterhyperpolarization amplitude decreased in cocaine vs. saline IVSA-trained rats on WD 45. SK channel activation by 1-EBIO (300 μm) increased the medium afterhyperpolarization amplitude and decreased the excitability of M2-L2 pyramidal neurons in cocaine IVSA-trained rats. Furthermore, intra-M2 injection of 1-EBIO on WD 45 attenuated cocaine-seeking. These experiments suggest that cocaine IVSA-training-induced persistent changes in M2-L2 pyramidal neurons' intrinsic excitability contributes to enhanced cocaine-seeking. Our results provide evidence targeting the SK channels in the superficial layer for M2 could be an important therapeutic approach for preventing cocaine relapse.
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    Propranolol Elicits Long Term Systemic Effects After Repetitive Mild Traumatic Brain Injury
    (2023-07) Smith, Jared Andre; Obukhov, Alexander; White, Fletcher; Hato, Takashi; Naugle, Kelly; Jin, Xiaoming; Truitt, William; Grimes, Jaison
    There are almost 2 million new traumatic brain injuries (TBIs) every year in the US. Of these, 80% of these can be classified as mild TBI, also known as concussions, that can lead to pronounced long term symptoms months and years after injury. The presence of post traumatic headaches (PTH) is the most common chronic side effect with prevalence of 47-95% of mTBI patients within a week of injury. Though the mechanisms after TBI leading to these headaches and other post mTBI side effects are poorly understood, recent studies have suggested the role of the immune system after injury plays a causal role in this process. Peripheral immune cells can travel to the brain after mTBI as a result of blood brain barrier dysfunction, sympathetic nervous signaling, and the release of inflammatory mediators. Recent studies have shown sympathetic activation after injury can result in IL-10 dependent systemic immunosuppressive state after mTBI. In this study we sought to limit sympathetic dependent immune alterations after injury by injecting the beta blocker propranolol directly after injury and investigating the immune changes in the blood, brain, spleen, and bone marrow of mTBI animals. Together, these data show mTBI causes immune genetic and pathway level changes at least one month after injury and that propranolol alters genes important for metabolism, cytokine signaling, epigenetic modification, innate, and adaptive immunity. We also find that propranolol reduces the presence of Ly6C+ and increases the presence of Ly6C- monocytes in the blood one month after injury; however, it leads to increased Ly6C+ monocyte presence in the spleen of mTBI mice. In conclusion, propranolol administration directly after mTBI leads to immune changes that may lead to long-term improvement in post TBI symptomology.
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    White Matter Microstructural Changes in the Cingulum Bundle of the Hippocampus in Alzheimer's Disease
    (2023-07) Hirschfeld, Lauren Rose; Risacher, Shannon L.; Saykin, Andrew J.; Block, Michelle L.; Wu, Yu-Chien; Nho, Kwangsik; Cao, Sha
    Alzheimer’s disease (AD), the most common dementia, is the seventh leading cause of death for adults in the US. Recent FDA-approved therapeutics targeting hallmark amyloid pathology may slow but do not yet halt the disease. Thus, it is likely that other factors, such as neurodegeneration, play a role in disease development. Compromised white matter (WM) microstructural integrity may be a promising biomarker for disease progression, as it has been demonstrated in both preclinical and clinical AD. WM microstructural changes can be estimated using diffusion tensor imaging (DTI), a noninvasive imaging modality which measures the diffusion of water molecules within brain tissue. First, literature describing a biological relationship between signaling pathways involved in both insufficient myelin-repair mechanisms and AD pathology was assessed to establish a foundation for studying WM in the context of AD. Then, DTI was used to assess possible WM microstructural changes of the cingulum bundle of the hippocampus (CBH), a disease-relevant region, in the KBASE cohort of older Korean adults on the AD continuum. Similar to White cohorts of European ancestry, DTI values corresponding to worsening WM microstructure occurred in a stepwise fashion across diagnostic stages and were associated with both cognition and amyloid deposition. In a sample of older adults from the Indiana Memory and Aging Study (IMAS), CBH microstructure was a sensitive biomarker of preclinical conversion from cognitively unimpaired status to mild cognitive impairment. Finally, a novel extension of the T1/T2 weighted ratio imaging method, thought to estimate myelin- related tissue integrity, yielded values in IMAS subjects reflective of previously observed DTI patterns. The complex interplay between amyloid, tau, and neurodegeneration in AD is not yet fully characterized. This research shows decreased integrity of WM microstructure in the CBH, which may be useful as a biomarker of early disease conversion to MCI.