Master of Science, Miami University, 2025, Microbiology

Chlamydomonas reinhardtii is a model alga used in the study of photosynthesis. One of the attractive attributes of C. reinhardtii is its ability to grow autotrophically, heterotrophically, or mixotrophically depending on the availability of light and acetate. Due to the advantages of mixotrophic growth, many studies of the photosynthetic stress response are conducted in acetate supplemented medium, while studies conducted under autotrophic growth conditions are limited. There is evidence that even under non- stressed conditions, mixotrophically grown cultures exhibit distinct photophysiology. To describe differences in the response to abiotic stress, C. reinhardtii growth and physiology during control and long-term stress (high light, high salt, combined high light/high salt) were compared under autotrophic versus mixotrophic conditions. Photochemistry, ATP:ADP ratios, and abundance of photosynthetic proteins were measured. Higher growth rates in mixotrophic vs. autotrophic-grown cultures were accompanied by retention of photosystem II activity and higher rates of photosystem I- driven cyclic electron flow. Under long-term stress, mixotrophic cultures exhibited reduced capacity for non-photochemical quenching and increased abundance of photosystem II. We conclude that mixotrophic growth conditions significantly impact the physiology and tolerance of C. reinhardtii to stress. Trophic mode should be considered when using C. reinhardtii to study the photosynthetic stress response.

Doctor of Philosophy in Regulatory Biology, Cleveland State University, 2024, College of Arts and Sciences

Eukaryotic initiation factor 2A (eIF2A) is a highly conserved 65 kDa eukaryotic protein that functions in minor initiation pathways, which affect the translation of only a subset of messenger ribonucleic acid (mRNAs), such as internal ribosome entry site (IRES)-containing mRNAs and/or mRNAs harboring upstream near cognate/non-AUG start codons. These non-canonical initiation events are important for regulation of protein synthesis during cellular development, differentiation and/or the integrated stress response. Selective eIF2A knockdown in cellular systems was shown to inhibit translation of such mRNAs, which rely on alternative initiation mechanisms for their translation. However, there existed a gap in our understanding of how eIF2A functions in mammalian systems in vivo (on the organismal level) and ex vivo (in cells). To address this question we have developed the eIF2A-total body knockout (KO) mouse model. Using this model, we presented evidence implicating eIF2A in the biology of aging and metabolic syndrome. We discovered that eIF2A-KO mice have reduced life span and that eIF2A plays an important role in maintenance of lipid homeostasis, the control of glucose tolerance and insulin resistance. We also showed the eIF2A KO affects male and female mice differently, suggesting that eIF2A may affect sex-specific pathways. The metabolic syndrome phenotype has three main etiological categories: obesity and disorders of adipose tissue (particularly increased size of adipocytes); glucose intolerance and insulin resistance; and a constellation of independent factors of hepatic, vascular, and immunologic origin and all of these features were observed in the eIF2A-KO mice. Increased adipocyte size is, in particular, well known to be positively correlated with impaired insulin sensitivity and glucose tolerance leading to metabolic syndrome. To specifically check whether the absence of eIF2A in adipose tissue is responsible for metabolic abnormalities observed in the tota (open full item for complete abstract)

Master of Science (MS), Wright State University, 2024, Microbiology and Immunology

Calcium signaling is a crucial regulator in many cellular processes, from immune response modulation to gene expression and apoptosis. Here, I have investigated the dynamics of calcium entry pathways, namely, store-operated calcium entry and store-independent calcium entry using a variety of cell types such as Jurkat T cells, HEK-293 cells and murine macrophages. Using fluorescence calcium imaging and selective inhibitor compounds, I have investigated the contributions of Orai and related calcium channels to calcium entry in both immune and non-immune cell types. The results demonstrate the pharmacological modulation of store-operated and store-independent calcium entry, showing the ability of compounds such as AE-19, AE-10, and EL-113 to inhibit calcium entry. Experiments involving membrane depolarization with elevated potassium concentrations confirmed that calcium entry through the plasma membrane is membrane potential-dependent. My results highlight the medical relevance of these channels, especially in the process of tubular aggregated myopathy, immune regulation and cancer. My studies may lead to therapeutics targeting calcium signaling pathways relevant to immune and muscular disorders.

Doctor of Nursing Practice , Case Western Reserve University, 0, School of Nursing

Background: Pulmonary embolism (PE) is a common cause of morbidity and mortality and continues to increase in overall incidence. Percutaneous Thromboembolectomy is a less invasive alternative therapeutic strategy that can be used for the treatment of acute pulmonary embolism (PE). This therapy is usually performed in patients with acute high-risk (massive) PE for whom thrombolysis is contraindicated or has failed and for whom surgical intervention is not available or is contraindicated. These patients are at extremely high risk for cardiopulmonary complications and are very challenging to care for from an anesthetic standpoint. Because it is a growing mode of therapy, CRNAs should be educated on how to appropriately care for these patients. Purpose: The purpose of this DNP project was to develop, implement, and evaluate an evidence-based educational module for anesthetic management of minimally invasive catheter-based removal of massive pulmonary embolism (Minimally-Invasive Pulmonary Thromboembolectomy) for CRNAs and SRNAs who practice in the General Adult ORs at a large academic quaternary care center. Methods: This DNP scholarly project was designed as a one-time clinical educational module on the current best practice guidelines for anesthetic management for the minimally invasive removal of massive PE for CRNAs and SRNAs. Keywords: pulmonary embolism, minimally-invasive pulmonary embolectomy, thromboembolectomy, Certified Registered Nurse Anesthetists, Student Registered Nurse Anesthetists

Doctor of Philosophy, The Ohio State University, 2024, Molecular, Cellular and Developmental Biology

Cardiovascular disease in the leading cause of death worldwide. While the etiology of heart disease is diverse, a well-documented unifying theme in the progression of heart disease is the development of cardiac fibrosis. Cardiac fibrosis, defined as excess collagen and extracellular matrix deposition in the heart, is mediated by the activation and differentiation of resident quiescent fibroblasts into myofibroblasts. Pathologic fibrotic remodeling in the heart occurs in response to persistent stress stimuli, as well as injury. Fibrotic remodeling leads to stiffening of the ventricles, impaired relaxation, and progressive impairment in cardiac function. While current standard-of-care medications have been vital in the treatment of heart diseases over the last several decades, there are currently there are currently no effective therapies in clinical use specifically targeting fibrosis in the heart. This highlights a significant need to identify novel therapeutics that target fibroblasts and fibrotic remodeling. Our group recently identified Sertad4 (Serta Domain Containing Protein 4) as a potential regulator of fibroblast activation in the heart. However, the function of Sertad4 is unknown. In this dissertation, I sought to investigate the role of Sertad4 in pathologic cardiac remodeling through a combination of in vitro and in vivo studies. To study the role of Sertad4 in vivo we utilized a reporter mouse and a global loss of function mouse. We also utilized three models of cardiac dysfunction: 1) myocardial infarction through coronary LAD ligation; 2) chronic angiotensin II/phenylephrine infusion through osmotic minipumps and 3) chronological aging as a model of diastolic dysfunction. We observed that Sertad4 is predominantly expressed in activated fibroblasts and its expression is significantly elevated in failing human and mouse hearts. Loss of Sertad4 in mice led to preserved cardiac function post-MI and decrease in cardiac hypertrophy and fibrosis. Using is (open full item for complete abstract)

Doctor of Philosophy, Case Western Reserve University, 2024, Physiology and Biophysics

Receptor protein tyrosine phosphatases γ and ζ (RPTPγ and RPTPζ) are integral transmembrane proteins pivotal in central nervous system (CNS) development and function, implicated in various physiological processes such as tumor suppression, neurodegeneration, and extracellular [CO2]o and [HCO3–]o sensing. While RPTPγ is expressed ubiquitously, RPTPζ exhibits preferential expression in the central nervous system (CNS). In this dissertation, I explore the distinct expression patterns of RPTPγ and RPTPζ splice variants in three different sources derived from wild-type (WT) laboratory C57BL/6 mice: mixed neuron-astrocyte hippocampal (HC) cultures, P0-P2 pup hippocampi, and adult hippocampi. Notably, RPTPγ and RPTPζ exhibit co-expression in the somata and processes of nearly all HC neurons across all preparations but are virtually absent from astrocytes. RPTPζ plays a crucial role in cellular adhesion and signaling during development, akin to its homolog RPTPγ. Additionally, RPTPζ has been associated with sensing extracellular CO2/HCO3– and modulating acid-base transport in the kidney. In this dissertation, I investigate whether RPTPζ similarly regulates acid-base disturbances in the brain. I examine the effects of extracellular metabolic acidosis (MAc) or respiratory acidosis (RAc) on the intracellular pH (pHi) physiology of hippocampal neurons and their adjacent astrocytes, co-cultured from WT or RPTPζ knockout (RPTPζ–/–) mice. It has been speculated that MAc and RAc lowers pHi because these challenges inhibit acid extruders or stimulate acid loaders. I now demonstrate that RPTPζ–/– neurons exhibit a decreased baseline pHi and increased vulnerability to MAc or RAc compared to WT neurons. Specifically, RPTPζ–/– neurons display heightened acidification rates during both MAc and RAc exposures, indicating a compromised ability to defend against pHo decreases. Similarly, RPTPζ–/– astrocytes in mixed culture of neuron and astrocyte exhibit dysregulated pHi responses to MAc (open full item for complete abstract)

PHD, Kent State University, 2024, College of Arts and Sciences / School of Biomedical Sciences

Successful infant feeding requires effective milk acquisition, followed by the transport of ingested material across the oral cavity and through the pharynx, ultimately culminating in esophageal peristalsis. Several elements that underlie the neural control of swallowing are underexplored, including the neurological relationships among different aspects of swallowing (oral, pharyngeal, and esophageal). The works described here aim to improve our understanding of the sensorimotor relationships that drive infant swallowing, primarily by stimulating specific areas with capsaicin. To begin, we use an animal model of superior laryngeal nerve lesion to assess the effects of oropharyngeal capsaicin administration on feeding physiology. Next, we analyze the impacts of esophageal afferents on upstream feeding behaviors using a model of simulated gastroesophageal reflux. Finally, we explore the role of mandibular afferents in infant feeding, and determine whether capsaicin administration can recover any deficits resulting from anesthetization of these afferents. All experiments were conducted using infant pigs, a validated model for the study of infant feeding. Common methodology across specific aims includes videofluoroscopy (to assess kinematics and feeding performance) and electromyography (to assess motor outputs to muscles of interest). These experiments ultimately shed light on the extent of brainstem sensorimotor integration across feeding behaviors. Additionally, the results of these studies provide insights into the mechanisms by which specific sensory signals are integrated during feeding. These insights are critical and will ultimately facilitate the design of targeted interventions for specific feeding pathophysiologies in infants.

Doctor of Philosophy (PhD), Ohio University, 2024, Molecular and Cellular Biology (Arts and Sciences)

The increasing prevalence of type 1 and type 2 diabetes has made studying insulin secretion and production an urgent necessity. Parathyroid hormone-related protein (PTHrP) is a multifunctioning protein that is expressed in many cell types including pancreatic beta cells. It consists of an N-terminus, a mid-region which includes a nuclear localization sequence (NLS), and a C-terminus. PTHrP has been shown to increase insulin secretion and beta-cell proliferation. However, the mechanism underlying its actions is still unclear. Furthermore, other studies have shown unique roles of the NLS and C-terminus of PTHrP, but these regions of PTHrP have not been studied in beta cells. In this study, full-length murine PTHrP (-36-139) was overexpressed in isolated mouse islets and a beta-cell line Ins-GLuc. There was an increase in insulin content of islets due to PTHrP overexpression and an increase in intracellular calcium levels at higher extracellular glucose concentrations of 12-28mM. Transfecting Ins-GLuc cells with full-length PTHrP caused an increase in insulin secretion in 28mM glucose; however, this effect was reversed when cells were transfected with PTHrP lacking the NLS (-36-67…95-139) or PTHrP lacking the NLS and C-terminus (-36-67). Further studies with the PTHrP ∆/∆ mouse model which lacks the NLS and C-terminus of PTHrP revealed that the mice were both hypoglycemic and hypoinsulinemic, with smaller islets than control mice, increased intracellular calcium response to glucose and reduced pancreatic glucagon content. Subsequent RNA sequencing studies with a murine beta-cell line MIN6 revealed a downregulation of ER stress-related genes and an upregulation of genes involved in amino acid biosynthesis after transfection with full-length human PTHrP (-36-141). The data from the RNA sequencing studies suggested that PTHrP increased insulin production by enhancing protein processing in the ER and protecting the cells from ER stress. Collectively, this study showed (open full item for complete abstract)

Master of Science, Miami University, 2024, Biology

Coordination between central pattern generators (CPG) is important in proper functioning of related rhythmic behaviors such as vocalization, breathing and locomotion. Despite their importance, the cellular mechanisms of inter-network coordination and how it impacts muscles remain largely unidentified. Using the small, well-characterized networks in the stomatogastric nervous system of the crab, Cancer borealis, I identify the role of a dual-network neuron in coordinating the feeding-related pyloric (“fast”: 1 Hz: filtering food) and gastric mill (“slow”: 0.1 Hz: chewing) CPG networks, during a unique modulatory state elicited by the neuropeptide Gly1-SIFamide. The dual-network neuron, LPG, rhythmically increases and decreases the frequency of the pyloric rhythm in time with different phases of the slower gastric mill rhythm. Using these findings, I show that this complex coordination pattern is distinctly translated into electrical responses of two behaviorally different muscles, innervated by LPG. Where the LPG innervated pyloric muscle follows LPG's dual-network activity, while the gastric mill muscle prioritizes gastric mill over pyloric activity. This study provides insight into a unique coordination mechanism that is funneled through a dual-network neuron, and how the muscles innervated by this neuron respond to and participate in the overall coordination of related but distinct behaviors.

Master of Science (MS), Ohio University, 2024, Exercise Physiology-Research (Health Sciences and Professions)

The 3-minute critical power test (3MT) was developed to measure critical power (CP) in a small amount of time. While considered a valid and reliable test, there is potential to improve the test itself with a consistent regulated warm-up (WU). It was hypothesized that the three WU conditions in this investigation would not affect CP, anaerobic work capacity (W'), VO2peak and peak power output (PPO). 7 trained male cyclists performed three 3MT's with a randomized 10-minute WU prior to testing: WU 1 (low intensity), WU 2 (moderate intensity) and WU 3 (severe intensity). There were no significant differences between WU conditions in CP, W', VO2peak and PPO. CP for each WU were: WU 1 (362.5 ± 17.3 W); WU 2 (365.5 ± 10.9 W); WU 3 (346.1 ± 13.6 W). These results show that these WU intensities did not influence CP significantly, but further research in WU and its influence on the 3MT is needed.

PhD, University of Cincinnati, 2024, Medicine: Systems Biology and Physiology

This dissertation investigates the dynamic behavior of the hypothalamic-pituitary-adrenal (HPA) axis in response to stress and its implications for Major Depressive Disorder (MDD). The primary aim is to determine whether HPA axis activity can serve as an objective biomarker for MDD diagnosis by combining mechanistic and data-driven modeling approaches. Chapter 1 introduces MDD and the HPA axis, emphasizing the need for accurate and objective diagnostic tools beyond subjective patient reporting. The HPA axis, a critical regulator of the stress response, is highlighted as a potential source of biomarkers. Statistical analyses of our Trier Social Stress Test (TSST) data and background related to mathematical modeling methods are provided. Chapter 2 details the development of VeVaPy, a Python platform designed to facilitate the verification and validation (V&V) of systems biology models. VeVaPy addresses current V&V process shortfalls for HPA axis models, ensuring they meet systems biology and pharmacology community standards. The framework includes four functional modules and is publicly available on GitHub, demonstrating its utility through the V&V of five selected HPA axis models. This chapter underscores the importance of robust V&V for credible model contributions and proposes best practices for model publication and usage. Chapter 3 explores Neural Ordinary Differential Equations (NODEs) for analyzing hormone dynamics during TSST. The NODE models replicated hormone changes in healthy individuals and MDD patients without prior knowledge of the stressor. Dynamic analysis revealed that stress effects are embedded in non-autonomous vector fields derived from the NODE model. These learned vector fields were then used as inputs to Convolutional Neural Networks (CNNs) for classification. The results show the potential of combining NODEs and CNNs to classify patients based on disease state, offering a preliminary step toward clinical applications using HPA axi (open full item for complete abstract)

PHD, Kent State University, 2024, College of Arts and Sciences / School of Biomedical Sciences

In nonhuman primates, the adrenal gland serves an important function in the stress response and as an additional source of steroid hormones: estrogens and testosterone. Therefore, the measurement of these adrenal steroids can provide valuable information regarding the relationship between stress and reproductive fitness. This research documented the hormonal maturation of the adrenal gland in small apes and explored how environmental, reproductive, and social changes influence adrenal hormone secretions in primates. The first objective was to determine the presence and pattern of adrenarche in the small apes. This objective collected cross-sectional fecal samples from 64 (35F, 29M) zoo-housed small apes and longitudinal fecal samples from 7 female zoo-housed small apes and measured dehydroepiandrosterone-sulfate (DHEAS) by enzyme immunoassay to determine the pattern of hormone secretion characteristic of the activation of the adrenal gland, or adrenarche. This study tested the effects of age, sex, and genus on fecal DHEAS levels using generalized linear mixed-effects models (GLMM). The results showed that age was positively correlated with a pre-pubertal increase in fecal DHEAS across all genera in the study (Hylobates spp., Hoolock spp., Nomascus spp., Symphalangus syndactylus) indicating that the small apes exhibit delayed adrenarche similar to the great apes. The second objective was to examine how reproductive state and social dominance impact fecal androgens and the glucocorticoid metabolite (GCM) to DHEAS ratio (GCM:DHEAS) in free-ranging Japanese macaques (Macaca fuscata) considering environmental factors (season and ambient temperature) and social behaviors (i.e., aggression, and affiliation) as potential variables influencing these steroid hormones. This objective measured fecal GCM and DHEAS in 354 samples by enzyme immunoassay in 11 female macaques (7 pregnant/lactating, 4 nonpregnant/nonlactating). Using GLMM, the results showed that pregnant and lactati (open full item for complete abstract)

Master of Science, The Ohio State University, 2024, Environmental Science

Blood-seeking mosquitoes primarily rely on thermal and chemical cues as they navigate towards hosts. Mosquitoes display specific preferences for target host temperature while avoiding harmful ambient temperature. This behavior known as thermotaxis is in part regulated by the nociceptor transient receptor potential ankyrin 1 (TRPA1), which is expressed in thermosensitive sensilla of mosquitoes. TRPA1 of female mosquitoes is known to detect both noxious temperatures and chemicals; when activated by these stimuli, TRPA1 triggers avoidance behaviors. Therefore, TRPA1 is considered a potential biochemical target for mosquito repellents and antifeedants. One aspect of TRPA1 channels from mosquitoes and other insects that has not been fully studied is the potential interactions between temperature and chemical agonists. In this study, I examined whether high ambient temperatures that activate Aedes aegypti TRPA1 (AaTRPA1) influence the sensitivity of the channel and behavior of mosquitoes to repellent TRPA1 chemical agonists. First, I expressed AaTRPA1 heterologously in Xenopus laevis oocytes and used whole-cell two-electrode voltage clamping to measure TRPA1 activity. I found that the electrophysiological response of AaTRPA1 to two chemical agonists (catnip oil, citronellal) was significantly reduced while the channel was activated by a thermal stimulus ( ~38°C). Moreover, in behavioral bioassays, I found that adult female Ae. aegypti were less repelled by catnip oil when exposed to a noxious temperature (~50°C). Collectively, my results suggest that TRPA1-agonizing repellents, such as catnip oil, may be less efficacious during extreme heat events, which are becoming more common as global climate change progresses.

Doctor of Philosophy, The Ohio State University, 2024, Molecular, Cellular and Developmental Biology

Pancreas is exposed to chronic stress for the duration of human life that can lead to extended inflammatory damage and risk towards development of different pancreatic diseases. Novel therapeutic interventions that could lessen inflammation and stress could significantly improve quality of life in patients. In this dissertation, the role of dietary metabolites in abrogating the progression of pancreatic diseases is explored. In the first part of the dissertation, chronic pancreatitis (CP), a fibro-inflammatory syndrome that develops upon persistent pathological responses to parenchymal stress, is studied. Complex dietary foods like soy and tomatoes have anti-inflammatory effects. Thus, we hypothesized that administration of a soy-tomato enriched diet can reduce inflammation and severity of CP. C57BL/6 mice were injected intraperitoneally with caeurlein to induce CP and administered a control or a soy-tomato enriched diet for 2 weeks. Mice fed a soy-tomato enriched diet had a significantly reduced level of inflammation and severity of CP compared to mice administered a control diet with reduced inflammatory factors and suppressive immune populations, improved total activity and health, and restored serum lipase and amylase levels. These pre-clinical results indicate that a soy-tomato enriched diet may be a novel treatment approach to reduce inflammation and pain in patients with CP. Next, the role of tomatidine, a small molecule tomato metabolite, in limiting pancreatic ductal adenocarcinoma (PDAC) was investigated. PDAC is an aggressive cancer with a five-year survival rate of 13%. The pancreas has a highly functioning endoplasmic reticulum (ER) and is subjected to ER stress. Activating Transcription Factor 4 (ATF4), a master regulator of cellular stress, is exploited by cancer cells to survive. We found evidence that high ATF4 expression correlates with worse overall survival in PDAC, and previous research indicated tomatidine was associated with inhibit (open full item for complete abstract)

Master of Science (MS), Bowling Green State University, 2024, Communication Disorders

The purpose of this study was to examine the interaction between cognitive demands during speech production and concurrent performance of a visuomotor tracking task. Participants performed a working memory task involving embedding a numerical response in a carrier phrase. To modulate cognitive load, participants performed two speech task variants with different degrees of mental tracking effort. For the low-demand variant, participants completed the carrier phrase by counting forward from one, a task that is relatively simple and considered automatic. For the high-demand variant, participants completed the carrier phrase by performing serial subtraction by three, requiring a modest amount of mental tracking effort. Both tasks were performed in isolation and while performing a concurrent visuomotor tracking task. Concurrent serial subtraction led to a reduction in visuomotor tracking accuracy, whereas counting forward did not affect tracking accuracy. Compared to counting forward, serial subtraction was associated with a decrease in speech intensity, lip opening and closing range, and lower lip opening and closing velocities. Compared to speaking insolation, participants exhibited a reduction in lower lip opening and closing velocities and utterance-to-utterance variability when performing the visuomotor tracking task. This pattern suggests that increasing cognitive demands, compounded by divided attention requirements, can affect processing and speech production.K

Doctor of Philosophy, Case Western Reserve University, 2024, Applied Mathematics

Physiological systems underlying vital behaviors, such as breathing, walking, and feeding, are controlled by closed-loop systems integrating central neural circuitry, biomechanics, and sensory feedback. The brain and body orchestration allows these motor systems to demonstrate crucial biological phenomena such as homeostasis, adaptability, and robustness. In this thesis, we investigate the role of sensory feedback in motor dynamics and control, based on an abstract model for motor pattern generation that combines central pattern generator (CPG) dynamics with a sensory feedback mechanism. Given the underdevelopment of control theory for limit cycle systems, we extend recently developed variational tools, which allow us to characterize the sensitivity of the systems to perturbations and changing conditions both within and outside the body. As concrete examples, we apply our methods to several closed-loop models with sensory feedback in place, including locomotion, ingestion, and respiration. Our analytic framework provides a mathematically grounded numerical quantification of the effects of a sustained perturbation on the rhythm performance and robustness, which is also broad enough to study control of oscillations in any nonlinear dynamical systems. Moreover, the observations we obtain from the examples provide important information for future work modeling neuro-motor rhythm generation and insights that have the potential to inform the design of control or rehabilitation systems.

Master of Science (MS), Wright State University, 2024, Physiology and Neuroscience

Organophosphate pesticides, such as Malathion, are commonly used in U.S. and foreign agricultural industries and homes so exposure is inevitable and unavoidable for some people (example: military personnel). Environmental exposure to organophosphate pesticides is an identified risk of neuropathy and neurodegeneration. This study investigates the morphological changes of muscle spindle mechanoreceptors in rat extensor digitorum longus (EDL) muscles in response to environmental Malathion exposure. Animals were exposed to low-dose Malathion 5 days a week for 4 weeks and muscle tissue was collected within 5 days of the last exposure. The EDL muscle was frozen and sectioned onto slides before immunohistochemistry was performed to identify the muscle spindle sensory neurons in the muscle. To analyze morphological changes confocal images were taken, and three parameters of the muscle spindles were measured: length, inter-rotational distance (IRD), and axonal width (AW). In this experiment, we found no significant differences between the Control/Saline and Malathion/Exposure groups. Expanding this research will help us understand the effects of pesticide exposure, even at low doses.

Doctor of Philosophy (PhD), Wright State University, 2024, Biomedical Sciences PhD

Myotonia Congenita (MC) is a rare, inherited ion channelopathy caused by a loss-of-function mutation in the CLCN1 gene. The resulting downregulation of the skeletal muscle chloride channel (ClC-1) results in hyperexcitable skeletal muscle fibers that fire action potentials involuntarily. Patients with MC suffer from debilitating stiffness due to myotonia, described clinically as delayed muscle relaxation following voluntary contraction. Skeletal muscle is a unique system we use in this study to advance our understanding of the pathophysiology underlying MC, and other channelopathies characterized by hyperexcitable cells (i.e., forms of epilepsy and cardiac arrhythmia). Furthermore, re-assessing what makes anti-myotonic drugs such as mexiletine efficacious, yet imperfect, may help us improve the quality of life of affected patients through optimizing treatment outcomes. The current therapeutic approach to treating myotonia is to block voltage-gated Na+ channels in skeletal muscle (Nav1.4) to reduce the fast-transient Na+ current responsible for action potential generation (NaT) via use-dependent block (Statland et al., 2012),(Lo Monaco et al., 2015). Two assumptions are made by the field when taking this approach. First, open channel blockers, those that increase their block of Na channels during repetitive firing, are the preferred type of drug. Second, the block of NaT is the mechanism underlying efficacy. We tested both hypotheses utilizing both transgenic and pharmacologic models of MC in mice. Techniques used included current clamp of the extensor digitorum longus (EDL) muscle and voltage clamp of the flexor digitorum brevis (FDB) muscle. We compared the efficacy of two open-state NaCh blockers; mexiletine and ranolazine, to a closed-state blocker, µ-conotoxin GIIIA (uCTX). We quantitated efficacy against myotonia and determined the optimal concentration of each drug using intracellular recordings from EDL fibers. We were surprised to find that open-channel b (open full item for complete abstract)

Doctor of Philosophy (Ph.D.), University of Dayton, 2024, Biology

Vertebrate freeze tolerance is an extraordinary phenomenon in which up to 70% of an animal's extracellular fluids are converted to ice while circulation, respiration, and neuronal function are all simultaneously suspended during freezing. Upon thawing, animals must tolerate the resumption of physiological function, restore intracellular fluid volumes, and repair injuries that occurred during freeze-thaw. The mechanisms that enable animal freeze tolerance vary by species and represent a myriad of biochemical, cellular, systems, and organismal strategies. Cope's gray treefrog Dryophytes (Hyla) chrysoscelis is a freeze tolerant anuran that repeatedly freezes and thaws each winter in part by utilizing a complex system of cryoprotectants including glycerol, glucose, and urea. Intracellular transport is facilitated by specialized aquaglyceroporin proteins that enable transmembrane movement of water and the cryoprotectants glycerol and urea during freeze-thaw. The physiological mechanisms that enable freeze tolerance in D. chrysoscelis are not entirely understood and cannot be explained by cryoprotectant accumulation alone. The aim of this dissertation is to explore the unknown physiological mechanisms of freeze tolerance in D. chrysoscelis by using an integrative perspective that incorporates all levels of biological organization. Novel experimental protocols were used to evaluate the ecophysiological effects of repeated freezing and thawing, characterize organismal responses to seasonal and cold acclimation, and determine the effects of cold acclimation and freeze-thaw cycles on membrane lipid composition using 1H-NMR analysis. The results from these studies emphasized the complexity of freeze tolerance in D. chrysoscelis and revealed several novel aspects of freeze tolerance in this species including dynamic blue and green dorsal coloration in frozen and thawing frogs, “freeze resistance” in a freeze tolerant vertebrate, evidence of seasonal (open full item for complete abstract)

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

Heart failure is a leading cause of death in the United States and worldwide. Heart failure is defined simply as the inability of the heart to pump a sufficient volume of blood to meet the metabolic needs of the body. This can occur because of impaired ability of the heart to relax and refill with blood (diastolic dysfunction) or because of impaired contractility (systolic dysfunction). Current standards of care for heart failure treat symptoms and can alleviate some symptoms, however, there are currently no approved therapies that directly improve heart function. Therefore, it is necessary to find alternative mechanisms to beneficially increase heart function and improve outcomes for heart failure patients. One key mechanism by which cardiac contraction and relaxation are regulated is by calcium binding to myofilament regulatory proteins. Cardiac contraction is initiated upon increased intracellular calcium. Calcium then binds to the thin filament protein troponin C (TnC), resulting in a conformational change that exposes myosin binding sites on actin allowing myosin cross bridge formation and the sarcomere can shorten. Relaxation is also an active process: following the decrease in intracellular calcium, the thin filament is actively returned to a blocking state, inhibiting actin-myosin interaction. A healthy heart is able to alter this process to increase its function through post-translational modifications. In the classic example of beta-adrenergic signaling, the activation of protein kinase A (PKA) results in the phosphorylation of many proteins within the myocyte including those that affect calcium transients and myosin cycling. Importantly, PKA also phosphorylates the inhibitory subunit of the troponin complex, troponin I (TnI), at serine (S) residues 23 and 24. This phosphorylation has been well characterized to change the responsiveness of the myofilament proteins to calcium, or to decrease calcium sensitivity, therefore accelerating relaxation to incr (open full item for complete abstract)