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Title: INVESTIGATING THE ROLE OF NEURONAL AGING IN FRAGILE X-ASSOCIATED TREMOR/ATAXIA SYNDROME.
Creator: Hencak, Katlin Marie, von Kalm, Laurence, Southwell, Amber, University of Central Florida
Abstract / Description: Fragile X-associated tremor/ataxia syndrome (FXTAS) is an X-linked late-onset neurodegenerative disorder caused by a noncoding trinucleotide repeat expansion in the FMR1 gene. This gene produces fragile x mental retardation protein (FMRP), an RNA binding protein whose targets are involved in brain development and synaptic plasticity. One of the proposed mechanisms of FXTAS pathogenesis is an RNA gain-of-function in which the repeat expansion causes toxic mRNA that sequesters important...
Show moreFragile X-associated tremor/ataxia syndrome (FXTAS) is an X-linked late-onset neurodegenerative disorder caused by a noncoding trinucleotide repeat expansion in the FMR1 gene. This gene produces fragile x mental retardation protein (FMRP), an RNA binding protein whose targets are involved in brain development and synaptic plasticity. One of the proposed mechanisms of FXTAS pathogenesis is an RNA gain-of-function in which the repeat expansion causes toxic mRNA that sequesters important proteins in the cell, interfering with their functions. Another suggested method of pathogenesis is through a mutant protein called FMRpolyG. This protein results from repeat-associated non-AUG (RAN) translation, in which the expanded repeats are translated where they otherwise would not be. This protein co-localizes with intranuclear inclusions and nuclear membrane proteins, causing disorganization of the nuclear lamina in FXTAS patient brain samples and neurons differentiated from FXTAS patient-derived induced pluripotent stem cells (iPSCs). iPSC technology involves reprogramming an adult somatic cell back to an embryonic-like state, allowing it to be differentiated into all cell types. A limit with iPSCs, though, is modeling late-onset disorders because the cells lose all age-related features during reprogramming. Progerin, a truncated form of the lamin A protein, has been used to age neurons differentiated from Parkinson Disease (PD) patient-derived iPSCs. Progerin-mediated aging was found to unmask PD-like phenotypes in those neurons, making it a promising technology for modeling late-onset disorders such as FXTAS. In this study, we investigated the link between the aging process and FXTAS pathogenesis in neurons differentiated from FXTAS patient-derived iPSCs with the use of progerin. Progerin transduction was successful in aging the FXTAS neurons. The presence of FMRpolyG was confirmed and an interaction with Lap2b was observed. In some neurons, there was also an observed interaction between FMRpolyG and progerin. Overall, this data suggests that there is an interaction between the mutant FMRpolyG protein and the nuclear membrane during aging, which may contribute to the cell death that causes neurodegeneration in FXTAS patients.
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Date Issued: 2019
Identifier: CFH2000554, ucf:45678
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFH2000554

Title: HYBRID ADULT NEURON CULTURE SYSTEMS FOR USE IN PHARMACOLOGICAL TESTING.
Creator: Edwards, Darin, Hickman, James, University of Central Florida
Abstract / Description: Neuronal culture systems have many applications, such as basic research into neuronal structure, function, and connectivity as well as research into diseases, conditions, and injuries affecting the brain and its components. In vitro dissociated neuronal systems have typically been derived from embryonic brain tissue, most commonly from the hippocampus of E18 rats. This practice has been motivated by difficulties in supporting regeneration, functional recovery and long-term survival of adult...
Show moreNeuronal culture systems have many applications, such as basic research into neuronal structure, function, and connectivity as well as research into diseases, conditions, and injuries affecting the brain and its components. In vitro dissociated neuronal systems have typically been derived from embryonic brain tissue, most commonly from the hippocampus of E18 rats. This practice has been motivated by difficulties in supporting regeneration, functional recovery and long-term survival of adult neurons in vitro. The overall focus of this dissertation research was to develop a dissociated neuronal culture system from human and animal adult brain tissue, one more functionally and developmentally correlative to the mature brain. To that end, this work was divided into five interrelated topics: development of an adult in vitro neuronal culture system comprised of electrically functional, mitotically stable, developmentally mature neurons from the hippocampus of adult rats; creation of stable two-cell neuronal networks for the study of synaptic communication in vitro; coupling of electrically active adult neurons to microelectrode arrays for high-throughput data collection and analysis; identification of inadequacies in embryonic neuronal culture systems and proving that adult neuronal culture systems were not deficient in similar areas; augmentation of the rat hippocampal culture system to allow for the culture and maintenance of electrically active human neurons for months in vitro. The overall hypothesis for this dissertation project was that tissue engineered in vitro systems comprised of neurons dissociated from mature adult brain tissue could be developed using microfabrication, defined medium formulations, optimized culture and maintenance parameters, and cell-cycle control. Mature differentiated glutamatergic neurons were extracted from hippocampal brain tissue and processed to purify neurons and remove tissue debris. Terminally differentiated rat hippocampal neurons recovered in vitro and displayed mature neuronal morphology. Extracellular glutamate in the culture medium promoted neuronal recovery of electrical function and activity. After recovery, essential growth factors in the culture medium caused adult neurons to reenter the cell cycle and divide multiple times. Only after reaching confluence did some neurons stop dividing. Strategies for inhibition of neuronal mitotic division were investigated, and manipulation of the cdk5 pathway was ultimately found to prevent division in vitro. Prevention of mitotic division as well as optimization of culture and maintenance parameters resulted in a neuronal culture system derived from adult rats in which the neuronal morphology, cytoskeleton and surface protein expression patterns, and electrical activity closely mirrored mature, terminally differentiated adult neurons in vivo. Improvements were also made to the growth surfaces on which neurons attached, regenerated, and survived long-term. Culture surfaces, in this case glass cover slips, were modified with the chemical substrate N-1 [3-(trimethoxysilyl) propyl]-diethylenetriamine (DETA) to create a covalently modified interface with exposed cell-adhesive triamine groups. DETA chemical surfaces were also further modified to create high-resolution patterns, useful in creating engineered two-cell networks of adult hippocampal neurons. Adult hippocampal neurons were also coupled to microelectrode array systems (MEAs) and recovered functionally, fired spontaneously, and reacted to synaptic antagonists in a manner consistent to adult neurons in vivo. Last, neurons from the brains of deceased Alzheimer's disease (AD) patients and from brain tissue excised during surgery for Parkinson's disease (PD), Essential Tremor (ET), and brain tumor were isolated and cultured, with these neurons morphological regenerating and electrically recovering in vitro.
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Date Issued: 2011
Identifier: CFE0004045, ucf:49127
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0004045

Title: MCP-1 AND APP INVOLVEMENT IN GLIAL DIFFERENTIATION AND MIGRATION OF NEUROPROGENITOR CELLS.
Creator: Vrotsos, Emmanuel, Sugaya, Kiminobu, University of Central Florida
Abstract / Description: Neuroprogenitor cells are an important resource because of their potential to replace damaged cells in the brain caused by trauma and disease. It is of great importance to better understand which factors influence the differentiation and migration of these cells. Previously it has been reported that neuroprogenitor cells undergoing apoptotic stress have increased levels of Amyloid precursor protein (APP) and increased APP expression results in glial differentiation. APP activity was also...
Show moreNeuroprogenitor cells are an important resource because of their potential to replace damaged cells in the brain caused by trauma and disease. It is of great importance to better understand which factors influence the differentiation and migration of these cells. Previously it has been reported that neuroprogenitor cells undergoing apoptotic stress have increased levels of Amyloid precursor protein (APP) and increased APP expression results in glial differentiation. APP activity was also shown to be required for staurosporine induced glial differentiation of neuroprogenitor cells. Monocyte chemoattractant protein-1 (MCP-1) is a chemokine that is expressed during inflammatory. The binding of MCP-1 to its chemokine receptor induces expression of novel transcription factor MCP-1 induced protein (MCPIP). MCPIP expression subsequently leads to cell death. Previous studies have shown that pro apoptotic factors have the ability to induce neural differentiation. Therefore, we investigated if MCPIP expression leads to differentiation of NT2 neuroprogenitor cells. Results showed that MCPIP expression increased glial fibrillary acid protein expression and also caused distinct morphological changes, both indicative of glial differentiation. Similar results were observed with MCP-1 treatment. Interestingly, APP expression decreased in response to MCPIP. Instead, we found APP activity regulates expression of both MCP-1 and MCPIP. Furthermore, inhibition of either p38 MAPK or JAK signaling pathways significantly reduced APP's effect on MCP-1 and MCPIP. These data demonstrate the role APP has in glial differentiation of NT2 cells through MCP-1/MCPIP signaling. It is possible that increased APP expression after CNS injury could play a ii role in MCP-1 production, possibly promoting astrocyte activation at injured site. We next investigated the effect that MCP-1 has on NT2 cell migration. Studies have shown that when neuroprogenitor cells are transplanted into the brain they migrate towards damaged areas, suggesting that these areas express factors that recruit migrating cells. Generally, after neuronal injury there is a neuroinflammatory response that results in increased chemokine production. We demonstrate that MCP-1 significantly induces the migration of NT2 neuroprogenitor cells. Activation of intracellular cyclic adenosine monophosphate (cAMP) or protein kinase C with forskolin and phorbol 12-myristate 13-acetate (PMA), respectively, was able to completely abolish the MCP-1 induced migration. Contrarily, neither extracellular signal-regulated kinase or p38 mitogen activated protein kinase was required for NT2 cells to respond to MCP-1. Interestingly, APP's ability to activate MCP-1 expression was shown to play a role in NT2 cell migration. We showed that NT2 cells expressing APP were capable of inducing migration of other neuroprogenitor cells. Utilizing a MCP-1 neutralizing antibody we discovered that APP induced migration was not caused solely by increased MCP-1 production. Interestingly, APP increased expression of several C-C chemokines: MCP-1, Regulated upon Activation, Normal T-cell Expressed, and Secreted (RANTES), and macrophage inflammatory protein alpha (MIP-1 alpha). This demonstrates the unique role APP has in regulating chemokine production, which directly affects cell migration. Taken together, this study provides us with a greater understanding of the mechanisms involved in both glial differentiation and migration of NT2 neuroprogenitor cells.
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Date Issued: 2009
Identifier: CFE0002517, ucf:47661
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0002517

Title: MICROSCOPIC ANALYSIS OF SYMPATHETIC AND PARASYMPATHETIC DISTRIBUTION, TERMINAL MORPHOLOGY, AND INTERACTION IN WHOLE-MOUNT ATRIA OF C57BL/6 MICE.
Creator: Harden, Scott, Cheng, Zixi (Jack), University of Central Florida
Abstract / Description: The sympathetic (SNS) and parasympathetic (PSNS) branches of the autonomic nervous system (ANS) innervate the heart, exerting excitatory and inhibitory influences (respectively) over cardiac functions (heart rate, AV conduction velocity, and contractility). However, the distribution and structure of SNS and PSNS innervation has not yet been well studied. Detailed characterization of the distributional organization and structural morphology of the SNS and PSNS in normal states is essential to...
Show moreThe sympathetic (SNS) and parasympathetic (PSNS) branches of the autonomic nervous system (ANS) innervate the heart, exerting excitatory and inhibitory influences (respectively) over cardiac functions (heart rate, AV conduction velocity, and contractility). However, the distribution and structure of SNS and PSNS innervation has not yet been well studied. Detailed characterization of the distributional organization and structural morphology of the SNS and PSNS in normal states is essential to the study of pathological autonomic remodeling. The present study utilized double immunohistochemical labeling techniques to examine tyrosine hydroxylase (TH) immunoreactive (IR) SNS and vesicular acetylcholine transporter (VAChT) IR PSNS axons and terminal structures in whole-mount atria of C57BL/6 mice. We found that: (1) The atria contain a dense network of ANS axons. TH-IR, VAChT-IR, and dual cholinergic/dopaminergic TH+VAChT-IR axons travel together in bundles on the epicardium before branching into differentiated terminal structures. (2) Parallel TH-IR and VAChT-IR axons often diverge from epicardial bundles and travel in parallel (less than 1μm apart) before forming terminal structures in the epicardium and myocardium. Such parallel SNS/PSNS axons interdigitize and have large alternating varicosities along their length adjacent to one other, suggesting possible antagonistic communication between both branches of the ANS at the prejunctional level. (3) Intrinsic cardiac ganglia (ICG) are targets for extrinsic sympathetic nerves which travel through ICG without forming large synaptic varicosities around cardiac principal neurons (PNs). (4) Small intensely fluorescent (SIF) cells (presumably chemoreceptors and/or interneurons) exist near SNS bundles, inside ICG, and in the epicardium unaccompanied by ganglia and nerve bundles. (5) The subpopulation of TH+VAChT-IR PNs within ICG form loose terminals in the atria and do not project to other PNs. (6) Both TH-IR and VAChT-IR axons innervate atrial vasculature. (7) TH-IR axons innervate fat pads adjacent to the heart. (8) SNS/PSNS parallelism is not exclusive to the atria. Similar structures exist in the esophagus, right ventricle, and small intestine. This study provides a novel and overall view of the innervation and interaction of the SNS and PSNS in the atria. This will underlie a foundation for future physiological, pharmacological, and anatomical studies of SNS/PSNS innervation, interaction, and remodeling in pathological states (such as aging, intermittent hypoxia and diabetes).
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Date Issued: 2009
Identifier: CFE0002561, ucf:47647
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0002561

Title: THE EFFECT OF CAFFEINE ON MIGRAINE HEADACHES.
Creator: Shimshoni, Deborah, Samsam, Mohtashem, University of Central Florida
Abstract / Description: As the most widely consumed drug around the globe, there is a vast array of contradicting research available on caffeine. One of the most debated and researched topics on caffeine is its effect on the brain. Meanwhile, the data on the neurological condition of migraine has information scattered throughout countless research articles and experiments. Although neither migraine or caffeine are completely understood by the medical world, this analysis attempts to give a more coherent...
Show moreAs the most widely consumed drug around the globe, there is a vast array of contradicting research available on caffeine. One of the most debated and researched topics on caffeine is its effect on the brain. Meanwhile, the data on the neurological condition of migraine has information scattered throughout countless research articles and experiments. Although neither migraine or caffeine are completely understood by the medical world, this analysis attempts to give a more coherent understanding of the relationship between the two. This is done by first understanding the known and theorized mechanisms of caffeine as well as the pathologies of migraine. Discussions on channelopathies, current migraine medications, and case studies will be presented. After much background research, we hypothesized that caffeine could excite neurons at physiological concentrations to the point of activation. This was tested by targeting the transcription factor cFos using immunocytochemistry in vitro. The protein cFos was identified due to its rapid translation�just 15 minutes after stimuli�to indicate activation. In addition to a control culture, three different caffeine concentrations were tested on the neurons: 50 micromoles� average plasma level after 1-2 cups of coffee consumption, 100 micromoles�average plasma level after 5-6 cups of coffee also believed to be the therapeutic amount to defend against neurological diseases such as Alzheimers Disease, and 250 micromoles�the average plasma level considered to be toxic in humans. Indeed, we saw a 53.8% increase in cFos expression in the neurons as 100 micromolar of caffeine was added and exposed to the cell cultures for 24 hours. In order to ensure the results obtained in this study were physiologically relevant in vivo, known toxic levels were tested for in vitro neurotoxicity. It was found in vitro that at the non toxic plasma concentrations of 50 micromolar and 100 micromolar of caffeine did not display cellular death as tested by Trypan Blue viability testing, Crystal Violet morphologies, and fleurojade immunochemistry that tests for degeneration. Each of these experiments identified a significant death increase as the toxic level of 250 micromoles of caffeine were utilized. This allowed us to theorize that the activation of neurons found in these experiments due to caffeine exposure would apply the same effect in vivo.
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Date Issued: 2016
Identifier: CFH2000087, ucf:45559
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFH2000087

Title: THE EFFECTS OF MATRIX METALLOPROTEINASE-9 ON CX3CL1 SHEDDING AND AXON RETRACTION.
Creator: Dobrie, Lauren A, Hawthorne, Alicia, University of Central Florida
Abstract / Description: Spinal cord injury (SCI) often leads to irreversible damage, and permanent paralysis inferior to the injury is common (Leibinger et al., 2013). Injury to the spinal cord occurs in two phases. In the first phase, components of the spinal cord are subject to mechanical trauma causing direct damage. In the second phase, damage spreads from the area of injury through molecular processes. Several studies have linked M1 "pro-inflammatory" macrophages to exacerbation of damage by inducing dieback of...
Show moreSpinal cord injury (SCI) often leads to irreversible damage, and permanent paralysis inferior to the injury is common (Leibinger et al., 2013). Injury to the spinal cord occurs in two phases. In the first phase, components of the spinal cord are subject to mechanical trauma causing direct damage. In the second phase, damage spreads from the area of injury through molecular processes. Several studies have linked M1 "pro-inflammatory" macrophages to exacerbation of damage by inducing dieback of dystrophic axons, but not healthy axons, through direct cellular contact. Several studies have identified the presence of macrophage subtypes at specific time. A literature review was conducted in order to summarize these findings (Busch, Horn, Silver, and Silver, 2009; Evans et al., 2014; Horn, Busch, Hawthorne, van Rooijen, and Silver, 2008; Kigerl et al., 2009; Shechter et al., 2013). Although the full mechanism behind the process of M1 macrophage-mediated dieback of dystrophic axons is unclear, matrix metalloproteinase-9 (MMP-9) produced by these macrophages has been shown to play a role. However, the specific interaction between MMP-9 and neurons is under investigation. The research described explores the relationship between MMP-9 and fractalkine (CX3CL1), a surface protein expressed by CNS neurons. SDS-PAGE and western blot were used to determine whether the presence of MMP-9 increases the cleavage of fractalkine at several time intervals. At a concentration of 300ng/ml, MMP-9 was not found to demonstrate cleavage of fractalkine.
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Date Issued: 2019
Identifier: CFH2000506, ucf:45636
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFH2000506

Title: CHARACTERIZATION OF AN ADVANCED NEURON MODEL.
Creator: Echanique, Christopher, Behal, Aman, University of Central Florida
Abstract / Description: This thesis focuses on an adaptive quadratic spiking model of a motoneuron that is both versatile in its ability to represent a range of experimentally observed neuronal firing patterns as well as computationally efficient for large network simulation. The objective of research is to fit membrane voltage data to the model using a parameter estimation approach involving simulated annealing. By manipulating the system dynamics of the model, a realizable model with linear parameterization (LP)...
Show moreThis thesis focuses on an adaptive quadratic spiking model of a motoneuron that is both versatile in its ability to represent a range of experimentally observed neuronal firing patterns as well as computationally efficient for large network simulation. The objective of research is to fit membrane voltage data to the model using a parameter estimation approach involving simulated annealing. By manipulating the system dynamics of the model, a realizable model with linear parameterization (LP) can be obtained to simplify the estimation process. With a persistently excited current input applied to the model, simulated annealing is used to efficiently determine the best model parameters that minimize the square error function between the membrane voltage reference data and data generated by the LP model. Results obtained through simulation of this approach show feasibility to predict a range of different neuron firing patterns.
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Date Issued: 2012
Identifier: CFH0004259, ucf:44958
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFH0004259

Title: End to End Brain Fiber Orientation Estimation Using Deep Learning.
Creator: Puttashamachar, Nandakishore, Bagci, Ulas, Shah, Mubarak, Rahnavard, Nazanin, Sundaram, Kalpathy, University of Central Florida
Abstract / Description: In this work, we explore the various Brain Neuron tracking techniques, one of the most significant applications of Diffusion Tensor Imaging. Tractography is a non-invasive method to analyze underlying tissue micro-structure. Understanding the structure and organization of the tissues facilitates a diagnosis method to identify any aberrations which can occurwithin tissues due to loss of cell functionalities, provides acute information on the occurrences of brain ischemia or stroke, the...
Show moreIn this work, we explore the various Brain Neuron tracking techniques, one of the most significant applications of Diffusion Tensor Imaging. Tractography is a non-invasive method to analyze underlying tissue micro-structure. Understanding the structure and organization of the tissues facilitates a diagnosis method to identify any aberrations which can occurwithin tissues due to loss of cell functionalities, provides acute information on the occurrences of brain ischemia or stroke, the mutation of certain neurological diseases such as Alzheimer, multiple sclerosis and so on. Under all these circumstances, accurate localization of the aberrations in efficient manner can help save a life. Following up with the limitations introduced by the current Tractography techniques such as computational complexity, reconstruction errors during tensor estimation and standardization, we aim to elucidate these limitations through our research findings. We introduce an End to End Deep Learning framework which can accurately estimate the most probable likelihood orientation at each voxel along a neuronal pathway. We use Probabilistic Tractography as our baseline model to obtain the training data and which also serve as a Tractography Gold Standard for our evaluations. Through experiments we show that our Deep Network can do a significant improvement over current Tractography implementations by reducing the run-time complexity to a significant new level. Our architecture also allows for variable sized input DWI signals eliminating the need to worry about memory issues as seen with the traditional techniques. The advantageof this architecture is that it is perfectly desirable to be processed on a cloud setup and utilize the existing multi GPU frameworks to perform whole brain Tractography in minutes rather than hours. The proposed method is a good alternative to the current state of the art orientation estimation technique which we demonstrate across multiple benchmarks.
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Date Issued: 2017
Identifier: CFE0007292, ucf:52156
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007292

Title: PATTERNED CELL CULTURES FOR HIGH THROUGHPUT STUDIES OF CELL ELECTROPHYSIOLOGY AND DRUG SCREENING APPLICATIONS.
Creator: Natarajan, Anupama, Hickman, James, University of Central Florida
Abstract / Description: Over the last decade, the field of tissue and bio-engineering has seen an increase in the development of in vitro high-throughput hybrid systems that can be used to understand cell function and behavior at the cellular and tissue levels. These tools would have a wide array of applications including for implants, drug discovery, and toxicology, as well as for studying cell developmental behavior and as disease models. Currently, there are a limited number of efficient, functional drug...
Show moreOver the last decade, the field of tissue and bio-engineering has seen an increase in the development of in vitro high-throughput hybrid systems that can be used to understand cell function and behavior at the cellular and tissue levels. These tools would have a wide array of applications including for implants, drug discovery, and toxicology, as well as for studying cell developmental behavior and as disease models. Currently, there are a limited number of efficient, functional drug screening assays in the pharmacology industry and studies of cell-surface interactions are complicated and invasive. Most cell physiology studies are performed using conventional patch-clamp techniques or random networks cultured on silicon devices such as Microelectrode Arrays (MEAs) and Field Effect transistors (FETs). The objective of this study was to develop high-throughput in vitro platforms that could be used to analyze cell function and their response to various stimuli. Our hypothesis was that by utilizing surface modification to provide external guidance cues for various cell types and by controlling the cell environment in terms of culture conditions, we could develop an in vitro hybrid platform for sensing and testing applications. Such a system would not only give information regarding the surface effects on the growth and behavior of cells for implant development applications, but also allow for the study of vital cell physiology parameters like conduction velocity in cardiomyocytes and synaptic plasticity in neuronal networks. This study outlines the development of these in vitro high throughput systems that have varied applications ranging from tissue engineering to drug development. We have developed a simple and relatively high-throughput method in order to test the physiological effects of varying chemical environments on rat embryonic cardiac myocytes in order to model the degradation effects of polymer scaffolds. Our results, using our simple test system, are in agreement with earlier observations that utilized a complex 3D biodegradable scaffold. Thus, surface functionalization with self-assembled monolayers combined with histological/physiological testing could be a relatively high throughput method for biocompatibility studies and for the optimization of the material/tissue interface in tissue engineering. Traditional multielectrode extracellular recording methods were combined with surface patterning of cardiac myocyte monolayers to enhance the information content of the method; for example, to enable the measurement of conduction velocity, refractory period after action potentials or to create a functional reentry model. Two drugs, 1-Heptanol, a gap junction blocker, and Sparfloxacin, a fluoroquinone antibiotic, were tested in this system. 1-Heptanol administration resulted in a marked reduction in conduction velocity, whereas Sparfloxacin caused rapid, irregular and unsynchronized activity, indicating fibrillation. As shown in these experiments, the patterning of cardiac myocyte monolayers increased the information content of traditional multielectrode measurements. Patterning techniques with self-assembled monolayers on microelectrode arrays were also used to study the physiological properties of hippocampal networks with functional uni-directional connectivity, developed to study the mono-synaptic connections found in the dentate gyrus. Results indicate that changes in synaptic connectivity and strength were chemically induced in these patterned hippocampal networks. This method is currently being used for studying long term potentiation at the cellular level. For this purpose, two cell patterns were optimized for cell migration onto the pattern as demonstrated by time lapse studies, and for supporting the best pattern formation and cell survival on these networks. The networks formed mature interconnected spiking neurons. In conclusion, this study demonstrates the development and testing of in vitro high-throughput systems that have applications in drug development, understanding disease models and tissue engineering. It can be further developed for use with human cells to have a more predictive value than existing complex, expensive and time consuming methods.
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Date Issued: 2010
Identifier: CFE0003384, ucf:48437
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0003384

Title: Characterization of neural cells derived from reelin-deficient schizophrenic patient iPS cells.
Creator: Roberts, Nicole, Sugaya, Kiminobu, Ebert, Steven, Masternak, Michal, University of Central Florida
Abstract / Description: Reelin is a large, extracellular glycoprotein that binds to several membrane receptors on neural stem cells (HNSCs), neural progenitor cells (NPCs), and neuroblasts of mammals to direct their migration. Previously, our lab established the presence of Reelin increased migration of wild-type fetal-derived HNSC's, both in vitro and in vivo. In addition, we demonstrated that Reelin protein treatment also increases the formation of radial glia via Notch-1 signaling, in vitro. Radial glia are...
Show moreReelin is a large, extracellular glycoprotein that binds to several membrane receptors on neural stem cells (HNSCs), neural progenitor cells (NPCs), and neuroblasts of mammals to direct their migration. Previously, our lab established the presence of Reelin increased migration of wild-type fetal-derived HNSC's, both in vitro and in vivo. In addition, we demonstrated that Reelin protein treatment also increases the formation of radial glia via Notch-1 signaling, in vitro. Radial glia are precursors to NPCs, as well as a scaffold for neuroblast migration during cortical lamination. Reelin has long been associated with Schizophrenia (SZ). Because post-mortem brains are limited to describing the end-point of the disease, heterozygous haplodeficient Reelin knock-out (Reeler) mice are used to model developmental aspects of SZ in vivo. However, SZ is a complex, polyfactoral disease with a myriad of dysfunctional pathways that may have unforeseen effects on Reelin signaling. K. Brennand et al. (2014) reported low Reelin mRNA expression and cellular characteristics mirroring the Reeler mouse in induced pluripotent stem (iPS) cell-derived NPCs and neurons from living SZ patients. Building upon this and our work with stem cells, here we consider Reelin's effects on migration of Reelin-deficient iPS cell-derived NPCs. Reelin treatment of consists of secreted Reelin from transfected human embryonic kidney 293 cells (HEK 293) with the pCRL RELN gene-containing plasmid created by G. D'Arcangelo (1997) and given to us by T. Curran. Using the metric of cellular migration, this is the first time it have been shown that SZ iNPCs are capable of receiving and reacting to extracellular Reelin. Due to our validation of this model, further work using iPS cell-derived neural cells can confidently be used for future disease modeling and drug discovery of Reelin-deficient SZ.
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Date Issued: 2018
Identifier: CFE0007361, ucf:52091
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007361

Title: Development of a Functional In Vitro 3D Model of the Peripheral Nerve.
Creator: Anderson, Wesley, Lambert, Stephen, Hickman, James, Fernandez-Valle, Cristina, Willenberg, Bradley, University of Central Florida
Abstract / Description: Peripheral neuropathies, affect approximately 20 million people in the United States and are often a complication of conditions such as diabetes that can result in amputation of affected areas such as the feet and toes. In vitro methodologies to facilitate the understanding and treatment of these disorders often lack the cellular and functional complexity required to accurately model peripheral neuropathies. In particular, they are often 2-D and functional readouts, such as electrical...
Show morePeripheral neuropathies, affect approximately 20 million people in the United States and are often a complication of conditions such as diabetes that can result in amputation of affected areas such as the feet and toes. In vitro methodologies to facilitate the understanding and treatment of these disorders often lack the cellular and functional complexity required to accurately model peripheral neuropathies. In particular, they are often 2-D and functional readouts, such as electrical activity, are limited to cell bodies thereby limiting the understanding of axonopathy which often characterizes these disorders. We have developed a functional 3-D model of peripheral nerves using a capillary alginate gel (Capgel(TM)), as a scaffold. We hypothesize that: 1) The unique microcapillary structure of Capgel(TM) allows for the modeling of the 3-D microstructure of the peripheral nerve, and 2) That axon bundling in the capillary allows for the detection of axonal electrical activity. In our initial studies, we demonstrate that culturing embryonic dorsal root ganglia (DRG) within the Capgel(TM) environment allows for the separation of cell bodies from axons and recreates many of the features of an in vivo peripheral nerve fascicle including myelinated axons and the formation of a rudimentary perineurium. To develop functionality for this model we have integrated the DRG Capgel(TM) culture with a microelectrode array to examine spontaneous activity in axon bundles, which we find demonstrates superiority to other widely used 2-D models of the same tissue. Furthermore, by analyzing the activity on individual electrodes, we were able to record action potentials from multiple axons within the same bundle indicating a functional complexity comparable to that observed in fascicles in vivo. This 3D model of the peripheral nerve can be used to study the functional complexities of peripheral neuropathies and nerve regeneration as well as being utilized in the development of novel therapeutics.
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Date Issued: 2018
Identifier: CFE0007150, ucf:52303
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007150

Title: Artificial Neuron using MoS2/Graphene Threshold Switching Memristor.
Creator: Kalita, Hirokjyoti, Roy, Tania, Sundaram, Kalpathy, Yuan, Jiann-Shiun, University of Central Florida
Abstract / Description: With the ever-increasing demand for low power electronics, neuromorphic computing has garnered huge interest in recent times. Implementing neuromorphic computing in hardware will be a severe boost for applications involving complex processes such as pattern recognition. Artificial neurons form a critical part in neuromorphic circuits, and have been realized with complex complementary metal(-)oxide(-)semiconductor (CMOS) circuitry in the past. Recently, insulator-to-metal-transition (IMT)...
Show moreWith the ever-increasing demand for low power electronics, neuromorphic computing has garnered huge interest in recent times. Implementing neuromorphic computing in hardware will be a severe boost for applications involving complex processes such as pattern recognition. Artificial neurons form a critical part in neuromorphic circuits, and have been realized with complex complementary metal(-)oxide(-)semiconductor (CMOS) circuitry in the past. Recently, insulator-to-metal-transition (IMT) materials have been used to realize artificial neurons. Although memristors have been implemented to realize synaptic behavior, not much work has been reported regarding the neuronal response achieved with these devices. In this work, we study the IMT in 1T-TaS2 and the volatile threshold switching behavior in vertical-MoS2 (v-MoS2) and graphene van der Waals heterojunction system. The v-MoS2/graphene threshold switching memristor (TSM) is used to produce the integrate-and-fire response of a neuron. We use large area chemical vapor deposited (CVD) graphene and MoS2, enabling large scale realization of these devices. These devices can emulate the most vital properties of a neuron, including the all or nothing spiking, the threshold driven spiking of the action potential, the post-firing refractory period of a neuron and strength modulated frequency response. These results show that the developed artificial neuron can play a crucial role in neuromorphic computing.
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Date Issued: 2018
Identifier: CFE0007203, ucf:52291
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007203

Title: Cell Printing: An Effective Advancement for the Creation of um Size Patterns for Integration into Microfluidic BioMEMs Devices.
Creator: Aubin, Megan, Hickman, James, Coffey, Kevin, Lambert, Stephen, University of Central Florida
Abstract / Description: The Body-on-a-Chip (BoaC) is a microfluidic BioMEMs project that aims to replicate major organs of the human body on a chip, providing an in vitro drug testing platform without the need to resort to animal model testing. Using a human model also provides significantly more accurate drug response data, and may even open the door to personalized drug treatments. Microelectrode arrays integrated with human neuronal or human cardiac cells that are positioned on the electrodes are essential...
Show moreThe Body-on-a-Chip (BoaC) is a microfluidic BioMEMs project that aims to replicate major organs of the human body on a chip, providing an in vitro drug testing platform without the need to resort to animal model testing. Using a human model also provides significantly more accurate drug response data, and may even open the door to personalized drug treatments. Microelectrode arrays integrated with human neuronal or human cardiac cells that are positioned on the electrodes are essential components for BoaC systems. Fabricating these substrates relies heavily on chemically patterned surfaces to control the orientation and growth of the cells. Currently, cells are plated by hand onto the surface of the chemically patterned microelectrode arrays. The cells that land on the cytophobic 2-[Methoxy(Polyethyleneoxy)6-9Propyl]trimethoxysilane (PEG) coating die and detach from the surface, while the cells that land on the cytophilic diethylenetriamine (DETA) coating survive and attach to the surface exhibiting normal physiology and function. The current technique wastes a significant amount of cells, some of which are extremely expensive, and is labor intensive. Cell printing, the process of dispensing cells through a 3D printer, makes it possible to pinpoint the placement of cells onto the microelectrodes, drastically reducing the number of cells utilized. Scaled-up manufacturing is also possible due to the automation capabilities provided by printing. In this work, the specific conditions for printing each cell type is unique, the printing of human motoneurons, human sensory neurons and human cardiac cells was investigated. The viability and functionality of the printed cells are demonstrated by phase images, immunostaining and electrical signal recordings. The superior resolution of cell printing was then taken one step further by successfully printing two different cell types in close proximity to encourage controlled innervation and communication.
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Date Issued: 2017
Identifier: CFE0007390, ucf:52074
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0007390

Title: GLIAL DIFFERENTIATION OF HUMAN UMBILICAL STEM CELLS IN 2D AND 3D ENVIRONMENTS.
Creator: Davis, Hedvika, Hickman, James, University of Central Florida
Abstract / Description: During differentiation stem cells are exposed to a range of microenvironmental chemical and physical cues. In this study, human multipotent progenitor cells (hMLPCs) were differentiated from umbilical cord into oligodendrocytes and astrocytes. Chemical cues were represented by a novel defined differentiation medium containing the neurotransmitter norepinephrine (NE). In traditional 2 dimensional (2D) conditions, the hMLPCs differentiated into oligodendrocyte precursors, but did not progress...
Show moreDuring differentiation stem cells are exposed to a range of microenvironmental chemical and physical cues. In this study, human multipotent progenitor cells (hMLPCs) were differentiated from umbilical cord into oligodendrocytes and astrocytes. Chemical cues were represented by a novel defined differentiation medium containing the neurotransmitter norepinephrine (NE). In traditional 2 dimensional (2D) conditions, the hMLPCs differentiated into oligodendrocyte precursors, but did not progress further. However, in a constructed 3 dimensional (3D) environment, the hMLPCs differentiated into committed oligodendrocytes that expressed MBP. When co-cultured with rat embryonic hippocampal neurons (EHNs), hMLPCs developed in astrocytes or oligodendrocytes, based on presence of growth factors in the differentiation medium. In co-culture, physical cues provided by axons were essential for complete differentiation of both astrocytes and oligodendrocytes. This study presents a novel method of obtaining glia from human MLPCs that could eliminate many of the difficulties associated with their differentiation from embryonic stem cells. In addition, it reveals the complex interplay between physical cues and biomolecules on stem cell differentiation.
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Date Issued: 2011
Identifier: CFE0003570, ucf:48894
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0003570

Title: The Effects of Phosphatidylserine on Reaction Time and Cognitive Function Following an Exercise Stress.
Creator: Wells, Adam, Hoffman, Jay, Fragala, Maren, Stout, Jeffrey, University of Central Florida
Abstract / Description: Phosphatidylserine (PS) is an endogenously occurring phospholipid that has been shown to have cognition and mood enhancing properties in humans, possibly through its role as an enzyme co-factor in cellular signal transduction. Specifically, PS has been identified as activator of classical isoforms of protein kinase C, an enzyme known to be involved in the growth and differentiation of neural cells, and is therefore thought to play a role in the protection of neurons.The purpose of this study...
Show morePhosphatidylserine (PS) is an endogenously occurring phospholipid that has been shown to have cognition and mood enhancing properties in humans, possibly through its role as an enzyme co-factor in cellular signal transduction. Specifically, PS has been identified as activator of classical isoforms of protein kinase C, an enzyme known to be involved in the growth and differentiation of neural cells, and is therefore thought to play a role in the protection of neurons.The purpose of this study was to examine the effects of supplementation with PS and caffeine on measures of cognition, reaction time and mood prior to and following an exercise stress. Twenty, healthy, resistance trained males (17) and females (3) (mean (&)#177; SD; age: 22.75 (&)#177; 3.27 yrs; height: 177.03 (&)#177; 8.44cm; weight: 78.98 (&)#177; 11.24kg; body fat%: 14.28 (&)#177; 6.6), volunteered to participate in this randomized, double-blind, placebo-controlled study. Participants were assigned to a PS group (400mg/day PS; 100mg/day caffeine, N=9) or PL (16g/day Carbs, N=11) delivered in the form of 4 candy chews identical in size, shape and color. Subjects performed an acute bout of full body resistance exercise, prior to (T1) and following 14 days of supplementation (T2). Measures of reaction time (Dynavision(&)#174; D2 Visuomotor Training Device), cognition (Serial Subtraction Test, SST), and mood (Profile of Mood States, POMS) were assessed immediately before and following resistance exercise in both T1 and T2. Data was analyzed using two-way ANCOVA and repeated measures ANOVA.Supplementation with 400mg PS and 100mg caffeine did not have a significant impact upon measures of reaction time or cognition between groups at baseline or following acute resistance exercise. However, there was a non-significant trend to the attenuation of fatigue between groups, following acute resistance exercise (p = 0.071). Interestingly, our data suggests that acute resistance exercise alone may improve cognitive function.Although more research is necessary regarding optimal dosage and supplementation duration, the current findings suggest that supplementation 400mg/day PS with 100mg/day caffeine may attenuate fatigue following acute resistance exercise. It is possible that the lack of significance may be the result of both an inhibition of the PS activated pathway and a withdrawal effect from caffeine.
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Date Issued: 2012
Identifier: CFE0004457, ucf:49325
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0004457

Title: Characterization of a Spiking Neuron Model via a Linear Approach.
Creator: Jabalameli, Amirhossein, Behal, Aman, Hickman, James, Haralambous, Michael, University of Central Florida
Abstract / Description: In the past decade, characterizing spiking neuron models has been extensively researched as anessential issue in computational neuroscience. In this thesis, we examine the estimation problemof two different neuron models. In Chapter 2, We propose a modified Izhikevich model withan adaptive threshold. In our two-stage estimation approach, a linear least squares method anda linear model of the threshold are derived to predict the location of neuronal spikes. However,desired results are not...
Show moreIn the past decade, characterizing spiking neuron models has been extensively researched as anessential issue in computational neuroscience. In this thesis, we examine the estimation problemof two different neuron models. In Chapter 2, We propose a modified Izhikevich model withan adaptive threshold. In our two-stage estimation approach, a linear least squares method anda linear model of the threshold are derived to predict the location of neuronal spikes. However,desired results are not obtained and the predicted model is unsuccessful in duplicating the spikelocations. Chapter 3 is focused on the parameter estimation problem of a multi-timescale adaptivethreshold (MAT) neuronal model. Using the dynamics of a non-resetting leaky integrator equippedwith an adaptive threshold, a constrained iterative linear least squares method is implemented tofit the model to the reference data. Through manipulation of the system dynamics, the thresholdvoltage can be obtained as a realizable model that is linear in the unknown parameters. This linearlyparametrized realizable model is then utilized inside a prediction error based framework to identifythe threshold parameters with the purpose of predicting single neuron precise firing times. Thisestimation scheme is evaluated using both synthetic data obtained from an exact model as well asthe experimental data obtained from in vitro rat somatosensory cortical neurons. Results show theability of this approach to fit the MAT model to different types of reference data.
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Date Issued: 2015
Identifier: CFE0005958, ucf:50803
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005958

Title: Development of human and rodent based in vitro systems toward better translation of bench to bedside in vivo results.
Creator: Berry, Bonnie, Hickman, James, Khaled, Annette, Lambert, Stephen, Sugaya, Kiminobu, University of Central Florida
Abstract / Description: Prospective medicinal compounds progress through multiple testing phases before becoming licensed drugs. Testing of novel compounds includes a preclinical phase where the potential therapeutic is tested in vitro and/or in animal models in vivo to predict its potential efficacy and/or toxicity in humans. The failure of preclinical models to accurately predict human drug responses can lead to potentially dangerous compounds being administered to humans, or potentially beneficial compounds being...
Show moreProspective medicinal compounds progress through multiple testing phases before becoming licensed drugs. Testing of novel compounds includes a preclinical phase where the potential therapeutic is tested in vitro and/or in animal models in vivo to predict its potential efficacy and/or toxicity in humans. The failure of preclinical models to accurately predict human drug responses can lead to potentially dangerous compounds being administered to humans, or potentially beneficial compounds being kept in development abeyance. Moreover, inappropriate choice in model organism for studying disease states may result in pushing forward inappropriate drug targets and/or compounds and wasting valuable time and resources in producing much-needed medications. In this dissertation, models for basic science research and drug testing are investigated with the intention of improving current preclinical models in order to drive drugs to market faster and more efficiently. We found that embryonic rat hippocampal neurons, commonly used to study neurodegenerative disease mechanisms in vitro, take 3-4 weeks to achieve similar, critical ion-channel expression profiles as seen in adult rat hippocampal cultures. We also characterized a newly-available commercial cell line of human induced pluripotent stem cell-derived neurons for their applicability in long-term studies, and used them to develop a more pathologically relevant model of early Alzheimer's Disease in vitro. Finally, we attempted to create an engineered, layered neural network of human neurons to study drug responses and synaptic mechanisms. Utilization of the results and methods described herein will help push forward the development of better model systems for translation of laboratory research to successful clinical human drug trials.
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Date Issued: 2015
Identifier: CFE0006261, ucf:51031
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0006261

Title: Nonlinear dynamic modeling, simulation and characterization of the mesoscale neuron-electrode interface.
Creator: Thakore, Vaibhav, Hickman, James, Mucciolo, Eduardo, Rahman, Talat, Johnson, Michael, Behal, Aman, Molnar, Peter, University of Central Florida
Abstract / Description: Extracellular neuroelectronic interfacing has important applications in the fields of neural prosthetics, biological computation and whole-cell biosensing for drug screening and toxin detection. While the field of neuroelectronic interfacing holds great promise, the recording of high-fidelity signals from extracellular devices has long suffered from the problem of low signal-to-noise ratios and changes in signal shapes due to the presence of highly dispersive dielectric medium in the neuron...
Show moreExtracellular neuroelectronic interfacing has important applications in the fields of neural prosthetics, biological computation and whole-cell biosensing for drug screening and toxin detection. While the field of neuroelectronic interfacing holds great promise, the recording of high-fidelity signals from extracellular devices has long suffered from the problem of low signal-to-noise ratios and changes in signal shapes due to the presence of highly dispersive dielectric medium in the neuron-microelectrode cleft. This has made it difficult to correlate the extracellularly recorded signals with the intracellular signals recorded using conventional patch-clamp electrophysiology. For bringing about an improvement in the signal-to-noise ratio of the signals recorded on the extracellular microelectrodes and to explore strategies for engineering the neuron-electrode interface there exists a need to model, simulate and characterize the cell-sensor interface to better understand the mechanism of signal transduction across the interface. Efforts to date for modeling the neuron-electrode interface have primarily focused on the use of point or area contact linear equivalent circuit models for a description of the interface with an assumption of passive linearity for the dynamics of the interfacial medium in the cell-electrode cleft. In this dissertation, results are presented from a nonlinear dynamic characterization of the neuroelectronic junction based on Volterra-Wiener modeling which showed that the process of signal transduction at the interface may have nonlinear contributions from the interfacial medium. An optimization based study of linear equivalent circuit models for representing signals recorded at the neuron-electrode interface subsequently proved conclusively that the process of signal transduction across the interface is indeed nonlinear. Following this a theoretical framework for the extraction of the complex nonlinear material parameters of the interfacial medium like the dielectric permittivity, conductivity and diffusivity tensors based on dynamic nonlinear Volterra-Wiener modeling was developed. Within this framework, the use of Gaussian bandlimited white noise for nonlinear impedance spectroscopy was shown to offer considerable advantages over the use of sinusoidal inputs for nonlinear harmonic analysis currently employed in impedance characterization of nonlinear electrochemical systems. Signal transduction at the neuron-microelectrode interface is mediated by the interfacial medium confined to a thin cleft with thickness on the scale of 20-110 nm giving rise to Knudsen numbers (ratio of mean free path to characteristic system length) in the range of 0.015 and 0.003 for ionic electrodiffusion. At these Knudsen numbers, the continuum assumptions made in the use of Poisson-Nernst-Planck system of equations for modeling ionic electrodiffusion are not valid. Therefore, a lattice Boltzmann method (LBM) based multiphysics solver suitable for modeling ionic electrodiffusion at the mesoscale neuron-microelectrode interface was developed. Additionally, a molecular speed dependent relaxation time was proposed for use in the lattice Boltzmann equation. Such a relaxation time holds promise for enhancing the numerical stability of lattice Boltzmann algorithms as it helped recover a physically correct description of microscopic phenomena related to particle collisions governed by their local density on the lattice. Next, using this multiphysics solver simulations were carried out for the charge relaxation dynamics of an electrolytic nanocapacitor with the intention of ultimately employing it for a simulation of the capacitive coupling between the neuron and the planar microelectrode on a microelectrode array (MEA). Simulations of the charge relaxation dynamics for a step potential applied at t = 0 to the capacitor electrodes were carried out for varying conditions of electric double layer (EDL) overlap, solvent viscosity, electrode spacing and ratio of cation to anion diffusivity. For a large EDL overlap, an anomalous plasma-like collective behavior of oscillating ions at a frequency much lower than the plasma frequency of the electrolyte was observed and as such it appears to be purely an effect of nanoscale confinement. Results from these simulations are then discussed in the context of the dynamics of the interfacial medium in the neuron-microelectrode cleft. In conclusion, a synergistic approach to engineering the neuron-microelectrode interface is outlined through a use of the nonlinear dynamic modeling, simulation and characterization tools developed as part of this dissertation research.
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Date Issued: 2012
Identifier: CFE0004797, ucf:49718
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0004797

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