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Real-time Measurement of Glial Progenitor Chemotactic Migration
Year of Dissertation:
Gliomas are the most commonly diagnosed form of central nervous system tumors, occurring primarily in adults. Like many malignant cancers, gliomas pathologically exhibit very aggressive spreading and lead to an average diagnosed survival expectancy of twelve months. This prognosis is due in large part to the uncontrolled division and migration of malignant tumor cells within healthy brain, which makes complete surgical resection impossible. Gliomas are known to contain numerous genotypic and phenotypic alterations that affect cell proliferation and survival. Previous research has indicated that both gliomas and their precursor cells exhibit distinct migration patterns in brain tissue, which may be induced by specific cytokines and their concentration gradients. Here, we investigated the migration of four brain tumor cell (BTCs) lines (U-87 MG; U-251 MG; Daoy; and XFMPDGF) and three RCAS-infected glial progenitor cell (GPCs) populations (GPCLacZ, GPCPDGF, and GPCkRas) toward various growth factors, including but not limited to: EGF, HGF/SF, PDGF-BB, and TGF-α. Mouse neural cultures of nestin expressing glial progenitor cells that have been engineered to display the Avian Leukosis Virus (ALV) target receptor, tv-a, were infected with the RCAS viral vectors RCAS-LacZ, RCAS-PDGF-B, and RCAS-kRas, and are referred to as Ntv-a glial progenitor cells (Ntv-a-GPCs). Upon successful infection, the Ntv-a-GPCs were capable of constitutively producing β-galactosidase, PDGF-B, and kRas, respectively. Additionally, when these viral vectors were injected into the brains of transgenic Ntv-a mouse pups, only the RCAS-PDGF-B viral titer induced gliomas (Dai et al, 2001; Holland et al, 2000). For the purpose of this work it is relevant to note that the Ntv-a cell lines used, with the exception of the XFMPDGF cells, were never injected into the mouse brain. Conversely, the XFMPDGF cell population represents an excised tumor from the brain of a ≤ 12-week-old Ntv-a transgenic mouse having the Ink4a-Arf -/- genetic background. After cranial injection with a RCAS-PDGF-B viral titer, tumors of the oligodendroglioma classification developed within 1-12 weeks in 100% of the injected mice (Tchougounova et al, 2007). We use these oligodendrogliomas as our mouse tumor model. Additionally, Daoy medulloblastomas were used to model the migration of the most common childhood human brain tumor, while the U-87 MG and U-251 MG were used to model glioblastomas, the most common adult brain tumor. The design and characterization of our stand-alone bridged µLane system demonstrated sustained steady-state concentration gradients over 2-3 days while enabling the diffusivity measurement of 0.82 ± 0.01 X 10-6 cm2 / s for Dextran molecules having a molecular weight similar to growth factors examined here. A modified version of this device was then used to examine the migratory response of human MB-derived Daoy, glioblastomas, and neonatal mouse glial progenitors, which has led to the characterization of differential invasion patterns that each cell line utilizes when exposed to various growth factor concentration gradients. This data is fundamental for understanding how BTCs and GPCs use cytokines to communicate with each other and alter cellular functions, specifically migration. Additionally, we demonstrate EGF-induced Akt activation in the migration of MB-derived Daoy cells, and the induced recruitment GPCs towards picomolar concentrations of both HGF and TGF-α. We suggest based on our results that glial progenitors consisting of varying genetic background can migrate in very distinct patterns and may contribute to the selectivity of glioma recruitment/or progenitor sorting.
THE ROLE OF GLUTAMATE IN AXONAL PHYSIOLOGY
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The information within the spinal cord and peripheral nervous system propagates along the nerves in the form of Compound Action Potentials (CAP). Although CAPs were always considered to be steady signals, with constant amplitude and velocity as determined by the conductive properties of the nerves our data show that exogenous glutamate increased the CAP. This increase in CAP was blocked after the addition of the general glutamate receptor antagonist kynurenic acid, the specific glutamate receptor antagonist MK801 and CNQX and prevented when these experiments were performed in a calcium-free medium. The goal of this thesis was to examine the changes in axonal physiology in response to electrical stimulation and to pharmacological manipulation. We found that high frequency stimulation, or addition of exogenous glutamate (100 µM) increases the amplitude of compound action potentials (CAPs) in sciatic nerve preparations. These results were further extended and supported by immunohistochemical experiments showing that axolemma contains glutamate receptors (NMDA, AMPA/kainate and mGluR2), the excitatory amino acid transporter responsible for glutamate uptake (Excitatory Amino Acid Transporter-EAAT), and voltage-gated sodium and calcium channels. Thus, the axolemma of peripheral nerves expresses several proteins important for neuronal communication and modulation of the membrane excitability. Apparently, these proteins embedded into the axonal membrane, can under the influence of electrical stimulation or exogenous glutamate change membrane permeability and ionic conductance leading to an increase in the amplitude of the compound action potentials observed in our experiments. Our results demonstrate of existence of axonal plasticity expressed as a change in the amplitude of the action potential following periods of changed activity accompanied by release of neurotransmitters. Therefore we suggest a mechanism of the process whereby electrical stimulation leads to increased axonal activity and subsequent release of glutamate that through activation of the glutamate receptors results in changes in the amplitude of CAPs. We term this phenomenon as axonal plasticity, which would represent one of the forms of neuronal plasticity. Neuronal plasticity is defined as a treatment-induced change in the neuronal response in spite of unchanged strength of the test stimulation. This observation was long described as a property of central synapses and thought to be the basis of learning (Malenka, 1994). Axonal plasticity , would constitute exclusive property of the axon and could contribute together with synaptic plasticity to modification of the efficiency of neuronal connections. This type of plasticity would be fundamentally different from the synaptic plasticity expressed in CNS in the form of Long-Term Potentiation-LTP (Bliss and Collingridge, 1993), Long-Term Depression-LTD (Dudek et al; 1992), and Spike Timing Dependent Plasticity (STDP) (Markram et al., 1997) which has been intensively investigated for last several decades. We assume that high frequency electrical stimulation induces the release of glutamate from stimulated axons. Subsequent increase in the extracellular glutamate concentration would be responsible for observed increase in CAP. Increase in the amplitude of CAP may be a result of: An increase in the number of activated axons (recruitment), 2) and/or increase in the amplitudes of individual potentials generated by single axons. The mechanisms responsible for each of these changes are very different. In the case of recruitment one can suggest paracrine action of glutamate which released from group of axons would enhance the CAP of their neighbors. The increase in the action potential generated by individual axon could be due to a change in the threshold of this individual axon. Our novel data together with published results clearly indicate that in spite of prevailing notion about "all-or-nothing" property of the action potential, axons and action potentials are capable of conveying the information in an analog manner (Clark and Hausser 2006). Presented results convincingly demonstrate that the amplitude of subsequently generated action potentials can change in a way correlated with the frequency of stimulation, or pharmacological treatment. In both cases the change occurred gradually with each evoked action potential slightly larger than its predecessor. This indicates that the effect was building step by step as the intraaxonal mechanisms have been recruited to contribute to the final effect. We have also observed reduced latency and increased area of CAP after glutamate application. The most obvious explanation for both phenomena would be a recruitment of additional, fast conducting axons which would shorten the latency and increase the area of CAP. Simultaneously this would increase the duration of the entire CAP, as slower conducting axons which contributed to CAP before the treatment would be activated as well.
The effects of estrogen on carrageenan-induced nociceptive behaviors and inflammatory mediators in ovariectomized female mice
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Epidemiological studies have shown that pain perception is sexually dimorphic; females tend to experience greater sensitivity to painful stimuli and more chronic pain compared to males. Researchers believe that this dichotomy is caused by the distinct endocrinological profile of females. 17beta -estradiol has been shown to attenuate inflammatory behaviors in both the formalin and carrageenan (Cg) models of inflammation. Research also shows that estrogen affects many inflammatory mediators, including proinflammatory cytokines and prostaglandins (PG). Estrogen plays an important, yet complicated role, in inflammation, and little is known about the specific biochemical mechanisms involved. The objective of this study is to determine if, similar to rats, estrogen attenuates Cg-induced thermal hyperalgesia by altering cytokine or PG release. To that end, female OVX mice were pretreated with various doses of estradiol and injected with 1% Cg. Paw withdrawal latency was recorded prior to, 1 hour, and 5 hours after Cg-treatment in response to a low, medium, and high heat stimuli. Additional animals were treated with indomethacin, a COX blocker. High doses of estradiol caused an increase in nociceptive responses prior to and subsequent to Cg administration. This increase in these pain behaviors was not directly caused by an increase in proinflammatory cytokine levels or a decrease in anti-inflammatory cytokines levels. However, estradiol caused increases in cytokine levels in the untreated paw. Furthermore, treatment with indomethacin caused an attenuation of hyperalgesia. Additionally, indomethacin negated the difference between estradiol- and vehicle-treated mice, indicating that estrogen may interact with prostanoid synthesis. This effect, however, was not seen in the Cg-treated paw, suggesting that estradiol may be increasing hyperalgesia via another pathway as well. Taken together, our results suggest that estrogen's hyperalgesic effects are partly mediated through cytokine up-regulation and prostanoid synthesis, but the main mechanism of action still needs further investigation.
Registered Sex Offenders in the Community: A Test of Agnew's General Strain Theory
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Over the past two decades, sexual offending and offenders have become a topic of interest among researchers, policymakers, and the public. Since the inception of Registration and Community Notification Laws (RCNLS), researchers have assessed the negative consequences associated with the laws and how they affect sex offenders in the community; however, no study has utilized a criminological framework to do so. Agnew's General Strain Theory, which should be able to account for all crime, suggests that when individuals do not achieve desired goals, have negative stimuli placed on them or positive stimuli taken away, they are more likely to engage in crime. These are conditioned by certain factors, such as coping strategies and self-esteem. This study will synthesize these two distinct fields of research to determine whether the negative consequences of RCNLSs lead to recidivism. In all, surveys were mailed to 4,500 sex offenders with (N=997) in Nebraska, (N=2086) and (N=1417) sex and violent offenders in Kansas and Montana, respectively. These states are similar in population and demographic aspects, though they differ in RCNLSs. Findings lend partial support to GST and suggest that, consistently, anger influences recidivism.
Infrastructure, Production, and Archive: American and Japanese Video Art Production of 1960s and 1970s
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Focusing a study on the infrastructure of artistic production and maintenance opens a space in which to examine the relationship between artistic inspiration and knowledge making, the occurrences and the writing of its history. In the case of the comparative study of the emergence of Japanese and American video art, common artistic technique employed may indicate motivations derived from the technical possibility of the video medium, while the study of infrastructure demonstrates how large-scale funding, formation of archives, the establishment of systems of distribution and channels of education affect the emergence and development of video art as a genre. This thesis analyzes the synchronous and productive rise of video art in the U.S. and Japan from mid-1960s through mid-1970s, while problematizing the systems of knowledge formation and cultural maintenance that produced a different reception of the histories. In reviewing the structure of cultural maintenance today, the digital field inspires an exploration of new avenues for models of infrastructure that is alternative to traditional institutional frameworks. The digital platform as a pedagogical tool, research resource, and discussion forum allows cross-linguistic understanding, networked scholarship, and open accessibility, suggesting an ideal method for knowledge making. Moreover, this thesis suggests the digital platform as a potential catalyst for a collaborative administration of archive and preservation of Japanese experimental film and video, proposing an alternative method of knowledge formation and cultural maintenance moving forward.
The Amphibious Public: A historical geography of municipal swimming and bathing New York City, 1870 - 2013
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Earth & Environmental Sciences
Since 1870, the city of New York has engaged in a project of building and maintaining enclosed sites for municipal bathing, including building floating `river baths' (1870 - 1942), indoor municipal baths (1901 - 1975), eleven enormous outdoor pools built with WPA funds (1936 - present), and outdoor pools of various sizes built under the Lindsay administration (1968 - present). This dissertation explores the changing rationale, over almost 150 years, for the municipal construction of public bathing places in New York City, and the ways in which the physical structures have taken on new social goals, meanings and ideals, both for patrons and for agents of municipal government over time. Each bathhouse and pool is a physical site that belongs to an infrastructural network, and is also bound up in its relationship to reigning ideas about what public space should encompass and for whom it should provide. Throughout, water has been attributed particular characteristics in order to mediate social life in public space, through programs of building, teaching and regulating. These are theorized in terms of public space and the public life that bring them together as a material, technological, symbolic whole. The municipal bathing project has resulted in corporeal publics over time, which produce public social life through the bodies of users, both real and ideal, through infrastructures that integrate materials, water, capital and political will. Contests over who belongs to the corporeal public and how it should be managed, based on race, gender and sexuality, class, and age, are mediated through shifting notions of hygiene and wellness in the urban setting. Research methods include archival research in New York City since 1870, including municipal records, other local archives, newspaper sources, and secondary histories; observation (and some participation) and interviews with the Harlem Honeys and Bears, an African-American senior citizen synchronized swim team; and comparative ethnography of outdoor pools in the summer, including extended participant observation at Kosciuszko Pool and McCarren Pool in Brooklyn, as well as interviews with Parks Department officials.
Building the New American Nation: The U.S. Army and Economic Development, 1787-1860
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This dissertation examines the Army's integral role in the early American political economy. Notwithstanding its small size, the Army proved to be a powerful instrument for promoting economic expansion and guiding the pattern and direction of development. The Army spurred development through two lines of activity: first, the traditional application of coercion and, second, by providing public goods that neither private actors nor state governments could supply. Considering the Army leads me to reconceptualize the early American state as a bifurcated entity: a state of the periphery, dominated by the Army, and a state of the center, in which the Army still influenced economic development but other public institutions also performed key development functions.
Performance Effects of Computer-Based Multitasking Behavior
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This research examines multitasking from the perspective of human-computer interaction (HCI). Multitasking is defined as the performance of multiple tasks concurrently. In a computer-based environment, users generally switch between multiple computer-based tasks either due to a personal decision to break from the current task or due to an external interruption, such as an electronic notification. This dissertation describes an in-depth empirical study, using a laboratory setting with different numeric, verbal, and visual computer-based tasks. Six hundred and thirty six subjects were randomly assigned into three conditions: discretionary multitasking, where participants were allowed to decide when and how often to switch tasks, forced multitasking, where participants were forced to switch tasks at certain allotted times, and non-multitasking, where participants performed the tasks sequentially and were not allowed to multitask. In order to investigate performance effectiveness (accuracy) and performance efficiency (productivity), participants' overall accuracy and productivity scores were compared across conditions. The results suggest that during difficult tasks, subjects who were forced to multitask had the lowest accuracy. In addition, those subjects in the forced multitasking condition who felt the primary task was difficult had lower accuracy than those who felt the task was easy. This was not true in the other two conditions. Receiving interruptions during a difficult task impacted not only their primary task, but their secondary tasks as well. In the discretionary multitasking condition, the more subjects decided to multitask, the lower their accuracy scores. In fact, an additional analysis revealed that high multitaskers not only performed worse than low and medium multitaskers in the discretionary condition, but actually had the worst performance than subjects in any other condition. Medium multitaskers, however, had the highest productivity scores. While multitasking in that case was considered the best in terms of efficiency, it was not true in terms of effectiveness. Therefore, discretionary multitasking gives the illusion of high performance. Furthermore, this study also explored why people chose to multitask and the impact that had on performance. The results of this study can assist HCI researchers in developing a more comprehensive understanding of user multitasking which can lead to better interface designs.
The Role of Striatal Neuropeptides on Glutamate and Methamphetamine-Induced Neurotoxicity in the Murine Brain
ABSTRACT THE ROLE OF STRIATAL NEUROPEPTIDES ON GLUTAMATE AND METHAMPHETAMINE-INDUCED NEUROTOXICITY IN THE MURINE BRAIN by Lauriaselle Afanador Adviser: Dr. Jesus A. Angulo
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The rising worldwide epidemic in addiction to methamphetamine (METH) and the well-documented neurological detriments it causes emphasizes the importance of elucidating the mechanisms by which METH causes widespread and prolonged damage. Also, METH's pathophysiology resembles a number of neurodegenerative diseases. Therefore a better understanding of the mechanisms involved would provide more effective therapeutic targets for the treatment of these neurological disorders.
METH toxicity is a complex interplay of various factors however a number of necessary components have been identified such as dopamine overflow (DA), glutamate signaling, and oxidative stress. Although METH-induced DA overflow is the initiating event, it is not the direct cause of damage. Oxidative stress is thought to be the mediator of METH toxicity and nitric oxide (NO) as a contributor.
We have found that substance P (SP) exacerbates METH-induced NO. Inhibition of SP signaling mitigated NO synthesis and conferred protection. Considering the role SP is playing in METH toxicity we wanted to investigate the role that other striatal neuropeptides play in these events, notably the inhibitory peptides neuropeptide Y (NPY) and somatostatin (SST).
We hypothesized that SP is augmenting NMDA signaling and thus magnifying NO production. Whereas NPY and SST would serve as a counteracting force thus dampening oxidative stress and conferring protection. Overall, our data demonstrated that SP does augment NMDA signaling as inhibition of the neurokinin-1 receptor (NK-1R) decreased NMDA-induced striatal cell loss. We found that SP was potentiating NMDA-induced NO production. Although the predominant source of NO was the inducible form of nitric oxide synthase (NOS).
In support of our hypothesis, NPY and SST proved to attenuate NO. Also, they were protective from METH-induced cell death although SST failed to protect DA terminals. However, an agonist for the NPY-Y2 receptor was successful in maintaining DA terminal viability. Of interest is that neither NPY nor SST modulated NMDA-induced NO or cell loss suggesting that their protective mechanism does not include modulation glutamate signaling within the striatum.
PUBLIC EDUCATION IN THE UNITED STATES: THE PRODUCTION OF A NORMATIVE CULTURAL LOGIC OF INEQUALITY THROUGH CHOICE
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"Public Education in the United States: The Production of a Normative Cultural Logic of Inequality Through Choice" is a historically informed ethnography that examines how choice emerged in the post-Brown v. Board of Education era as a key principle of reform and management in public education and became central to how rights, freedom, and citizenship were structured, constrained, and imagined. Scholarship within education studies has identified choice as a predicament of neoliberalism as privatization. Yet as my research examines, the inequalities associated with privatization mechanisms like charter schools are an exaggerated indexical representation of a much deeper and older problem. My research extends the historical trajectory through which we understand neoliberal restructuring and traces the ideological and material contours of a post-Brown realignment between the state, the structuring of rights, and the market--a realignment that extends beyond the realm of the private. This dissertation is based on eighteen months of ethnographic fieldwork in New York City's Community School District (CSD) 3, one of the most racially and economically diverse districts in the nation's largest school system. CSD 3 is also one of the most segregated districts in New York City and one of the districts that provides the most choice-based programs and policies. In my ethnography, I examine the ways that low-income and middle-income parents navigate and negotiate selecting a public non-charter elementary school for their child. I trace how situated claims to universal rights as choices facilitate the continuance of a tiered citizenship and the production of what I term a "normative cultural logic of inequality." My research interrogates how this logic narrates inequality in education as resulting from "bad" yet "fair" choices that are qualified by a lack of individual initiative, informed decisions, and capacities of parental care. My findings suggest that rather than these explanations, the differential accumulation of living in what Ruth Wilson Gilmore has termed the "forgotten" or "abandoned places" of a racial state are central to understanding how similar desires---of wanting the very best for one's child---result in very different outcomes.