• The acetylcholine binding protein of Lymnaea stagnalis as a biosensor and model for ligand gated ion channel proteins

      Harms-Smyth, Abraham Edward (2008-12)
      The structural similarity of the Acetylcholine Binding Protein (AChBP) from Lymnaea stagnalis to the extracellular ligand binding domain of Ligand Gated Ion Channel (LGIC) receptors suggests that the AChBP could be used to mimic cys-loop ligand-gated ion channel (LGIC) receptors. An LGIC mimic could be used as a sensor molecule in a range of biotechnology applications including high throughput drug screening as well as in vivo and in vitro sensing of biologically active compounds. It could also be used as a lead molecule for engineering novel proteins with binding characteristics similar to non-acetylcholine receptor LGIC's. The soluble AChBP is easily expressed and purified and can be produced in reasonably large amounts. This thesis explores the potential for using the AChBP and related proteins as biosensors by evaluating their action on three key medium and high throughput systems: Scintillation Proximity Assay (SPA), Surface Plasmon Resonance (SPR), and Microcantilevers (MC). As a preliminary step to developing a 5-HT₃R-ligand binding protein, by altering the ligand specificity of the AChBP, the interaction of 5-HT₃R ligands with the AChBP is also evaluated. The work presented in this thesis contributes to improved methods of drug design and testing, and to a better understanding of LGIC structure.
    • Addressing feasibility of using microdialysis to monitor adenosine in Arctic ground squirrel (Spermophilus parryii)

      Chi, Bongchu (2005-12)
      Both hibernation and anapyrexia are important physiological adaptations. In addition, there is a strong relationship between survival rate and hypothermia during metabolic trauma. Evidence shows that adenosine might be one of the mediators, both of hibernation and anapyrexia. Adenosine is an inhibitory neuromodulator that suppresses neuronal activity when energy stores are low. This inhibitory activity makes adenosine a good candidate for neuroprotection via the suppression of neural activity during metabolic trauma. The main objective of this study was to investigate the feasibility of using microdialysis to monitor extracellular adenosine in the brains of Arctic ground squirrels (AGS), Spermophilus parryii, throughout hibernation. To investigate this feasibility, the basal level of adenosine concentration in AGS was collected using an in vivo microdialysis method. Samples then were analyzed by high performance liquid chromatography (HPLC). Also, the effects of adding an adenosine transporter inhibitor, S-(p-Nitrobenzyl)-6-thioinosine (NBTI), on extracellular adenosine concentration and body temperature (Tb) of AGS were observed. The results of this study indicate that it is not feasible to observe extracellular adenosine concentrations in AGS during hibernation using these techniques.
    • Air quality degradation in Alaska wilderness areas due to emission changes

      Tran, Trang T.; Cahill, Catherine; Webley, Peter; Newby, Gregory; Simpson, William (2013-08)
      The increasing trends in aerosol concentrations observed by the Interagency Monitoring of Protected Visual Environments (IMPROVE) network in the wilderness areas along the Gulf of Alaska during low insolation periods and in Denali National Park and Preserve (Denali NP) during high insolation periods have raised the concerns about air quality degradation and visibility impairment in these pristine areas. This dissertation aims to investigate the reason for those observed increases in aerosol concentrations in Alaska wilderness areas by performing a series of simulation sets with the Weather Research and Forecasting model coupled with Chemistry (WRF-Chem). These simulation sets use the same meteorological conditions but change the emission scenarios. The model evaluation analysis showed that WRF-Chem performed well in simulating meteorological conditions over Alaska and the North Pacific under both low and high insolation conditions. Performance skill-scores of the WRF-Chem model in simulating aerosol concentrations for the coastal monitoring sites along the Gulf of Alaska were consistent with state-of-the-science air-quality model performance. During low insolation periods, domestic and international ship emissions were the most important contributors to aerosol concentrations in the coastal regions along the Gulf of Alaska. The increases/decreases in ship emissions led to subsequent increases/decreases in aerosol concentrations in the coastal areas along the Gulf of Alaska during low insolation periods. During high insolation periods, in Interior Alaska, the contributions of local wildfire emissions to aerosol concentrations were notable even during the weak Alaska fire activity scenario. Under the strong Alaska fire activity scenario, local wildfire emissions were the dominant source of aerosols in Interior Alaska. The increases in Alaskan wildfire emissions led to significant increases in aerosol concentrations in Interior Alaska. During both low and high insolation periods, Japanese anthropogenic and Siberian wildfire emissions were not important contributors to total aerosol concentrations in all regions of Alaska. Overall in the wilderness areas along the Gulf of Alaska, the increases in aerosol concentrations observed during low insolation periods stemmed from increases in domestic and international ship emissions in the North Pacific. In contrast, the increases in aerosol concentrations observed at Denali NP during high insolation periods stemmed from increases in Alaskan wildfire emissions.
    • Analysis of structure and function of the serotonin type-3 receptor using site directed mutagenesis, structure activity relationship and chimeric constructs

      Suryanarayanan, Asha; Schulte, Marvin K.; Taylor, Barbara; Drew, Kelly; Kuhn, Thomas B. (2005-05)
      The serotonin type-3 receptor (5-HT₃R) is a cation conducting ligand gated ion channel that mediates fast synaptic transmission. The 5-HT₃R belongs to the Cys loop superfamily of ligand gated ion channels that also includes the nicotinic acetylcholine, glycine and GABAa receptors. The 5-HT₃R has been implicated in several processes such as emesis, gastrointestinal motility, drug abuse, alcoholism and nociception. Studies involving the ligand-binding domain will thus aid in development of new drugs that modulate these physiological and pathophysiological processes. The ligand-binding site of this receptor is comprised of six putative loops, viz. loop A-F. The focus of this thesis was to study the interactions of both agonists and antagonists with the 5- HT₃R. Interactions of two agonists, 5-HT and mCPBG, with the loop C region of the receptor were studied employing biochemical and receptor modeling studies. These studies identify novel determinants of 5-HT and mCPBG interactions with the 5-HT3 receptor. Similar studies involving granisetron, a competitive 5-HT₃R antagonist also reveal novel amino acids that interact with this antagonist. In order to further understand antagonist interactions with this receptor, the approach of structure activity relationship (SAR) studies was also employed to study the functional group interactions of lerisetron, a novel 5-HT₃R antagonist. Taken together with data from loops A, B, D and E, these data reveal an emerging picture of ligand interactions with the 5-HT₃R.
    • Analysis of trinitrophenylated adenosine and inosine using capillary electrophoresis-laser induced fluorescence detection and gamma-cyclodextrin

      Stephen, Terilyn Koehler Lawson; Green, Thomas K.; Duffy, Lawrence K.; Drew, Kelly L. (2016-05)
      Adenosine (Ado) and adenine ribonucleotides are essential in cell metabolism and energy production, cellular signaling, and DNA and RNA synthesis. The biosynthesis of these molecules takes place in both the intracellular and extracellular space via transphosphorylation reactions catalyzed by several distinct kinase enzymes like adenylate kinase. Several analytical detection methodologies have been developed to monitor these molecules in biological tissue, including both liquid chromatography (LC) and capillary electrophoresis (CE) techniques. However, many of these methodologies are limited by separation resolution and sample injection volume requirements. This thesis presents a novel capillary electrophoresis-laser induced fluorescence detection (CE-LIF) method with high separation power to analyze Ado and Inosine (Ino), a metabolite of Ado, by derivatization with 2,4,6-trinitrobenzenesulfonic acid to form fluorescent trinitrophenylated complexes of Ado (TNP-Ado) and Ino (TNP-Ino). The development and validation of the CE-LIF method, optimization of the trinitrophenylation reaction, and fluorescence enhancement of TNP-Ado and TNP-Ino with γ-cyclodextrin will be discussed. Detection limits were 1.6 μM for Ado and 4 μM for Ino in rat brain tissue. Large-volume sample stacking (LVSS) was employed to further enhance the sensitivity of the CE-LIF method, with detections limits of 310 nM and 159 nm for Ado and Ino, respectively. The CE-LIF method offers promise for the analysis of Ado, Ino and potentially other adenine ribonucleotides in small volume generating biological experiments like in vivo microdialysis and single cell metabolomics.
    • The application of cavity ring-down spectroscopy to determine nitrate radical concentrations in the atmosphere

      Dick, Eric Michael (2002-05)
      The nitrate radical, NO₃, is a potent atmospheric oxidant. Current NO₃ measurement methods average NO₃ concentrations over several kilometer pathlengths. Recent modeling studies predict that NO₃'s distribution is vertically inhomogenous, causing previous NO₃ observations to not be representative of its atmospheric chemistry. In-situ measurements offer a test of these modeling predictions and a better understanding of NO₃ chemistry. In this thesis, we describe construction of an instrument capable of detecting NO₃ in-situ. This sensor is based upon cavity ring-down spectroscopy. Our initial work demonstrated that CRDS could sensitively detect NO₃. We then built and tested a field prototype during June 2001, successfully detecting NO₃ in the field. CRDS observations were compared to path averaged NO₃ observations. Similar time behavior of both signals indicated that the techniques were observing NO₃. A consistent difference in signal levels indicated that the two techniques were not sampling a homogeneous air mass.
    • The application of synthesized transition metal compounds as paint additives for the preparation of a self-decontaminating coating

      Giles, Spencer Lawrence; Rasley, Brian; Green, Thomas; Wynne, James (2011-08)
      The synthesis of transition metal compounds has been performed to create reactive additives that can be incorporated into commercial grade paints for the purpose of developing a continuous self-decontaminating coating. The coatings were designed to allow for the decontamination of persistent chemical warfare agents: sulfur mustard and organophosphate nerve agents. Since the persistence of these chemicals on surfaces due to their low volatility and long hydrolysis half lives could possibly lead to involuntary exposures, a continuous self-decontaminating coating would provide an alternative approach to eliminate residual chemical warfare agents without further treatment to the painted surface. The synthesized additives include metal oxide nanoparticles,polyoxometallates,andmetalfunctionalizedcyclodextrins. Coatings were formulated with 1% w/w synthesized additives, and tested for their ability to decontaminate chemical warfare agent simulants of sulfur mustard and organophosphate nerve agents. The Dawson structure polyoxometallate α2- K8P2W17O61(Ni2+ ∙OH2)∙17H2O coatings exhibited percent reductions of three of the four simulants above 30% with a high of 53% reduction of Malathion in 24 hours. The nickel functionalized gamma cyclodextrin coatings exhibited percent reductions for all simulants above 28% with a high of 61% reduction of Malathion in 24 hours. Overall, coatings enhanced with additives were tested and produced painted surfaces capable of decontaminating chemical warfare agent simulants.
    • Assessment of total mercury and methyl mercury in selected subsistence fish in Western Alaska

      Zhang, Xiaoming; Duffy, Lawrence K.; Kelly, John J.; Naidu, Satyanarayan (2001-08)
      Total Hg (THg) and methylmercury (MeHg) were examined in muscle and liver samples of salmon species (Chinook: Oncorhynchus tshawytscha; Chum: O. keta; Sockeye: O. nerka; Coho: O. kisutch) and freshwater fish species (Pike: Esox lucius; Grayling: Thymallus arcticus; Whitefish: Caregonus nelsoni) collected in 1999 and 2000 from the Western Alaska rivers (Yukon, Kuskokwim, Nushagak and Kvichak). The THg in salmon muscles has a mean value of 62 ng/g (ww). In Pike muscles, THg has a mean value of 879 ng/g. The mean concentrations of THg in Grayling and Whitefish muscle are 153 ng/g and 32 ng/g respectively. In salmon muscle and liver the MeHg levels constitute 77% and 62% of the THg levels, respectively. In Pike muscle the MeHg levels constitute 100% of the THg levels. A significant correlation between Hg levels and fish length was found. Calculated consumption limits indicate that children may consume 0.05-1.5 kg of fish per month, depending on the species consumed. The study suggests that, from 1979 to 1998, nearly 21 kg of MeHg was transported by Sockeye salmon to the Alaskan rivers of the Bering Sea east coast.
    • Atp-Dependent Chromatin Remodeling Complexes In Xenopus Development

      Brown, Elvin E.; Krebs, Jocelyn E.; Drew, Kelly (2010)
      A central question in the study of vertebrate development is how to account for the exquisite interplay of genes within differentiating cells and of groups of cells as they create the organs of the vertebrate embryo. Recently it has become clear that gene regulation by epigenetic processes adds a formerly unappreciated level of complexity to the regulatory network of development. One form of epigenetic gene regulation is embodied in ATP-dependent chromatin remodeling complexes, which use the energy of ATP hydrolysis to alter the interactions of DNA and histones. Chromatin remodeling complexes can both promote and repress expression of a gene at the appropriate time and place in vertebrate development. The list of their known roles in development is long and growing. Here I have studied the developmental role of CHRAC17, a subunit of the CHRAC and ATAC complexes, by visualizing its expression and by ablating CHRAC17 function in Xenopus laevis embryos. Whole mount in situ hybridization localized CHRAC17 expression to the neural tube, cranial placodes, and myotomes. Loss of CHRAC17 function following injection of embryos with CHRAC17-specific morpholino oligonucleotides resulted in abnormal development in the neural tube, eyes, notochord, and pharyngeal pouches, underlining the critical importance of CHRAC17 function in Xenopus development. Similarly, ablating the function of CHD4, the ATPase motor of the NuRD chromatin remodeling complex, resulted in severe developmental abnormalities in early Xenopus development.
    • The behavior of tellurium during copper ore processing at the American Smelting and Refining Company (Tucson, AZ)

      Josephson, Amy E.; Hayes, Sarah M.; Newberry, Rainer J.; Trainor, Thomas P. (2016-08)
      Essentially all tellurium (Te), an element used in solar panels and other high technology devices, is recovered as a byproduct of copper mining. Recent increases in demand have sparked questions of long-term supplies of Te (crustal abundance ~3 μg∙kg-1). As part of a larger study investigating Te resources, this project examines the behavior of Te during Cu ore mining, smelting, and refining at the American Smelting and Refining Company (Tucson, AZ) as a first step toward optimizing Te recovery. Mass balance calculations estimate that only 4 ± 1% of the Te in the ore reports to the Cu anodes, while 60 ± 30%, 0.8 ± 0.2% and 5.8 ± 0.4% is lost in the tailings, slag, and dust, respectively. The uncertainties reported are the standard deviation of analytical measurements, but due to heterogeneity of Te distribution in the ore, the actual uncertainty is likely higher. Microprobe data shows that Te in the concentrate is mainly present as telluride minerals, but substitution into sulfides most likely also occurs. X-ray fluorescence (XRF) mapping showed that Te is collocated with S in the raw anode slimes, pressed anode slimes, and doré furnace soda slag. X-ray absorption spectroscopy (XAS) was used to examine Te speciation in anode slimes. It was found that Te oxidizes during the Cu ore smelting process, with 44% Te4+ in the flash furnace SO₂ filter. Te also showed 32% Te4+ in the raw and pressed anode slimes. The doré furnace soda slag and dust filter showed the most oxidation of Te at 57% Te4+ and 60% Te6+ respectively. These results indicate several points in the extraction process that could be examined further to determine if additional Te might be recovered from the overall process.
    • Benzene as an indoor air pollutant in Fairbanks, Alaska

      Ricker, Janet Elaine (2000-08)
      Benzene is a known carcinogen found in gasoline, automobile exhaust, cigarette smoke, and organic solvents. Previous studies suggest that sources of chemicals within the home are the major factors influencing personal exposure to benzene. Indoor air was sampled for benzene in order to determine the concentrations present in Fairbanks homes, and to indentify what factors might be associated with higher concentrations. Sampling sites were limited to homes with attached garages and with no smokers in the household. A wide range of benzene concentrations was observed in the eight homes sampled. The highest concentration was about 70 ppbv, and the concentration in most homes was at or above 4 ppbv. The primary source of benzene appears to be gasoline, most likely from small engines (such as lawnmowers) stored in the attached garage. More sites, including homes with attached garages that contain various numbers of small engines, need to be sampled to confirm these conclusions.
    • Bioanalytical Development Of Charged Cyclodextrin Capillary Electrokinetic Chromatography And Microperfusion Sampling To Study Endogenous D-Serine And L-Glutamate Efflux In Brain

      Kirschner, Daniel L.; Greene, Thomas (2009)
      A multitude of studies have revealed specific biological mechanisms that contribute to D-amino acid action and regulation in the mammalian central nervous system. The remarkable increase in our understanding of D-amino acid function and distribution in mammals is in many ways a result of the development of sensitive enantioselective separation strategies that allow for quantification in real biological samples. In capillary electrokinetic chromatography (cEKC) the most powerful chiral resolving agents are anionic cyclodextrins (CDs), yet these have not previously been investigated for chiral bioanalysis of amino acids. The focus of this dissertation research was to investigate for the first time the feasibility of and application of anionic cyclodextrins as resolving agents in bioanalytical chiral separations of amino acids. This dissertation encompasses (1) the development of a new bioanalytical separation utilizing capillary electrophoresis laser induced fluorescence (CE-LIF) with sulfated-beta-cyclodextrin for analysis of D-serine (D-ser) and L-glutamate (L-glu) in mammalian brain, (2) the first synthesis and characterization of 6 members of a new family of single isomer sulfoalkyl cyclodextrins, (3) initial studies on chiral analysis of amino acids using single isomer sulfoalkyl CDs, and (4) development and application of a novel microperfusion sampling approach for acute brain slices and coupling of this method to the developed chiral CE-LIF for studying magnitude and timing of D-ser and L-glu efflux from acute hippocampus in response to modeled cerebral ischemia. The results of these studies demonstrate that (1) anionic CDs are powerful chiral selectors for amino acids and can be applied for sensitive bioanalysis of D-amino acids including D-ser, D-glu, and D-asp in brain samples; (2) single isomer sulfoalkyl CDs can be synthesized by regioselective reaction chemistry; (3) single isomer sulfoalkyl CDs are excellent resolving agents for amino acid analysis and may be valuable for bioanalytical chiral applications; and (4) microperfusion sampling coupled to CE-LIF can be used to analyze dynamic changes in the magnitude and timing of neurochemical efflux from single acute hippocampus slices exposed to modeled ischemia. Results of these latter studies suggest that D-ser and L-glu efflux occurs simultaneously in acute hippocampus with similar timing but differing magnitudes.
    • Biochemical study of a PSI-LHCI complex and molecular study of fcp genes in the diatom Cylindrotheca fusiformis

      Zhang, Hua; Plumley, Gerard; Boyer, Bert; Duffy, Lawrence K. (2002-08)
      A photosystem I-light-harvesting complex I (PSI-LHCI) was isolated from oxygen-evolving thylakoids of the diatom Cylindrotheca fusiformis. The circular dichroism (CD) spectrum of the complex resembled the photosystem I (PSI) complex of green plants. A single 77K fluorescence emission was observed at 715 nm. The excitation spectrum confirmed that both chlorophyll c and carotenoids were energetically coupled to chlorophyll a. The complex contained PSI reaction center proteins (PsaA/B, the PSI accessory subunit PsaC, and nine light-harvesting complex (LHC) apoproteins including an 18kD and a 17.5kD protein. Photosystem II core polypeptides were not detected by immuno- or silver staining. Taken together, the CD, fluorescence, and protein data indicate that at least nine LHC apoproteins can be specifically associated with PSI in this diatom. Twenty fcp gene sequences that encode fucoxanthin-chlorophyll a/c light-harvesting proteins (FCPs) and three encoded proteins that are similar to a 17.5kD FCP, but it was not possible to conclusively confirm that any of these genes encode proteins associated LHCI.
    • Biologically relevant secondary metabolites of Vaccinium uliginosum: bioassay-directed natural products identification of anti-neuroninflammatory agents in the Alaska bog blueberry

      McGill, Colin (2010-05)
      Dietary blueberry supplementation has demonstrated numerous health benefits including improved learning and memory in aging and neurodegenerative models, neuroprotection from ischemic events, anti-diabetic properties, and modulation of multiple inflammatory cascades. Despite previous research on antioxidant components prevalent in blueberries, no adequate explanation for a molecular mechanism for the benefits of blueberry supplementation has been proposed. Vaccinium uliginosum, the Alaska bog blueberry, possesses higher concentrations of antioxidant components than commercial varietals, and exhibits a greater oxygen radical scavenging capacity, making it an excellent candidate for the identification of biologically relevant secondary metabolites. An approach of bioassay-directed natural products identification was utilized to identify compounds in the Alaska bog blueberry responsible for the inhibition of both a magnesium-dependent neutral sphingomyelinase and NADPH oxidase in TNF-[alpha]-induced SH-SY5Y human neuroblastomas. Five relevant metabolites were identified: ß-sitosterol (1), ursolic acid (2), 3-0-(4-hydroxyphenylcarboxylic acid) 4-0-(ß-D-glucopyranosyl) gallic acid (3), malic acid (4), and 2,3-dihydroxybutane-1,2,3,4-tetracarboxylic acid (5). Neither compounds 3 or 5 had been previously described as a natural product in the literature. The identification of these compounds in the Alaska bog blueberry provides new explanations as to the benefits of blueberry consumption and offers new avenues of research for nutraceutical treatment of neuroinflammation.
    • Biophysical characterization of class II major histocompatibility complex (MHCII) molecules

      Osan, Jaspreet Kaur; Ferrante, Andrea; Kuhn, Thomas; Podlutsky, Andrej; Chen, Jack (2020-05)
      Class II Major Histocompatibility Complex (MHCII) molecules are transmembrane glycoproteins expressed on the surface of antigen-presenting cells (APCs). APCs engulf pathogens and digest pathogenic proteins into peptides, which are loaded onto MHCII in the MHCII compartment (MIIC) to form peptide-MHCII complexes (pMHCII). These pMHCII are then presented to CD4+ T cells on the surface of APCs to trigger an antigen-specific immune response against the pathogens. HLA-DM (DM), a non-classical MHCII molecule, plays an essential role in generating kinetically stable pMHCII complexes which are presented to CD4+ T cells. When a few peptides among the pool of the peptide repertoire can generate the efficient CD4+ T cell response, such peptides are known as immunodominant. The selection of immunodominant epitopes is essential to generate effective vaccines against pathogens. The mechanism behind immunodominant epitope selection is not clearly understood. My work is focused on investigating various factors that help in the selection of immunodominant epitopes. For this purpose, peptides derived from H1N1 influenza hemagglutinin protein with known CD4+ T cell responses have been used. We investigated the role of DM-associated binding affinity in the selection of immunodominant epitopes. Our analysis showed that the presence of DM significantly reduces the binding affinity of the peptides with low CD4+ T cell response and inclusion of DM-associated IC50 in training MHCII algorithms may improve the binding prediction. Previous studies have shown that there is an alternate antigen presentation depending on antigen protein properties. Here, we showed that the immunodominant epitope presentation is dependent on the pH and length of the peptides. To study the MHCII in its native form, we assembled full-length MHCII in a known synthetic membrane model known as nanodiscs. We noted that, based on the lipid composition, assembly of the MHCII differs. Preliminary binding studies with this tool showed that there might be a difference in the binding based on the type of the nanodisc. Collectively, our results showed that the immunodominant epitope selection is a complex process that is driven by various biochemical features.
    • Central Nervous System Regulation Of Metabolic Suppression In Arctic Ground Squirrels

      Jinka, Tulasi Ram; Drew, Kelly L. (2010)
      The main focus of this dissertation is central nervous system regulation of metabolic suppression in hibernating mammals in general, and the Arctic ground squirrel (Urocitellus parryii) as a model for seasonal hibernation. Hibernation is a unique physiological, morphological, and behavioral adaptation to overcome the periods of resource limitation. Metabolic suppression seen in torpor during hibernation has several biomedical applications. A multitude of studies have revealed the role of the central nervous system in regulating hibernation, including a role for neurotransmitters and neuromodulators. Previous studies have shown that the neuromodulator adenosine mediates altered thermoregulation during induction of torpor in facultative hibernators, but it is not clear how adenosine influences torpor in seasonal hibernators. The main focus of the current project was to test the hypothesis that a seasonal change in purinergic signaling is necessary for the onset of spontaneous torpor in the Arctic ground squirrel. My dissertation reports that adenosine meets all of the necessary requirements for an endogenous mediator of torpor in the hibernating Arctic ground squirrel. A progressive increase in sensitivity to adenosine A 1 receptors mediated signaling defines the seasonal transition into the hibernation phenotype. I show that adenosine A1 receptor activation is necessary and sufficient to induce torpor in the Arctic ground squirrel. Glutamate is an excitatory neurotransmitter which is widely studied in hibernation research. My dissertation demonstrates that N-methyl-D-aspartate type glutamate receptors, located in the periphery or circumventricular organs, are involved in inducing arousal from torpor in the hibernating Arctic ground squirrel. This dissertation also presents evidence that dietary restriction sensitizes adenosine A1receptors in rats through an increase in surface expression in thermoregulatory regions of the brain (hypothalamus). This contributes to the decline in body temperature and respiratory rate in animals subjected to a restricted diet, which mimics a torpor-like effect.
    • Ceramide Metabolism Regulates A Neuronal Nadph Oxidase Influencing Neuron Survival During Inflammation

      Barth, Brian M. (2009)
      Inflammation is a major component of acute and chronic pathologies of the central nervous system, including psychiatric disorders. Microglia respond to pathogens, injury, and toxins by secreting inflammatory mediators including pro-inflammatory cytokines in an event known as neuroinflammation. This thesis research investigated a link between neuroinflammation and oxidative stress, and ultimately neurodegeneration. The cytokine tumor necrosis factor alpha was shown to stimulate a neuronal NADPH oxidase (NOX), specifically by stimulating the production of ceramide and ceramide-1-phosphate via Mg 2+-neutral sphingomyelinase (Mg2+-nSMase) and ceramide kinase. Intriguingly, glucosylceramide blocked NOX activation, linking ceramide neutralization directly to a decline in oxidative stress. Most importantly, NOX activity interfered with actin and sphingosine kinase-1 via oxidation, demonstrating a positive and detrimental feedback mechanism that impedes neuronal survival pathways. Interestingly, crude extracts from wild Alaskan bog blueberries showed the ability to interfere with Mg2+-nSMase, demonstrating a specific neuroprotective property of the berry. Altogether, this thesis research defined a key neuronal pathway linking inflammation to oxidative stress via ceramide metabolism, potentially allowing for future therapeutic development to improve neuronal function and survival.
    • Characterization And Interpretation Of Volcanic Activity At Redoubt, Bezymianny And Karymsky Volcanoes Through Direct And Remote Measurements Of Volcanic Emissions

      Lopez, Taryn M.; Cahill, Catherine; Dehn, Jonathan; Newberry, Rainer; Simpson, William; Werner, Cynthia (2013)
      Surface measurements of volcanic emissions can provide critical insight into subsurface processes at active volcanoes such as the influx or ascent of magma, changes in conduit permeability, and relative eruption size. In this dissertation I employ direct and remote measurements of volcanic emissions to characterize activity and elucidate subsurface processes at three active volcanoes around the North Pacific. The 2009 eruption of Redoubt Volcano, Alaska, produced elevated SO2 emissions that were detected by the Ozone Monitoring Instrument (OMI) satellite sensor for over three months. This provided a rare opportunity to characterize Redoubt's daily SO2 emissions and to validate the OMI measurements. Order of magnitude variations in daily SO2 mass were observed, with over half of the cumulative SO2 emissions released during the explosive phase of the eruption. Correlations among OMI daily SO2 mass, tephra mass and acoustic energies during the explosive phase suggest that OMI data may be used to infer eruption size and explosivity. From 2007 through 2010 direct and remote measurements of volcanic gas composition and flux were measured at Bezymianny Volcano, Kamchatka, Russia. During this period Bezymianny underwent five explosive eruptions. Estimates of passive and eruptive SO2 emissions suggest that the majority of SO2 is released passively. Order of magnitude variations in total volatile flux observed throughout the study period were attributed to changes in the depth of gas exsolution and separation from the melt at the time of sample collection. These findings suggest that exsolved gas composition may be used to detect magma ascent prior to eruption at Bezymianny Volcano. Karymsky Volcano, Kamchatka, Russia, is a dynamic volcano which exhibited four end-member activity types during field campaigns in 2011 and 2012, including: discrete ash explosions, pulsatory degassing, gas jetting, and explosive eruption. These activity types were characterized quantitatively and uniquely distinguished using a multiparameter dataset based on infrasound, thermal imagery, and volcanic emissions. These observations suggest a decoupling between exsolved volatiles and magma at depth. I propose that variations in magma degassing depth influence the flux and proportions of decompression- and crystallization-induced degassing, as well as conduit permeability, can explain the variations in volcanic activity.
    • Characterization of host-pathogen interactions in two model pathogens: Francisella tularensis and simian virus SV40

      Smith, Lisa K.; Hueffer, Karsten; Khun, Thomas; Runstadler, Jonathan (2012-05)
      We sought to elucidate the molecular mechanisms of host-pathogen interaction. The bacterium Francisella tularensis and simian virus SV40 represent two ideal model systems. Francisella tularensis is a facultative intracellular bacterium known to dampen the host immune response to infection. The Francisella pathogenicity island (FPI) encodes a cluster of 19 genes essential for full virulence and the observed change in immune response. We investigated the role of two FPI encoded proteins, PdpC and PdpD, on immune response. While both proteins affect a change, the effect of PdpD is more pronounced, and appears to play a role in modulation of host immune responses. SV40 is a DNA polyoma virus that targets GM1 receptors for entry into cells. The GM I receptor is localized to cholesterol-rich membrane microdomains, termed lipid rafts. Disruption of lipid rafts using the cholesterol chelator methyl-P-cyclodextrin prevents SV40 entry into cells. We investigated whether natural product alternatives would similarly disrupt lipid raft integrity and prevent viral entry. The triterpenoid ursolic acid, present in many plants, has been shown to possess antimicrobial properties and was used to treat cells prior to infection with SV40. We found ursolic acid to have no effect on the viral infectivity of SV40.
    • Characterization of the adenosine A1 receptor in summer and winter Arctic ground squirrels

      Carlson, Zachary A.; Drew, Kelly; Rasley, Brian; Kuhn, Thomas (2014-12)
      Hibernation is an adaptation that allows the Arctic ground squirrel (Urocitellus parryii) to survive the harsh arctic winter. Recently the activation of the Adenosine A1 receptor (A₁AR) has been shown to be necessary for entrance into hibernation during the winter but not summer season. In the current study we characterize the A₁AR in the forebrain, hippocampus and hypothalamus of summer and winter AGS. We also tested the hypothesis that increased A₁AR agonist efficacy is responsible for increased sensitization of the A₁AR during the winter season. The resulting ³⁵S-GTPγS binding data indicated an increase in agonist potency during the winter season in all three brain regions. A plausible explanation of our results is that increased potency in the forebrain during the winter season is due to an increase in efficacy as indicated by a greater number of receptors in the high affinity state. In addition ³⁵S-GTPγS binding, [³H] DPCPX saturation and competition assays establish for the first time pharmacological characteristics such as EC⁵⁰, Kd, Kilo and Kihi in AGS brain.