Charles Loren Buck, Ph.D. - US grants

Because of their high latitude distribution, arctic ground squirrels are naturally exposed to the most extreme seasonal changes in day length and temperature of any hibernator. Arctic ground squirrels have unique adaptations that allow them to thrive under arctic conditions. The proposed work will resolve important questions in hibernation physiology using integrated approaches including respirometry for measuring rates of metabolism and respiratory exchange, stable isotope analysis for assessing changes in metabolic fuel use and body composition, and genomic approaches for a molecular understanding of hibernation in extreme conditions. This project seeks to 1) elucidate which metabolic fuels (fat, carbohydrate, protein) arctic ground squirrels draw from during arctic hibernation, 2) quantify the expression of genes for enzymes involved in metabolic fuel selection, 3) identify lower thermal limits to hibernation at sub-zero body temperatures, and 4) continue monitoring body and hibernacula temperatures in a natural population on the North Slope of Alaska. These field measures will be related to changes in body condition over winter, annual timing of hibernation and other life history events, and effects on reproduction and persistence of this population in the context of climate change. It is anticipated that arctic ground squirrels will hibernate in ambient temperatures as low as -40 degrees C, and that they will increase reliance on protein to fuel metabolism with decreasing ambient temperatures. At low temperatures, regulation of metabolic fuel selection by arctic ground squirrels will be accomplished via increased expression of genes that promote protein metabolism and gluconeogenesis (synthesis of sugar). Because hibernators have been used as models for cerebral ischemia (reduced blood flow to the brain), traumatic head injury, and hypothermia, investigations of the physiology and genomics of hibernation have biomedical implications. Moreover, this research will help us better understand organismal responses to a changing climate in the sensitive arctic ecosystem. This project is interdisciplinary and will provide training, in both field and lab settings, to 1 post-doc, 2 PhD students and undergraduates in molecular, isotopic, physiological and ecological techniques. It is international with a collaborating scientist from the Shanghai Institutes of Biology in China. Results of this research will be disseminated through publications and presentations at regional, national and international meetings and local interpretive displays.

0923571: MRI Acquisition: Detecting and monitoring changes in the arctic using stable isotope techniques

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5).

The University of Alaska at Anchorage has been awarded a grant to purchase two stable isotope mass spectrometers plus a tunable diode laser system to quantify the processes of change in high latitudes. The instruments will be used to measure the natural abundance of isotopes of carbon, nitrogen, hydrogen and oxygen (C-13, N-15, H-2 (deuterium), O-18), in studies involving plants, soils, water, and animals. The instruments are also capable of measuring samples that have been enriched with these isotopes, in labeling experiments designed to study plant water sources and field metabolic rates of small mammals. The intellectual merit and the scientific motivation for acquiring this instrumentation package derives from the fact that the Arctic is changing rapidly, and there is a great need for using tools that are integrative in time and space, and tools that can unravel the complexity of linkages between the biosphere-atmosphere and aquatic systems and the intricate nature of food webs. The instruments will facilitate quantification of the consequences of vegetation change, of shifting patterns in the field metabolic rates of small mammals, of the water sources of vegetation as permafrost melts, quantification of precipitation geochemistry at local to regional scales, and food web ecology of marine mammals. In addition to the impact on the research productivity of numerous scientists in Alaska, the new instruments will support the emerging needs of NEON (National Ecological Observatory Network) in Alaska, the International Polar Year, and citizen science with the Anchorage Waterways Council. This award will also allow the enhanced educational preparation of students in state-of-the-art instrumentation, including the training of undergraduates, graduate students, and Alaska Native students in the Alaska Natives Science and Engineering Program (ANSEP). Information on the study can be obtain in this website http://www.uaa.alaska.edu/enri/usnip/index.cfm

DESCRIPTION (provided by applicant): The recent dramatic increase and geographic differences in frequency of reproductive diseases are likely influenced by changes in the environment, including perchlorate exposure. Perchlorate (ClO4-) is a persistent, chlorinated water-soluble contaminant that is pervasive in the United States. As a toxicant, perchlorate poses a major risk to human health through ingestion of contaminated water, food, and breast milk. Perchlorate is a known endocrine disruptor that competitively inhibits iodide uptake at the Sodium-Iodide Symporter (NIS) in the thyroid, thus hindering thyroid hormone synthesis. Studies demonstrate, however, that perchlorate exposure masculinizes both female and male stickleback fish (Gasterosteus aculeatus), leading to hermaphroditic females and males with testicular hypertrophy, results that are not predicted by a simple, direct thyroid- disruption mechanism. The goal of this project is to reconcile the dominant paradigm of perchlorate action - exclusively by disruption of NIS in the thyroid - with masculinization of behavior, physiology, and morphology in stickleback. The project's goal is to identify previously unsuspected pathways by which perchlorate may impact human reproductive health. Our working hypothesis is that perchlorate disrupts gonadal development by acting independently of the thyroid. Aim 1 will determine whether all observed phenotypic responses to perchlorate exposure in stickleback are mediated by the thyroid by rescuing thyroid hormone levels in perchlorate-exposed fish. Aim 2 will define the functional roles of NIS and NIS-paralogs in disruption of gonadal development by perchlorate using in situ hybridization to localize mRNA (Aim 2a), loss-of-function experiments to knock down expression of NIS and NIS-paralogs with morpholino anti-sense oligonucleotides and induced mutations using zinc finger nucleases (Aim 2b), and gain-of-function experiments by over- expressing the NIS and NIS-paralogs (Aim 2c). Aim 3 will determine the mechanism by which perchlorate alters sex differentiation using whole genome transcription profiling to determine which genes are early responders to perchlorate exposure, which are likely to be downstream genes, and whether responding genes are related to thyroid or gonad development. Quantitative PCR (qPCR) and in situ hybridization will verify expression profiling results. Significance. The proposed experiments will identify molecular and physiological pathways by which perchlorate disrupts gonadal development, whether solely via NIS in the thyroid or by other mechanisms. Because perchlorate is a pervasive contaminant in the U.S., our proposed work has direct implications for human health, particularly regarding thyroid diseases and disorders of sexual development.

The Earth's light-dark (LD) cycle is the strongest cue for entraining circadian rhythms and is considered the primary driver for the emergence and evolution of endogenous clocks. In polar regions, however, photoperiod exhibits extreme annual variation culminating with the sun remaining above or below the horizon for extended periods. Without a defined LD cycle, some arctic residents lose daily organization of behavior and physiology, and it has been hypothesized that the molecular clockwork that drives circadian rhythms may be weak or absent in arctic vertebrates. This study tests the hypothesis that persistence of circadian organization in some arctic mammals is adaptive because it allows organisms to minimize thermoregulatory costs. Specifically, the investigators will examine persistence, entrainment, and function of circadian rhythms in the arctic ground squirrel (AGS) during the continuous daylight of the active season and the continuous dark of hibernation. Four research objectives span from molecular and neurobiological mechanisms to physiological, behavioral and ecological adaptation: (1) determine when circadian rhythms are exhibited during the annual cycle of free-living AGS and establish if the onset of rhythmicity in spring coincides with first exposure to light, (2) determine if circadian rhythms persist in the master clock of AGS during hibernation by measuring patterns of clock gene expression in the suprachiasmatic nuclei (SCN) of the brain, (3) determine if AGS entrain circadian rhythms to daily changes in light quality or intensity, and (4) investigate patterns of re-entrainment and the function of rhythms in the arctic summer by experimentally phase-shifting free-living ground squirrels and measuring metabolic costs of nocturnal activity. Whether circadian rhythms persist during hibernation is contentious. Transcription and translation are thought to be globally suppressed during deep torpor and the only study to examine clock-gene expression within the SCN during multi-day torpor found no evidence of oscillations. However, circadian body temperature rhythms have been observed in some studies of captive ground squirrels during torpor and timing of arousals has been hypothesized to be controlled by a circadian clock. Persistence of oscillations in clock-gene expression in the SCN would support existence of tissue-specific mechanisms for translational control of a subset of genes relevant to survival during hibernation. In contrast, expression of daily rhythms may not be compatible with hibernation; SCN function and or output may be inhibited to prolong torpor. This study integrates research and learning by (1) training post-docs and students from three universities in ecophysiological studies in the laboratory and field, (2) inclusion of Alaska Natives in research through participation in the Alaska Native Science and Engineering Program, and (3) increasing K-12 students' engagement in research by participating in Teacher Research Experience programs. Results of research will be disseminated locally in classroom presentations and nationally through the University of Alaska media relations team.

DESCRIPTION (provided by applicant): This community-based participatory research project investigates exposures to two classes of emerging endocrine-disrupting chemicals (EDCs) with the Yupik people of St. Lawrence Island (SLI) in the Alaskan Arctic. The Arctic acts as a cold trap and is a hemispheric sink for persistent organic pollutants (POPs) that are transported through a well-documented process known as global distillation via prevailing atmospheric and oceanic currents from warmer regions. The Arctic is significant as an indicator region for long-range transport of legacy chemicals-a term applied to those POPs that have been banned or restricted-as well as emerging chemicals of concern. Arctic wildlife and people are also exposed to high levels of POPs from local sources, such as military toxics, solid waste dumps, and household dust. Exposure to POPs from both distant and local sources is a trend in the Arctic that is likely to increase due to increased global use and production of EDCs and climate warming. The purpose of this project is to initiate research partnerships that work in collaboration with the two Yupik villages of SLI to assess multiple exposure routes of two emerging EDCs-polybrominated diphenyl ethers (PBDEs) and perfluorinated compounds (PFCs). The project will assess exposures to PBDEs and PFCs in surface waters through analyses of contaminant levels and biomarkers for xenobiotic chemicals in the threespine stickleback fish. The research team will also analyze household dust for PBDEs and PFCs. Because the Yupik people of SLI depend on the harvest of wild foods to sustain them and their way of life, the research team will analyze levels of PBDEs and PFCs in traditional foods which are likely a major exposure pathway due to the biomagnification of POPs in marine mammals and fish that are critical components of the Yupik diet. This study will include a human biomonitoring component in order to assess levels of PBDEs and PFCs in human blood serum in relation to measures of thyroid health. The aim of this exposure assessment is to provide information, ownership of data, and training for the people of SLI so that they can plan and participate in public health actions to reduce environmental health risks. Finally, the research team collaborates with the leadership, elders, and youth of SLI to develop measures to prevent and mitigate environmental exposures through community educational programs and public policy actions, including community-based research institutes for college credit, health fairs for all community members, and workshops for health care providers.

Project Summary/Abstract The objectives of this community-based participatory research (CBPR) project are to investigate exposures, endocrine effects, and mechanisms of developmental disruption associated with legacy contaminants and emerging flame retardant chemicals in two Yupik communities on St. Lawrence Island (SLI) in arctic Alaska. The Arctic is subject to atmospheric deposition of globally-distilled persistent organic pollutants (POPs), acting as a hemispheric sink for POPs that are transported from lower latitudes. Thus, the Arctic is significant as an indicator region and contains some of the most highly contaminated animals and people in the world. This study addresses a primary public health concern of the people of SLI by focusing on the levels and effects of legacy and emerging contaminants on the development of children in an arctic indigenous population that is vulnerable, underserved, and experiences significant health disparities. Other studies have shown that young children are more highly exposed than adults. Using innovative and minimally invasive techniques, we will assess exposures of children to polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs), and emerging halogenated and non-halogenated organophosphate flame retardants. We will quantify PCBs and flame retardant chemicals in household dust, known to be an important exposure route. We will assess relationships among contaminant levels and evidence of health disruption via transcriptomics and endocrinology. We will use chemical concentrations in household dust and utilization of a subsistence diet as determined by stable isotope analysis to assess exposure pathways of these compounds. In order to understand the mechanistic basis of developmental disruption, and to have a reference for interpretation of human data, we will monitor patterns of gene expression, endocrinology, and histology of our resident fish model, the stickleback, from both contaminated and reference sites. Collectively, we will increase our understanding of routes of exposure, endocrine disruption, and effects on the transcriptome of Yupik children exposed to high levels of PCBs and flame retardant chemicals. This study provides an opportunity to investigate the levels of PBDEs and emerging flame retardants in nails and blood in relation to health outcomes of arctic indigenous children for the first time. Our CBPR project will also empower SLI communities with the knowledge and tools they need to address important health disparities in their communities. Our results will inform public health interventions and improve health outcomes of arctic children broadly. Furthermore, discovery of bioindicators relevant to early detection of developmental disruption will enable early intervention and improve health outcomes. Importantly, we will build capacity through our CBPR approach, public health interventions, and policy outreach, which will mitigate future exposure of SLI children to toxic chemicals.

Our previous community-based participatory research at Northeast Cape (NEC) on St. Lawrence Island (SLI) found elevated levels of polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs), and mercury (Hg) in sediments and biota within the watershed at the formerly used defense (FUD) site. We found elevated OCPs and PCBs in serum of the SLI people due to both long-range transport and military-derived sources, with the highest levels of PCBs in people who have traditional and familial connections with NEC, including subsistence. Concentrations of persistent organic pollutants (POPs) in our resident fish model, the stickleback, closely mirror concentrations in the blood serum of SLI residents, indicating their efficacy as sentinel species on SLI. Despite extensive remediation at NEC, short-lived lower trophic level fish in the Suqitughneq (Suqi) River remain contaminated with PCBs, OCPs, and Hg originating from the FUD site, at levels that exceed EPA fish consumption guidelines for cancer risk. Elevated contaminant levels and disrupted health in Suqi River fish indicate potential health threats for residents and that site remediation is incomplete. Our overarching goal is to advance scientific understanding of the exposure pathways and long-term human health consequences associated with contaminant exposure from FUD sites and inform interventions necessary to protect the health and long-term well-being of the people of SLI as they re-establish their traditional community at NEC. In Aim 1, we propose to build on our prior discoveries and continue our collaboration with the communities of SLI to investigate potential exposure pathways and biological impacts of persistent contamination associated with the FUD site at NEC on SLI. We will analyze PCBs, OCPs, and Hg in the water of the Suqi and Tapisaggak rivers, as well as in traditional foods and air samples to assess ingestion and inhalation as potential exposure pathways. We will build on work with stickleback as a sentinel species to determine biological effects of contaminants on endocrine function and organ-specific histopathology. In Aim 2, we will characterize and quantify body burden of contaminants, and linkages to health outcomes, in people associated with NEC. In Aim 3, we will inform decisions and interventions to protect the health of the people of SLI and enable re- establishment of the traditional community at NEC. We will provide information that will lead to improved remediation, provide data and traditional knowledge to inform public health consultations and assessments, and develop a community-based public health action plan and interventions to protect health, ensure equity in decision-making, and restore the NEC community. This study will have local and circumpolar arctic implications for Indigenous communities. Locally, we will provide data and implement actions necessary for re- establishing the community at NEC. Given that thousands of such Cold War military sites exist throughout the Arctic, often in close proximity to Indigenous communities, our project may serve as a model for environmental and health monitoring and policy action by other Arctic Indigenous Peoples.