Vanderbilt University Medical Center 41 articles published in JoVE Medicine Enhanced Communication of Tumor Margins Using 3D Scanning and Mapping Carly Fassler1, Alexis Miller1, Kayvon Sharif1, Kavita Prasad1, Marina Aweeda1, Jim Lewis2, Michael C. Topf1,3 1Department of Otolaryngology - Head and Neck Surgery, Vanderbilt University Medical Center, 2Department of Pathology, Vanderbilt University Medical Center, 3Vanderbilt University School of Engineering A novel method for 3D scanning and virtual mapping of cancer resections is proposed with the goal of improving communication among the multidisciplinary cancer care team. Biology Human Pseudoislet System for Synchronous Assessment of Fluorescent Biosensor Dynamics and Hormone Secretory Profiles Tiffany M. Richardson*1, Yasminye D. Pettway*1, John T. Walker1, Heather A. Nelson2, Matthew Ishahak3, Gregory Poffenberger2, Radhika Aramandla2, Conrad Reihsmann2, Ashutosh Agarwal3, Alvin C. Powers1,2,4, Marcela Brissova2 1Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, 2Division of Diabetes, Endocrinology and Metabolism, Department of Medicine, Vanderbilt University Medical Center, 3Department of Biomedical Engineering, University of Miami, 4VA Tennessee Valley Healthcare System This protocol describes a method for the synchronous acquisition and co-registration of intracellular signaling events and the secretion of insulin and glucagon by primary human pseudoislets using the adenoviral delivery of a cyclic adenosine monophosphate (cAMP) biosensor, a cAMP difference detector in situ (cADDis), and a microperifusion system. Medicine Gastric Point of Care Ultrasound in Adults: Image Acquisition and Interpretation Eric R. Heinz1, Omar Al-Qudsi2, David L. Convissar3, Marianne D. David1, Jennifer E. Dominguez2, Stephen Haskins4, Christina Jelly5, Anahi Perlas6, Anita N. Vincent1, Yuriy S. Bronshteyn2 1Department of Anesthesiology and Critical Care Medicine, George Washington University, 2Department of Anesthesiology, Duke University School of Medicine, Duke University Health System, 3Department of Anesthesiology, Massachusetts General Hospital, 4Department of Anesthesiology, Critical Care & Pain Management, Hospital for Special Surgery, 5Department of Anesthesiology, Vanderbilt University Medical Center, 6Department of Anesthesia and Pain Management, Toronto Western Hospital, University Health Network This protocol introduces two methods for image acquisition in gastric ultrasonography. Additionally, tips are provided for interpreting this information to assist in medical decision-making. Immunology and Infection Murine Fecal Isolation and Microbiota Transplantation Jeanne A. Ishimwe1, Jianyong Zhong2,3, Valentina Kon2, Annet Kirabo1 1Division of Clinical Pharmacology, Department of Medicine Vanderbilt University Medical Center and Department of Molecular Physiology and Biophysics, Vanderbilt University, 2Division of Pediatric Nephrology Department of Pediatrics, Vanderbilt University Medical Center, 3Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center The goal here is to outline a protocol to investigate the mechanisms of dysbiosis in cardiovascular disease. This paper discusses how to aseptically collect and transplant murine fecal samples, isolate intestines, and use the "Swiss-roll" method, followed by immunostaining techniques to interrogate changes in the gastrointestinal tract. Editorial Methods For Studying Osteoenergetics And Metabolism Elizabeth Rendina-Ruedy1, Courtney M. Karner2 1Department of Medicine, Clinical Pharmacology, Vanderbilt University Medical Center, 2Department of Internal Medicine, Charles and Jane Pak Center for Mineral Metabolism and Clinical Research, University of Texas Southwestern Medical Center Medicine Modified Technique for the Use of Neonatal Murine Hearts in the Langendorff Preparation Matthew B. Barajas1, Richard J. Levy2 1Department of Anesthesiology, Vanderbilt University Medical Center, 2Department of Anesthesiology, Columbia University Medical Center The present protocol describes aortic cannulation and retrograde perfusion of the ex-vivo neonatal murine heart. A two-person strategy, using a dissecting microscope and a blunted small gauge needle, permits reliable cannulation. Quantification of longitudinal contractile tension is achieved using a force transducer connected to the apex of the left ventricle. Biology Using Real-Time Cell Metabolic Flux Analyzer to Monitor Osteoblast Bioenergetics Shobana Jayapalan1,2, Ananya Nandy1,2, Elizabeth Rendina-Ruedy1,2,3 1Vanderbilt Center for Bone Biology, Division of Clinical Pharmacology, Vanderbilt University Medical Center, 2Department of Medicine, Division of Clinical Pharmacology, Vanderbilt University Medical Center, 3Department of Molecular Physiology and Biophysics, Vanderbilt University Real-time cell metabolic flux assay measures the oxygen consumption rate and extracellular acidification rate, which corresponds to mitochondrial and glycolytic adenosine triphosphate production, using pH and oxygen sensors. The manuscript explains a method to understand the energy status of osteoblasts and the characterization and interpretation of the cellular bioenergetic status. Biology Development of a Cell Co-Culture Model to Mimic Cardiac Ischemia/Reperfusion In Vitro Zhu Li1, Matthew J. W. Hampton1, Matthew B. Barajas1, Matthias L. Riess1,2,3 1Department of Anesthesiology, Vanderbilt University Medical Center, 2Anesthesiology, TVHS VA Medical Center, 3Department of Pharmacology, Vanderbilt University Spatial distance is a key parameter in assessing hypoxia/reoxygenation injury in a co-culture model of separate endothelial and cardiomyocyte cell layers, suggesting, for the first time, that optimizing the co-culture spatial environment is necessary to provide a favorable in vitro model for testing the role of endothelial cells in cardiomyocyte protection. Bioengineering A Large Animal Model for Pulmonary Hypertension and Right Ventricular Failure: Left Pulmonary Artery Ligation and Progressive Main Pulmonary Artery Banding in Sheep Rei Ukita1, John W. Stokes1, W. Kelly Wu1, Jennifer Talackine1, Nancy Cardwell1, Yatrik Patel1, Clayne Benson5, Caitlin T. Demarest1, Erika B. Rosenzweig3, Keith Cook2, Emily J. Tsai4, Matthew Bacchetta1,6 1Department of Thoracic Surgery, Vanderbilt University Medical Center, 2Department of Biomedical Engineering, Carnegie Mellon University, 3Department of Pediatrics, Columbia University Medical Center, Columbia University Irving Medical Center, 4Department of Medicine, Columbia University Medical Center, Columbia University Irving Medical Center, 5Department of Anesthesiology, Vanderbilt University Medical Center, 6Department of Biomedical Engineering, Vanderbilt University This manuscript describes the surgical technique and experimental approach to develop severe right ventricular pressure overload to model their adaptive and maladaptive phenotypes. Biology Direct Detection of Isolevuglandins in Tissues Using a D11 scFv-Alkaline Phosphatase Fusion Protein and Immunofluorescence Cassandra Warden*1, Alan J. Simmons*2, Lejla Pasic3, Ashley Pitzer4,6, Sean S. Davies4, Justin H. Layer5, Raymond L. Mernaugh3, Annet Kirabo4,6 1Vanderbilt Eye Institute, Vanderbilt University Medical Center, 2Department of Cell and Developmental Biology, Vanderbilt University, 3Department of Biochemistry, Vanderbilt University, 4Division of Clinical Pharmacology, Department of Medicine, Vanderbilt University Medical Center, 5Division of Hematology and Oncology, Indiana University School of Medicine, 6Department of Molecular Physiology and Biophysics, Vanderbilt University This article provides a detailed methodology for the measurement of isolevuglandins in tissues by immunofluorescence using alkaline phosphatase-conjugated ScFv D11 antibody. Hypertension models in both mice and humans are used to explain the step-by-step procedures and fundamental principles associated with isolevuglandin measurement in tissue samples. Cancer Research Automated Dissection Protocol for Tumor Enrichment in Low Tumor Content Tissues Charles A. Havnar1, Oliver Zill2, Jeff Eastham1, Jeffrey Hung1, Manana Javey3, Emmanuel Naouri3, Jennifer Giltnane1, Justin M. Balko4, Andrew Wallace2, Nicolas Lounsbury2, Daniel Oreper2, Sarajane Saturnio1, G-Y Yang5, Amy A. Lo1 1Departments of Research Pathology, Genentech, 2Bioinformatics & Computational Biology, Genentech, 3Roche Sequencing Solutions, Hacienda Drive, 4Department of Medicine, Vanderbilt University Medical Center, Medical Center Drive, 5Department of Pathology, Northwestern University, Feinberg School of Medicine Digital annotation with automated tissue dissection provides an innovative approach to enriching tumor in low tumor content cases and is adaptable to both paraffin and frozen tissue types. The described workflow improves accuracy, reproducibility and throughput and could be applied to both research and clinical settings. Developmental Biology Establishing a High Throughput Epidermal Spheroid Culture System to Model Keratinocyte Stem Cell Plasticity Yvon Woappi1,2, Geraldine Ezeka3, Justin Vercellino4, Sean M. Bloos5, Kim E. Creek6, Lucia Pirisi1 1Department of Pathology, Microbiology and Immunology, University of South Carolina School of Medicine, 2 Here we describe a protocol for the systematic cultivation of epidermal spheroids in 3D suspension culture. This protocol has wide-ranging applications for use in a variety of epithelial tissue types and for the modeling of several human diseases and conditions. Chemistry Automated Sample Multiplexing by using Combined Precursor Isotopic Labeling and Isobaric Tagging (cPILOT) Albert B. Arul1, Renã A.S. Robinson1,2,3,4 1Department of Chemistry, Vanderbilt University, 2Vanderbilt Memory & Alzheimer's Center, Vanderbilt University Medical Center, 3Vanderbilt Brain Institute, Vanderbilt University Medical Center, 4Vanderbilt Institute of Chemical Biology, Vanderbilt University Medical Center Combined precursor isotopic labeling and isobaric tagging (cPILOT) is an enhanced sample multiplexing strategy that is capable of increasing the number of samples that can be analyzed simultaneously with available isobaric tags. Incorporation of a robotic platform has greatly increased experimental throughput, reproducibility, and quantitative accuracy. Medicine A Modified Two Kidney One Clip Mouse Model of Renin Regulation in Renal Artery Stenosis Mohammad Saleem1, Pierina Barturen-Larrea1, Luz Saavedra1, Jose A. Gomez1 1Department of Medicine/Clinical Pharmacology Division, Vanderbilt University Medical Center A modified 2 kidney 1 clip (2K1C) Goldblatt mouse model was developed using polyurethane tubing to initiate renal artery stenosis, inducing an increase in renin expression and kidney injury. Here, we describe a detailed procedure of preparing and placing the cuff onto the renal artery to generate a reproducible and consistent 2K1C mouse model. Medicine Halogenated Agent Delivery in Porcine Model of Acute Respiratory Distress Syndrome via an Intensive Care Unit Type Device Raiko Blondonnet1,2, Bertille Paquette1,2, Jules Audard1,2, Ridvan Guler1,2, François-Xavier Roman1,2, Ruoyang Zhai2, Corinne Belville2, Loïc Blanchon2, Thomas Godet1, Emmanuel Futier1,2, Jean-Etienne Bazin1, Jean-Michel Constantin3, Vincent Sapin2,4, Matthieu Jabaudon1,2,5 1Department of Perioperative Medicine, CHU Clermont-Ferrand, 2GReD, CNRS, INSERM, Université Clermont Auvergne, 3GRC 29, AP-HP, DMU DREAM, Department of Anesthesiology and Critical Care, Pitié-Salpêtrière Hospital, Sorbonne University, 4Department of Biochemistry and Molecular Genetics, CHU Clermont-Ferrand, 5Division of Allergy, Pulmonary, and Critical Care Medicine, Vanderbilt University Medical Center We describe a model of hydrochloric acid-induced acute respiratory distress syndrome (ARDS) in piglets receiving sedation with halogenated agents, isoflurane and sevoflurane, through a device used for inhaled intensive care sedation. This model can be used to investigate the biological mechanisms of halogenated agents on lung injury and repair. Neuroscience An Implantable System For Chronic In Vivo Electromyography David Zealear1, Yike Li1, Shan Huang1 1Department of Otolaryngology, Vanderbilt University Medical Center Presented here is a protocol for the manufacturing of an implantable system for in vivo chronological recording of evoked and spontaneous electromyographic potentials. The system is applied to the investigation of reinnervation of laryngeal muscles following nerve injury. Biology Isolation and Characterization of Adult Cardiac Fibroblasts and Myofibroblasts Meiling Melzer1, David Beier1, Pampee P. Young1,2, Sarika Saraswati1 1Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, 2American Red Cross, National Headquarters Obtaining a pure population of fibroblasts is crucial to studying their role in wound repair and fibrosis. Described here is a detailed method to isolate fibroblasts and myofibroblasts from uninjured and injured mouse hearts followed by characterization of their purity and functionality by immunofluorescence, RTPCR, fluorescence-assisted cell sorting, and collagen gel contraction. Developmental Biology Analysis of Non-Human Primate Pancreatic Islet Oxygen Consumption Joseph M. Elsakr1, Charles Deeter2, Valerie Ricciardi3, Maureen Gannon1,4,5,6 1Department of Molecular Physiology and Biophysics, Vanderbilt University, 2Agilent Technologies, 3Department of Biological Sciences, Vanderbilt University, 4Department of Veterans Affairs Tennessee Valley, 5Department of Medicine, Vanderbilt University Medical Center, 6Department of Cell and Developmental Biology, Vanderbilt University This protocol demonstrates the accurate and reproducible measurement of oxygen consumption in non-human primate pancreatic islets. The islet loading techniques and coating of the microplate provide a framework for efficient measurement of respiration in other types of cultured spheroids. Cancer Research Studying Normal Tissue Radiation Effects using Extracellular Matrix Hydrogels Steven M Alves1, Tian Zhu1, Anastasia Shostak1, Ninna S. Rossen2, Marjan Rafat1,3,4 1Department of Chemical and Biomolecular Engineering, Vanderbilt University, 2Department of Radiation Oncology, Stanford University, 3Depattment of Biomedical Engineering, Vanderbilt University, 4Department of Radiation Oncology, Vanderbilt University Medical Center This protocol presents a method for decellularization and subsequent hydrogel formation of murine mammary fat pads following ex vivo irradiation. Bioengineering Growth and Characterization of Irradiated Organoids from Mammary Glands Benjamin C. Hacker1, Javier D. Gomez1, Carlos A. Silvera Batista1, Marjan Rafat1,2,3 1Department of Chemical and Biomolecular Engineering, Vanderbilt University, 2Department of Biomedical Engineering, Vanderbilt University, 3Department of Radiation Oncology, Vanderbilt University Medical Center Organoids developed from mouse mammary glands were irradiated and characterized to assess epithelial traits and interactions with immune cells. Irradiated organoids can be used to better evaluate cell-cell interactions that may lead to tumor cell recruitment in irradiated normal tissue. Developmental Biology Analysis of Cardiac Chamber Development During Mouse Embryogenesis Using Whole Mount Epifluorescence Zhentao Zhang1,2,3, Young-Jae Nam1,2,3 1Department of Medicine, Division of Cardiovascular Medicine, Vanderbilt University Medical Center, 2Department of Cell and Developmental Biology, Vanderbilt University, 3Vanderbilt Center for Stem Cell Biology, Vanderbilt University We present the protocols to examine mouse heart development using whole mount epifluorescent microscopy on mouse embryos dissected from ventricular specific MLC-2v-tdTomato reporter knock-in mice. This method allows us to directly visualize each stage of the ventricular formation during mouse heart development without labor-intensive histochemical methods. Immunology and Infection Isolation and Adoptive Transfer of High Salt Treated Antigen-presenting Dendritic Cells Justin P. Van Beusecum1, Liang Xiao1, Natalia R. Barbaro1, David M. Patrick1, Annet Kirabo1 1Division of Clinical Pharmacology, Department of Medicine, Vanderbilt University Medical Center Here, we present a protocol to isolate dendritic cells from murine spleens by magnetic cell sorting and subsequent adoptive transfer into naïve mice. The model of high-salt activated dendritic cells was chosen to explain the step-by-step procedures of adoptive transfer and flow cytometry. Biochemistry Use of Electron Paramagnetic Resonance in Biological Samples at Ambient Temperature and 77 K Hanan B. Elajaili1, Laura Hernandez-Lagunas1, Kalina Ranguelova2, Sergey Dikalov3, Eva Nozik-Grayck1 1Cardiovascular Pulmonary Research Laboratories and Pediatric Critical Care Medicine, Department of Pediatrics, University of Colorado Anschutz Medical Campus, 2Bruker BioSpin Corp, 3Department of Medicine, Division of Clinical Pharmacology, Vanderbilt University Medical Center Electron paramagnetic resonance (EPR) spectroscopy is an unambiguous method to measure free radicals. The use of selective spin probes allows for detection of free radicals in different cellular compartments. We present a practical, efficient method to collect biological samples that facilitate treating, storing, and transferring samples for EPR measurements. Medicine Transdermal Measurement of Glomerular Filtration Rate in Mice Lauren Scarfe1,2, Daniel Schock-Kusch3, Lorenzo Ressel4, Jochen Friedemann3, Yury Shulhevich3, Patricia Murray2, Bettina Wilm2, Mark de Caestecker1 1Division of Nephrology, Department of Medicine, Vanderbilt University Medical Center, 2Department of Cellular and Molecular Physiology, University of Liverpool, 3MediBeacon GmbH, 4Department of Veterinary Pathology and Public Health, Institute of Veterinary Science, University of Liverpool Here we describe a protocol to measure glomerular filtration rate (GFR) in conscious, freely moving mice using a transdermal GFR monitor. Medicine Fat-Water Phantoms for Magnetic Resonance Imaging Validation: A Flexible and Scalable Protocol Emily C. Bush1, Aliya Gifford2, Crystal L. Coolbaugh1, Theodore F. Towse1,3,4, Bruce M. Damon1,5,6,7, E. Brian Welch1,5 1Vanderbilt University Institute of Imaging Science (VUIIS), Vanderbilt University Medical Center, 2Department of Biomedical Informatics, Vanderbilt University Medical Center, 3Department of Physical Medicine and Rehabilitation, Vanderbilt University Medical Center, 4Department of Biomedical Sciences, Grand Valley State University, 5Department of Radiology & Radiological Sciences, Vanderbilt University Medical Center, 6Department of Biomedical Engineering, Vanderbilt University, 7Department of Molecular Physiology and Biophysics, Vanderbilt University The purpose of this work is to describe a protocol for creating a practical fat-water phantom that can be customized to produce phantoms with varying fat percentages and volumes. Biochemistry Purification and Reconstitution of TRPV1 for Spectroscopic Analysis Francisco J. Sierra-Valdez1, Richard A. Stein2, Phanindra Velissety1,3, Valeria Vasquez1, Julio F. Cordero-Morales1 1Department of Physiology, University of Tennessee Health Science Center, 2Department of Molecular Physiology and Biophysics, Vanderbilt University Medical Center, 3CuriRX, Inc. This article describes specific methods to obtain biochemical quantities of detergent-solubilized TRPV1 for spectroscopic analysis. The combined protocols provide biochemical and biophysical tools that can be adapted to facilitate structural and functional studies for mammalian ion channels in a membrane-controlled environment. Neuroscience Optimization of Laser-Capture Microdissection for the Isolation of Enteric Ganglia from Fresh-Frozen Human Tissue Aaron A. May-Zhang1, Karen K Deal1, E. Michelle Southard-Smith1 1Vanderbilt University Medical Center The goal of this protocol is to obtain high-integrity RNA samples from enteric ganglia isolated from unfixed, freshly-resected human intestinal tissue using laser capture microdissection (LCM). This protocol involves preparing flash-frozen samples of human intestinal tissue, cryosectioning, ethanolic staining and dehydration, LCM, and RNA extraction. Neuroscience FM Dye Cycling at the Synapse: Comparing High Potassium Depolarization, Electrical and Channelrhodopsin Stimulation Danielle L. Kopke1, Kendal Broadie2 1Department of Biological Sciences, Vanderbilt University, 2Departments of Biological Sciences, Pharmacology, Cell and Developmental Biology, Kennedy Center for Research on Human Development, Vanderbilt University and Medical Center Synaptic vesicle (SV) cycling is the core mechanism of intercellular communication at neuronal synapses. FM dye uptake and release are the primary means of quantitatively assaying SV endo- and exocytosis. Here, we compare all the stimulation methods to drive FM1-43 cycling at the Drosophila neuromuscular junction (NMJ) model synapse. Biology Hydrodynamic Renal Pelvis Injection for Non-viral Expression of Proteins in the Kidney Lauren E. Woodard1,2,3, Richard C. Welch2, Felisha M. Williams2, Wentian Luo2, Jizhong Cheng3, Matthew H. Wilson1,2,3 1Department of Veterans Affairs, Tennessee Valley Healthcare System, 2Departments of Medicine and Pharmacology, Vanderbilt University Medical Center, 3Department of Medicine, Baylor University College of Medicine This protocol describes a method to inject plasmid DNA into the mouse kidney via the renal pelvis to produce transgene expression specifically in the kidney. Biochemistry Expression, Purification, and Antimicrobial Activity of S100A12 Emmanuel Jackson1, Saffron Little1, Dana S. Franklin1, Jennifer A. Gaddy2,3, Steven M. Damo1,4 1Department of Life and Physical Sciences, Fisk University, 2Tennessee Valley Healthcare Systems, U. S. Dept. of Veterans Affairs, 3Department of Medicine - Division of Infectious Diseases, Vanderbilt University Medical School, 4Department of Biochemistry, Vanderbilt University Here, we present a method to express and purify S100A12 (calgranulin C). We describe a protocol to measure its antimicrobial activity against the human pathogen H. pylori. Immunology and Infection Intracellular Staining and Flow Cytometry to Identify Lymphocyte Subsets within Murine Aorta, Kidney and Lymph Nodes in a Model of Hypertension Fanny Laroumanie1, Bethany L. Dale2, Mohamed A. Saleh3, Meena S. Madhur1 1Department of Medicine, Division of Clinical Pharmacology, Vanderbilt University Medical Center, 2Department of Molecular Physiology and Biophysics, Vanderbilt University, 3Department of Pharmacology and Toxicology, Faculty of Pharmacy, Mansoura University This article provides detailed methodology to identify and quantify functional T lymphocyte subsets present within murine kidney, aorta and lymph nodes by intracellular staining and flow cytometry. The model of angiotensin II induced hypertension was chosen to explain, step-by-step, the procedures and fundamental principles of flow cytometry and intracellular staining. Developmental Biology Effective Isolation of Functional Islets from Neonatal Mouse Pancreas Chen Huang1, Guoqiang Gu1 1Department of Cell and Development Biology, Vanderbilt University Medical School We describe a protocol herein for isolating intact islets from neonatal mice. Pancreata were partially digested with collagenase, followed by washing and hand picking. 20 - 80 islet clusters can be obtained per pancreas from newly born mice, which are suitable for several islet studies. Bioengineering Sustained Administration of β-cell Mitogens to Intact Mouse Islets Ex Vivo Using Biodegradable Poly(lactic-co-glycolic acid) Microspheres Raymond C. Pasek1, Taylor E. Kavanaugh2, Craig L. Duvall2, Maureen A. Gannon1,3 1Department of Medicine, Vanderbilt University Medical Center, 2School of Engineering, Vanderbilt University, 3Department of Veterans Affairs, Tennessee Valley Health Authority Here, we present methodology to generate and administer compound of interest-loaded poly(lactic-co-glycolic acid) (PLGA) microspheres to intact mouse islets in culture with subsequent immunofluorescence analysis of β-cell proliferation. This method is suitable for determining the efficacy of candidate β-cell mitogens. Biology Scanning Electron Microscopy of Macerated Tissue to Visualize the Extracellular Matrix Matthew K. Stephenson1, Sean Lenihan2,3, Roman Covarrubias2,3, Ryan M. Huttinger2,3, Richard J. Gumina2,3, Douglas B. Sawyer4, Cristi L. Galindo2,3 1Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, 2Department of Medicine, Vanderbilt University Medical Center, 3Division of Cardiovascular Medicine, Vanderbilt University Medical Center, 4Cardiovascular Institute, Maine Medical Center Shown here is a method for visualizing extracellular matrix ultrastructure in decellularized cardiac tissues. Biology High Throughput Danio Rerio Energy Expenditure Assay Savannah Y. Williams1, Benjamin J. Renquist2 1Department of Molecular Physiology and Biophysics, Vanderbilt University Medical Center, 2School of Animal and Comparative Biomedical Sciences, University of Arizona Zebrafish are an important model organism for the study of energy homeostasis. By utilizing a NADH2 sensitive redox indicator, alamar Blue, we have developed an assay that measures the metabolic rate of zebrafish larvae in a 96 well plate format and can be applied to drug or gene discovery. Behavior Testing Sensory and Multisensory Function in Children with Autism Spectrum Disorder Sarah H. Baum1, Ryan A. Stevenson2, Mark T. Wallace3 1Vanderbilt Brain Institute, Vanderbilt University Medical Center, 2Department of Psychology, University of Toronto, 3Department of Hearing and Speech Sciences, Vanderbilt University We describe how to implement a battery of behavioral tasks to examine the processing and integration of sensory stimuli in children with ASD. The goal is to characterize individual differences in temporal processing of simple auditory and visual stimuli and relate these to higher order perceptual skills like speech perception. Medicine Human Brown Adipose Tissue Depots Automatically Segmented by Positron Emission Tomography/Computed Tomography and Registered Magnetic Resonance Images Aliya Gifford1, Theodore F. Towse2, Ronald C. Walker3, Malcolm J. Avison4, E. Brian Welch3 1Chemical and Physical Biology Program, Vanderbilt University, 2Department of Physical Medicine and Rehabilitation, Vanderbilt University School of Medicine, 3Radiology & Radiological Sciences, Vanderbilt University Medical Center, 4Department of Pharmacology, Vanderbilt University The method presented here uses 18F-Fluorodeoxyglucose (18F-FDG) positron emission tomography/computed tomography (PET-CT) and fat-water separated magnetic resonance imaging (MRI), each scanned following 2 hr exposure to thermoneutral (24 °C) and cold conditions (17 °C) in order to map brown adipose tissue (BAT) in adult human subjects. Medicine Polyacrylamide Gels for Invadopodia and Traction Force Assays on Cancer Cells Rachel J. Jerrell1, Aron Parekh1 1Department of Otolaryngology, Vanderbilt University Medical Center Mechanical rigidity in the tumor microenvironment plays a crucial role in driving malignant behavior by increasing invadopodia activity and actomyosin contractility. Using polyacrylamide gels (PAAs), invadopodia and traction force assays can be utilized to study the invasive and contractile properties of cancer cells in response to matrix rigidity. Biology Reconstitution Of β-catenin Degradation In Xenopus Egg Extract Tony W. Chen*1, Matthew R. Broadus*1, Stacey S. Huppert2, Ethan Lee1,3 1Department of Cell and Developmental Biology and Program in Developmental Biology, Vanderbilt University Medical Center, 2 A method is described for analyzing protein degradation using radiolabeled and luciferase-fusion proteins in Xenopus egg extract and its adaptation for high-throughput screening for small molecule modulators of protein degradation. Biology Cytological Analysis of Spermatogenesis: Live and Fixed Preparations of Drosophila Testes Poojitha Sitaram1, Sarah Grace Hainline1, Laura Anne Lee1 1Department of Cell and Developmental Biology, Vanderbilt University Medical Center Methods for isolating and preparing Drosophila testes samples (live and fixed) for imaging by phase-contrast and fluorescence microscopy are described herein. Medicine Primary Orthotopic Glioma Xenografts Recapitulate Infiltrative Growth and Isocitrate Dehydrogenase I Mutation J. Geraldo Valadez1, Anuraag Sarangi1, Christopher J. Lundberg1, Michael K. Cooper1,2,3 1Department of Neurology, Vanderbilt University Medical Center, 2Vanderbilt Ingram Cancer Center, Vanderbilt University Medical Center, 3Neurology Service, Veteran Affairs TVHS Malignant gliomas constitute a heterogeneous group of highly infiltrative glial neoplasms with distinct clinical and molecular features. Primary orthotopic xenografts recapitulate the histopathological and molecular features of malignant glioma subtypes in preclinical animal models.
