Heinrich Heine University Dusseldorf View Institution's Website 13 articles published in JoVE Medicine Location, Dissection, and Analysis of the Murine Stellate Ganglion Katharina Scherschel1,2,3, Hanna Bräuninger2,4, Klara Glufke2, Christiane Jungen2,4,5, Nikolaj Klöcker3, Christian Meyer1,2,3 1Division of Cardiology, EVK Düsseldorf, cNEP, cardiac Neuro- and Electrophysiology Research Consortium, 2DZHK (German Centre for Cardiovascular Research), 3Institute of Neural and Sensory Physiology, Medical Faculty, Heinrich Heine University Düsseldorf, 4Clinic for Cardiology, University Heart & Vascular Centre, University Hospital Hamburg-Eppendorf, 5Department of Cardiology, Leiden University Medical Center Pathophysiological changes in the cardiac autonomic nervous system, especially in its sympathetic branch, contribute to the onset and maintenance of ventricular arrhythmias. In the present protocol, we show how to characterize murine stellate ganglia to improve the understanding of the underlying molecular and cellular processes. Neuroscience Imaging of Intracellular ATP in Organotypic Tissue Slices of the Mouse Brain using the FRET-based Sensor ATeam1.03YEMK Rodrigo Lerchundi*1, Karl W. Kafitz*1, Marcel Färfers1, Felix Beyer2, Na Huang1, Christine R. Rose1 1Institute of Neurobiology, Heinrich Heine University Düsseldorf, 2Department of Neurology, Düsseldorf University Hospital We describe a protocol for cell-type specific expression of the genetically encoded FRET-based sensor ATeam1.03YEMK in organotypic slice cultures of the mouse forebrain. Furthermore, we show how to use this sensor for dynamic imaging of cellular ATP levels in neurons and astrocytes. Neuroscience Optogenetic Entrainment of Hippocampal Theta Oscillations in Behaving Mice Franziska Bender1,2, Tatiana Korotkova2,3, Alexey Ponomarenko1,2 1Systems Neurophysiology Research Group, Institute of Clinical Neuroscience and Medical Psychology, Medical Faculty, Heinrich Heine University Düsseldorf, 2Behavioural Neurodynamics Group, Leibniz Institute for Molecular Pharmacology (FMP)/ NeuroCure Cluster of Excellence, 3Neuronal Circuits and Behavior Research Group, Max Planck Institute for Metabolism Research We describe the use of optogenetics and electrophysiological recordings for selective manipulations of hippocampal theta oscillations (5-10 Hz) in behaving mice. The efficacy of the rhythm entrainment is monitored using local field potential. A combination of opto- and pharmacogenetic inhibition addresses the efferent readout of hippocampal synchronization. Immunology and Infection Intraperitoneal Glucose Tolerance Test, Measurement of Lung Function, and Fixation of the Lung to Study the Impact of Obesity and Impaired Metabolism on Pulmonary Outcomes Katharina Dinger1, Jasmine Mohr1, Christina Vohlen1,2, Dharmesh Hirani1, Eva Hucklenbruch-Rother2, Regina Ensenauer3, Jörg Dötsch2, Miguel A. Alejandre Alcazar1,2 1Translational Experimental Pediatrics, Experimental Pulmonology, Department of Pediatrics and Adolescent Medicine, University of Cologne, 2Department of Pediatrics and Adolescent Medicine, University of Cologne, 3Division of Experimental Pediatrics and Metabolism, University Children's Hospital, Heinrich Heine University Düsseldorf The incidence of obesity is rising and increases the risk of chronic lung diseases. To establish the underlying mechanisms and preventive strategies, well-defined animal models are needed. Here, we provide three methods (glucose-tolerance-test, body plethysmography, and lung fixation) to study the effect of obesity on pulmonary outcomes in mice. Medicine The Inverted Heart Model for Interstitial Transudate Collection from the Isolated Rat Heart Kezhe Tan*1,2, Zhaoping Ding*1,2, Bodo Steckel2, Sonja Hartwig3, Stefan Lehr3, Xiaoming Deng1, Jürgen Schrader2 1Department of Anesthesiology, Changhai Hospital, Second Military Medical University, 2Department of Molecular Cardiology, Heinrich Heine University, 3Department of Clinical Biochemistry and Pathobiochemistry, German Diabetes Center, Heinrich Heine University This protocol describes a method to collect cardiac interstitial fluid from the isolated, perfused rat heart. To physically separate interstitial transudate from coronary venous effluent perfusate, the Langendorff perfused heart is inverted, and the transudate (interstitial fluid) formed on the cardiac surface is collected using a soft latex cap. Genetics Genetic Manipulation of the Plant Pathogen Ustilago maydis to Study Fungal Biology and Plant Microbe Interactions Kristin Bösch*1,2, Lamprinos Frantzeskakis*1, Miroslav Vraneš3, Jörg Kämper3, Kerstin Schipper1,2, Vera Göhre1,2,4 1Institute for Microbiology, Heinrich-Heine University Düsseldorf, 2Bioeconomy Science Center (BioSC), 3Department of Genetics, Institute of Applied Biosciences, Karlsruhe Institute of Technology, 4Cluster of Excellence in Plant Sciences (CEPLAS), Heinrich-Heine University Düsseldorf We describe a robust gene replacement strategy to genetically manipulate the smut fungus Ustilago maydis. This protocol explains how to generate deletion mutants to investigate infection phenotypes. It can be extended to modify genes in any desired way, e.g., by adding a sequence encoding a fluorescent protein tag. Neuroscience Double-barreled and Concentric Microelectrodes for Measurement of Extracellular Ion Signals in Brain Tissue Nicole Haack1, Simone Durry1, Karl W. Kafitz1, Mitchell Chesler2, Christine R. Rose1 1Institute of Neurobiology, Heinrich Heine University Düsseldorf, 2Departments of Physiology and Neuroscience, New York University School of Medicine We demonstrate the fabrication, calibration and properties of two types of ion-selective microelectrodes (double-barreled and concentric) for measurement of ion concentrations in brain tissue. These are then used in the mouse hippocampal slice preparation to show that excitatory activity changes both extracellular potassium and sodium concentrations. Biology An Innovative Method for Exosome Quantification and Size Measurement Arash Mehdiani1, Anatol Maier1, Antonio Pinto1, Mareike Barth1, Payam Akhyari1, Artur Lichtenberg1 1Research Group Experimental Surgery, Department of Cardiovascular Surgery, Medical Faculty, Heinrich-Heine-University Dusseldorf A method for isolation of exosomes from whole blood and further analysis by nanoparticle tracking using a semi-automatic instrument is presented in this article. The presented technology provides an extremely sensitive method for visualizing and analyzing particles in liquid suspension. Neuroscience Multi-photon Intracellular Sodium Imaging Combined with UV-mediated Focal Uncaging of Glutamate in CA1 Pyramidal Neurons Christian Kleinhans*1, Karl W. Kafitz*1, Christine R. Rose1 1Institute of Neurobiology, Heinrich Heine University Düsseldorf We describe the combination of focal UV-induced photo-activation of neuro-active compounds with whole-cell patch-clamp and multi-photon imaging of intracellular sodium transients in dendrites and spines of hippocampal neurons in acute tissue slices of the mouse brain. Neuroscience Generation of Topically Transgenic Rats by In utero Electroporation and In vivo Bioluminescence Screening Sandra Vomund1, Tamar Sapir2, Orly Reiner2, Maria A. de Souza Silva3, Carsten Korth1 1Department of Neuropathology, Medical School Düsseldorf, 2Department of Molecular Genetics, Weizmann Institute for Science, 3Center of Behavioral Neuroscience, University of Düsseldorf Genes can be manipulated during development of the cortex or the hippocampus of the rat via in utero electroporation (IUE) at E16, to enable fast and targeted modifications in neuronal connectivity for later studies of behavior or neuropathology in adult animals. Postnatal in vivo imaging for control of IUE success is performed by bioluminescence of activating co-transfected luciferase. Biology Generation, Purification, and Characterization of Cell-invasive DISC1 Protein Species Verian Bader1, Philipp Ottis1, Martin Pum2, Joseph P. Huston2, Carsten Korth1 1Department of Neuropathology, Medical School Düsseldorf, Germany, 2Center of Behavioral Neuroscience, University of Düsseldorf The generation, purification and cell invasion of intracellular, cytoplasmic full length DISC1 protein aggresomes from cell cultures and of a labeled, multimeric recombinant DISC1 protein fragment in E. coli are described. Cell invasiveness is shown for recipient cells in cell culture and for neurons in vivo after stereotactical brain inoculation. Immunology and Infection Prediction of HIV-1 Coreceptor Usage (Tropism) by Sequence Analysis using a Genotypic Approach Saleta Sierra1, Rolf Kaiser1, Nadine Lübke1, Alexander Thielen2, Eugen Schuelter1, Eva Heger1, Martin Däumer3, Stefan Reuter4, Stefan Esser5, Gerd Fätkenheuer6, Herbert Pfister1, Mark Oette7, Thomas Lengauer2 1Institute of Virology, University of Cologne, 2Max Planck Institute for Informatics, 3Institute for Immune genetics, 4Department of Gastroenterology, Hepatology and Infectiology, University of Duesseldorf, 5Department of Dermatology, University of Essen, 6Department of Internal Medicine, University of Cologne, 7Augustinerinnen Hospital The prediction of the coreceptor usage of HIV-1 is required for the administration of a new class of antiretroviral drugs, i.e. coreceptor antagonists. It can be performed by sequence analysis of the env gene and subsequent interpretation through an internet based interpretation system (geno2pheno[coreceptor]). Biology Fast and Sensitive Colloidal Coomassie G-250 Staining for Proteins in Polyacrylamide Gels Nadine Dyballa1, Sabine Metzger1 1Biological Medical Research Center (BMFZ), University Duesseldorf This video shall popularize a colloidal Coomassie G-250 staining protocol according to Kang et al. for the detection of average 4 ng protein in gels. The staining is completed within 2 hours and without any effort. We routinely use Kang's protocol for analytical purposes in gel-based proteomics.