Practical Methods in Cardiovascular Research

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Stefan Dhein

Friedrich Wilhelm Mohr Mario Delmar

Practical Methods in Cardiovascular Research

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Practical Methods in Cardiovascular Research

Stefan Dhein (Editor)

Friedrich Wilhelm Mohr (Editor) Mario Delmar (Editor)

With 33 Tables and 290 Figures

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Library of Congress Control Number: 2004105045

ISBN 3-540-40763-4 Springer Berlin Heidelberg New York

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Prof. Dr. Stefan Dhein Klinik für Herzchirurgie Universität Leipzig Herzzentrum Strümpellstraße 39 04289 Leipzig Germany

Prof. Dr. Friedrich Wilhelm Mohr Klinik für Herzchirurgie Universität Leipzig Herzzentrum Strümpellstraße 39 04289 Leipzig Germany

Mario Delmar, MD PhD Department of Pharmacology SUNY Upstate Medical University 766 Irving Avenue

Syracuse NY 13210 USA

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Preface ^V 0.00

Preface

Cardiovascular disease is among the most common causes of death in Western indus- trialized countries. Although a number of diagnostic and therapeutic possibilities exist today, many problems remain unsolved. This implies the urgent need for future cardiovascular research. The present book gives an overview of a number of available techniques most often used in cardiovascular sciences, especially for investigators coming from other fields of research.

The book is divided into three sections. In the first part, the focus is on techniques used for in vivo experimentation. This field has regained great interest, given the availability of knock-out models which now await in-vivo testing. The book describes the most commonly used procedures for anesthesia and analgesia in laboratory animals as well as „classical“ isolated organ techniques, including electrophysiological methods now – again – in common use.

In the second part of the book, we move from the organ and tissue level to the cellular domain. Topics covered include electrophysiology, histology and cell culture techniques as well as biochemical methods such as HPLC, protein biochemistry and binding assays. This part closes with a brief chapter on how to register and analyze concentration response curves giving briefly the mathematics and basic analysis techniques. In the final part, the focus narrows further into molecular procedures such as surface plasmon resonance, nuclear magnetic resonance, RNA methods and DNA arrays. Finally, a chapter is devoted to the molecular aspects of transgenic mice tech- nology as well as an overview of procedures and regulations applicable to housing and transfer of transgenic animals.

The novelty of this book is in the description of practical protocols and trouble- shooting procedures. It is our intention to provide a basic guide for investigators in- terested in applying these techniques in their own laboratories. The book also aims to close the gap between the integrative in-vivo and organ-based methods and the re- ductionist cellular and molecular approaches. We wish all researchers every success with their experiments and that this book may help them start their way toward new frontiers.

Stefan Dhein Mario Delmar

Friedrich Wilhelm Mohr

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Table of Contents ^VII 0.00

Table of Contents

In-Vivo Techniques

1 Models of Cardiovascular Diseases

1.1 Anaesthesia (Inhalative or i.v. Narcosis) of Laboratory Animals . . . . 5 Katja Schneider

1.2 Extracorporeal Circulation . . . 26 Cris W. Ullmann

1.3 Open-chest Models of Acute Myocardial Ischemia and Reperfusion . . . 37 Kai Zacharowski, Thomas Hohlfeld and Ulrich K.M. Decking

1.4 Models of Chronic Ischemia and Infarction . . . 58 Nicolas Doll, Heike Aupperle, Fabian Stahl and Thomas Walther

1.5 Models on Left Ventricular Hypertrophy (LVH) . . . 72 Thomas Walther and Volkmar Falk

1.6 Experimental Models of Heart Failure . . . 83 Volkmar Falk, Jens Garbade and Thomas Walther

1.7 In-Vivo Models of Arrhythmias: a Canine Model of Sudden Cardiac Death . . . . 111 George E. Billman

1.8 In-Vivo Models of Atrial Fibrillation . . . 129 Ulrich Schotten, Yuri Blaauw and Maurits Allessie

In-Vitro Techniques

2 Isolated Organs

2.1 The Langendorff Heart . . . 155 Stefan Dhein

2.2 The Working Heart . . . 173 Marc W. Merx and Jürgen Schrader

2.3 Isolated Papillary Muscles . . . 190 Stefan Dhein

2.4 Isolated Vessels . . . 198 Rudolf Schubert

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0 VIII Table of Contents

3 Electrophysiological Techniques

3.1 Optical Techniques for the Recording of Action Potentials . . . 215 Helmut A. Tritthart

3.2 Recording Action Potentials Using Voltage-Sensitive Dyes . . . 233 Vladimir G. Fast

3.3 Epicardial Mapping in Isolated Hearts – Use in Safety Pharmacology, Analysis of Torsade de Pointes Arrhythmia

and Ischemia-Reperfusion-Related Arrhythmia . . . 256 Stefan Dhein

3.4 Voltage-Clamp and Patch-Clamp Techniques . . . 272 Hans Reiner Polder, Martin Weskamp, Klaus Linz, Rainer Meyer

3.5 L-Type Calcium Channel Recording . . . 324 Uta C. Hoppe, Mathias C. Brandt, Guido Michels and Michael Lindner

3.6 Recording Cardiac Potassium Currents with the Whole-Cell

Voltage Clamp Technique . . . 355 Erich Wettwer and Ursula Ravens

3.7 Recording the Pacemaker Current If . . . 381 Uta C. Hoppe, Fikret Er and Mathias C. Brandt

3.8 Recording Gap Junction Currents . . . 397 Ardawan J. Rastan and Stefan Dhein

3.9 Recording Monophasic Action Potentials . . . 417 Paulus Kirchhof, Larissa Fabritz and Michael R. Franz

3.10 Heart on a Chip – Extracellular Multielectrode Recordings

from Cardiac Myocytes in Vitro . . . 432 Ulrich Egert and Thomas Meyer

4 Histological Techniques

4.1 Immunohistochemistry for Structural and Functional Analysis

in Cardiovascular Research . . . 457 Wilhelm Bloch, Yüksel Korkmaz and Dirk Steinritz

4.2 Classical Histological Staining Procedures in Cardiovascular Research . . . 485 Wilhelm Bloch and Yüksel Korkmaz

4.3 Practical Use of in situ Hybridisation and RT in situ PCR

in Cardiovascular Research . . . 500 Yüksel Korkmaz, Dirk Steinritz and Wilhelm Bloch

4.4 Practical Use of Transmission Electron Microscopy for Analysis

of Structures and Molecules in Cardiovascular Research . . . 525 Wilhelm Bloch, Christian Hoffmann, Eveline Janssen and Yüksel Korkmaz

4.5 DAF Technique for Real-Time NO Imaging in the Human Myocardium . . . 546 Dirk Steinritz and Wilhelm Bloch

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Table of Contents ^IX 0.00

5 Cell Culture Techniques

5.1 Isolation and Culture of Adult Ventricular Cardiomyocytes . . . 557 Klaus-Dieter Schlüter and Hans Michael Piper

5.2 Culture of Neonatal Cardiomyocytes . . . 568 Aida Salameh and Stefan Dhein

5.3 Culture of Embryoid Bodies . . . 577 Cornelia Gissel, Dirk Nierhoff, Bernd Fleischmann, Jürgen Hescheler

and Agapios Sachinidis

5.4 Culturing and Differentiation of Embryonic and Adult Stem Cells

for Heart Research and Transplantation Therapy . . . 592 Marcel AG van der Heyden and Henk Rozemuller

5.5 Preparation of Endothelial Cells from Micro- and Macrovascular Origin . . . 610 Saskia C. Peters, Anna Reis and Thomas Noll

5.6 In Vitro Cultivation of Vascular Smooth Muscle Cells . . . 630 Daniel G. Sedding and Ruediger C. Braun-Dullaeus

5.7 Engineering Heart Tissue for In Vitro and In Vivo Studies . . . 640 Wolfram-Hubertus Zimmermann, Ivan Melnychenko, Michael Didié,

Ali El-Armouche and Thomas Eschenhagen

6 Biochemical and Analytical Techniques

6.1 High-Performance Liquid Chromatography (HPLC) . . . 661 Veronika R. Meyer

6.2 Protein Biochemistry . . . 686 Aida Salameh, Stefan Lehr, Jürgen Weiss and Peter Rösen

6.3 Receptor and Binding Studies . . . 723 Peter Hein, Martin C. Michel, Kirsten Leineweber, Thomas Wieland,

Nina Wettschureck and Stefan Offermanns

6.4 Recording and Analysing Concentration-Response Curves . . . 784 Stefan Dhein

6.5 Measurement of NO and Endothelial Function in Cardiovascular Research . . . 799 Renate Roesen, Anke Rosenkranz, Dirk Taubert and Reinhard Berkels

6.6 Biomechanical Stimulation of Vascular Cells In Vitro . . . 813 Henning Morawietz and Andreas Schubert

6.7 Measurement of Function and Regulation of Adrenergic Receptors . . . 829 Marc Brede, Melanie Philipp and Lutz Hein

6.8 Measurement of Function and Regulation

of Muscarinic Acetylcholine Receptors . . . 848 Björn Kaiser and Chris J. van Koppen

6.9 Cytometry in Cardiovascular Research . . . 863 Attila Tárnok

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0 X Table of Contents

Molecular Biology in Cardiovascular Research

7 Molecular Biology Techniques

7.1 Real-Time PCR Analysis of Cardiac Samples . . . 891 Dalton A. Foster and Steven M. Taffet

7.2 Use of cDNA Arrays to Explore Gene Expression

in Genetically Manipulated Mice and Cell Lines . . . 907 Dumitru A. Iacobas, Sandra Iacobas and David C. Spray

7.3 Using Antibody Arrays to Detect Protein-Protein Interactions . . . 916 Heather S. Duffy, Ionela Iacobas, David C. Spray and Anthony W. Ashton

7.4 Surface Plasmon Resonance as a Method to Study the Kinetics

and Amplitude of Protein-Protein Binding . . . 936 Brian D. Lang, Mario Delmar and Wanda Coombs

7.5 How to Solve a Protein Structure by Nuclear Magnetic Resonance –

The Connexin43 Carboxyl Terminal Domain . . . 948 Paul L. Sorgen

7.6 Managing a Transgenic Mouse Colony: A Guide for the Novice . . . 959 Robert H. Quinn and Karen L. Vikstrom

Appendix

Colour Plates . . . 979 Index . . . 997

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List of Abbreviations ^XI 0.00

List of Abbreviations

AP action potential

1D one-dimensional

2D two-dimensional

3D three-Dimensional 5-HT serotonin α1AR α1-Adrenoceptor

AADC aromatic L-amino acid decarboxylase ABC avidin-biotin peroxidase complex AC adenylyl cyclase

Ach acetylcholine ACT activated clotting time AEC 3-amino-9-ethylcarbazole AF atrial fibrillation

AMI acute myocardial infarction

Ao Aorta

APHIS Agriculture Animal and Plant Health Inspection Service

Ara-C cytosine-β-D-arabinofuranoside ARI activation-recovery-interval AT-1 angiotensin type 1 receptor AT-2 angiotensin type 2 receptor

B0 the static magnetic field β3-AR β3-adrenoceptors BCIP 5-bromo-4-chloro-3-

indolylphosphate BCL basic cycle length BDM 2,3-butanedione-monoxime

BM bone marrow

BrdU 5-bromo-2-desoxyuridine BSA bovine serum albumin BSA biotin-streptavidin BSS balanced salt saline BTP breakthrough-points

C capacitance

CABG coronary artery bypass grafting CARB carbachol

CARD catalyzed reporter deposition CCD charge-coupled device CDC Centers for Disease Control

CF coronary flow

CHO Chinese hamster ovary cells CK creatine kinase

CK/CKMB cCreatine kinase/cardiospecific CK CK-MB cardiospecific CK

CN cellulose nitrate

CNBD cyclic nucleotide-binding domain CNS crystallography & NMR systems CO cardiac output

COMT catechol-O-methyltransferase COS cells kidney cells from the monkey

Cercopithecus aethiops CPB cardiopulmonary bypass CS cell signaling

CSD cell cycle-shape-differentiation CV superior caval vein

Cx43 connexin43

CY cytoskeletal

DA diacetate

DAB diaminobenzidine tetrahydro- chloride

DAFs diaminofluoresceins DAF-T triazole-fluoresceins Daun daunorubicin

DβH dopamine β-hydroxylase dd double-distilled

DDSA dodecenyl succinic anhydride DEPC diethylpyrocarbonate

DMEM Dulbecco’s modified Eagle medium DMEM-F12 Dulbecco’s Modified Medium DMP tri(dimethylaminomethyl)phenol DMSO dimethylsulfoxide

Dox doxorubicin

dP/dt_max maximum rate of left ventricular pressure development dP/dt_min maximum relaxation velocity dP/dt_min minimum rate of pressure

development

DSEVC discontinuous single electrode voltage clamp amplifier DSS Dahl salt sensitive rat DTT dithiothreitol

EBNA-LP Epstein-Barr virus nuclear antigen- leader protein

EBs embryoid bodies EC embryonal carcinoma EC endothelial cells ECG electrocardiogram

ECL HRP enhanced chemiluminescence horseradish peroxidase EDC 1-Ethyl 3-3-dimethylaminopropyl

carbodiimide EDP end-diastolic pressure EDTA ethylenediaminetetraacetate EGF epidermal growth factor

EGFP enhanced green fluorescent protein EnMet Energy and Metabolism

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0 XII List of Abbreviations

eNOS endothelial nitric oxide synthase EPC endothelial progenitor cells

Epi epirubicin

ERP effective refractory period

ES embryonic stem

ESI electrospray-ionisation ESTs expressed sequence tags ET-1 endothelin-1

ET_AR endothelin-1 receptor subtype A

EtOH ethanol

FACS fluorescence activated cell sorter FCS foetal calf serum

FISH fluorescence in situ hybridisation FITC- fluorescein-isothiocyanate- FOC force of contraction FP field potentials

Gab Grb-associated binder GAP GTPase-activating protein GFP green fluorescent protein GPCR G protein coupled receptor

HBSS Hank’s balanced salt solution HCN channels hyperpolarisation-activated cyclic

nucleotide-gated cation channels HE hematoxylin-eosin

HEK human embryonic kidney cells HEPA-filter high efficiency particulate air filter HEPES-Na 2-[4-(2-Hydroxyethyl)-1-piperazinyl]-

ethanesulphonic acid sodium salt

HR Heart rate

HRP horseradish peroxidase HSC haematopoietic stem cells HSQC ¹H¹⁵N-heteronuclear single quantum

coherence

HW heart weight

IACUC Institutional Animal Care and Use Committees

IC intracoronary

ICM inner cell mass

IMAC immobilized metal ion affinity chromatography

IMDM Iscove’s modified Dulbecco´s medium

IP intraperitoneal

IPPV intermittent positive pressure ventilation

ITO indium-tin oxide

IUPAC The International Union of Pure and Applied Chemistry

IV intravenous

JAE junction-adhesion-extracellular matrix

Kv-channels voltage-gated potassium channels

LA left atrium

LAD left anterior descending coronary artery

LCX left circumflex artery LDH lactate dehydrogenase LIF leukemia inhibitory factor LV left ventricle

LVDP left ventricular developed pressure LVEF left ventricular ejection fraction LVP left ventricular pressure

MACS magnetic activated cell sorting MALDI matrix-assisted-laser-desorption-

ionisation MAO monoamine oxidase MAP mean arterial pressure MAP mitogen activated protein MAP monophasic action potentials MAPC multipotent adult progenitor cell

Mb myoglobin

MBF myocardial blood flow MEA microelectrode arrays MEF mouse embryonic fibroblast MEM non-essential amino acids MHC myosin heavy chain MI myocardial infarction MLP muscle LIM-domain protein MNC mononucleated cells MOI multiplicity of infection MRI magnetic resonance imaging MS mass spectrometry MS/MS tandem mass spectrometry MSC mesenchymal stem cells

NA noradrenalin

NAT nordadrenaline transporter NBT nitro blue tetrazolium chloride NCS newborn calf serum NHS N-hydroxysuccinimide NMR nuclear magnetic resonance NO nitric oxide

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List of Abbreviations ^XIII 0.00

NOE nuclear overhauser effect

NOS NO-synthases

Ntg non-transgenic

OG organelle genetics

PAP peroxidase anti-peroxidase PAR1 thrombin activated protease receptor

type 1

PBS phosphate buffered saline PCNA proliferating nuclear antigen PCR polymerase chain reaction PD proportional distance PDA photodiode arrays

PDEA (2-(2-pyridinylthio)ethanamine hydrochloride)

PDGF platelet derived growth factor

PE phycoerythrin

PE phenylephrine

PEEP positive end expiratory pressure PEG polyethylene glycol

PEI polyethylenimine

PenStrep penicillin/streptomycin solution PET positron emission

PFA para-formaldehyde P-I proportional integral PIPES piperazine-N’-N’-bis[2-ethane-

sulfonic acid

PKA cAMP-dependent protein kinase PL intrapleural

PLC-β β-isoforms of phospholipase C PLD phospholipase D

PMMA polymethylmethacrylate PNMT phenylethanolamine-N-methyl-

transferase PPM parts per million

PQ atrioventricular conduction time PSD post source decay

PTCA percutaneous transluminal coronary angioplasty

PTK protein tyrosine kinase PTP protein tyrosine phosphatase PVDF polyvinylidendifluoride

QTc frequency corrected QT time

R resistance

RA right atrium RCA right coronary artery

RCX circumflex artery

REV relative estimated (transcription) variability

RGS regulators of G protein signalling ROS reactive oxygen species

RP reverse-phase

rpm revolutions per minute

RT room temperature

RT reverse transcription RT_1/2 relaxation halftime RU response units

S/N signal-to-noise ratio SAN sino-atrial node

SC subcutaneous

SCX strong cation exchange SECV single electrode voltage clamp sGC soluble guanyl cyclase SHR spontaneously hypertensive rats SMC smooth muscle cell

SM-MHC smooth muscle myosin heavy chain SNR signal-to-noise ratio

SO Symmetrical Optimum SP side population SP systolic pressure SPF specific-pathogen-free SPR surface plasmon resonance SSC standard saline citrate synthase L-NNA

T tesla

T1 transport into the cell T2 transport within the cell TAT total activation time

TB Tris buffer

TBS Tris-buffered saline

TBST 150 mM NaCl, 25 mM Tris-HCl, 0.05%

Tween20, pH 7.5 TD trypsin-diluted TdP torsade de pointes TE expiratory ventilation time TEA tetraethylammonium aspartate TEVC two-electrode voltage clamp

Tg transgenic

TH tyrosine hydroxylase TI inspiratory ventilation time TMR transmyocardial laser

revascularisation TPP time to peak pressure TR transcriptional

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0 XIV List of Abbreviations

Tris trishydroxymethylaminomethane TSA tyramide signal amplification TSP 3-(Trimethylsilyl)- Propionic acid-D4 tTA tetracycline-controlled transactivation TTX tetrodotoxin

USDA United States Department of Agri- culture

VDCC voltage dependent calcium channel V_L longitudinal ventricular propagation

velocity

VMAT vesicular monoamine transporter V_rev reversal potential

VT transverse ventricular propagation velocity

VWF von Willebrand factor

WPW Wolff-Parkinson-White syndrome

WT wildtype

YAC yeast artificial chromosome

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List of Contributors ^XV 0.00

List of Contributors

Allessie, Maurits A.

University Maastricht Department of Physiology PO Box 616

6200 MD Maastricht The Netherlands

Ashton, Anthony W.

Albert Einstein College of Medicine

Department of Neuroscience Bronx, New York 10461 USA

Aupperle, Heike Universität Leipzig Herzzentrum Klinik für Herzchirurgie Strümpellstraße 39 04289 Leipzig Germany

Berkels, Reinhard Universität zu Köln Institut für Pharmakologie Gleueler Straße 24 50931 Köln Germany

Billman, George E.

The Ohio State University Department of Physiology and Cell Biology 304 Hamilton Hall 1645 Neil Ave.

Columbus OH 43210-1218 USA

Blaauw, Yuri University Maastricht Department of Physiology PO Box 616

Bloch, Wilhelm

Deutsche Sporthochschule Köln Institut für Kreislaufforschung und Sportmedizin

Carl-Diem-Weg 6 50933 Köln Germany

Brandt, Mathias C.

Universität zu Köln Klinik für Innere Medizin III Joseph-Stelzmann Straße 9 50924 Köln

Germany

Braun-Dullaeus, Ruediger C.

Technische Universität Dresden Klinik für Innere Medizin Fetscherstraße 76 01307 Dresden Germany

Brede, Marc

Bayerische Julius-Maximilians- Universität Würzburg Institut für Pharmakologie und Toxikologie Versbacher Straße 9 97078 Würzburg Germany

Coombs, Wanda

SUNY Upstate Medical University Department of Pharmacology 766 Irving Ave.

Decking, Ulrich

Heinrich-Heine-Universität Universitätsklinikum Institut für Herz- und Kreislauf- physiologie

Universitätsstraße 1 40225 Düsseldorf Germany

Delmar, Mario

SUNY Upstate Medical University Department of Pharmacology 766 Irving Ave.

Dhein, Stefan Universität Leipzig Herzzentrum Klinik für Herzchirurgie Strümpellstraße 39 04289 Leipzig Germany

Didié, Michael

Universitätsklinikum Hamburg- Eppendorf

Zentrum für Experimentelle Medizin

Institut für Experimentelle und Klinische Pharmakologie Martinistraße 52

20246 Hamburg Germany

Doll, Nicolas Universität Leipzig Herzzentrum Klinik für Herzchirurgie Strümpellstraße 39 04289 Leipzig Germany

Duffy, Heather S.

Egert, Ulrich Universität Freiburg Neurobiologie und Biophysik Institut für Biologie III Schänzlestraße 1 79104 Freiburg Germany

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0 XVI List of Contributors

El-Armouche, Ali

Er, Fikret Universität zu Köln Klinik für Innere Medizin III Joseph-Stelzmann-Straße 9 50924 Köln

Germany

Eschenhagen, Thomas Universitätsklinikum Hamburg- Eppendorf

Fabritz, Larissa

Universitätsklinikum Münster Medizinische Klinik und Poliklinik C

Kardiologie und Angiologie und Institut für Arteriosklerose- forschung

Albert-Schweitzer-Straße 33 48149 Münster

Germany

Falk, Volkmar Universität Leipzig Herzzentrum Klinik für Herzchirurgie Strümpellstraße 39 04289 Leipzig Germany

Fast, Vladimir G.

University of Alabama at Birmingham

1670 University Blvd., VH B126 Birmingham, AL 35294 USA

Fleischmann, Bernd

Rheinische-Friedrich-Wilhelms- Universität Bonn

Institut für Physiologie I Argelander Straße 2a 53115 Bonn Germany

Franz, Michael Georgetown University and Veterans Administrations Medical Centers

Departments of Cardiology and Pharmacology 50 Irving Street, NW Washington DC 20422 USA

Foster, Dalton A.

SUNY Upstate Medical University Department of Microbiology and Immunology

750 E. Adams St Syracuse, NY 13210 USA

Garbade, Jens Universität Leipzig Herzzentrum Klinik für Herzchirurgie Strümpellstraße 39 04289 Leipzig Germany

Gissel, Cornelia Universität zu Köln Institut für Neurophysiologie Robert-Koch-Straße 39 50931 Köln

Germany

Hein, Lutz

Hein, Peter

Hescheler, Jürgen Universität zu Köln Institut für Neurophysiologie Robert-Koch-Straße 39 50931 Köln

Germany

van der Heyden, Marcel A.G.

University Medical Center Department of Medical Physiology

Universiteitsweg 100 3584 CG Utrecht The Netherlands

Hoffmann, Christian Universität zu Köln Zentrum Anatomie Institut I für Anatomie Joseph-Stelzmann-Straße 9 50931 Köln

Germany

Hohlfeld, Thomas Universität Düsseldorf Institut für Pharmakologie Moorenstraße 5 40225 Düsseldorf Germany

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List of Contributors ^XVII 0.00

Hoppe, Uta C.

Universität zu Köln Klinik für Innere Medizin III Joseph-Stelzmann Str. 9 50924 Köln

Germany

Iacobas, Dumitru A.

Iacobas, Ionela Albert Einstein College of Medicine

Iacobas, Sandra Albert Einstein College of Medicine

Janssen, Eveline Universität zu Köln Zentrum Anatomie Institut I für Anatomie Joseph-Stelzmann-Straße 9 50931 Köln

Germany

Kaiser, Björn

Universitätsklinikum Essen Institut für Pharmakologie Hufelandstraße 55 45122 Essen Germany

Kirchhof, Paulus

Universitätsklinikum Münster Medizinische Klinik und Poliklinik C

Albert-Schweitzer-Straße 33 48129 Münster

Germany

van Koppen, Chris J.

Universitätsklinikum Essen Institut für Pharmakologie Hufelandstraße 55 45122 Essen Germany

Korkmaz, Yüksel Universität zu Köln Zentrum Anatomie Institut I für Anatomie Joseph Stelzmann Straße 9 50931 Köln

Germany

Lang, Brian D.

Biacore Inc.,

Piscataway, New Jersey USA

Lehr, Stefan

Heinrich-Heine-Universität Düsseldorf

Institut für Klinische Biochemie und Pathobiochemie Diabetes Forschungszentrum Leibniz-Zentrum für Diabetes- Forschung

Auf’m Hennekamp 65 40225 Düsseldorf Germany

Leineweber, Kirsten Universitätsklinikum Essen Institut für Pathophysiologie Hufelandstraße 55 45147 Essen Germany

Lindner, Michael Universität zu Köln Klinik für Innere Medizin III Joseph-Stelzmann-Straße 9 50924 Köln

Germany

Linz, Klaus Grünenthal GmbH Safety Pharmacology Zieglerstraße 6 52078 Aachen Germany

Melnychenko, Ivan

Merx, Marc W.

Universitätsklinikum Aachen Medizinische Klinik 1 Kardiologie und Pneumologie Pauwelstraße 30

52074 Aachen Germany

Meyer, Rainer

Rheinische-Friedrich-Wilhelms- Universität Bonn

Institut für Physiologie II Wilhelmstraße 31 53111 Bonn Germany

Meyer, Thomas Multi Channel Systems MCS GmbH

Aspenhaustraße 21 72770 Reutlingen Germany

Meyer, Veronika R.

EMPA St. Gallen Eidgenössische Materialprüfungs- und Forschungsanstalt Metrologie in der Chemie Lerchenfeldstrasse 5 9014 St. Gallen Switzerland

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XVIII

0 List of Contributors

Michels, Guido Universität zu Köln Klinik für Innere Medizin III Joseph-Stelzmann-Straße 9 50924 Köln

Germany

Michel, Martin C.

University of Amsterdam Academic Medical Center Dept. Pharmacology

& Pharmacotherapy Meibergdreef 15 1105 AZ Amsterdam The Netherlands

Morawietz, Henning Technische Universität Dresden Medizinische Klinik

und Poliklinik III Bereich Gefäßendothel/

Mikrozirkulation Fetscherstraße 74 01307 Dresden Germany

Nierhoff, Dirk Universität zu Köln Institut für Neurophysiologie Robert-Koch-Straße 39 50931 Köln

Germany

Noll, Thomas

Justus-Liebig-Universität Giessen Physiologisches Institut Aulweg 129

35392 Giessen Germany

Offermanns, Stefan Universität Heidelberg Pharmakologisches Institut Im Neuenheimer Feld 366 69120 Heidelberg Germany

Peters, Saskia C.

Philipp, Melanie Universität Zürich Institut für Zoologie Winterthurerstrasse 190 8057 Zürich

Switzerland

Piper, Hans Michael

Polder, Hans Reiner npi electronic GmbH Hauptstraße 96 71732 Tamm Germany

Quinn, Robert H.

SUNY Upstate Medical University Department of Laboratory Animal Resources 750 East Adams Street Syracuse, NY 13210 USA

Rastan, Ardawan J.

Universität Leipzig Klinik für Herzchirurgie Herzzentrum Strümpellstraße 39 04289 Leipzig Germany

Ravens, Ursula

Technische Universität Dresden Institut für Pharmakologie und Toxikologie Fetscherstraße 74 01307 Dresden Germany

Reis, Anna

Rösen, Peter

Institut für Klinische Biochemie und Pathobiochemie Leibniz-Zentrum für Diabetes- Forschung

Rösen, Renate Universität zu Köln Institut für Pharmakologie Gleueler Straße 24 50931 Köln Germany

Rosenkranz, Anke Universität zu Köln Institut für Pharmakologie Gleueler Straße 24 50931 Köln Germany

Rozemuller, Henk Jordan Laboratory University Medical Center Department of Heamatology Universiteitsweg 100 3584 CG Utrecht The Netherlands

Sachinidis, Agapios Universität zu Köln Institut für Neurophysiologie Robert-Koch-Straße 39 50931 Köln

Germany

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List of Contributors ^XIX 0.00

Salameh, Aida Universität Leipzig Klinik I für Innere Medizin Johannisallee 32 04103 Leipzig Germany

Schlüter, Klaus-Dieter Justus-Liebig-Universität Giessen

Physiologisches Institut Aulweg 129

Schneider, Katja Universität Leipzig Herzzentrum Klinik für Herzchirurgie Strümpellstraße 39 04289 Leipzig

Schotten, Ulrich University Maastricht Department of Physiology PO Box 616

Schrader, Jürgen Heinrich-Heine-Universität Düsseldorf

Institut für Physiologie Postfach 10 10 07 40001 Düsseldorf Germany

Schubert, Andreas Universität Leipzig Herzzentrum Russenstraße 19 04289 Leipzig Germany

Schubert, Rudolf Universität Rostock Institut für Physiologie Postfach 10 08 88 18055 Rostock Germany

Sedding, Daniel G.

Justus-Liebig-Universität Giessen Medizinische Klinik I

Innere Medizin/Kardiologie Klinikstraße 36

Sorgen, Paul. L.

University of Nebraska Medical Center

Department of Biochemistry and Molecular Biology Omaha, NE 68198 USA

Spray, David C.

Stahl, Fabian Universität Leipzig Herzzentrum Klinik für Herzchirurgie Strümpellstraße 39 Germany 04289 Leipzig

Steinritz, Dirk Universität zu Köln Zentrum Anatomie Institut I für Anatomie Joseph-Stelzmann-Straße 9 50931 Köln

Germany

Taffet, Steven M.

SUNY Upstate Medical University Department of Microbiology and Immunology

750 E. Adams St Syracuse, NY 13210 USA

Tárnok, Attila Universität Leipzig Herzzentrum

Klinik für Kinderkardiologie Strümpellstraße 39 04289 Leipzig Germany

Taubert, Dirk Universität zu Köln Institut für Pharmakologie Gleueler Straße 24 50931 Köln Germany

Tritthart, Helmut A.

Karl-Franzens-Universität Graz Institut für Medizinische Physik und Biophysik

Harrachgasse 21 8010 Graz Austria

Ullmann, Cris W.

Universität Leipzig Herzzentrum Klinik für Herzchirurgie Strümpellstraße 39 04289 Leipzig Germany

Vikstrom, Karen L.

SUNY Upstate Medical University Department of Laboratory Animal Resources 750 East Adams Street Syracuse, NY 13210 USA

Walther, Thomas Universität Leipzig Herzzentrum Klinik für Herzchirurgie Strümpellstraße 39 04289 Leipzig Germany

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0 XX List of Contributors

Weiß, Jürgen

Institut für Klinische Biochemie und Pathobiochemie Leibniz-Zentrum für Diabetes- Forschung

Weskamp, Martin npi electronic GmbH, Hauptstraße 96, 71732 Tamm Germany

Wettschureck, Nina Universität Heidelberg Pharmakologisches Institut Im Neuenheimer Feld 366 69120 Heidelberg Germany

Wettwer, Erich

Technische Universität Dresden Institut für Pharmakologie und Toxikologie Fetscherstraße 74 01307 Dresden Germany

Wieland, Thomas Universität Heidelberg Fakultät für Klinische Medizin Mannheim

Institut für Pharmakologie und Toxikologie Maybachstraße 14–16 68169 Mannheim Germany

Zacharowski, Kai Heinrich-Heine-Universität Düsseldorf

Universitätsklinikum Klinik für Anästhesiologie und Intensivmedizin Moorenstraße 5 40225 Düsseldorf Germany

Zimmermann, Wolfram- Hubertus