Wir danken Dr. Helio Salgado, Renata Lataro und Mauro de Oliveira, School of Medicine der Ribeirão Preto, Universität von Sao Paulo und Dr. Ben Janssen, Cardiovascular Research Institute Maastricht, Maastricht University, für die großzügige Unterstützung während der Setup-Prozess. Diese Arbeit wurde von Fondecyt Gewährung N ° 11090114 DEZA unterstützt

Protokoll wurde von der Ethikkommission der Facultad de Medicina Clinica Alemana-Universidad del Desarrollo zugelassen. I. Vorbereitung Dobutamin Infusion <ol><li> Man löst 10 mg Dobutamin in 20 ml steriles destilliertes Wasser, um eine Stammlösung von 500 ug / ml Dobutamin erhalten. Aliquotieren und bei -20 ° C. Diese Lösung kann für mindestens 3 Monate verwendet werden. </li><li> Auftauen eines Aliquots Dobutamin Stammlösung bei Raumtemperatur. </li><li> Verdünnen Dobutamin Stammlösung in steriler 0,9% NaCl, um Dobutamin funktionierende Lösung, die Konzentration wird anhand der Formel: Dobutamin (pg / ml) = Körpergewicht x 0,2. </li><li> Füllen Sie eine 1 ml Spritze 29Gx1 / 2 &quot;mit Dobutamin funktionierende Lösung. </li><li> Legen Sie die Nadel der Spritze in einen 20 cm PE-10 Röhre. </li><li> Passen Sie die Spritze in die Infusionspumpe nach den Anweisungen des Herstellers. </li><li> Richten Sie die Rampe Infusion in einem Schritt-Für-Schritt-Format mit einer Steigerung von 10 ul / min für jeden Schritt, für 6 Schritten. </li></ol> II. Vorbereiten Pressure Sensor <ol><li> Um Signaldrift, tauchen Drucksensor in sterilem Wasser bei 37 ° C zu minimieren für mindestens 15 min. Nicht einweichen den Katheter mehr als 0,5 cm tief, um zu verhindern, dass hydrostatische Druck wirkt sich auf die Drucksensor. </li><li> Elektronisch den Drucksensor kalibrieren, bei 25 und 100 mmHg. Elektrischen Eingang (V) mit Drucksignal (mmHg) umgewandelt. </li><li> Stellen Sie die Abtastrate von 2 k / s und nutzen Sie den Filter Tiefpass mit einer Grenzfrequenz von 100 Hz. Stellen Sie den Druck auf Null mmHg. </li><li> Markieren Sie den Katheter 15 mm von der Spitze. Entfernt, um das Herz von der Einleitungsstelle erreicht wurde geschätzt durch Echokardiographie, Erfassen der Anwesenheit des Katheters in den linken Ventrikel. </li></ol> III. Vorbereiten Maus für Katheterisierung <ol><li> Gewicht C57BL / 6 männlichen Mäusen, 30-32 Wochen alt. </li><li> Inject intraperitoneal 60 ug / kg Ketamin und 4 ug / kg Xylazin 1. Hinweis: Andere Anästhetika könnte verwendet werden, zum Beispiel: 350 - 450 ug / kg avertin, 50 ug / kg Pentobarbital oder 1,5 - 2% Isofluran 2-3. </li><li> Rasur am Hals mit einem elektrischen Rasierer. </li><li> Setzen Sie den narkotisierten Maus in Rückenlage auf einer vorgewärmten isothermen Heizplatte. Sichern Sie ihre Gliedmaßen mit Lochstreifen. </li><li> Führen Sie einen Zeh Prise vollständige Sedierung bestätigen. </li><li> Führen Sie eine rektale Sonde, um die Körpertemperatur zu überwachen. Mit vaselinized Sonde wird empfohlen. </li><li> Wenn Körpertemperatur von 37 unterscheidet ° C ± 0,5 ° C beträgt, wird es über die Heizplatte. </li><li> Setz Maus nahe der Schnauze Sauerstoffversorgung. </li><li> Platzieren Sie die Maus Halsbereich unter dem Stereomikroskop. </li></ol> IV. Data Acquisition <ol><li> In der LabChartPro 7-Software, wählen Sie einen Kanal für Druck Registrierung und einen Kanal für die Herzfrequenz (HR)Anmeldung. Für letztere, wählen Sie die Option &quot;Zyklische Messungen&quot; und Setup-Messung als Rate. </li><li> Für den Druckkanal den Maßstab: 0 bis 150 mmHg. </li><li> Für HR-Kanal eingestellt Skalenbereich: 200 bis 600 bpm. </li><li> Drücken Sie die Starttaste, um die Registrierung zu beginnen. </li><li> Kommentare einfügen angibt Verfahren durchgeführt werden, zum Beispiel: Anästhesie Verabreichung von Dobutamin Infusion, Dobutamin Konzentration starten, atmende ändert. </li></ol> V. Herzkatheter 4,5 <ol><li> Durchführen einer kleinen Einschnitt auf der rechten Seite in der Nähe des Kiefers. Mit Schere trennen die Haut-Muskel-Bindegewebe. </li><li> Führe eine longitudinale Dissektion (1,5 - 2 cm) auf der rechten Seite der Luftröhre. Trennen die Bindegewebe, Fett und Muskeln mit gekrümmten Zange, um die rechte Arteria carotis in der Nähe der Luftröhre freizulegen. </li><li> Platzieren eines Expanders im Tier rechts um die Halsschlagader freizulegen. PulsierendeDruck durch das Herz erzeugt erleichtert die Identifizierung der Arterie. Die Halsschlagader, die dunkel rot ist, ist auf der rechten Seite. </li><li> Trennen Sie die Arterie von benachbarten Geweben mit gebogenen Pinzette. Der Vagusnerv, der eine weiße Faden ähnelt, liegt entlang der Arterie. </li><li> Schneiden Sie ein 20 cm Stück 6/0 Seidenfaden und &quot;double&quot; ist. </li><li> Führen Sie das &quot;doppelte&quot; Faden unter der Arterie von links nach rechts. Den Faden abschneiden, um getrennte Enden zu erhalten. </li><li> Pass ein Drittel Gewinde (10 cm) unterhalb der Arterie. </li><li> Tie einen festen Knoten in dem Faden in der Nähe des Kopfes positioniert ist, und eine lose ein in der mehr distale Gewinde. </li><li> Tie eine lose Knoten in der Mitte Gewinde, und fixieren das rechte Ende des mittleren Faden nach Heizkissen mit einem Papierband. </li><li> Halten Halsschlagader durch Fallenlassen feuchter steriler 0,9% NaCl. Trocknen Sie überschüssige Flüssigkeit mit Wattestäbchen. </li><li> Dehnen Sie den Unterfaden mit einer Gefäßklemme Klemme. </li><li> Fixieren Sie die Position des hemostat Schere durch pTippen Sie die Haut des Bauches, Stretching den Oberfaden, um den Blutfluss zu verschließen. Überprüfen Sie, ob das Bindegewebe um die Arterie entfernt wurde. Die Arterie sollte voller Blut und beraubt Puls. Verhindern Fäden aus ein Drehmoment Kraft auf die Arterie. </li><li> Machen Sie einen Querschnitt nick in der Nähe der Unterseite der Arterie mit einem Vannas Mikro-Schere. Blutstropfen wird verschüttet werden. </li><li> Legen Sie den Katheter in die Halsschlagader. Achten Sie darauf, die gesamte Drucksensor einzuführen. Stellen Sie sicher, dass es keinen Blutverlust. </li><li> Schieben Sie vorsichtig den mittleren Faden Knoten, um den Katheter in Position zu halten. Nicht zu viel zu komprimieren, ist der Drucksensor sehr instabil. </li><li> Lassen Sie die hemostat Schere aus dem Tier Bauch. </li><li> Halten Sie den Katheter mit der Hand und drücken Sie den mittleren Faden, um den Blutverlust zu vermeiden. Hinweis: Arterie sollte voller Blut. </li><li> Starten Sie die Aufnahme Drucksignalen. </li><li> Wenn der Katheter im Inneren ist, derarteriellen Druck Signal schwankt von 60 bis 70 auf 100-120 mmHg. Die Form des Drucksignals ist in Abbildung 1.A. Hinweis gezeigt: wenn Sie interessiert sind, zu diesem Zeitpunkt können Sie arteriellen Druck aufnehmen, wenn das Signal für mindestens 5 min stabil. HR-Werte wurden aus der Druckwellenformen Prüfung eines Intervalls von 30 Sekunden des Aufzeichnungssignals erhalten. Es ist möglich, auch eine EKG Verfahren zur direkten Messung von HR, entsprechend den Untersuchungen Ziele. </li><li> Schieben Sie den Katheter bis zu einer Änderung in der Form des Drucksignals (Abbildung 1.B) zu beobachten. Sobald der Katheter im Inneren des linken Ventrikels, schwankt das Drucksignal von 0 bis 100 bis 120 mmHg. Wenn es schwierig ist, den Katheter gleiten kann, einzuklemmen tierischen Brust mit zwei Fingern. </li><li> Kontinuierlich zu steuern Atemfrequenz, Körpertemperatur, Anästhesie und Druck Signal. Alle von ihnen sollte stabil bleiben. </li></ol> VI. Die Infusion von Dobutamin <ol><li&gt; Für Jugularvene Kanülisier unbedingt abziehen des Fettgewebes um die Vene, um dessen Perforation zu verhindern. Das chirurgische Verfahren zum Venenverschluss ist ähnlich dem Verfahren für Halsschlagader gezeigt. </li><li> Einführung eines PE-10 Rohr in die Vene. Bestätigen Sie, dass der Blutfluss nicht blockiert rückwärts bewegt den Kolben der Spritze. </li><li> Dobutamin Infusion beginnt mit 10 ul / min und enden mit 60 ul / min. In jedem Schritt wird die Infusionsrate für 2 min 6 gehalten. </li><li> Nach dem letzten Dobutamin Dosis, einschläfern das Tier mit einer Überdosis der Anästhesie. </li></ol> VII. Data Analysis <ol><li> Für die Datenanalyse, wählen Sie den Abschnitt der aufgezeichneten Daten, die Sie interessieren. Achten Sie darauf, um ein Zeitintervall überlegen, wo Drucksignal ist stabil. </li><li> Wählen Sie das Setup-Symbol in der Blood Pressure Modul. Geben Sie die gewählte Art der Drucksignal. </li><li> Automatisch die LabChartPro 7 Software zeigt Mittelwert, Maximumund Minimalwerte für HR, systolischer Druck (P max), der diastolische Druck (P min), enddiastolische Druck (EDP), maximal Überdruck Entwicklung (dP / dtmax) und maximaler Unterdruck Entwicklung (dP / dt min). Darüber hinaus können auf der Herzparametern Druckspur darstellen. </li></ol>

<ol>
	<li>Hart, C. Y., Burnett, J. C., Redfield, M. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+avertin+versus+xylazine-ketamine+anesthesia+on+cardiac+function+in+normal+mice.">Effects of avertin versus xylazine-ketamine anesthesia on cardiac function in normal mice.</a> Am. J. Physiol. Heart Circ. Physiol. 281, H1938-H1945 (2001).</li><li>Janssen, B. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+anesthetics+on+systemic+hemodynamics+in+mice.">Effects of anesthetics on systemic hemodynamics in mice.</a> Am. J. Physiol. Heart Circ. Physiol. 287, H1618-H1624 (2004).</li><li>Lorenz, J. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+practical+guide+to+evaluating+cardiovascular,+renal,+and+pulmonary+function+in+mice.">A practical guide to evaluating cardiovascular, renal, and pulmonary function in mice.</a> Am. J. Physiol. Regul. Integr. Comp. Physiol. 282, R1565-R1582 (2002).</li><li>Lorenz, J. N., Robbins, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measurement+of+intraventricular+pressure+and+cardiac+performance+in+the+intact+closed-chest+anesthetized+mouse.">Measurement of intraventricular pressure and cardiac performance in the intact closed-chest anesthetized mouse.</a> Am. J. Physiol. 272, H1137-H1146 (1997).</li><li>Nemoto, S., DeFreitas, G., Carabello, B. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cardiac+catheterization+technique+in+a+closed-chest+murine+model.">Cardiac catheterization technique in a closed-chest murine model.</a> Contemp. Top. Lab Anim. Sci. 42, 34-38 (2003).</li><li>Daniels, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impaired+cardiac+functional+reserve+in+type+2+diabetic+db/db+mice+is+associated+with+metabolic,+but+not+structural,+remodelling.">Impaired cardiac functional reserve in type 2 diabetic db/db mice is associated with metabolic, but not structural, remodelling.</a> Acta Physiol. (Oxf). 200, 11-22 (2010).</li><li>De Celle, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-term+structural+and+functional+consequences+of+cardiac+ischaemia-reperfusion+injury+in+vivo+in+mice.">Long-term structural and functional consequences of cardiac ischaemia-reperfusion injury in vivo in mice.</a> Exp. Physiol. 89, 605-615 (2004).</li><li>Huntgeburth, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transforming+growth+factor+beta+oppositely+regulates+the+hypertrophic+and+contractile+response+to+beta-adrenergic+stimulation+in+the+heart.">Transforming growth factor beta oppositely regulates the hypertrophic and contractile response to beta-adrenergic stimulation in the heart.</a> PLoS One. 6, e26628 (2011).</li><li>Reddy, A. K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cardiac+function+in+young+and+old+Little+mice.">Cardiac function in young and old Little mice.</a> J. Gerontol. A. Biol. Sci. Med. Sci. 62, 1319-1325 (2007).</li><li>Christoffersen, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cardiac+lipid+accumulation+associated+with+diastolic+dysfunction+in+obese+mice.">Cardiac lipid accumulation associated with diastolic dysfunction in obese mice.</a> Endocrinology. 144, 3483-3490 (2003).</li></ol>

Keine Interessenskonflikte erklärt.

Cardiac Stresstest Dobutamin induziert und überwacht durch Herzkatheter ist aufwändig. Dennoch, nachdem das Protokoll hier zu beschreiben und mit einer kurzen Zeit der Ausbildung ist es möglich, bis sechs hämodynamischen Parameter in einem einzigen Experiment, dass dauert ca. 1 Stunde zu beurteilen. Die kritischen Schritte des Protokolls hier präsentiert sind die Kanülierungen der Blutgefäße. Bezüglich der Kanülierung Halsschlagader, sollte der Einschnitt durchgeführt tief genug s...

<table border="1"><tbody><tr><td></td><td></td><td></td><td> Reagenzien </td></tr><tr><td> PE-50/10 </td><td> Warner Instruments </td><td> 64-0752 </td><td></td></tr><tr><td> Seidenfaden 6/0 HR17 </td><td> Tagum </td><td> SN0713K </td><td></td></tr><tr><td> Xylacin 20 mg / ml </td><td> Laboratorio Centrovet </td><td></td><td></td></tr><tr><td> Ketamin 100 mg / ml </td><td> Drag Pharma </td><td></td><td></td></tr><tr><td> Natriumchlorid 0,9% </td><td> Lab Sanderson SA </td><td></td><td></td></tr><tr><td> Dobutamin Hydrochlorid </td><td> Sigma-Aldrich </td><td> D0676 </td><td></td></tr><tr><td> Spritze U-100 Insulin 29G x ½ &quot; </td><td> Terumo Medical Co. </td><td></td><td></td></tr><tr><td> Dissektionszangen Micro 11,5 cm Style 7 </td><td> LawtonMedizintechnik </td><td> 09-0959 </td><td></td></tr><tr><td> Graefe Pinzette Cvd 0.7mm 7cm </td><td> Lawton Medizintechnik </td><td> 62-0263 </td><td></td></tr><tr><td> Clamps Dieffenbach Bulldog Cl Str. 38 mm </td><td> Lawton Medizintechnik </td><td> 60-010 </td><td></td></tr><tr><td> Vannas Schere 8 cm Str Fh </td><td> Lawton Medizintechnik </td><td> 63-1400 </td><td></td></tr><tr><td></td><td></td><td></td><td> Ausrüstung </td></tr><tr><td> SPR-671 Mikro-Tip-Katheter Druck </td><td> Millar Instrumenten </td><td> 840-6719 </td><td></td></tr><tr><td> PCU-2000 Pressure Control Unit </td><td> Millar Instrumenten </td><td> 880-0129 </td><td></td></tr><tr><td> PowerLab 4/30 </td><td> ADInstruments Pty Ltd </td><td> ML866 </td><td></td></tr><tr><td> LabChartPro 7 </td><td> ADInstruments Pty Ltd </td><td> MLU260 / 7 </td><td></td></tr><tr><td> Legato200 Infusionspumpe </td><td> KdScientific </td><td> KD-KDS210P </td><td></td></tr><tr><td> TCAT-2LV Temperaturregler und isotherme Heizplatte </td><td> PhysiTemp Instruments Inc. </td><td></td><td></td></tr><tr><td> Medical Sauerstoffversorgung </td><td> Indura </td><td></td><td></td></tr><tr><td> Rektalsonde </td><td> ADInstruments Pty Ltd </td><td> MLT1404 </td><td></td></tr><tr><td> Trinokular Mikroskop, axiale Beleuchtung </td><td> LW Scientific </td><td> Z2B-TRI-ETNE, ILP-1502-LTS1, ILP-1502-DGGF </td><td></td></tr></tbody></table>

cardiac stress test induced by dobutamine and monitored by cardiac catheterization in mice

Dobutamin ist ein β-adrenergen Agonisten mit einem höheren Affinität für den Rezeptor im Herzen (β 1) als für Rezeptoren in den Arterien (β 2) ausgedrückt. Wenn systemisch verabreicht, erhöht es Herzbedarf. So entlarvt Dobutamin anormalen Rhythmus oder ischämische Bereiche potentielles Risiko Herzinfarkt. Überwachung der Herzfunktion während einer Belastungs-Test kann entweder durch ecocardiography oder Herzkatheter durchgeführt werden. Letzteres ist eine invasive, aber genauer und informative Technik, die der ehemalige. Cardiac Stress-Test induziert durch Dobutamin und überwacht durch Herzkatheter wie hier beschrieben durchgeführt ermöglicht, in einem einzigen Experiment die Messung der folgenden hämodynamischen Parameter: Herzfrequenz (HR), systolischer Druck, diastolischer Druck, end-diastolische Druck, maximal Überdruck Entwicklung (dP / dtmax) und die maximale negative Presseure Entwicklung (dp / dt MIN), an der Basislinie und unter Bedingungen steigender Dosen von Dobutamin. Wie zu erwarten, bei normalen Mäusen beobachteten wir eine Dobutamin dosisabhängigen Anstieg der HR, dP / dt max und dP / dt min. Darüber hinaus bei der höchsten getesteten Dosis (12 ng / g / min) der kardiale Dekompensation der fettreichen Diät-induzierten adipösen Mäusen war entlarvt.

Der arterielle Druck wird durch Signal systolischen und diastolischen Druck definiert. Wenn der Drucksensor innerhalb des linken Ventrikels ist, dessen Druck (LVP) Wellenform wird durch einen Abfall auf Null des diastolischen Drucks und dem Auftreten des linken Ventrikels vor Vorhofkontraktion Kontraktion (Abbildung 1). An Ausgangszustand hatte Ketamin-Xylazin anästhesiert normalen Mäusen HR von 280 ± 24, P max von 107 ± 8, P min von 5 ± 1, EDP nach 14 ± 2, dP / dt max</...

Watch this Scientific Journal Video about Cardiac Stress Test Induced by Dobutamine and Monitored by Cardiac Catheterization in Mice at JoVE.com

Cardiac Stress Test Induced by Dobutamine and Monitored by Cardiac Catheterization in Mice

Wir beschreiben das Protokoll eine kardiale Stresstest Dobutamin induziert und überwacht durch Herzkatheter bei normalen Mäusen durchzuführen. Auch zeigen wir ihre Anwendung auf subklinische Herzerkrankungen in fettreichen Diät-induzierten adipösen Mäusen zu entlarven.

Cardiac Stress Test durch Dobutamin induziert und Überwacht durch Herzkatheter in Mäuse

Dobutamine is a &beta;-adrenergic agonist with an affinity higher for receptor expressed in the heart (&beta;1) than for receptors expressed in the ...

cardiac-stress-test-induced-dobutamine-monitored-cardiac

Research

JoVE Journal

Medicine

17.9K Aufrufe.  Clínica Alemana Universidad del Desarrollo. Wir beschreiben das Protokoll eine kardiale Stresstest Dobutamin induziert und überwacht durch Herzkatheter bei normalen Mäusen durchzuführen. Auch zeigen wir ihre Anwendung auf subklinische Herzerkrankungen in fettreichen Diät-induzierten adipösen Mäusen zu entlarven.

Serie: Cardiac Stress Test Induced by Dobutamine and Monitored by Cardiac Catheterization in Mice

Assessment of Right Ventricular Structure and Function in Mouse Model of Pulmonary Artery Constriction by Transthoracic Echocardiography

Emerging clinical data support the notion that RV dysfunction is critical to the pathogenesis of cardiovascular disease and heart failure1-3. Moreover, the RV is significantly affected in pulmonary diseases such as pulmonary artery hypertension (PAH). In addition, the RV is remarkably sensitive to cardiac pathologies, including left ventricular (LV) dysfunction, valvular disease or RV infarction4. To understand the role of RV in the pathogenesis of cardiac diseases, a reliable and noninvasive method to access the RV structurally and functionally is essential.
A noninvasive trans-thoracic echocardiography (TTE) based methodology was established and validated for monitoring dynamic changes in RV structure and function in adult mice. To impose RV stress, we employed a surgical model of pulmonary artery constriction (PAC) and measured the RV response over a 7-day period using a high-frequency ultrasound microimaging system. Sham operated mice were used as controls. Images were acquired in lightly anesthetized mice at baseline (before surgery), day 0 (immediately post-surgery), day 3, and day 7 (post-surgery). Data was analyzed offline using software.
Several acoustic windows (B, M, and Color Doppler modes), which can be consistently obtained in mice, allowed for reliable and reproducible measurement of RV structure (including RV wall thickness, end-diastolic&#160;and end-systolic dimensions), and function (fractional area change, fractional shortening, PA peak velocity, and peak pressure gradient) in normal mice and following PAC.
Using this method, the pressure-gradient resulting from PAC was accurately measured in real-time using Color Doppler mode and was comparable to direct pressure measurements performed with a Millar high-fidelity microtip catheter. Taken together, these data demonstrate that RV measurements obtained from various complimentary views using echocardiography are reliable, reproducible and can provide insights regarding RV structure and function. This method will enable a better understanding of the role of RV cardiac dysfunction.

Right ventricle (RV) dysfunction is critical to the pathogenesis of cardiovascular disease, yet limited methodologies are available for its evaluation. Recent advances in ultrasound imaging provide a noninvasive and accurate option for longitudinal RV study. Herein, we detail a step-by-step echocardiographic method using a murine model of RV pressure overload.

Beurteilung der rechtsventrikulären Struktur und Funktion in Maus-Modell der Lungenarterie Verengung durch transthorakale Echokardiographie

Emerging clinical data support the notion that RV dysfunction is critical to the pathogenesis of cardiovascular disease and heart failure1-3. Moreover, ...

Forschung

Medizin

Noninvasive Assessment of Cardiac Abnormalities in Experimental Autoimmune Myocarditis by Magnetic Resonance Microscopy Imaging in the Mouse

Myocarditis is an inflammation of the myocardium, but only ~10% of those affected show clinical manifestations of the disease. To study the immune events of myocardial injuries, various mouse models of myocarditis have been widely used. This study involved experimental autoimmune myocarditis (EAM) induced with cardiac myosin heavy chain (Myhc)-&#945; 334-352 in A/J mice; the affected animals develop lymphocytic myocarditis but with no apparent clinical signs. In this model, the utility of magnetic resonance microscopy (MRM) as a non-invasive modality to determine the cardiac structural and functional changes in animals immunized with Myhc-&#945; 334-352 is shown. EAM and healthy mice were imaged using a 9.4 T (400 MHz) 89 mm vertical core bore scanner equipped with a 4 cm millipede radio-frequency imaging probe and 100 G/cm triple axis gradients. Cardiac images were acquired from anesthetized animals using a gradient-echo-based cine pulse sequence, and the animals were monitored by respiration and pulse oximetry. The analysis revealed an increase in the thickness of the ventricular wall in EAM mice, with a corresponding decrease in the interior diameter of ventricles, when compared with healthy mice. The data suggest that morphological and functional changes in the inflamed hearts can be non-invasively monitored by MRM in live animals. In conclusion, MRM offers an advantage of assessing the progression and regression of myocardial injuries in diseases caused by infectious agents, as well as response to therapies.

This study demonstrates the successful establishment of magnetic resonance microscopy imaging as a non-invasive tool to assess the cardiac abnormalities in mice affected with autoimmune myocarditis. The data indicate that the technique can be used to monitor the disease-progression in live animals.

Nicht-invasive Beurteilung der Herzfehlbildungen in experimentellen autoimmunen Myokarditis von Magnetic Resonance Imaging Microscopy in der Maus

Myocarditis is an inflammation of the myocardium, but only ~10% of those affected show clinical manifestations of the disease. To study the immune events ...

Early Detection of Drug-Induced Renal Hemodynamic Dysfunction Using Sonographic Technology in Rats

The kidney normally functions to maintain hemodynamic homeostasis and is a major site of damage caused by drug toxicity. Drug-induced nephrotoxicity is estimated to contribute to 19- 25% of all clinical cases of acute kidney injury (AKI) in critically ill patients. AKI detection has historically relied on metrics such as serum creatinine (sCr) or blood urea nitrogen (BUN) which are demonstrably inadequate in full assessment of nephrotoxicity in the early phase of renal dysfunction. Currently, there is no robust diagnostic method to accurately detect hemodynamic alteration in the early phase of AKI while such alterations might actually precede the rise in serum biomarker levels. Such early detection can help clinicians make an accurate diagnosis and help in in decision making for therapeutic strategy. Rats were treated with Cisplatin to induce AKI. Nephrotoxicity was assessed for six days using high-frequency sonography, sCr measurement and upon histopathology of kidney. Hemodynamic evaluation using 2D and Color-Doppler images were used to serially study nephrotoxicity in rats, using the sonography. Our data showed successful drug-induced kidney injury in adult rats by histological examination. Color-Doppler based sonographic assessment of AKI indicated that resistive-index (RI) and pulsatile-index (PI) were increased in the treatment group; and peak-systolic velocity (mm/s), end-diastolic velocity (mm/s) and velocity-time integral (VTI, mm) were decreased in renal arteries in the same group. Importantly, these hemodynamic changes evaluated by sonography preceded the rise of sCr levels. Sonography-based indices such as RI or PI can thus be useful predictive markers of declining renal function in rodents. From our sonography-based observations in the kidneys of rats that underwent AKI, we showed that these noninvasive hemodynamic measurements may consider as an accurate, sensitive and robust method in detecting early stage kidney dysfunction. This study also underscores the importance of ethical issues associated with animal use in research.

Early stage hemodynamic dysfunction is critical to the development of kidney disease. Yet, detection methodologies are limited. Recent advances in sonography provide a noninvasive, accurate option for early detection of kidney injury. This study outlines a step-by-step, sonographic methodology for detecting kidney dysfunction using a drug-induced nephrotoxicity rat model.

Früherkennung von Medikamenten-induzierten Nieren hämodynamischen Dysfunktion Mit sonographische Technologie bei Ratten

The kidney normally functions to maintain hemodynamic homeostasis and is a major site of damage caused by drug toxicity. Drug-induced nephrotoxicity is ...

Cardiac Catheterization in Mice to Measure the Pressure Volume Relationship: Investigating the Bowditch Effect

Animal models that mimic human cardiac disorders have been created to test potential therapeutic strategies. A key component to evaluating these strategies is to examine their effects on heart function. There are several techniques to measure in vivo cardiac mechanics (e.g., echocardiography, pressure/volume relations, etc.). Compared to echocardiography, real-time left ventricular (LV) pressure/volume analysis via catheterization is more precise and insightful in assessing LV function. Additionally, LV pressure/volume analysis provides the ability to instantaneously record changes during manipulations of contractility (e.g., &#946;-adrenergic stimulation) and pathological insults (e.g., ischemia/reperfusion injury). In addition to the maximum (+dP/dt) and minimum (-dP/dt) rate of pressure change in the LV, an accurate assessment of LV function via several load-independent indexes (e.g., end systolic pressure volume relationship and preload recruitable stroke work) can be attained. Heart rate has a significant effect on LV contractility such that an increase in the heart rate is the primary mechanism to increase cardiac output (i.e., Bowditch effect). Thus, when comparing hemodynamics between experimental groups, it is necessary to have similar heart rates. Furthermore, a hallmark of many cardiomyopathy models is a decrease in contractile reserve (i.e., decreased Bowditch effect). Consequently, vital information can be obtained by determining the effects of increasing heart rate on contractility. Our and others data has demonstrated that the neuronal nitric oxide synthase (NOS1) knockout mouse has decreased contractility. Here we describe the procedure of measuring LV pressure/volume with increasing heart rates using the NOS1 knockout mouse model.

This article describes the measurement of murine left ventricular function via pressure/volume analysis at different heart rates.

Herzkatheter in Mäuse, um den Druck Volumen Relationship Maßnahme: Untersuchung der Bowditch Effect

Animal models that mimic human cardiac disorders have been created to test potential therapeutic strategies. A key component to evaluating these ...

Myocardial Infarction in Neonatal Mice, A Model of Cardiac Regeneration

Myocardial infarction induced by coronary artery ligation has been used in many animal models as a tool to study the mechanisms of cardiac repair and regeneration, and to define new targets for therapeutics. For decades, models of complete heart regeneration existed in amphibians and fish, but a mammalian counterpart was not available. The recent discovery of a postnatal window during which mice possess regenerative capabilities has led to the establishment of a mammalian model of cardiac regeneration. A surgical model of mammalian cardiac regeneration in the neonatal mouse is presented herein. Briefly, postnatal day 1 (P1) mice are anesthetized by isoflurane and placed on an ice pad to induce hypothermia. After the chest is opened, and the left anterior descending coronary artery (LAD) is visualized, a suture is placed around the LAD to inflict myocardial ischemia in the left ventricle. The surgical procedure takes 10-15 min. Visualizing the coronary artery is crucial for accurate suture placement and reproducibility. Myocardial infarction and cardiac dysfunction are confirmed by triphenyl-tetrazolium chloride (TTC) staining and echocardiography, respectively. Complete regeneration 21 days post myocardial infarction is verified by histology. This protocol can be used to as a tool to elucidate mechanisms of mammalian cardiac regeneration after myocardial infarction.

This protocol describes a highly reproducible model of cardiac regeneration by surgical induction of myocardial infarction in the left ventricle of postnatal day 1 mice. The method involves induction of hypothermic anesthesia and ligation of the left anterior descending coronary artery.

Myokardinfarkts bei neugeborenen Mäusen, ein Modell von Cardiac Regeneration

Myocardial infarction induced by coronary artery ligation has been used in many animal models as a tool to study the mechanisms of cardiac repair and ...

A Model of Glaucoma Induced by Circumlimbal Suture in Rats and Mice

The circumlimbal suture is a technique for inducing experimental glaucoma in rodents by chronically elevating intraocular pressure (IOP), a well-known risk factor for glaucoma. This protocol demonstrates a step-by-step guide on this technique in Long Evans rats and C57BL/6 mice. Under general anesthesia, a "purse-string" suture is applied on the conjunctiva, around the equator and behind the limbus of the eye. The fellow eye serves as an untreated control. Over the duration of our study, which was a period of 8 weeks for rats and 12 weeks for mice, IOP remained elevated, as measured regularly by rebound tonometry in conscious animals without topical anesthesia. In both species, the sutured eyes showed electroretinogram features consistent with preferential inner retinal dysfunction. Optical coherence tomography showed selective thinning of the retinal nerve fiber layer. Histology of the rat retina in cross-section found reduced cell density in the ganglion cell layer, but no change in other cellular layers. Staining of flat-mounted mouse retinae with a ganglion cell specific marker (RBPMS) confirmed ganglion cell loss. The circumlimbal suture is a simple, minimally invasive and cost-effective way to induce ocular hypertension that leads to ganglion cell injury in both rats and mice.

Chronic ocular hypertension is induced by applying a circumlimbal suture in rats and mice, leading to functional and structural deterioration of the retinal ganglion cells consistent with glaucoma.

Ein Modell des Glaukoms induziert durch Circumlimbal Naht bei Ratten und Mäusen

The circumlimbal suture is a technique for inducing experimental glaucoma in rodents by chronically elevating intraocular pressure (IOP), a well-known ...

Isolation of Atrial Myocytes from Adult Mice

The electrophysiological properties of atrial myocytes importantly affect overall cardiac function. Alterations in the underlying ionic currents responsible for the action potential can cause pro-arrhythmic substrates that underlie arrhythmias, such as atrial fibrillation, which are highly prevalent in many conditions and disease states. Isolating adult mouse atrial cardiomyocytes for use in patch-clamp experiments has greatly advanced our knowledge and understanding of the cellular electrophysiology in the healthy atrial myocardium and in the setting of atrial pathophysiology. In addition, studies using genetic mouse models have elucidated the role of a vast array of proteins in regulating atrial electrophysiology. Here we provide a detailed protocol for the isolation of cardiomyocytes from the atrial appendages of adult mice using a combination of enzymatic digestion and mechanical dissociation of these tissues. This approach consistently and reliably yields isolated atrial cardiomyocytes that can then be used to characterize cellular electrophysiology by measuring action potentials and ionic currents in patch-clamp experiments under a number of experimental conditions.

This protocol is used to isolate single atrial cardiomyocytes from the adult mouse heart using a chunk digestion approach. This approach is used to isolate right or left atrial myocytes that can be used to characterize atrial myocyte electrophysiology in patch-clamp studies.

Isolierung von Atrial Myozyten von erwachsenen Mäusen

The electrophysiological properties of atrial myocytes importantly affect overall cardiac function. Alterations in the underlying ionic currents ...

Biventricular Assessment of Cardiac Function and Pressure-Volume Loops by Closed-Chest Catheterization in Mice

Assessment of cardiac function is essential to conduct cardiovascular and pulmonary-vascular preclinical research. Pressure-volume loops (PV loops) generated by recording both pressure and volume during cardiac catheterization are vital when assessing both systolic and diastolic cardiac function. Left and right heart function are closely related, reflected in ventricular interdependence. Thus, recording biventricular function in the same animal is important to get a complete assessment of cardiac function. In this protocol, a closed chest approach to cardiac catheterization consistent with the way catheterization is performed in patients is adopted in mice. While challenging, the closed chest strategy is a more physiological approach, because opening the chest results in major changes in preload and afterload that create artifacts, most notably a fall in systemic blood pressure. While high-resolution echocardiography is used to assess rodents, cardiac catheterization is invaluable, particularly when assessing diastolic pressures in both ventricles.
Described here is a procedure to perform invasive, closed chest, sequential left and right ventricular pressure-volume (PV) loops in the same animal. PV loops are acquired using&#160;admittance technology with a mouse pressure-volume catheter and pressure-volume system acquisition. The procedure is described, beginning with the neck dissection, which is required to access the right jugular vein and the right carotid artery, to the insertion and positioning of the catheter, and finally the data acquisition. Then, the criteria required to ensure the acquisition of high-quality PV loops are discussed. Finally, the analysis of the left and right ventricular PV loops and the different hemodynamic parameters available to quantify systolic and diastolic ventricular function are briefly described.

Presented here is a protocol to assess biventricular heart function in mice by generating pressure-volume (PV) loops from the right and left ventricle in the same animal using closed chest catheterization. The focus is on the technical aspect of surgery and data acquisition.

Biventrikuläre Beurteilung der Herzfunktion und der Druck-Volumen-Schleifen durch Closed-Thorax-Katheterisierung bei Mäusen

Assessment of cardiac function is essential to conduct cardiovascular and pulmonary-vascular preclinical research. Pressure-volume loops (PV loops) ...

Cardiac Response to &#946;-Adrenergic Stimulation Determined by Pressure-Volume Loop Analysis

Determination of the cardiac function is a robust endpoint analysis in animal models of cardiovascular diseases in order to characterize effects of specific treatments on the heart. Due to the feasibility of genetic manipulations the mouse has become the most common mammalian animal model to study cardiac function and to search for new potential therapeutic targets. Here we describe a protocol to determine cardiac function&#160;in vivo using pressure-volume loop measurements and analysis during basal conditions and under &#946;-adrenergic stimulation by intravenous infusion of increasing concentrations of isoproterenol. We provide a refined protocol including ventilation support taking into account the positive end-expiratory pressure to ameliorate negative effects during open-chest measurements, and potent analgesia (Buprenorphine) to avoid uncontrollable myocardial stress evoked by pain during the procedure. All together the detailed description of the procedure and discussion about possible pitfalls enables highly standardized and reproducible pressure-volume loop analysis, reducing the exclusion of animals from the experimental cohort by preventing possible methodological bias.

Cardiac Response to &amp;#946;-Adrenergic Stimulation Determined by Pressure-Volume Loop Analysis

Here we describe a cardiac pressure-volume loop analysis under increasing doses of intravenously infused isoproterenol to determine the intrinsic cardiac function and the &#946;-adrenergic reserve in mice. We use a modified open-chest approach for the pressure-volume loop measurements, in which we include ventilation with positive end-expiratory pressure.

Kardiale Reaktion auf β-adrenerge Stimulation durch Druck-Volumen-Schleifenanalyse bestimmt

Determination of the cardiac function is a robust endpoint analysis in animal models of cardiovascular diseases in order to characterize effects of ...

A Large Animal Model for Acute Kidney Injury by Temporary Bilateral Renal Artery Occlusion

Acute kidney injury (AKI) is associated with higher risk for morbidity and mortality post-operatively. Ischemia-reperfusion injury (IRI) is the most common cause of AKI. To mimic this clinical scenario, this study presents a highly reproducible large animal model of renal IRI in swine using temporary percutaneous bilateral balloon-catheter occlusion of the renal arteries. The renal arteries are occluded for 60 min by introducing the balloon-catheters through the femoral and carotid artery and advancing them into the proximal portion of the arteries. Iodinated contrast is injected in the aorta to assess any opacification of the kidney vessels and confirm the success of the artery occlusion. This is furtherly confirmed by the flattening of the pulse waveform at the tip of the balloon catheters. The balloons are deflated and removed after 60 min of bilateral renal artery occlusion, and the animals are allowed to recover for 24 h. At the end of the study, plasma creatinine and blood urea nitrogen significantly increase, while eGFR and urine output significantly decrease. The need for iodinated contrast is minimal and does not affect renal function. Bilateral renal artery occlusion better mimics the clinical scenario of perioperative renal hypoperfusion, and the percutaneous approach minimizes the impact of the inflammatory response and the risk of infection seen with an open approach, such as a laparotomy. The ability to create and reproduce this clinically relevant swine model eases the clinical translation to humans.

This study presents a highly reproducible large animal model of renal ischemia-reperfusion injury in swine using temporary percutaneous bilateral balloon-catheter occlusion of the renal arteries for 60 min and reperfusion for 24 h.

Ein Großtiermodell für akute Nierenverletzungen durch vorübergehenden bilateralen Nierenarterienverschluss

Acute kidney injury (AKI) is associated with higher risk for morbidity and mortality post-operatively. Ischemia-reperfusion injury (IRI) is the most ...