Ringraziamo il Dott. Helio Salgado, Renata Lataro e Mauro de Oliveira, Facoltà di Medicina di Ribeirão Preto, Università di San Paolo e il dottor Ben Janssen, Cardiovascular Research Institute di Maastricht, Università di Maastricht, per la generosa assistenza durante il processo di configurazione. Questo lavoro è stato sostenuto da FONDECYT concessione n ° 11090114 di DSC

Protocollo è stato approvato dal Comitato Etico di Medicina Clinica Alemana Facultad de-Universidad del Desarrollo. I. Preparazione infusione di dobutamina <ol><li> Sciogliere 10 mg di dobutamina in 20 ml di acqua distillata sterile, per ottenere una soluzione stock di 500 ug / ml dobutamina. Aliquotare e conservare a -20 ° C. Questa soluzione può essere utilizzata per almeno 3 mesi. </li><li> Scongelare una aliquota di soluzione di dobutamina a temperatura ambiente. </li><li> Diluire la soluzione stock in dobutamina sterile 0,9% NaCl, per ottenere dobutamina soluzione di lavoro, la cui concentrazione è calcolata utilizzando la formula: dobutamina (pg / ml) = peso corporeo x 0,2. </li><li> Riempire una siringa da 1 ml 29Gx1 / 2 &quot;con dobutamina soluzione di lavoro. </li><li> Inserire l&#39;ago della siringa in un cm 20 PE-10 tubo. </li><li> Regolare la siringa nella pompa per infusione seguendo le istruzioni del produttore. </li><li> Impostare la rampa di infusione in una fase-Al passaggio formato con un aumento di 10 microlitri / min per ogni passo, per 6 passaggi. </li></ol> II. Preparazione Sensore di pressione <ol><li> Per minimizzare la deriva del segnale, sensore di pressione immergere in acqua sterile a 37 ° C per almeno 15 min. Non bagnare il catetere più di 0,5 cm di profondità, per evitare che la pressione idrostatica colpisce il sensore di pressione. </li><li> Elettronicamente calibrare il sensore di pressione 25 e 100 mmHg. Elettrico di ingresso (Volt) viene convertito in segnale di pressione (mmHg). </li><li> Impostare la frequenza di campionamento di 2 k / s e usare il filtro passa basso con un cut-off a 100 Hz. Segnale di pressione impostato a zero mmHg. </li><li> Segna da 15 mm dalla punta del catetere. Distante per raggiungere il cuore dal punto di introduzione è stato stimato mediante ecocardiografia, rilevando la presenza del catetere nel ventricolo sinistro. </li></ol> III. Preparazione del mouse per il cateterismo <ol><li> Peso C57BL / 6 topi maschi, settimane di età 30-32. </li><li> Iniettare per via intraperitoneale 60 mg / kg di ketamina e 4 mg / kg xilazina 1. Nota: altri anestetici potrebbero essere utilizzati, ad esempio: 350-450 mg / kg avertin, 50 ug / kg pentobarbital o 1,5 - 2% 2-3 isoflurano. </li><li> Shave il collo con un rasoio elettrico. </li><li> Posizionare il mouse anestetizzato in posizione supina su una piastra riscaldata riscaldamento isotermico. Fissare le sue membra con nastro di carta. </li><li> Eseguire un pizzico punta per confermare sedazione completa. </li><li> Inserire delicatamente una sonda rettale per controllare la temperatura del corpo. Utilizzando la sonda vaselinized è raccomandato. </li><li> Se la temperatura del corpo diversa da 37 ° C ± 0,5 ° C, regolarlo mediante la piastra riscaldante. </li><li> Mettere muso mouse vicino l&#39;apporto di ossigeno. </li><li> Posizionare la regione del collo del mouse con lo stereomicroscopio. </li></ol> IV. Acquisizione dei dati <ol><li> Nel software 7 LabChartPro, selezionare un canale per la registrazione della pressione e un canale per la frequenza cardiaca (HR)registrazione. Per questi ultimi, selezionare l&#39;opzione &quot;Misurazioni cicliche&quot; e misura impostato come frequenza. </li><li> Per il canale di pressione impostano la scala: da 0 a 150 mmHg. </li><li> Per HR canale impostato Scala: da 200 a 600 bpm. </li><li> Premere il tasto di avvio per iniziare la registrazione. </li><li> Inserire i commenti che indicano le procedure eseguite, ad esempio: la somministrazione dell&#39;anestesia, l&#39;inizio della infusione di dobutamina, dobutamina concentrazione, la respirazione cambia. </li></ol> V. cateterismo cardiaco 4,5 <ol><li> Eseguire una piccola incisione sul lato destro vicino alla mascella. Con le forbici separare la pelle-muscolare del tessuto connettivo. </li><li> Eseguire una dissezione longitudinale (1,5 - 2 cm) sul lato destro della trachea. Separare il tessuto connettivo, grasso e muscolare con pinze curve, in modo da esporre la carotide destra vicino alla trachea. </li><li> Inserire un espansore nel lato destro animale per esporre la carotide. Pulsatilepressione generata dal cuore facilita l&#39;identificazione dell&#39;arteria. La vena giugulare, che è rosso scuro, è sulla destra. </li><li> Separare l&#39;arteria dai tessuti adiacenti con pinze curve. Il nervo vago, che assomiglia a un filo bianco, si trova lungo l&#39;arteria. </li><li> Tagliare un pezzo 20 cm 6/0 filo di seta e &quot;doppio&quot; di esso. </li><li> Passare il filo &quot;doppio&quot; sotto l&#39;arteria da sinistra a destra. Tagliare il filo, in modo da ottenere le estremità separate. </li><li> Passa un terzo filo (10 cm) sotto l&#39;arteria. </li><li> Un nodo stretto nel thread posizionato vicino alla testa, e uno sciolto nel filo più distale. </li><li> Fare un nodo sciolto nel thread mezzo, e fissare l&#39;estremità destra del filo centrale per il pad di riscaldamento con un nastro di carta. </li><li> Tenere carotide umido facendo cadere sterile 0,9% NaCl. Asciugare l&#39;eccesso di liquido con bastoncini di cotone. </li><li> Tendere il filo inferiore con una fascetta pinza emostatica. </li><li> Fissare la posizione della forbice hemostat da pinching la pelle dell&#39;addome, estende il filo superiore, per occludere il flusso sanguigno. Verificare che il tessuto connettivo intorno all&#39;arteria è stato rimosso. L&#39;arteria dovrebbe essere pieno di sangue e privo di pulsazioni. Impedire fili da produrre una forza di coppia sulla arteria. </li><li> Realizzare una sezione trasversale nick vicino al fondo della arteria con un micro-Vannas forbice. Gocce di sangue sarà versato. </li><li> Inserire il catetere nell&#39;arteria carotide. Assicurarsi di introdurre l&#39;intero sensore di pressione. Verificare che non vi è perdita di sangue. </li><li> Regolare delicatamente il nodo centrale filo, per mantenere il catetere in posizione. Non comprimere troppo, il sensore di pressione è molto fragile. </li><li> Rilasciare le forbici hemostat dall&#39;addome animale. </li><li> Tenere il catetere con la mano e spingere il filo centrale, al fine di evitare la perdita di sangue. Nota: arteria dovrebbe essere pieno di sangue. </li><li> Avviare segnali di pressione di registrazione. </li><li> Quando il catetere è all&#39;interno, lasegnale di pressione arteriosa oscilla 60-70 a 100-120 mmHg. La forma del segnale di pressione è mostrato nella Figura 1.A. Nota: se siete interessati a, a questo punto nel tempo è possibile registrare la pressione arteriosa se ​​il segnale è stabile per almeno 5 min. HR valori sono stati ottenuti dalle forme d&#39;onda di pressione considerando un intervallo di 30 secondi di segnale di registrazione. È possibile utilizzare anche un metodo ECG per la misura diretta di HR, in accordo con gli obiettivi di indagine. </li><li> Spingere delicatamente il catetere fino ad osservare una variazione della forma del segnale di pressione (Figura 1 B). Una volta che il catetere è all&#39;interno del ventricolo sinistro, il segnale di pressione oscilla da 0 a 100-120 mmHg. Se è difficile far scorrere il catetere, pizzicare petto animale con due dita. </li><li> Continuo controllo della frequenza respiratoria, temperatura corporea, livello di anestesia e segnale di pressione. Tutti loro dovrebbero rimanere stabili. </li></ol> VI. L&#39;infusione di Dobutamina <ol><li&gt; Per incannulazione della vena giugulare assicurarsi di staccare il tessuto adiposo intorno alla vena, per evitare la perforazione. La procedura chirurgica per occlusione venosa è simile al procedimento mostrato per carotide. </li><li> Introdurre un PE-10 tubo nella vena. Verificare che il flusso di sangue non è bloccato spostando all&#39;indietro lo stantuffo della siringa. </li><li> Infusione di dobutamina inizia con 10 microlitri / min e finire con 60 microlitri / min. In ogni passo, la velocità di infusione viene mantenuta per 2 min 6. </li><li> Dopo la dose di dobutamina ultimo, l&#39;eutanasia l&#39;animale con una overdose di anestesia. </li></ol> VII. Analisi dei dati <ol><li> Per l&#39;analisi dei dati, scegliere la sezione dei dati registrati di tuo interesse. Essere sicuri di prendere in considerazione un intervallo di tempo in cui il segnale di pressione è stabile. </li><li> Selezionare l&#39;icona di installazione nel modulo di pressione sanguigna. Indicare il tipo selezionato di segnale di pressione. </li><li> Automaticamente le 7 display software LabChartPro dire, massimavalori minimi e per HR, pressione sistolica (P max), la pressione diastolica (P min), pressione telediastolica (EDP), massimo sviluppo a pressione positiva (dP / dtmax) e la massima pressione negativa sviluppo (dP / dt min). Inoltre, i parametri cardiaci può essere raffigurato sulla traccia di pressione. </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>

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Stress test cardiaco indotto da dobutamina e monitorato da cateterismo cardiaco è laborioso. Tuttavia, secondo il protocollo qui descritto e con un breve tempo di formazione, è possibile valutare sei parametri emodinamici in un singolo esperimento che durano circa un&#39;ora. Le fasi critiche del protocollo qui presentati sono i cannulazioni di vasi sanguigni. Per quanto riguarda la cannulazione della carotide, l&#39;incisione effettuata deve essere sufficientemente profonda per rompere i ...

<table border="1"><tbody><tr><td></td><td></td><td></td><td> Reagenti </td></tr><tr><td> PE-50/10 </td><td> Warner Instruments </td><td> 64-0752 </td><td></td></tr><tr><td> Filo di seta 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> Ketamina 100 mg / ml </td><td> Trascina Pharma </td><td></td><td></td></tr><tr><td> Cloruro di sodio 0,9% </td><td> Lab Sanderson SA </td><td></td><td></td></tr><tr><td> Cloridrato Dobutamina </td><td> Sigma-Aldrich </td><td> D0676 </td><td></td></tr><tr><td> Siringa insulina U-100 29G x ½ &quot; </td><td> Terumo Medical Co. </td><td></td><td></td></tr><tr><td> Pinze per dissezione Micro Style 11,5 cm 7 </td><td> LawtonMedizintechnik </td><td> 09-0959 </td><td></td></tr><tr><td> Graefe Pinze Cvd 0,7 millimetri 7 centimetri </td><td> Lawton Medizintechnik </td><td> 62-0263 </td><td></td></tr><tr><td> Morsetti Dieffenbach bulldog Cl Str 38 millimetri </td><td> Lawton Medizintechnik </td><td> 60-010 </td><td></td></tr><tr><td> Forbici Vannas 8 centimetri Str Fh </td><td> Lawton Medizintechnik </td><td> 63-1400 </td><td></td></tr><tr><td></td><td></td><td></td><td> Attrezzatura </td></tr><tr><td> SPR-671 Mikro-Tip pressione catetere </td><td> Millar strumenti </td><td> 840-6719 </td><td></td></tr><tr><td> PCU-2000 Pressure Control Unit </td><td> Millar strumenti </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 pompa di infusione </td><td> KdScientific </td><td> KD-KDS210P </td><td></td></tr><tr><td> TCAT-2LV Regolatore di temperatura e la piastra di riscaldamento isotermico </td><td> PhysiTemp strumenti Inc. </td><td></td><td></td></tr><tr><td> Dotazioni di ossigeno medica </td><td> Indura </td><td></td><td></td></tr><tr><td> Sonda rettale </td><td> ADinstruments Pty Ltd. </td><td> MLT1404 </td><td></td></tr><tr><td> Microscopio trinoculare, illuminazione assiale </td><td> LW scientifico </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

Dobutamina è un β-adrenergico con un&#39;affinità maggiore per il recettore espresso nel cuore (β 1) rispetto ai recettori espressi nelle arterie (β 2). Quando somministrato per via sistemica, aumenta la domanda cardiaca. Così, dobutamina smaschera ritmo anormale o aree ischemiche potenzialmente a rischio di infarto. Monitoraggio della funzione cardiaca durante una prova da sforzo può essere eseguita sia ecocardiografia o cateterizzazione cardiaca. Quest&#39;ultima è una tecnica invasiva ma più accurate e informative che il primo. Test di stress cardiaco indotto da dobutamina e monitorato da cateterismo cardiaco realizzato come qui descritto consente, in un singolo esperimento, la misurazione dei seguenti parametri emodinamici: frequenza cardiaca (HR), la pressione sistolica, pressione diastolica, pressione telediastolica, massima pressione positiva sviluppo (dP / dtmax) e la massima stampa negativaure sviluppo (dP / dt min), in condizioni basali e in dosi crescenti di dobutamina. Come previsto, in topi normali, abbiamo osservato una dobutamina aumento dose-correlato in HR, dP / dt max e dP / dt min. Inoltre, alla massima dose testata (12 ng / g / min), lo scompenso cardiaco di alto contenuto di grassi topi obesi indotta dalla dieta è stato smascherato.

Il segnale di pressione arteriosa è definita da pressione sistolica e diastolica. Quando il sensore di pressione è all&#39;interno del ventricolo sinistro, la sua pressione (LVP) della forma d&#39;onda è caratterizzata da una caduta a zero della pressione diastolica e l&#39;aspetto della contrazione atriale sinistra prima contrazione ventricolare (Figura 1). In condizioni originarie, topi anestetizzati ketamina-xilazina normali aveva HR di 280 ± 24, P max di 107 ± 8, P min di ...

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

Descriviamo il protocollo di eseguire un test di stress cardiaco indotto da dobutamina e monitorato da cateterizzazione cardiaca in topi normali. Inoltre ci mostra la sua applicazione per smascherare subclinica cardiopatia in alto contenuto di grassi topi obesi indotta dalla dieta.

Stress Test cardiaca indotta da dobutamina e monitorato da cateterizzazione cardiaca nei topi

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 Views.  Clínica Alemana Universidad del Desarrollo. Descriviamo il protocollo di eseguire un test di stress cardiaco indotto da dobutamina e monitorato da cateterizzazione cardiaca in topi normali. Inoltre ci mostra la sua applicazione per smascherare subclinica cardiopatia in alto contenuto di grassi topi obesi indotta dalla dieta.

Video: 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.

Valutazione del ventricolo destro Struttura e funzione in modello murino di polmonare Arteria Costrizione da ecocardiografia transtoracica

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

Ricerca

Medicina

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.

La valutazione non invasiva della Cardiac Anomalie in Experimental Autoimmune miocardite da Risonanza Magnetica Imaging Microscopy nel 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 ...

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.

La diagnosi precoce del farmaco-indotta renale emodinamica erettile Utilizzando ecografica Technology di ratti

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.

Cateterizzazione cardiaca nei topi per misurare il rapporto pressione del volume: Indagare l&#39;effetto Bowditch

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.

Infarto del miocardio in topi neonati, un modello di rigenerazione cardiaca

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.

Un modello del Glaucoma indotto da sutura Circumlimbal nei ratti e nei topi

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.

Isolamento dei miociti atriale dai topi adulti

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.

Valutazione biventricolare della funzione cardiaca e delle anse pressione-volume mediante cateterismo a torace chiuso nei topi

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.

Risposta cardiaca alla stimolazione β-adrenergica determinata dall'analisi del ciclo pressione-volume

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.

Un modello animale di grandi dimensioni per lesioni renali acute da occlusione temporanea bilaterale dell'arteria renale

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