This work was supported in part by: Fundaci&#243;n S&#233;neca, Agencia de Ciencia y Tecnolog&#237;a, Regi&#243;n de Murcia, Spain (JT) (Grant number: 11935/PI/09) and the University of Reading, United Kingdom (AG, GB) (Central Funding). The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.

<ol>
	<li>Pogwizd, S. M., Bers, D. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rabbit+models+of+heart+disease.">Rabbit models of heart disease.</a> Drug Discovery Today Disease Models. 5, 185-193 (2008).</li><li>Gandolfi, F., 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=Large+animal+models+for+cardiac+stem+cell+therapies.">Large animal models for cardiac stem cell therapies.</a> Theriogenology. 75, 1416-1425 (2011).</li><li>Harding, J., Roberts, R. M., Mirochnitchenko, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Large+animal+models+for+stem+cell+therapy.">Large animal models for stem cell therapy.</a> Stem Cell Research &amp; Therapy. 4, 23 (2013).</li><li>Chong, J. J., Murry, C. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cardiac+regeneration+using+pluripotent+stem+cells--progression+to+large+animal+models.">Cardiac regeneration using pluripotent stem cells--progression to large animal models.</a> Stem Cell Research. 13, 654-665 (2014).</li><li>Del, M. F., Mynett, J. R., Sugden, P. H., Poole-Wilson, P. A., Harding, S. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Subcellular+mechanism+of+the+species+difference+in+the+contractile+response+of+ventricular+myocytes+to+endothelin-1.">Subcellular mechanism of the species difference in the contractile response of ventricular myocytes to endothelin-1.</a> Cardioscience. 4, 185-191 (1993).</li><li>Sahn, D. J., DeMaria, A., Kisslo, J., Weyman, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recommendations+regarding+quantitation+in+M-mode+echocardiography:+results+of+a+survey+of+echocardiographic+measurements.">Recommendations regarding quantitation in M-mode echocardiography: results of a survey of echocardiographic measurements.</a> Circulation. 58, 1072-1083 (1978).</li><li>Thomas, W. P., 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=Recommendations+for+standards+in+transthoracic+two-dimensional+echocardiography+in+the+dog+and+cat.+Echocardiography+Committee+of+the+Specialty+of+Cardiology,+American+College+of+Veterinary+Internal+Medicine.">Recommendations for standards in transthoracic two-dimensional echocardiography in the dog and cat. Echocardiography Committee of the Specialty of Cardiology, American College of Veterinary Internal Medicine.</a> Journal of Veterinary Internal Medicine. 7, 247-252 (1993).</li><li>Lang, R. 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=Recommendations+for+cardiac+chamber+quantification+by+echocardiography+in+adults:+an+update+from+the+American+Society+of+Echocardiography+and+the+European+Association+of+Cardiovascular+Imaging.">Recommendations for cardiac chamber quantification by echocardiography in adults: an update from the American Society of Echocardiography and the European Association of Cardiovascular Imaging.</a> European Heart Journal Cardiovascular Imaging. 16, 233-270 (2015).</li><li>Talavera, 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=An+Upgrade+on+the+Rabbit+Model+of+Anthracycline-Induced+Cardiomyopathy:+Shorter+Protocol,+Reduced+Mortality,+and+Higher+Incidence+of+Overt+Dilated+Cardiomyopathy.">An Upgrade on the Rabbit Model of Anthracycline-Induced Cardiomyopathy: Shorter Protocol, Reduced Mortality, and Higher Incidence of Overt Dilated Cardiomyopathy.</a> BioMed Research International. 2015, 465342 (2015).</li><li>Borkowski, R., Karas, A. Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sedation+and+anesthesia+of+pet+rabbits.">Sedation and anesthesia of pet rabbits.</a> Clinical Techniques in Small Animal Practice. 14, 44-49 (1999).</li><li>Cantwell, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ferret,+rabbit+and+rodent+anesthesia.">Ferret, rabbit and rodent anesthesia.</a> The Veterinary Clinics of North America. Exotic Animal Practice. 4, 169-191 (2001).</li><li>Giraldo, 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=Percutaneous+intramyocardial+injection+of+amniotic+membrane-derived+mesenchymal+stem+cells+improves+ventricular+function+and+survival+in+non-ischaemic+cardiomyopathy+in+rabbits.">Percutaneous intramyocardial injection of amniotic membrane-derived mesenchymal stem cells improves ventricular function and survival in non-ischaemic cardiomyopathy in rabbits.</a> European Heart Journal. 36, 149 (2015).</li><li>Giraldo, 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=Allogeneic+amniotic+membrane-derived+mesenchymal+stem+cell+therapy+is+cardioprotective,+restores+myocardial+function,+and+improves+survival+in+a+model+of+anthracycline-induced+cardiomyopathy.">Allogeneic amniotic membrane-derived mesenchymal stem cell therapy is cardioprotective, restores myocardial function, and improves survival in a model of anthracycline-induced cardiomyopathy.</a> European Journal of Heart Failure. 19, 594 (2017).</li><li>Bellenger, N. G., 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=Comparison+of+left+ventricular+ejection+fraction+and+volumes+in+heart+failure+by+echocardiography,+radionuclide+ventriculography+and+cardiovascular+magnetic+resonance;+are+they+interchangeable?.">Comparison of left ventricular ejection fraction and volumes in heart failure by echocardiography, radionuclide ventriculography and cardiovascular magnetic resonance; are they interchangeable?.</a> European Heart Journal. 21, 1387-1396 (2000).</li><li>Flachskampf, F. 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=Cardiac+Imaging+to+Evaluate+Left+Ventricular+Diastolic+Function.">Cardiac Imaging to Evaluate Left Ventricular Diastolic Function.</a> Journal of the American College of Cardiology Cardiovascular Imaging. 8, 1071-1093 (2015).</li></ol>

The authors have nothing to disclose.

We have described a protocol for the echocardiographic examination of cardiac function parameters in the rabbit, representing a large preclinical model1,2,3. The step by step methodology described herein should be considered guidance, which with a complementary study of the basic principles of echocardiography, and a basic knowledge of ultrasound imaging, will help the researcher to obtain, through practice and complementary and...

Longitudinal evaluation of cardiac function in large animal models is a robust research methodology commonly used for the assessment of the effects of novel therapies for treating ischemic and non-ischemic cardiomyopathy. Amongst the several cardiovascular imaging techniques available for preclinical research, echocardiography has been used extensively because of its non-invasive and portable characteristics. In experienced hands, echocardiography is also a very reproducible imaging technique to study cardiac anatomy as well as systolic and diastolic function of the heart.
Large preclinical animal models such as pigs, dogs and rabbits, are paramount for preclinical translational research1,2,3. Indeed, the potential benefit of novel therapies such as cardiac regenerative medicine in the setting of cardiomyopathy requires extensive hypothesis testing in large preclinical models before they can be considered for human use2,4. Compared to other large preclinical models, the rabbit model offers some advantages, including its low maintenance cost, which is comparable to that of mice and rats. However, in contrast to mice and rats, the Ca+2 transport system and cardiac electrophysiology are similar in rabbits as those of humans, and those of other large animal models such as dogs and pigs, thus increasing the translational potential of the rabbit model1,5. Therefore, the rabbit, as a large experimental preclinical model, has an exceptional balance of cost and reproducibility for preclinical translational research.
The rabbit has the additional benefit of its amenability for echocardiographic imaging using clinical ultrasound units routinely used in human and veterinary patients, thus taking advantage of the superiority of harmonic imaging and state-of-the-art technology. For this, sector transducers (also known as phase array) of relatively high frequency (up to 12 MHz), such as those used in neonatal/pediatric cardiology, are preferred. Echocardiographic examination in the rabbit preclinical model allows the complete evaluation of systolic and diastolic function using multiple views and different modes available in modern echocardiographic units (e.g. continuous wave Doppler (CWD), pulsed-wave Doppler (PWD), and Tissue Doppler imaging (TDI)).
Echocardiography is an operator-dependent technique and therefore requires extensive training and core knowledge of the technique in accord with international guidelines. Part of this training can be facilitated with the visualization of videos explaining in detail how different echocardiographic views can be obtained. The achievement of high competency in echocardiographic imaging, as well as development of a standardized protocol and correct technique, are essential to minimize the influence of the operator and to generate reliable quantitative data, as required in rigorous scientific research.
Some considerations are necessary regarding the system and laboratory setup used for echocardiography in rabbits and other large animal models. For a standard transthoracic echocardiographic evaluation of cardiac function, the ultrasound system must include the following modalities: bi-dimensional mode (B-mode or 2D), motion mode (M-mode), color Doppler, as well as CWD, PWD and TDI. Moreover, the machine should have full cardiac analysis and measurement software installed, as well as sufficient internal hard drive space to store enough high quality digital still images and video loops for offline analysis. Some systems use linear array transducers; however, for the best imaging of the heart, phased array sector transducers with a small scan head diameter are preferred, because these allow an easier passage of the ultrasound waves through the narrow intercostal spaces. For rabbits, we use relatively high frequency transducers (up to 12 MHz). The position of the animal for imaging is of utmost importance to acquire good quality images. Thus, both right and left lateral recumbent positions are recommended to obtain all standard imaging planes during an echocardiographic examination. For this, a table with a notch that coincides with the cardiac area of the chest is advisable (Figure 1A). This notched table facilitates the access with the transducer to the area of the chest that will be scanned, and therefore allows free mobility of the hand of the operator whist maintaining the best scanning position of the animal. Positioning the animal in a lateral recumbent position results in a fall of the heart towards the transducer and elevation of the lungs, as well as widening the access window of the ultrasound beam through the intercostal spaces, thus improving overall imaging quality (Figure 1A). The echocardiographic examination should be performed in a blinded fashion and following the guidelines of the Echocardiography Committee of the American College of Veterinary Internal Medicine and the American Society of Echocardiography/European Association for Cardiovascular Imaging6,7,8.
Part of our scientific team is associated with the Cardiology Service of a Veterinary Teaching Hospital that attends daily to veterinary patients (e.g. dogs and cats), for which it has the relevant training and accreditation in veterinary cardiology and echocardiography, and its different imaging modalities, as well as extensive experience in imaging different sizes of animal patients and thoracic conformations with this technique. In addition, we commonly use echocardiography for longitudinal evaluation of cardiac function in a rabbit model of cardiomyopathy induced by anthracyclines9. Here, we describe a step by step echocardiography protocol for evaluation of cardiac function using a clinical ultrasound unit in a large preclinical model such as the rabbit. This protocol is adapted for current international guidelines8, and includes practical recommendations based on our own experiences in clinical and experimental settings.

<table border="0" cellpadding="0" cellspacing="0" style="width:915px;" width="914">
	<tbody>
		<tr height="20">
			<td height="20" style="height:20px;width:327px;">Bluesensor</td>
			<td style="width:135px;">Medicotest</td>
			<td style="width:187px;">13BY1062</td>
			<td style="width:267px;">Disposable adhesive ECG lectrodes</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Domtor (Medetomidine)</td>
			<td style="width:135px;">Esteve</td>
			<td style="width:187px;">CN 570686.3</td>
			<td>Veterinary prescription is necessary</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:327px;">HD11 XE Ultrasound System</td>
			<td style="width:135px;">Philips</td>
			<td style="width:187px;">10670267</td>
			<td>Echocardiography system.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:327px;">Heating Pad</td>
			<td style="width:135px;">Solac</td>
			<td style="width:187px;">CT8632</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Imalgene (Ketamine)</td>
			<td style="width:135px;">Merial</td>
			<td style="width:187px;">RN 9767</td>
			<td>Veterinary prescription is necessary</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Omnifix-F 1 ml syringe</td>
			<td style="width:135px;">Braun</td>
			<td style="width:187px;">9161406V</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:327px;">S12-4</td>
			<td style="width:135px;">Philips</td>
			<td style="width:187px;">B01YgG</td>
			<td>4-12 MHz phase array transducer</td>
		</tr>
		<tr height="41">
			<td height="41" style="height:41px;">Ultrasound Transmision Gel (Aquasone)</td>
			<td style="width:135px;">Parker laboratories Inc.</td>
			<td style="width:187px;">N 01-08</td>
			<td></td>
		</tr>
	</tbody>
</table>

transthoracic echocardiographic examination in the rabbit model

Large animal models such as the rabbit are valuable for translational preclinical research. Rabbits have a similar cardiac electrophysiology compared to that of humans and that of other large animal models such as dogs and pigs. However, the rabbit model has the additional advantage of lower maintenance costs compared to other large animal models. The longitudinal evaluation of cardiac function using echocardiography, when appropriately implemented, is a useful methodology for preclinical assessment of novel therapies for heart failure with reduced ejection fraction (e.g. cardiac regeneration). The correct use of this non-invasive tool requires the implementation of a standardized examination protocol following international guidelines. Here we describe, step by step, a detailed protocol supervised by veterinary cardiologists for performing echocardiography in the rabbit model, and demonstrate how to correctly obtain the different echocardiographic views and imaging planes, as well as the different imaging modes available in a clinical echocardiography system routinely used in human and veterinary patients.

Parasternal long axis view of the heart
Figure 5A shows an imaging plane of the right PSLAX view where the 4 chambers of the heart are clearly distinguished. You can identify in this view the right ventricle (RV), tricuspid valve (TV), IVS, LV, FW, as well as the mitral valve (MV). When the apex is clearly visible on the left side of the image in this view and the LV is not foreshor...

Watch this Scientific Journal Video about Transthoracic Echocardiographic Examination in the Rabbit Model at JoVE.com

Transthoracic Echocardiographic Examination in the Rabbit Model

Here we describe, step by step, a detailed protocol for performing echocardiography in the rabbit model. We show how to correctly obtain the different echocardiographic views and imaging planes, as well as the different imaging modes available in a clinical echocardiography system routinely used in human and veterinary patients.

Large animal models such as the rabbit are valuable for translational preclinical research. Rabbits have a similar cardiac electrophysiology compared to ...

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12.3K Views.  University of Reading. Here we describe, step by step, a detailed protocol for performing echocardiography in the rabbit model. We show how to correctly obtain the different echocardiographic views and imaging planes, as well as the different imaging modes available in a clinical echocardiography system routinely used in human and veterinary patients.

Video: Transthoracic Echocardiographic Examination in the Rabbit Model

Transthoracic Echocardiography in Mice

In recent years, murine models have become the primary avenue for studying the molecular mechanisms of cardiac dysfunction resulting from changes in gene expression. Transgenic and gene targeting methods can be used to generate mice with altered cardiac size and function,1-3 and as a result, in vivo techniques are needed to evaluate their cardiac phenotype. Transthoracic echocardiography, pulse wave Doppler (PWD), and tissue Doppler imaging (TDI) can be used to provide dimensional measurements of the mouse heart and to quantify the degree of cardiac systolic and diastolic performance. Two-dimensional imaging is used to detect abnormal anatomy or movements of the left ventricle, whereas M-mode echo is used for quantification of cardiac dimensions and contractility.4,5 In addition, PWD is used to quantify localized velocity of turbulent flow,6 whereas TDI is used to measure the velocity of myocardial motion.7 Thus, transthoracic echocardiography offers a comprehensive method for the noninvasive evaluation of cardiac function in mice.

Transthoracic echocardiography offers a noninvasive method for the evaluation of cardiac function in mice. A combination of ultrasound and Doppler imaging modalities can be used to obtain dimensional measurements of the heart and intracardiac blood flow, which together provide an assessment of cardiac systolic and diastolic performance.

In recent years, murine models have become the primary avenue for studying the molecular mechanisms of cardiac dysfunction resulting from changes in gene ...

Microsurgical Venous Pouch Arterial-Bifurcation Aneurysms in the Rabbit Model: Technical Aspects

For ruptured human cerebral aneurysms endovascular embolization has become an equivalent alternative to aneurysm clipping.1 However, large clinical trials have shown disappointing long-term results with unacceptable high rates of aneurysm recanalization and delayed aneurysm rupture.2 To overcome these problems, animal experimental studies are crucial for the development of better endovascular devices.3-5
Several animal models in rats, rabbits, canines and swine are available.6-8 Comparisons of the different animal models showed the superiority of the rabbit model with regard to hemodynamics and comparability of the coagulation system and cost-effectiveness.9-11 
The venous pouch arterial bifurcation model in rabbits is formed by a venous pouch sutured into an artificially created true bifurcation of both common carotid arteries (CCA). The main advantage of this model are true bifurcational hemodynamics.12 The major drawbacks are the sofar high microsurgical technical demands and high morbidity and mortality rates of up to 50%.13 These limitations have resulted in less frequent use of this aneurysm model in the recent years. These shortcomings could be overcome with improved surgical procedures and modified peri- and postoperative analgetic management and anticoagulation.14-16 Our techniques reported in this paper demonstrate this optimized technique for microsurgical creation of arterial bifurcation aneurysms.

An optimized technique for the microsurgical creation of arterial bifurcation aneurysms mimicking bifurcation human cerebral aneurysms is described. A venous pouch is sutured into an artificially created true bifurcation of both common carotid arteries. Facilitated microsurgical techniques and aggressive postoperative anticoagulation and analgesia lead to minimized morbidity rates and high aneurysm patency rates.

For ruptured human cerebral aneurysms endovascular embolization has become an equivalent alternative to aneurysm clipping.1 However, large clinical trials ...

Fetal Echocardiography and Pulsed-wave Doppler Ultrasound in a Rabbit Model of Intrauterine Growth Restriction

Fetal intrauterine growth restriction (IUGR) results in abnormal cardiac function that is apparent antenatally due to advances in fetoplacental Doppler ultrasound and fetal echocardiography. Increasingly, these imaging modalities are being employed clinically to examine cardiac function and assess wellbeing in utero, thereby guiding timing of birth decisions. Here, we used a rabbit model of IUGR that allows analysis of cardiac function in a clinically relevant way. Using isoflurane induced anesthesia, IUGR is surgically created at gestational age day 25 by performing a laparotomy, exposing the bicornuate uterus and then ligating 40-50% of uteroplacental vessels supplying each gestational sac in a single uterine horn. The other horn in the rabbit bicornuate uterus serves as internal control fetuses. Then, after recovery at gestational age day 30 (full term), the same rabbit undergoes examination of fetal cardiac function. Anesthesia is induced with ketamine and xylazine intramuscularly, then maintained by a continuous intravenous infusion of ketamine and xylazine to minimize iatrogenic effects on fetal cardiac function. A repeat laparotomy is performed to expose each gestational sac and a microultrasound examination (VisualSonics VEVO 2100) of fetal cardiac function is performed. Placental insufficiency is evident by a raised pulsatility index or an absent or reversed end diastolic flow of the umbilical artery Doppler waveform. The ductus venosus and middle cerebral artery Doppler is then examined. Fetal echocardiography is performed by recording B mode, M mode and flow velocity waveforms in lateral and apical views. Offline calculations determine standard M-mode cardiac variables, tricuspid and mitral annular plane systolic excursion, speckle tracking and strain analysis, modified myocardial performance index and vascular flow velocity waveforms of interest. This small animal model of IUGR therefore affords examination of in utero cardiac function that is consistent with current clinical practice and is therefore useful in a translational research setting.

We describe examination of fetal cardiac function with contemporary functional fetal echocardiography and fetoplacental Doppler ultrasound using the VisualSonics VEVO 2100 microultrasound in a surgically induced model of intrauterine fetal growth restriction in a rabbit.

Fetal intrauterine growth restriction (IUGR) results in abnormal cardiac function that is apparent antenatally due to advances in fetoplacental Doppler ...

Echocardiographic Assessment of the Right Heart in Mice

Transgenic and toxic models of pulmonary arterial hypertension (PAH) are widely used to study the pathophysiology of PAH and to investigate potential therapies. Given the expense and time involved in creating animal models of disease, it is critical that researchers have tools to accurately assess phenotypic expression of disease. Right ventricular dysfunction is the major manifestation of pulmonary hypertension. Echocardiography is the mainstay of the noninvasive assessment of right ventricular function in rodent models and has the advantage of clear translation to humans in whom the same tool is used. Published echocardiography protocols in murine models of PAH are lacking.
In this article, we describe a protocol for assessing RV and pulmonary vascular function in a mouse model of PAH with a dominant negative BMPRII mutation; however, this protocol is applicable to any diseases affecting the pulmonary vasculature or right heart. We provide a detailed description of animal preparation, image acquisition and hemodynamic calculation of stroke volume, cardiac output and an estimate of pulmonary artery pressure.

This article provides a protocol for the echocardiographic assessment of right ventricular size and pulmonary hypertension in mice. Applications include phenotype determination and serial assessment in transgenic and toxin-induced mouse models of cardiomyopathy and pulmonary vascular disease.

Transgenic and toxic models of pulmonary arterial hypertension (PAH) are widely used to study the pathophysiology of PAH and to investigate potential ...

Ultrasound-guided Transthoracic Intramyocardial Injection in Mice

Murine models of cardiovascular disease are important for investigating pathophysiological mechanisms and exploring potential regenerative therapies. Experiments involving myocardial injection are currently performed by direct surgical access through a thoracotomy. While convenient when performed at the time of another experimental manipulation such as coronary artery ligation, the need for an invasive procedure for intramyocardial delivery limits potential experimental designs. With ever improving ultrasound resolution and advanced noninvasive imaging modalities, it is now feasible to routinely perform ultrasound-guided, percutaneous intramyocardial injection. This modality efficiently and reliably delivers agents to a targeted region of myocardium. Advantages of this technique include the avoidance of surgical morbidity, the facility to target regions of myocardium selectively under ultrasound guidance, and the opportunity to deliver injectate to the myocardium at multiple, predetermined time intervals. With practiced technique, complications from intramyocardial injection are rare, and mice quickly return to normal activity on recovery from anesthetic. Following the steps outlined in this protocol, the operator with basic echocardiography experience can quickly become competent in this versatile, minimally invasive technique.

Echocardiography-guided percutaneous intramyocardial injection represents an efficient, reliable, and targetable modality for the delivery of gene transfer agents or cells into the murine heart. Following the steps outlined in this protocol, the operator can quickly become competent in this versatile, minimally invasive technique.

Murine models of cardiovascular disease are important for investigating pathophysiological mechanisms and exploring potential regenerative therapies. ...

The Rabbit Blood-shunt Model for the Study of Acute and Late Sequelae of Subarachnoid Hemorrhage: Technical Aspects

Early brain injury and delayed cerebral vasospasm both contribute to unfavorable outcomes after subarachnoid hemorrhage (SAH). Reproducible and controllable animal models that simulate both conditions are presently uncommon. Therefore, new models are needed in order to mimic human pathophysiological conditions resulting from SAH.
This report describes the technical nuances of a rabbit blood-shunt SAH model that enables control of intracerebral pressure (ICP). An extracorporeal shunt is placed between the arterial system and the subarachnoid space, which enables examiner-independent SAH in a closed cranium. Step-by-step procedural instructions and necessary equipment are described, as well as technical considerations to produce the model with minimal mortality and morbidity. Important details required for successful surgical creation of this robust, simple and consistent ICP-controlled SAH rabbit model are described.

The experimental intracranial pressure-controlled blood shunt subarachnoid hemorrhage (SAH) model in the rabbit combines the standard procedures &#8212; subclavian artery cannulation and transcutaneous cisterna magna puncture, which enables close mimicking of human pathophysiological conditions after SAH. We present step-by-step instructions and discuss key surgical points for successful experimental SAH creation.

Early brain injury and delayed cerebral vasospasm both contribute to unfavorable outcomes after subarachnoid hemorrhage (SAH). Reproducible and ...

Echocardiographic and Histological Examination of Cardiac Morphology in the Mouse

An increasing number of genetically modified mouse models has become available in recent years. Moreover, the number of pharmacological studies performed in mice is high. Phenotypic characterization of these mouse models also requires the examination of cardiac function and morphology. Echocardiography and magnetic resonance imaging (MRI) are commonly used approaches to characterize cardiac function and morphology in mice. Echocardiographic and MRI equipment specialized for use in small rodents is&#160;expensive and requires a dedicated space. This protocol describes cardiac measurements in mice using a clinical echocardiographic system with a 15 MHz human vascular probe. Measurements are performed on anesthetized adult mice. At least three image sequences are recorded and analyzed for each animal in M-mode in the parasternal short-axis view. Afterwards, cardiac histological examination is performed, and cardiomyocyte diameters are determined on hematoxylin-eosin- or wheat germ agglutinin (WGA)-stained paraffin sections. Vessel density is determined morphometrically after Pecam-1 immunostaining. The protocol has been applied successfully to pharmacological studies and different genetic animal models under baseline conditions, as well as after experimental myocardial infarction by the permanent ligation of the left anterior descending coronary artery (LAD). In our experience, echocardiographic investigation is limited to anesthetized animals and is feasible in adult mice weighing at least 25 g.

Echocardiographic examination is frequently used in mice. Expensive high-resolution ultrasound devices have been developed for this purpose. This protocol describes an affordable echocardiographic procedure combined with histological morphometric analyses to determine cardiac morphology.

An increasing number of genetically modified mouse models has become available in recent years. Moreover, the number of pharmacological studies performed ...

Echocardiographic Measurement of Right Ventricular Diastolic Parameters in Mouse

Diastolic dysfunction is a prominent feature of right ventricular (RV) remodeling associated with conditions of pressure overload. However, the RV diastolic function is rarely quantified in experimental studies. This might be due to technical difficulties in the visualization of the RV in the apical four-chamber view in rodents. Here we describe two positions facilitating the visualization of the apical four-chamber view in mice to assess the RV diastolic function.
The apical four-chamber view is enabled by tilting the mouse fixation platform to the left and caudally (LeCa) or to the right and cranially (RiCr). Both positions provide images of comparable quality. The results of the RV diastolic function obtained from two positions are not significantly different. Both positions are comparably easy to perform. This protocol can be incorporated into published protocols and enables detailed investigations of the RV function.

Here we describe and compare two positions for obtaining the apical four-chamber view in mice. These positions enable the quantification of the right ventricular function, provide comparable results, and can be used interchangeably.

Diastolic dysfunction is a prominent feature of right ventricular (RV) remodeling associated with conditions of pressure overload. However, the RV ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

Isolated Lung Perfusion System in the Rabbit Model

The isolated lung perfusion system has been widely used in pulmonary research, contributing to elucidate the lungs&#39; inner workings, both micro and macroscopically. This technique is useful in the characterization of pulmonary physiology and pathology by measuring metabolic activities and respiratory functions, including interactions between circulatory substances and the effects of inhaled or perfused substances, as in drug testing. While in vitro methods involve the slicing and culturing of tissues, the isolated ex vivo lung perfusion system allows to work with a complete functional organ making possible the study of a continuous physiological function while recreating ventilation and perfusion. However, it should be noted that the effects of the absence of central innervation and lymphatic drainage still have to be fully assessed. This protocol aims to describe the assembly of the isolated lung apparatus, followed by the surgical extraction and cannulation of lungs and heart from experimental lab animals, as well as to display the perfusion technique and signal processing of data. The average viability of the isolated lung ranges between 5-8 h; during this period, the pulmonary capillary permeability increases, causing edema and lung injury. The functionality of preserved pulmonary tissue is measured by the capillary filtration coefficient (Kfc), used to determine the extent of pulmonary edema through time.

The isolated rabbit lung preparation is a gold standard tool in pulmonary research. This publication aims to describe the technique as developed for the study of physiological and pathological mechanisms involved in airway reactivity, lung preservation, and preclinical research in lung transplantation and pulmonary edema.

The isolated lung perfusion system has been widely used in pulmonary research, contributing to elucidate the lungs&#39; inner workings, both micro and ...