Name: Contractility Measurements on Isolated Papillary Muscles for the Investigation of Cardiac Inotropy in Mice (Video) | JoVE Journal
Uploaded: 2015-09-17T15:00:00.000+00:00
Duration: 6 min 22 s
Description: Learn to isolate murine left ventricular papillary muscles for in vitro cardiac contractility studies using field electric pulse stimulation. This method enables precise evaluation of cardiac inotropy under controlled physiological conditions.

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This work was supported by the Deutsche Forschungsgemeinschaft (KFO 196 &#8220;Signaltransduktion bei adaptativen und maladaptiven kardialen Remodelling-Prozessen&#8221;, FR 1638/1-2) and by the DZHK (German Centre for Cardiovascular Research, a part of the German Centres of Health Research, which is a BMBF (German Ministry of Education and Research) initiative).

NOTE: The basic steps of the isolation procedure are shown in Figure&#160;1. All steps are described in detail in the following protocol. Papillary muscle isolation, mounting in organ bath chamber, acquisition and analysis is performed in a consecutive and compulsory timescale.
All animal experiments were performed in accordance with German legislation on protection of animals and were approved by the Ethics Review Board of University of Heidelberg.
1. Preparation of Instrumentation
<ol>
	<li>For contractility measurements use a multichambered organ bath setup. Components of an ex vivo contractility organ bath setup are shown in Figure 2. 
	NOTE: To minimize the amount of time between the preparation of the papillary muscle and the beginning of the experimental protocol, set up equipment and prepare buffers prior to collecting experimental tissue.This provides the greatest amount of time that the tissue remains viable to conduct experiments.</li>
	<li>Start up computer and data acquisition equipment. Turn on organ bath heat (preset to 32 &#176;C) and allow units to preset temperature.</li>
	<li>Prepare the acquisition equipment. Start recording data as mentioned in the software manual.</li>
	<li>Check and conduct calibration (if necessary) on each channel and zero all channels to standardize the electric signal supplied from the respective force transducer (associated with that channel) to a 2-g force supplied by a certified weight.</li>
	<li>Turn on gas supply (95% O2/5% CO2) to organ bath chambers. Continuously gas all chambers during the entire protocol measurements.</li>
</ol>
2. Preparation of Buffers and Physiological Solutions
<ol>
	<li>Prepare a Krebs-Henseleit buffer solution to achieve final concentrations of 119 mM NaCl; 25 mM NaHCO3; 4.6 mM KCl; 11mM glucose; 1 mM Na-pyruvate; 1.5 mM CaCl2; 1.64 mM MgSO4; 1.18 mM KH2PO4 and adjust pH to 7.4 (see Table 1 Physiological solution).</li>
	<li>Prepare separate Krebs-Henseleit buffer solution to use during isolation as described in step 2.1 with additional 30mM 2,3-Butanedione monoxime (BDM) and 50 IE Heparin/ml. Cool solution to 4&#176;C (see Table 1 Preparation solution). For ischemia simulation, prepare ischemia-solution as described in Table 1.</li>
	<li>For the protocol of the Ca2+-dependent contractility, add Ca2+ to the Krebs-Henseleit buffer solution to requested final concentration. 
	NOTE: prepare these solutions directly before its usage and gass with carbogen to avoid precipitation of Calciumcarbonate. Prewarm solution to 32&#176;C before using.</li>
</ol>
3. Preparation of the Gassing Tube and Dissection Dish used during the Papillary Excision Procedure
<ol>
	<li>Connect a soft gassing tube (external diameter 2-4 mm) to the gas connection (100% oxygen, carbogen alternatively) with expert working.</li>
	<li>Create a closed ring of the gassing tube fitting inside the dissection dish. Puncture the gassing tube several times that a continuous bubbling is assured. This gassing tube can be used several times.</li>
	<li>Prepare dissection dish with a silicon elastomer at the bottom (0.5 cm thick), so that pins can be stuck into it. This dish can be used several times.</li>
</ol>
4. Isolation of Left Anterior Papillary Muscle from Mice
NOTE: Before starting the isolation of the papillary muscle, check that the gas lines are clear of blockages.
<ol>
	<li>Prepare organ bath setup as mentioned in the protocol section 1-3 before starting the preparation of the papillary muscles. Prepare all solutions at the day of the experiment.</li>
	<li>Sacrifice the mouse (approximately 8-12 weeks old) by cervical dislocation according to expert working and institutional guidelines.</li>
	<li>Fix the animal in dorsal position by fixing forepaws and hind paws on a dissection board, open the thorax lateral on both sides by cutting through the ribs with sharp bone scissors, then cut the diaphragm with a transversal cut. Remove the pericardium. Now the heart and the aorta are accessible.</li>
	<li>Fix the heart with blunt forceps on the vascular truncus close to the heart and separate the heart quickly with scissors from the lungs and the surrounding tissue.</li>
	<li>Transfer the beating heart to a Petri dish filled with cooled preparation solution gassed with oxygen (step 2.2). Allow the heart to beat and stimulate heart contractions softly via touching the cardiac apex with forceps.</li>
	<li>As soon the heart is completely exsanguinous, transfer it to the dissection dish filled with cooled preparation solution (step 2.2) and gassed with O2. From this step on use a stereomicroscope.</li>
	<li>Fix the heart with a small pin through the right ventricle so that the heart is seen from dorsal (right ventricle is situated on the right side from the operator&#8217;s view).</li>
	<li>Using microsurgical scissors separate both atria on the atrioventricular level, cut connecting tissue from the ventricles and discard. Perform a cut through the ventricular wall from the AV-valve-level to the apex of the heart avoiding any mechanical pressure.</li>
	<li>Open the ventricle and fix the left-sided free wall with forceps, thus having a look at both left ventricular papillary muscles.</li>
	<li>Slightly cut away the ventricular tissue lateral on both sides of the left anterior papillary muscle preserving a part of the valvular sail on the papillary muscle preparation and avoid touching or stretching the papillary muscle as much as possible. Dissect the remaining ventricular wall tissue from the papillary muscle.</li>
	<li>Attach silk threads (7/0, metric 0.5) on both sides of the papillary muscle preparation, one on the valvular sail and one on the muscular part. Fix the preparation in the organ bath chamber filled with organ bath solution and gassed with carbogen, as temperature 32 &#176;C is suggested.</li>
</ol>
5. Equilibration and Stimulation of the Papillary Muscle
<ol>
	<li>Stimulate the papillary muscle preparation with rectangular pulses (duration of 2 ms and current of 100 mA) at stimulation frequency of 1 Hz immediately after the fixation in the organ bath.</li>
	<li>Ensure a frequent change of the organ bath solution, either by continuous change or by frequent manual change (every 5 min) according to the setup of the organ bath type used.</li>
	<li>Gradually increase the pretension until maximal twitch force in reached. Maximal twitch force is reached when the increase of the pretension is not followed by a further augmentation in the twitch force.</li>
	<li>After 45-60 min of equilibration, start the experimental protocol.</li>
</ol>
6. Suggested Experimental Protocols for Contractility Measurements
NOTE: The experimental protocol outlined below comprises standard maneuvers to characterize cardiac contractility under physiological and pathophysiological conditions. In the representative section we describe these protocols in detail also showing representative results (see also Table 2).
<ol>
	<li>Recording and analysis of the experimental protocols
	<ol>
		<li>For recording, use a suitable software with adequate temporal resolution (acquisition rate &#8805; 1kHz).</li>
		<li>Analyze parameters including twitch force amplitude (height), time to peak tension (TTP) and half maximal relaxation time (R50/TFall, respectively) as mentioned in the software program (as example Chart 5.5 of ADInstruments, see Figure 4).</li>
	</ol>
	</li>
</ol>

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<li>Bluhm, W. F., Kranias, E. G., Dillmann, W. H., Meyer, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Phospholamban%3A+a+major+determinant+of+the+cardiac+force-frequency+relationship%5BTitle%5D)+AND+%22Am+J+Physiol+Heart+Circ+Physiol%22%5BJournal%5D)">Phospholamban: a major determinant of the cardiac force-frequency relationship.</a> Am J Physiol Heart Circ Physiol. 278 (1), H249-H255 (2000).</li>
<li>Redel, A., Baumgartner, W., Golenhofen, K., Drenckhahn, D., Golenhofen, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Mechanical+activity+and+force-frequency+relationship+of+isolated+mouse+papillary+muscle%3A+effects+of+extracellular+calcium+concentration%2C+temperature+and+contraction+type%5BTitle%5D)+AND+%22Pflugers+Arch%22%5BJournal%5D)">Mechanical activity and force-frequency relationship of isolated mouse papillary muscle: effects of extracellular calcium concentration, temperature and contraction type.</a> Pflugers Arch. 445 (2), 297-304 (2002).</li>
<li>Uhl, S., Mathar, I., Vennekens, R., Freichel, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Adenylyl+cyclase-mediated+effects+contribute+to+increased+Isoprenaline-induced+cardiac+contractility+in+TRPM4+deficient+mice%5BTitle%5D)+AND+%22JMCC%22%5BJournal%5D)">Adenylyl cyclase-mediated effects contribute to increased Isoprenaline-induced cardiac contractility in TRPM4 deficient mice.</a> JMCC. 74, 307-317 (2014).</li>
<li>Allen, D. G., Jewell, B. R., Wood, E. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Studies+of+the+contractility+of+mammalian+myocardium+at+low+rates+of+stimulation%5BTitle%5D)+AND+%22J+Physiol%22%5BJournal%5D)">Studies of the contractility of mammalian myocardium at low rates of stimulation.</a> J Physiol. 254 (1), 1-17 (1976).</li>
<li>Pieske, B., Maier, L. S., Schmidt-Schweda, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Sarcoplasmic+reticulum+Ca2%2B+load+in+human+heart+failure%5BTitle%5D)+AND+%22Basic+Res+Cardiol%22%5BJournal%5D)">Sarcoplasmic reticulum Ca2+ load in human heart failure.</a> Basic Res Cardiol. 97, Suppl 1. 163-171 (2002).</li>
<li>Koch-Weser, J., Blinks, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+Influence+of+the+Interval+between+Beats+on+Myocardial+Contractility%5BTitle%5D)+AND+%22Pharmacol+Rev%22%5BJournal%5D)">The Influence of the Interval between Beats on Myocardial Contractility.</a> Pharmacol Rev. 15, 601-652 (1963).</li>
<li>Bocalini, D. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Myocardial+remodeling+after+large+infarcts+in+rat+converts+post+rest-potentiation+in+force+decay%5BTitle%5D)+AND+%22Arq+Bras+Cardiol%22%5BJournal%5D)">Myocardial remodeling after large infarcts in rat converts post rest-potentiation in force decay.</a> Arq Bras Cardiol. 98 (3), 243-251 (2012).</li>
<li>Juggi, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Effect+of+ischemia-reperfusion+on+the+post-rest+inotropy+of+isolated+perfused+rat+heart%5BTitle%5D)+AND+%22J+Cell+Mol+Med%22%5BJournal%5D)">Effect of ischemia-reperfusion on the post-rest inotropy of isolated perfused rat heart.</a> J Cell Mol Med. 6 (4), 621-630 (2002).</li>
<li>Lakatta, E. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Beyond+Bowditch%3A+the+convergence+of+cardiac+chronotropy+and+inotropy%5BTitle%5D)+AND+%22Cell+Calcium%22%5BJournal%5D)">Beyond Bowditch: the convergence of cardiac chronotropy and inotropy.</a> Cell Calcium. 35 (6), 629-624 (2004).</li>
<li>Taylor, D. G., Parilak, L. D., LeWinter, M. M., Knot, H. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Quantification+of+the+rat+left+ventricle+force+and+Ca2%2B+-frequency+relationships%3A+similarities+to+dog+and+human%5BTitle%5D)+AND+%22Cardiovasc+Res%22%5BJournal%5D)">Quantification of the rat left ventricle force and Ca2+ -frequency relationships: similarities to dog and human.</a> Cardiovasc Res. 61 (1), 77-86 (2004).</li>
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<li>Rossman, E. I., Petre, R. E., Chaudhary, K. W., Piacentino, V. 3rd, Janssen, P. M., Gaughan, J. P., Houser, S. R., Margulies, K. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Abnormal+frequency-dependentresponses+represent+the+pathophysiologic+signature+of+contractile+failure+inhuman+myocardium%5BTitle%5D)+AND+%22JMCC%22%5BJournal%5D)">Abnormal frequency-dependentresponses represent the pathophysiologic signature of contractile failure inhuman myocardium.</a> JMCC. 36 (1), 33-42 (2004).</li>
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The authors declare that they have no competing financial interests.

In this manuscript we describe a method to investigate contractility of murine papillary muscle in vitro which can be used to answer several scientific questions related to heart physiology and pathology in mice as well as to support the analysis of transgenic lines and the discovery of new pharmaceutical approaches to treat heart dysfunctions. We illustrate the use of this method to assess physiological, pathological and pharmacological properties of cardiac muscle contractility (see Fig 3). Ad...

The investigation of proteins like ion channels referring their role for cardiac contractility is essential to discover different pathomechanisms and to establish new therapeutic strategies for cardiac diseases such as ischemia and heart failure.
Contractile function of mammalian cardiomyocytes is known to be modulated by various ion channels, transporters and other proteins. Action potential evoked activation of voltage dependent sarcolemmal L-type Ca2+ channels leads to Ca2+ influx from extracellular space and subsequently to Ca2+-induced Ca2+ release (CICR)1, which triggers cellular contraction2. Ca2+-signaling plays a central role in cardiac contractility and adaptation to physiological or pathological stress. Catecholamines activate cardiac &#946;-adrenergic receptors, thus stimulating adenylyl cyclase (AC) which synthesizes cAMP. Being activated, protein kinase A (PKA) phosphorylates different intracellular and membrane associated proteins like L-type Ca2+ channels, phospholamban and ryanodine receptors resulting in modification of Ca2+ transients and cardiac contractility1,3,4. cAMP is degraded by phosphodiesterase (PDE). Activation of Gs-coupled receptors other than &#946;-adrenoceptors also leads to accumulation of cAMP.
The technique of contractility measurements in isolated ventricular muscle strips is well established for larger mammalian species5-8. Based on the possibility of gene targeting in mice it is important to establish methods to analyze murine cardiac physiology. However, existing data about the physiological properties of isolated muscle preparations in mice differ depending on experimental conditions9-12.
The described method is used to analyze cardiac contractility of left ventricular papillary muscle preparations in vitro. Investigation of cardiac contractility is performed in the absence of influences modifying cardiac contractility in vivo, like blood pressure, neurohumoral stimulation and physical or metabolic stress. The beating rate of the contracting muscle preparation can be rigorously defined and changed arbitrarily. Twitch force can be analyzed in the context of specific stimuli such as calcium concentration, beating frequency or temperature. In addition, this method can be used to investigate different signaling pathway components and to compare cardiac performance of genetically modified mouse models by controlling experimental conditions mentioned above.

<table><tbody><tr><td>Sodium chloride</td><td>Sigma-Aldrich</td><td>S7653</td><td></td></tr><tr><td>Sodium bicarbonate</td><td>Sigma-Aldrich</td><td>S5761&amp;nbsp;</td><td></td></tr><tr><td>Potassium chloride</td><td>Sigma-Aldrich</td><td>P9333</td><td></td></tr><tr><td>Glucose</td><td>Sigma-Aldrich</td><td>D9434&amp;nbsp;</td><td></td></tr><tr><td>Sodium pyruvate</td><td>Sigma-Aldrich</td><td>P5280&amp;nbsp;</td><td></td></tr><tr><td>Calcium chloride dihydrate</td><td>Sigma-Aldrich</td><td>223506</td><td></td></tr><tr><td>Magnesium sulfate heptahydrate</td><td>Sigma-Aldrich</td><td>230391</td><td></td></tr><tr><td>Potassium phosphate monobasic</td><td>Sigma-Aldrich</td><td>P 5655</td><td></td></tr><tr><td>2,3-Butanedione monoxime</td><td>Sigma-Aldrich</td><td>B0753</td><td></td></tr><tr><td>3-Isobutyl-1-methylxanthine</td><td>Sigma-Aldrich</td><td>I5879</td><td>Hazard statement H 302, solve in DMSO</td></tr><tr><td>Dimethyl sulfoxide (DMSO)</td><td>Sigma-Aldrich</td><td>D2650</td><td></td></tr><tr><td>Isoprenaline hydrochloride</td><td>Sigma-Aldrich</td><td>I5627</td><td>Hazard statement H 315-H319-H335</td></tr><tr><td>Sodium Heparine 250.000 IE/10 ml</td><td>ratiopharm</td><td>PZN 3874685</td><td></td></tr><tr><td>Histamine dihydrochloride</td><td>Sigma-Aldrich</td><td>H7250</td><td>Hazard statement H 315-H 317-H319- H334-H335</td></tr></tbody></table>

contractility measurements on isolated papillary muscles for the investigation of cardiac inotropy in mice

Contractility Measurements on Isolated Papillary Muscles for the Investigation of Cardiac Inotropy in Mice

Papillary muscle isolated from adult mouse hearts can be used to study cardiac contractility during different physiological/pathological conditions. The contractile characteristics can be evaluated independently of external influences such as vascular tonus or neurohumoral status. It depicts a scientific approach between single cell measurements with isolated cardiac myocytes and in vivo studies like echocardiography. Thus, papillary muscle preparations serve as an excellent model to study cardiac physiology/pathophysiology and can be used for investigations like the modulation by pharmacological agents or the exploration of transgenic animal models. Here, we describe a method of isolating the murine left anterior papillary muscle to investigate cardiac contractility in an organ bath setup. In contrast to a muscle strip preparation isolated from the ventricular wall, the papillary muscle can be prepared in toto without damaging the muscle tissue severely. The organ bath setup consists of several temperature-controlled, gassed and electrode-equipped organ bath chambers. The isolated papillary muscle is fixed in the organ bath chamber and electrically stimulated. The evoked twitch force is recorded using a pressure transducer and parameters such as twitch force amplitude and twitch kinetics are analyzed. Different experimental protocols can be performed to investigate the calcium- and frequency-dependent contractility as well as dose-response curves of contractile agents such as catecholamines or other pharmaceuticals. Additionally, pathologic conditions like acute ischemia can be simulated.

The protocol of this manuscript for contractility measurements of isolated murine papillary muscle preparations is tuned to optimal conditions to achieve reproducible experimental results under physiological conditions. To define optimal experimental conditions we performed pilot experiments varying organ bath temperature and extracellular calcium concentration (see also12). The protocol described here was performed with an extracellular calcium concentration of 1.5 mM and a temperature of 32 °C.
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Murine left ventricular papillary muscle can be used to investigate cardiac contractility in vitro. This article describes in detail the isolation and experimental protocols to study cardiac contractile characteristics.

Papillary muscle isolated from adult mouse hearts can be used to study cardiac contractility during different physiological/pathological conditions. The ...

The overall goal of this procedure is to isolate healthy, intact papillary muscles for the study of in vitro cardiac contractile function in mice. This is accomplished by first isolating the heart, followed by interruption of the contractions, by immersion in a cardioplegic solution to avoid ischemic damage. In the second step, the intact left ventricular anterior papillary muscle is isolated and equipped with fixation sutures.The muscle is then mounted onto the forced transducer and transferred into the organ bath chamber. In the final step, the papillary muscle is stimulated with field electric pulses to induce papillary muscle contractions in vitro. Ultimately, the effects of various physiological and pathological conditions on the contractility of the cardiac muscle can be transmitted to the forced transducer and recorded.This method can be used to analyze the cardiac contractility of left ventricular popularity muscles in vitro in the absence of neuro humeral and vascular influences to allow the direct evaluation of the cardiac contractile. Properties of interest. Papillary muscle preparation serve as excellent models for studying cardiac physiology and pathophysiology, allowing the investigation of the effects of pharmacological agents on cardiac contractility.We first had the idea for this method when we found a cardiovascular phenotype in mice deficient for a specific calcium activated, non-selective ion channel leading to the discovery of specific inotropic alterations in the beta AIC signaling pathway. Begin by placing an eight to 12 week old mouse in the dorsal position on a dissection board. Next, using sharp bone scissors, open the thorax laterally on both sides of the chest and cut through the diaphragm.Then using blunt forceps, grab the heart on the vascular chunks and use scissors to separate the heart quickly from the lungs and the surrounding tissue. Transfer the beating heart to a Petri dish filled with cooled preparation solution, gassed with oxygen and gently touch the cardiac apex with forceps to stimulate heart contractions. As soon as it is completely ex sanguineous, transfer the heart to the dissection dish and place a small pin through the right ventricle to fix the heart in the dorsal position.Now, transfer the dish under a stereo microscope and use microsurgical scissors to separate both of the atria on the atrial ventricular level. Turn the heart about 180 degrees and make a cut through the left ventricular wall from the atrial ventricular valve level to the apex of the heart. Taking care to avoid any mechanical pressure on the heart.Then rotate the heart back. Then open the left ventricle and fix the left-sided free wall with forceps exposing both of the left ventricular papillary muscles. Carefully cut the ventricular tissue laterally on both sides of the left anterior papillary muscle.Taking care to preserve a part of the valvular SA on the papillary muscle preparation. Then dissect the remaining ventricular wall tissue from the papillary muscle and attach one silk thread at the muscular part with two knots and one at the valvular sail of the papillary muscle. Preparation with one knot.Fix the preparation in a previously prepared organ bath chamber. Then immediately after the fixation, stimulate the tissue with two millisecond rectangular pulses at a current of 100 milliamps and a stimulation frequency of one hertz. And start the recording.Changing the solution as appropriate for the type of organ bath used. Gradually increase the mechanical pretension over a period of 45 to 60 minutes until the increase of the pretension is not followed by a further augmentation in the twitch force, and then initiate the desired downstream experimental analysis. Cardiac contractility can be characterized by a variety of parameters, including the twitch force, time to peak and half maximal relaxation.The correlation between the beating rate and the twitch force is calculated by the force frequency relationship. For example, in this experiment, the stimulation frequency was increased in a stepwise fashion from 0.1 to five hertz leading to changes in the cardiac contractility. The post rest potentiation protocol illustrates the alterations of the twitch force Amplitude observed after a previous stimulation followed by a rest period of a defined duration.By varying the calcium concentration of the physiological solution in the organ bath, the calcium sensitivity of the contractile apparatus can be evaluated to assess the inotropic effects mediated by the activation of beta recepts. The isolated papillary muscle can also be treated with isoprene as demonstrated in this representative experiment to simulate ischemia. In vitro, the muscles are exposed to a glucose and pyruvate free extracellular solution bubbled with 95%nitrogen and 5%carbon dioxide leading to a decline of the twitch force and the development of ischemic contracture as illustrated in the graph.Once mastered the dissection and equilibration of one popularity, muscle preparation can be completed in about 19 minutes, even with taking the time to carefully isolate and mount the preparation without introducing mechanical or metabolic damage Following this procedure. Other methods like sharp electrode measurements, enzyme linked immunoassays and western blood analysis can be performed in order to answer additional questions After its development. This technique is a crucial approach for research in cardiac physiology and pharmacology, and for investigations of the cardiac phenotype in various transgenic rodent models.

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JoVE Journal

Biology

Transgenic mice have been extremely useful in ascribing physiological function to genes. As such, research in general, and functional studies of airway, in particular, have undergone a remarkable shift toward murine models. Here we provide protocols for in vitro trachea constriction studies to evaluate smooth muscle function in murine airway.

In vitro Measurements of Tracheal Constriction Using Mice

Individual cardiomyocytes from wild type and mutant mice can be isolated from the heart in order to study their contractility and calcium transients. This allows characterization of the contribution of cellular dysfunction to heart dysfunction from any cause.

Isolation and Physiological Analysis of Mouse Cardiomyocytes

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

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

This paper describes a protocol that assesses the changes of myofilament Ca2+ sensitivity during contraction in isolated cardiac myocytes from rat heart. Together with cardiac electrophysiology, systolic/diastolic cytosol Ca2+ levels and contraction/relaxation, this measurement is imperative in underpinning the mechanisms mediating cardiac excitation-contraction coupling in healthy and diseased hearts.

Assessment of Myofilament Ca2+ Sensitivity Underlying Cardiac Excitation-contraction Coupling

This protocol provides a detailed description of the echocardiographic approach for comprehensive phenotyping of heart and heart valve function in mice.

Echocardiographic Approaches and Protocols for Comprehensive Phenotypic Characterization of Valvular Heart Disease in Mice

We present the methodology for the isolation of murine myocytes and how to obtain voltage or calcium traces simultaneously with sarcomere shortening traces using fluorescence photometry with simultaneous digital cell geometry measurements.

Optical Imaging of Isolated Murine Ventricular Myocytes

Presented here is a protocol to study the coronary microcirculation in living murine heart tissue by ex vivo monitoring of the arterial perfusion pressure and flow that maintains the pressure, as well as vascular tree components including the capillary beds and pericytes, as the septal artery is cannulated and pressurized.

Dynamic Measurement and Imaging of Capillaries, Arterioles, and Pericytes in Mouse Heart

A set of protocols are presented that describe the measurement of contractile function via sarcomere length detection along with calcium (Ca2+) transient measurement in isolated rat myocytes. The application of this approach for studies in animal models of heart failure is also included.

Analysis of Cardiac Contractile Dysfunction and Ca2+ Transients in Rodent Myocytes

This protocol describes how to measure contractility in adult human primary cardiomyocytes from donor hearts with the MyoBLAZER system, a reliable platform for assessing drug-induced changes in contractility during preclinical development.

Measurement of Heart Contractility in Isolated Adult Human Primary Cardiomyocytes

Rapid myocardial and cardiac relaxation is essential for normal physiology. Mechanical relaxation mechanisms are now known to be dependent on strain rate. This protocol provides an overview of the acquisition and analysis of experiments to further study the mechanical control of relaxation.

Mechanical Control of Relaxation Using Intact Cardiac Trabeculae

Learn to isolate murine left ventricular papillary muscles for in vitro cardiac contractility studies using field electric pulse stimulation. This method enables precise evaluation of cardiac inotropy under controlled physiological conditions.

Contractility Measurements on Isolated Papillary Muscles for the Investigation of Cardiac Inotropy in Mice

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