Name: contractility measurements on isolated papillary muscles for the investigation of cardiac inotropy in mice
Uploaded: 2015-09-17T15:00:00
Description: Watch this Scientific Journal Video about Contractility Measurements on Isolated Papillary Muscles for the Investigation of Cardiac Inotropy in Mice at JoVE.com

这项工作是由德意志研究联合会（B＆K 196“Signaltransduktion北adaptativen UND maladaptiven kardialen改建-Prozessen”，FR 1638 / 1-2）以及由DZHK（德国中心心血管研究，健康研究的德国中心的部分支持，这是一个BMBF（德国教育和研究部的）行动）。

注：分离过程的基本步骤示于图1中的所有步骤都在下面详细描述的协议。乳头肌隔离，安装在器官浴室，采集和分析是在一个连续的和强制性的时间表进行。 所有的动物实验均按照有关保护动物的有德国法律进行，并批准了海德堡大学的伦理审查委员会。 1.准备仪表<ol><li>对于收缩测量使用多节器官浴设置。 离体收缩器官浴中设置?...

<ol>
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H., Meyer, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phospholamban:+a+major+determinant+of+the+cardiac+force-frequency+relationship.">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/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanical+activity+and+force-frequency+relationship+of+isolated+mouse+papillary+muscle:+effects+of+extracellular+calcium+concentration,+temperature+and+contraction+type.">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/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adenylyl+cyclase-mediated+effects+contribute+to+increased+Isoprenaline-induced+cardiac+contractility+in+TRPM4+deficient+mice.">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/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Studies+of+the+contractility+of+mammalian+myocardium+at+low+rates+of+stimulation.">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/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sarcoplasmic+reticulum+Ca2++load+in+human+heart+failure.">Sarcoplasmic reticulum Ca2+ load in human heart failure.</a> Basic Res Cardiol. 97, 163-171 (2002).</li><li>Koch-Weser, J., Blinks, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Influence+of+the+Interval+between+Beats+on+Myocardial+Contractility.">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/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Myocardial+remodeling+after+large+infarcts+in+rat+converts+post+rest-potentiation+in+force+decay.">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/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+ischemia-reperfusion+on+the+post-rest+inotropy+of+isolated+perfused+rat+heart.">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/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Beyond+Bowditch:+the+convergence+of+cardiac+chronotropy+and+inotropy.">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/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantification+of+the+rat+left+ventricle+force+and+Ca2++-frequency+relationships:+similarities+to+dog+and+human.">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><li>Schmidt, U., Hajjar, R. J., Gwathmey, J. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+force-interval+relationship+in+human+myocardium.">The force-interval relationship in human myocardium.</a> J Card Fail. 1 (4), 311-321 (1995).</li><li>Rossman, E. I., Petre, R. E., Chaudhary, K. W., Piacentino, V. 3. r. d., Janssen, P. M., Gaughan, J. P., Houser, S. R., Margulies, K. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Abnormal+frequency-dependentresponses+represent+the+pathophysiologic+signature+of+contractile+failure+inhuman+myocardium.">Abnormal frequency-dependentresponses represent the pathophysiologic signature of contractile failure inhuman myocardium.</a> JMCC. 36 (1), 33-42 (2004).</li><li>Moran, A. E., Forouzanfar, M. H., Roth, G. A., Mensah, G. A., Ezzati, M., Murray, C. J., Naghavi, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Temporal+trends+in+ischemic+heart+disease+mortality+in+21+world+regions,+1980+to+2010:+the+Global+Burden+of+Disease+2010+stud.">Temporal trends in ischemic heart disease mortality in 21 world regions, 1980 to 2010: the Global Burden of Disease 2010 stud.</a> Circulation. 129 (14), 1483-1492 (1980).</li><li>Lee, J. A., Allen, D. 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在这个手稿，我们描述的方法来研究的鼠的乳头肌收缩力在体外可被用来回答有关心脏生理和病理小鼠几个科学问题以及支持的转基因品系的分析和新药物的方法的发现治疗心脏功能障碍。我们示出了使用这种方法来评估的心肌收缩性生理，病理和药理性质（参照图3）。这里未示出的其它应用包括使用不同的药理学试剂的细胞内途径的评价（参见12）。 </p...

像离子通道参照它们的作用为心脏收缩蛋白的调查是必不可少的发现不同的病理机制，并建立新的治疗策略用于心脏疾病，如局部缺血和心脏衰竭。 哺乳动物心肌收缩功能已知通过各种离子通道，转运和其他蛋白质被调制。动作电位诱发激活电压依赖性肌膜L型钙通道引出的Ca 2+内流 ，从细胞外空间，并随后到的 Ca 2+ -诱导的 Ca 2+释放（CICR）1，这将触发蜂窝收缩2。钙 -信号起着心脏收缩力和适应方面发挥核心作用的生理或病理压力。儿茶酚胺激活心脏β肾上腺素能受体，从而刺激腺苷酸环化酶（AC）的合成营。被激活，proteiÑ ​​激酶A（PKA）的磷酸化细胞内的不同和膜相关蛋白质如L-型Ca 2+通道 ，磷蛋白和兰尼碱受体导致的钙瞬变和心脏收缩1,3,4修饰。的cAMP通过磷酸二酯酶（PDE）的降解。 G s - 偶联受体比β肾上腺素能受体其它的活化也导致的cAMP蓄积。 心室肌细胞肌条收缩测量的技术已经非常成熟的大型哺乳动物物种5-8。基于基因的可能性靶向小鼠建立的方法来分析鼠心脏生理是重要的。然而，大约在小鼠中分离肌肉制剂的生理特性的现有数据的差异取决于实验条件9-12。 所描述的方法被用来分析左心室乳头肌的预心脏收缩parations 体外 。在没有影响改变心肌收缩力在体内 ，如血压，神经体液刺激和生理或代谢应激进行心肌收缩力的研究。承包肌肉制剂的跳动速率可以严格定义，改变随意。抽搐力能够在特定刺激，如钙浓度，跳动频率或温度的情况下进行分析。此外，这种方法可用于研究不同信号传导途径的组件和通过控制上述实验条件下进行比较的转基因小鼠模型的心脏的性能。

<table border="0" cellpadding="0" cellspacing="0">
	<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&#160;</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&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium pyruvate</td>
			<td>Sigma-Aldrich</td>
			<td>P5280&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Calcium chloride dihydrate</td>
			<td>Sigma-Aldrich</td>
			<td align="right">223506</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnesium sulfate heptahydrate</td>
			<td>Sigma-Aldrich</td>
			<td align="right">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>Forskolin</td>
			<td>Sigma-Aldrich</td>
			<td>F3917</td>
			<td>Hazard statement H312, solve in DMSO</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/10ml</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

乳头肌从成年小鼠心脏分离，可用于在不同的生理/病理条件下，研究心脏收缩。独立的外部影响，如血管紧张或神经体液状态的收缩特性可以被评估。它描述了单细胞测量孤立心肌细胞之间， 像超声心动图体内研究科学的方法。因此，乳头肌制剂作为一个很好的模型来研究心脏生理/病理生理学和可用于像由药剂的调制或转基因动物模型的探索调查。这里，我们描述分离鼠左前乳头肌以调查在器官浴设置心脏收缩的方法。在对比一个肌肉条准备从心室壁分离，乳头肌可以全盘而不损伤肌肉Tissu酒店制备Ë严重。器官浴设置包括几个温度控制，毒气和电极配备器官浴室。将分离的乳头肌固定在器官浴室和电刺激。诱发抽搐力是使用压力传感器和参数，如抽搐力的振幅和抽搐动力学分析记录。不同的实验方案，可以执行以调查收缩剂如儿茶酚胺或其他药物的碳酸钙和频率相关的收缩以及剂量 - 反应曲线。此外，像急性缺血病理状况可以模拟。

本手稿的孤立鼠的乳头肌收缩制剂测量值的协议被调谐到最佳条件，以实现在生理条件下可再现的实验结果。为了确定最佳的实验条件下，我们进行试点实验，不同的器官浴温度和细胞外钙离子浓度（参见12）。这里描述的协议用的1.5mM的胞外钙浓度和32℃的温度下进行。 基底收缩特性要抽搐力振幅表征基底收缩性质，参数，抽搐率，达峰时间（?...

Watch this Scientific Journal Video about Contractility Measurements on Isolated Papillary Muscles for the Investigation of Cardiac Inotropy in Mice at JoVE.com

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

鼠左心室乳头肌可用于研究在体外心脏收缩。本文详细介绍了隔离和实验方案，研究心脏收缩的特点。

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