Cardiomyocyte culture — an update on the in vitro cardiovascular model and future challenges

Parameswaran, Sreejit; Kumar, Sujeet; Verma, Rama Shanker; Sharma, Rajendra K.

doi:10.1139/cjpp-2013-0161

Cited by 37 publications

(36 citation statements)

References 198 publications

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“…Methods that preserve as closely as possible the in vivo integrity and function of the myocytes during 1 to 2 weeks of culture are still undergoing improvements. 223,231,232 Nevertheless, with current culture methods, specific morphological and functional alterations do occur. Myocytes become more rounded as opposed to square-edged, rod-shaped appearance, and show a decreased T-tubule density, as well as specific changes in ion channels and ion currents.…”

Section: Metabolism Of Isolated Cardiac Cells and Cell Linesmentioning

confidence: 99%

Assessing Cardiac Metabolism

Taegtmeyer¹,

Young²,

Lopaschuk³

et al. 2016

Circulation Research

233

147

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In a complex system of interrelated reactions, the heart converts chemical energy to mechanical energy. Energy transfer is achieved through coordinated activation of enzymes, ion channels, and contractile elements, as well as structural and membrane proteins. The heart’s needs for energy are difficult to overestimate. At a time when the cardiovascular research community is discovering a plethora of new molecular methods to assess cardiac metabolism, the methods remain scattered in the literature. The present statement on “Assessing Cardiac Metabolism” seeks to provide a collective and curated resource on methods and models used to investigate established and emerging aspects of cardiac metabolism. Some of those methods are refinements of classic biochemical tools, whereas most others are recent additions from the powerful tools of molecular biology. The aim of this statement is to be useful to many and to do justice to a dynamic field of great complexity.

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Section: Metabolism Of Isolated Cardiac Cells and Cell Linesmentioning

confidence: 99%

Assessing Cardiac Metabolism

Taegtmeyer¹,

Young²,

Lopaschuk³

et al. 2016

Circulation Research

233

147

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“…However, the lack of other potentially important regulatory subunits or intracellular pathways could be a real disadvantage. Another possibility is to reduce the expression of the channel protein in cell culturing which is a rather difficult process in cardiomyocytes (Parameswaran et al 2013). Carefully planned voltage-protocols together with the suppression of other currents could be used in native cardiac cells to isolate the examined current.…”

Section: Introductionmentioning

confidence: 99%

9–Anthracene carboxylic acid is more suitable than DIDS for characterization of calcium-activated chloride current during canine ventricular action potential

Váczi

Hegyi

Ruzsnavszky

et al. 2014

Naunyn-Schmiedeberg's Arch Pharmacol

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Understanding the role of ionic currents in shaping the cardiac action potential (AP) has great importance as channel malfunctions can lead to sudden cardiac death by inducing arrhythmias. Therefore, researchers frequently use inhibitors to selectively block a certain ion channel like 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS) and 9-anthracene carboxylic acid (9-AC) for calcium-activated chloride current (I Cl(Ca) ). This study aims to explore which blocker is preferable to study I Cl (Ca) . Whole-cell voltage-clamp technique was used to record I Ca,L, I Ks, I Kr and I K1 , while action potentials were measured using sharp microelectrodes. DIDS-(0.2 mM) and 9-AC-sensitive (0.5 mM) currents were identical in voltage-clamp conditions, regardless of intracellular Ca 2+ buffering. DIDS-sensitive current amplitude was larger with the increase of stimulation rate and correlated well with the rate-induced increase of calcium transients. Both drugs increased action potential duration (APD) to the same extent, but the elevation of the plateau potential was more pronounced with 9-AC at fast stimulation rates. On the contrary, 9-AC did not influence either the AP amplitude or the maximal rate of depolarization (V max ), but DIDS caused marked reduction of V max . Both inhibitors reduced the magnitude of phase-1, but, at slow stimulation rates, this effect of DIDS was larger. All of these actions on APs were reversible upon washout of the drugs. Increasing concentrations of 9-AC between 0.1 and 0.5 mM in a cumulative manner gradually reduced phase-1 and increased APD. 9-AC at 1 mM had no additional actions upon perfusion after 0.5 mM. The halfeffective concentration of 9-AC was approximately 160 μM with a Hill coefficient of 2. The amplitudes of I Ca,L, I Ks, I Kr and I K1 were not changed by 0.5 mM 9-AC. These results suggest that DIDS is equally useful to study I Cl(Ca) during voltageclamp but 9-AC is superior in AP measurements for studying the physiological role of I Cl(Ca) due to the lack of sodium channel inhibition. 9-AC has also no action on other ion currents (I Ca,L , I Kr , I Ks , I K1 ); however, I Ca,L tracings can be contaminated with I Cl(Ca) when measured in voltage-clamp condition.

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“…Specifically, hPSC-CMs can model myocardial physiology in vitro (7). hPSCCMs may be better in vitro models of contractility than neonatal or mature murine primary CMs, because they can be maintained in culture longer (2) and because the sarcomeric contractile machinery differs between human and murine CMs (8,9).…”

mentioning

confidence: 99%

Contractility of single cardiomyocytes differentiated from pluripotent stem cells depends on physiological shape and substrate stiffness

Ribeiro

Ang

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

405

456

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Single cardiomyocytes contain myofibrils that harbor the sarcomerebased contractile machinery of the myocardium. Cardiomyocytes differentiated from human pluripotent stem cells (hPSC-CMs) have potential as an in vitro model of heart activity. However, their fetallike misalignment of myofibrils limits their usefulness for modeling contractile activity. We analyzed the effects of cell shape and substrate stiffness on the shortening and movement of labeled sarcomeres and the translation of sarcomere activity to mechanical output (contractility) in live engineered hPSC-CMs. Single hPSC-CMs were cultured on polyacrylamide substrates of physiological stiffness (10 kPa), and Matrigel micropatterns were used to generate physiological shapes (2,000-μm 2 rectangles with length:width aspect ratios of 5:1-7:1) and a mature alignment of myofibrils. Translation of sarcomere shortening to mechanical output was highest in 7:1 hPSC-CMs. Increased substrate stiffness and applied overstretch induced myofibril defects in 7:1 hPSC-CMs and decreased mechanical output. Inhibitors of nonmuscle myosin activity repressed the assembly of myofibrils, showing that subcellular tension drives the improved contractile activity in these engineered hPSC-CMs. Other factors associated with improved contractility were axially directed calcium flow, systematic mitochondrial distribution, more mature electrophysiology, and evidence of transverse-tubule formation. These findings support the potential of these engineered hPSC-CMs as powerful models for studying myocardial contractility at the cellular level.contractility | sarcomeres | cardiomyocyte | stem cell | single cell

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Cardiomyocyte culture — an update on the in vitro cardiovascular model and future challenges

Cited by 37 publications

References 198 publications

Assessing Cardiac Metabolism

Assessing Cardiac Metabolism

9–Anthracene carboxylic acid is more suitable than DIDS for characterization of calcium-activated chloride current during canine ventricular action potential

Contractility of single cardiomyocytes differentiated from pluripotent stem cells depends on physiological shape and substrate stiffness

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