Electrical, contractile, and ultrastructural properties of adult rat and guinea-pig ventricular myocytes in long-term primary cultures

1989 ◽  
Vol 67 (7) ◽  
pp. 740-750 ◽  
Author(s):  
M. Horackova ◽  
C. Mapplebeck
Keyword(s):  
Guinea Pig ◽  
Cultured Cells ◽  
Ventricular Myocytes ◽  
Contractile Activity ◽  
Primary Cultures ◽  
Morphological Characteristics ◽  
Ultrastructural Organization ◽  
Adult Rat ◽  
Intercellular Contacts

The electrical, contractile, and morphological characteristics of ventricular myocytes isolated from adult rat and guinea-pig hearts and maintained in cultures for 7–24 days are described. These cultured cells form different networks, depending on the species, when plated at certain density and maintained under specific conditions; the cells within the networks appear to be electrically coupled. Their resting and action potentials, their contractile activity (shortenings), and their pharmacological responses qualitatively resemble those of freshly isolated myocytes. Cultured cells from both species exhibit near-normal ultrastructural organization of sarcomeres, myofilaments, and mitochondria, as well as formation of intercellular contacts, or gap junctions. These data indicate that cultured adult rat and guinea-pig myocardial cells that make intercellular contacts possess electrical, contractile, and ultrastructural properties and responses to pharmacological agents similar to those of the respective adult myocardial tissues and the functionally intact freshly isolated cells from which these cultures are prepared. Thus, this study indicates that long-term cultures (7–24 days) of networked cardiac myocytes could be used as a valuable experimental model in various investigations of excitation–contraction coupling in cardiac muscle.Key words: cultured adult cardiomyocytes, contractility, electrical activity, ultrastructure, long-term primary cultures.

Possible role of Na+–Ca2+ exchange in the regulation of contractility in isolated adult ventricular myocytes from rat and guinea pig

1989 ◽  
Vol 67 (12) ◽  
pp. 1525-1533 ◽  
Author(s):  
M. Horackova
Keyword(s):  
Guinea Pig ◽  
Action Potentials ◽  
Ventricular Myocytes ◽  
Contractile Activity ◽  
Strong Dependence ◽  
Tonic Component ◽  
Adult Rat ◽  
Phasic Component ◽  
Single Ventricular Myocytes ◽  
Rat Myocytes

Action potentials and developed contractions of externally unloaded single ventricular myocytes isolated from adult rat and guinea pig hearts were recorded by means of an optical system for recording contractile activity during regular stimulation by microelectrodes. Under control conditions, the shortenings (twitches) in the rat myocytes were fully inhibited by 0.1 μM ryanodine, but they were rather insensitive to the Ca2+ blocker 0.2–0.5 μM nifedipine. In contrast, the contractions of the isolated guinea pig ventricular myocytes were greatly suppressed by 0.2–0.5 μM nifedipine (to <30%), while they were only slightly reduced by 1 μM ryanodine. When the Na+ gradient was decreased by reducing [Na]o or by elevating [Na]i in the presence of veratridine, the twitch contractions were increased in both species. The effect of reduced [Na]o on twitch contractions was not affected by ryanodine in either type of myocytes, while nifedipine still fully abolished the twitches in the guinea pig cells, indicating a strong dependence of guinea pig contractions on Ca2+ influx. On the other hand, the effect of a reduced Na gradient by veratridine was more complex; the usual twitch (phasic component) was increased and it was followed by a second (tonic) component which relaxed only after the repolarization of the action potential. While the phasic component was decreased by nifedipine and ryanodine in the usual way (as in the controls), the sustained contractions (lasting up to several seconds) were ryanodine and nifedipine insensitive. Furthermore, the cardiomyocytes of both species exposed to strontium in place of external calcium still exhibited all the effects observed when reducing the Na+ gradient. These data indicated that Sr2+ transport may occur via various routes similar to those of Ca2+ transport, including the Na+–Sr2+ exchange. In addition, in the presence of SrCl2 the rat myocytes exhibited longer durations of action potentials and their contractions became insensitive to ryanodine (like the guinea pig cells). It is concluded that Na+–Ca2+ exchange probably does not directly contribute in a quantitative fashion to the activation of contractile activity under normal conditions, but when the Na+ gradient is decreased, especially by increasing [Na]i, the contractions could be increased severalfold via this mechanism in both the rat and guinea pig cardiomyocytes.Key words: excitation–contraction coupling, isolated cardiac myocytes, nifedipine, ryanodine, veratridine, reduced [Na]o, epinephrine, SrCl2.

Remodelling of Ca2+ handling organelles in adult rat ventricular myocytes during long term culture

2010 ◽  
Vol 49 (3) ◽  
pp. 427-437 ◽  
Author(s):  
Karin Hammer ◽  
Sandra Ruppenthal ◽  
Cedric Viero ◽  
Anke Scholz ◽  
Ludwig Edelmann ◽  
...  
Keyword(s):  
Ventricular Myocytes ◽  
Long Term Culture ◽  
Rat Ventricular Myocytes ◽  
Adult Rat

Characteristics of papillary collecting duct cells in primary culture

1988 ◽  
Vol 255 (6) ◽  
pp. F1160-F1169 ◽  
Author(s):  
R. F. Husted ◽  
M. Hayashi ◽  
J. B. Stokes
Keyword(s):  
Cultured Cells ◽  
Collecting Duct ◽  
Short Circuit ◽  
Primary Cultures ◽  
Calf Serum ◽  
Morphological Characteristics ◽  
Short Circuit Current ◽  
Papillary Collecting Duct ◽  
Collecting Duct Cells ◽  
Na Uptake

We examined the electrophysiological and Na+ transport characteristics of rat papillary collecting duct (PCD) cells grown in primary cultures. Grown as monolayers on polycarbonate filters, the cells displayed similar morphological characteristics to native epithelia. They also bound Dolichus biflorus lectin, a property shared by native cells. Monolayers developed a peak electrical resistance of 100-200 omega.cm2 and a transmonolayer voltage of less than 2 mV. Similar values were measured in the perfused, native PCD of the same species as well as PCD cells cultured from rabbit and bovine kidneys. Hamster cells did not readily develop confluent monolayers under the same conditions. Exposure of the cultured cells to 10% fetal calf serum for 24 h caused the Na+ uptake across the apical membrane to double, an effect not reproduced by indomethacin, insulin, vasopressin, aldosterone, dexamethasone, or hexamethylene bisacetamide (an inducer of differentiation). Amiloride (1 mM) inhibited Na+ uptake by 50-80%. The measured short-circuit current did not correlate with Na+ uptake and was clearly dissociated by exposure to serum. The results suggest that there is more than one mechanism of ion transport by the rat PCD.

NRG-1-induced cardiomyocyte hypertrophy. Role of PI-3-kinase, p70S6K, and MEK-MAPK-RSK

1999 ◽  
Vol 277 (5) ◽  
pp. H2026-H2037 ◽  
Author(s):  
Ragavendra R. Baliga ◽  
David R. Pimental ◽  
You-Yang Zhao ◽  
William W. Simmons ◽  
Mark A. Marchionni ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
Ventricular Myocytes ◽  
Primary Cultures ◽  
Mitogen Activated Protein Kinase ◽  
Neonatal Rat ◽  
Cardiomyocyte Hypertrophy ◽  
S6 Kinase ◽  
Adult Rat ◽  
Mitogen Activated Protein ◽  
Ribosomal S6 Kinase

Neuregulins are a family of growth-promoting peptides known to be important in neural and mesenchymal tissue development. Targeted disruption of neuregulin (NRG)-1 or one of two of its cognate receptors, ErbB2 or ErbB4, results in embryonic lethality because of failure of the heart to develop. Although expression of NRGs and their receptors declines after midembryogenesis, both ErbB2 and ErbB4 are present in cardiac myocytes, and NRG-1 expression remains inducible in primary cultures of coronary microvascular endothelial cells from adult rat ventricular muscle. In neonatal rat ventricular myocytes, a soluble NRG-1, recombinant human glial growth factor-2, increased [3H]phenylalanine uptake and induced expression of atrial natriuretic factor (ANF) and sarcomeric F-actin polymerization. The effect of NRG-1 on [3H]phenylalanine uptake and sarcomeric F-actin polymerization was maximal at 20 ng/ml but declined at higher concentrations. NRG-1 activated p42/p44 mitogen-activated protein kinase (MAPK) [extracellular signal-regulated kinase (ERK)-2/ERK1] and ribosomal S6 kinase (RSK)-2 (90-kDa ribosomal S6 kinase), both of which could be inhibited by the MAPK/ERK kinase-1 antagonist PD-098059. NRG-1 also activated 70-kDa ribosomal S6 kinase, which was inhibited by either rapamycin or wortmannin. Activation of these pathways exhibited the same “biphasic” response to increasing NRG-1 concentrations. Wortmannin and LY-294002 blocked sarcomeric F-actin polymerization but not [3H]phenylalanine uptake or ANF expression, whereas PD-098059 consistently blocked both [3H]phenylalanine uptake and ANF expression but not actin polymerization. In contrast, rapamycin inhibited [3H]phenylalanine uptake and F-actin polymerization but not ANF expression. Thus NRG-ErbB signaling triggers multiple nonredundant pathways in postnatal ventricular myocytes.

Primary Culture of Strial Marginal Cells of Guinea Pig Cochlea: Growth, Morphologic Features, and Characterization

1991 ◽  
Vol 100 (12) ◽  
pp. 999-1006 ◽  
Author(s):  
Josué Achouche ◽  
An H. Wu ◽  
Der S. Liu ◽  
Patrice Tran Ba Huy
Keyword(s):  
Guinea Pig ◽  
Primary Culture ◽  
Cultured Cells ◽  
Stria Vascularis ◽  
Primary Cultures ◽  
Suitable Model ◽  
Type I ◽  
Cellular Mechanisms ◽  
Electron Microscopic ◽  
Marginal Cells

To further investigate the cellular mechanisms involved in the formation of endolymph, primary cultures of marginal cells of guinea pig were established. Minute explants obtained by mechanical dissociation of stria vascularis were plated on collagen type I precoated impermeable substrate in serum-free, hormone-supplemented medium. A confluent layer of epithelial-like cells was obtained within 2 weeks. The cultured cells formed domes, demonstrating that they retain some of their transepithelial properties. Polarization was also suggested by electron microscopic observation of apical microvilli and tight junctions. Immunohistochemical methods revealed that the cultured cells coexpressed cytokeratin and vimentin, demonstrating their epithelial origin, although some degree of dedifferentiation occurred. Thus, a primary culture of marginal cells can be established that may be a suitable model for an in-depth investigation of the function of the marginal cells.

Electrical activity of rat myocardial cells in culture: La3+-induced alterations

1979 ◽  
Vol 237 (1) ◽  
pp. C87-C95 ◽  
Author(s):  
M. C. Kitzes ◽  
M. W. Berns
Keyword(s):  
Electrical Activity ◽  
Cultured Cells ◽  
Contractile Activity ◽  
Membrane Potentials ◽  
Neonatal Rat ◽  
Complete Disappearance ◽  
Myocardial Cells ◽  
Adult Rat ◽  
Cells In Culture ◽  
Ventricular Cells

Intracellular analysis of neonatal rat (1–2 day old) ventricular cells in culture shows that contracting myocardial cells exhibit an array of different patterns of spontaneous electrical activity. Resting membrane potentials varied between -40 mV and -98 mV. Our results indicate that some cultured cells show resting membrane potentials, overshoot, and total spike amplitude values comparable to those normally found in neonatal and adult rat heart. A low ratio of pacemaker (40%) to nonpacemaker cells (60%) and low incidence of hyperpolarizing after-potentials (35%) were found. La3+application (0.1–4.0 mM) induced progressive cell depolarization, concomitant diminution in discharge frequency, and marked alteration of action potential configuration. A parallel decline in frequency and strength of rhythmic contractions was observed. Abolition of contractility occurred only in association with depolarization and complete disappearance of action potentials. Recovery of electrical and contractile activity followed medium replacement. Our results indicate that La3+ does not act as a specific excitation-contraction (E-C) uncoupler in the cultured cells but has multiple effects upon their normal electrical characteristics.

Cardiac myocytes release leukocyte-stimulating factors

1995 ◽  
Vol 269 (3) ◽  
pp. H980-H987 ◽  
Author(s):  
K. D. Massey ◽  
R. M. Strieter ◽  
S. L. Kunkel ◽  
J. M. Danforth ◽  
T. J. Standiford
Keyword(s):  
Cardiac Myocytes ◽  
Ventricular Myocytes ◽  
Oxidant Stress ◽  
Interleukin 1 ◽  
Primary Cultures ◽  
Biologically Active ◽  
Enzyme Linked Immunosorbent Assay ◽  
Adult Rat ◽  
Inflammatory Protein ◽  
Cellular Source

The production of cytokines directly from cardiac myocytes has not been previously demonstrated and could represent an important mechanism and site of intervention in ischemia and reperfusion injuries. Macrophage inflammatory protein-2 (MIP-2) and monocyte chemotactic protein (MCP) are chemotactic cytokines (chemokines) that stimulate polymorphonuclear leukocytes (PMNs) and monocytes, respectively. Endothelium has been implicated as being a major cellular source of leukocyte-activating factors. We hypothesized that the myocardial cells may also play an important role in producing chemokines independently of endothelium. Primary cultures of adult rat ventricular myocytes were prepared. Cultured myocytes were stimulated with either interleukin 1 (IL-1), tumor necrosis factor (TNF), or lipopolysaccharide (LPS). MIP-2 and MCP mRNA were expressed in adult rat myocytes following stimulation. Our studies indicate that ventricular myocytes expressed chemokine mRNA and protein in both a dose- and time-dependent fashion. MIP-2 and MCP release, determined by enzyme-linked immunosorbent assay, was biologically active, accounting for approximately 40% of the PMN and monocyte chemotactic activity produced by these cells. These results suggest that cardiac myocytes may directly recruit activated leukocytes into areas of injury. Such a recruiting process could underlie the migration of leukocytes into areas of oxidant stress and play a role in development of reperfusion injury of myocardium.

scholarly journals Myocyte Response to Stimulation of Receptors and Ion Channels

1990 ◽  
Vol 18 (4a) ◽  
pp. 454-463 ◽  
Author(s):  
James D. Marsh
Keyword(s):  
Calcium Channels ◽  
Prolonged Exposure ◽  
Cultured Cells ◽  
Ventricular Myocytes ◽  
Binding Studies ◽  
Down Regulation ◽  
Intact Cells ◽  
Adult Rat ◽  
Dependent Process ◽  
Ventricular Cells

Ventricular myocytes dissociated from adult rat heart and cultured chick embryo ventricular cells were utilized to examine mechanisms by which neurotransmitters, hormones, and ontogeny modulate expression and function of β-adrenergic receptors and L-type calcium channels. Either freshly dissociated cells or cultured cells were studied by an optical-video system to characterize contractility and, in some instances, by a microspectrofluorimeter to determine [Ca2+]i as reported by fura 2. Ligand binding studies in intact cells and membranes were conducted with receptor and ion channel antagonists and agonists. Exposure of intact cells to isoproterenol produced contractile de-sensitization, loss of high affinity receptors from the sarcolemma and closely coupled decline in hormone-sensitive adenylate cyclase activity. Desensitization was by a microfilament-dependent process. Down-regulation depended upon microtubular function. During development of the chick heart, there was an increase in number of dihydropyridine binding sites, taken as a measure of number of L-type calcium channels, at a time when sensitivity to [Ca2+]o and to Bay k 8644 declined. Thyroid hormone was capable of up-regulating L-type calcium channels. Prolonged exposure to a β-adrenergic agonist produced coordinate down-regulation of β-receptors and calcium channels. Down-regulation was a cAMP-dependent process. Thus, the β-adrenergic receptor and a distal component of the effector-response coupling system, the L-type calcium channel, can be regulated independently and in concert by physiologically and pathophysiologically important mechanisms.

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