Structure of myofibrils at extra-junctional membrane attachment sites in cultured cardiac muscle cells

The composition and organization of myofibrils at extra-junctional membrane attachment sites in cultured neonatal rat cardiac muscle cells were analysed by immunofluorescence and electron microscopy. When myofibril terminals attached to the cell membrane via focal contacts at regions of the sarcolemma that lacked intercalated discs, they appeared to be non-striated and resembled thick actin cables. Although the non-striated terminals contained actin, myosin and alpha-actinin, the proteins were not organized into recognizable sarcomeres at the light microscopic level. Analysis of the structure of the terminals in the electron microscope confirmed that the usual sarcomeric organization and attachments to the sarcolemma were markedly modified. The non-striated myofibril terminals differed in structure from both stress fibres in non-muscle cells and stress fibre-like structures present in embryonic heart cells in culture. Non-striated myofibril terminals attached to the cell membrane by lateral contact with extra-junctional electron-dense membrane plaques rather than by insertion by their ends into the fascia adherens. It is proposed that the structure and composition of membrane-attachment points for myofibrils may have an influence on the structure, organization or stability of contractile elements in cardiac muscle.

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Choline Permeability in Cardiac Muscle Cells of the Cat

The Journal of General Physiology ◽

10.1085/jgp.55.5.602 ◽

1970 ◽

Vol 55 (5) ◽

pp. 602-619 ◽

Cited By ~ 11

Author(s):

S. Bosteels ◽

A. Vleugels ◽

E. Carmeliet

Keyword(s):

Cell Membrane ◽

Cardiac Muscle ◽

Paper Chromatography ◽

Heart Muscle ◽

Muscle Cells ◽

Heart Cells ◽

Ventricular Muscle ◽

Cardiac Cell ◽

Cardiac Muscle Cells ◽

Rapid Phase

Permeability of the cardiac cell membrane to choline ions was estimated by measuring radioactive choline influx and efflux in cat ventricular muscle. Maximum values for choline influx in 3.5 and 137 mM choline were respectively 0.56 and 9 pmoles/cm2·sec. In 3.5 mM choline the intracellular choline concentration was raised more than five times above the extracellular concentration after 2 hr of incubation. In 137 mM choline, choline influx corresponded to the combined loss of intracellular Na and K ions. Paper chromatography of muscle extracts indicated that choline was not metabolized to any important degree. The accumulation of intracellular choline rules out the existence of an efficient active pumping mechanism. By measuring simultaneously choline and sucrose exchange, choline efflux was analyzed in an extracellular phase, followed by two intracellular phases: a rapid and a slow one. Efflux corresponding to the rapid phase was estimated at 16–45 pmoles/cm2·sec in 137 mM choline and at 1.3–3.5 pmoles/cm2·sec in 3.5 mM choline; efflux in 3.5 mM choline was proportional to the intracellular choline concentration. The absolute figures for unidirectional efflux were much larger than the net influx values. The data are compared to Na and Li exchange in heart cells. Possible mechanisms for explaining the choline behavior in heart muscle are discussed.

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Coaggregation and formation of a joint myocardial tissue by embryonic mammalian and avian heart eel Is

Development ◽

10.1242/dev.59.1.263 ◽

1980 ◽

Vol 59 (1) ◽

pp. 263-279

Author(s):

Asish C. Nag ◽

Mei Cheng ◽

Christopher J. Healy

Keyword(s):

Cardiac Muscle ◽

Tritiated Thymidine ◽

Muscle Cells ◽

Intercellular Adhesion ◽

Heart Cells ◽

Embryonic Cells ◽

Cardiac Muscle Cells ◽

Electron Microscopic Autoradiography ◽

Stage Of Development ◽

Mixed Aggregates

Intercellular adhesion and tissue reconstruction from homologous dissociated embryonic cells from two species were studied. Dissociated 12-day-old embryonic rat heart cells and 50-h-old embryonic chick heart cells were labeled with tritiated thymidine and allowed to aggregate in Erlenmeyer flasks during rotation culture on a gyratory water-bath shaker. The cultures were continued for 72 h. Cell aggregates were examined microscopically for evidence of contractility and subsequently processed at intervals between 1 and 72 h for transmission electron microscopic autoradiography. Rat and chick hearts used in this study appeared comparable in their stage of development and cellular composition. With the exception of mature blood cells and some fibroblastic non-muscle cells, all chick cardiac muscle cells were labeled with tritiated thymidine. As the cultures continued, aggregates increased in size by continuous accretion of cells and joining of small clusters. The cells within these mixed aggregates exhibited synchronous contractility from 1 h until the cultures were terminated. Most of the aggregation in both control and mixed aggregates was completed within 24 h. Control aggregates consisted of cells from a single species, either chick or rat. Approximately 18 % of labeled chick cardiac muscle cells established intercellular contacts and junctions with unlabeled rat cardiac muscle cells after 6 h of culture. This increased to 72 % after 72 h in culture. The junctions observed between chick and rat cardiac muscle cells were desmosomes and hemidesmosomes. Approximately 13 % of the cardiac muscle cells of one species either chick or rat were found scattered within clusters which contained .15 % of the cells from the other species after 72 h of culture. These scattered cells did not form junctions with monospecific cell groups. The implications of these intercellular adhesions between the same and different species are discussed, bearing two hypotheses in mind: (1) Moscona and collaborators, (2) Burdickand Steinberg. This study suggests that cellular as well as species identity plays an important role in the determination of intercellular adhesion among the embryonic cells from different species.

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Effects of hypoxia on cardiac growth in neonatal rat

American Journal of Physiology-Legacy Content ◽

10.1152/ajplegacy.1976.231.5.1445 ◽

1976 ◽

Vol 231 (5) ◽

pp. 1445-1450 ◽

Cited By ~ 37

Author(s):

M Hollenberg ◽

N Honbo ◽

AJ Samorodin

Keyword(s):

Body Weight ◽

Left Ventricle ◽

Cardiac Muscle ◽

Neonatal Period ◽

Vascular Endothelial Cells ◽

Muscle Cells ◽

Left Ventricular ◽

Neonatal Rat ◽

Low Oxygen ◽

Cardiac Muscle Cells

To determine whether low oxygen environments enhance cardiac cell division in the neonatal period, newborn rat pups were reared for 21 days in 12-15% oxygen. Left ventricle and right ventricle weights were 30 and 180% greater than controls matched for body weight (P less than 0.001) as were left ventricle/body weight ratios (3.68+/-0.26 vs. 2.99+/-0.05 mg LV/g body wt,P less than 0.001). Left ventricular total DNA and DNA concentration was 95 and 48% greater than controls (P less than 0.001). Autoradiography confirmed that this increase in ventricular DNA resulted from an increased rate of division of cardiac muscle cells, fibroblast, and vascular endothelial cells. When [3H]thymidine was injected on day ), autoradiographs prepared on day 21 reflected an increased dilution of label in hypoxic rats consistent with enhanced proliferation. The labeling index and grains per nucleus of ventricular muscle cells was 25% (P less than 0.01) and 20% (P less than 0.02) less than controls, Thus, hypoxic stress applied early in the neonatal period augments the rate of division and ultimate number of cardiac muscle cells. Whether this enhancement results from a primary effect of oxygen or from secondary hemodynamic factors remains unknown.

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Serum-free, chemically defined medium to evaluate the direct effects of growth factors and inhibitors on proliferation and function of neonatal rat cardiac muscle cells in culture

In Vitro Cellular & Developmental Biology ◽

10.1007/bf02623629 ◽

1989 ◽

Vol 25 (7) ◽

pp. 601-606 ◽

Cited By ~ 26

Author(s):

Takahiko Suzuki ◽

Miyuki Ohta ◽

Hiroyoshi Hoshi

Keyword(s):

Growth Factors ◽

Cardiac Muscle ◽

Muscle Cells ◽

Defined Medium ◽

Neonatal Rat ◽

Chemically Defined Medium ◽

Direct Effects ◽

Cardiac Muscle Cells ◽

Serum Free ◽

And Function

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Translation of heart preproenkephalin mRNA and secretion of enkephalin peptides from cultured cardiac myocytes

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.1992.263.5.h1560 ◽

1992 ◽

Vol 263 (5) ◽

pp. H1560-H1566 ◽

Cited By ~ 11

Author(s):

J. P. Springhorn ◽

W. C. Claycomb

Keyword(s):

Cardiac Muscle ◽

Muscle Cells ◽

Neonatal Rat ◽

Ribonuclease Protection Assay ◽

Cardiac Muscle Cells ◽

Adult Rat ◽

Rabbit Reticulocyte Lysate System ◽

Ribonuclease Protection ◽

Carboxy Terminal

Rat ventricular cardiac muscle has previously been shown to contain exceptionally high levels of preproenkephalin mRNA (ppEnk mRNA). We have recently determined that the level of ppEnk mRNA is developmentally and hormonally regulated in rat ventricular cardiac muscle tissue and in cultured myocytes (J. P. Springhorn and W. C. Claycomb. Biochem. J. 258: 73-77, 1989). We demonstrate in the current study that heart ppEnk mRNA is structurally identical at the 5' end to brain ppEnk mRNA using a ribonuclease protection assay and that heart ppEnk mRNA can be translated in vitro using a rabbit reticulocyte lysate system. In vitro synthesized preproenkephalin peptides were immunoprecipitated with a polyclonal antibody directed to the carboxy-terminal seven amino acids of preproenkephalin. We have also established by radioimmunoassay that enkephalin-containing peptides are secreted from cultured neonatal and adult rat ventricular cardiac muscle cells. This secretion is linear with respect to time and can be stimulated by phorbol 12-myristate 13-acetate (PMA) and adenosine 3',5'-cyclic monophosphate (cAMP). It was determined by column chromatography that cAMP induced neonatal rat ventricular cardiac muscle cells to secrete Met5-enkephalin-Arg6-Phe7, whereas PMA plus 3-isobutyl-1-methylxanthine induced adult rat ventricular cardiac muscle cells to secrete Met5-enkephalin. These studies establish that ventricular heart muscle ppEnk mRNA can be translated and that enkephalin peptides are secreted from ventricular cardiac muscle cells.

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Assembly and remodelling of myofibrils and intercalated discs in cultured neonatal rat heart cells

Journal of Cell Science ◽

10.1242/jcs.86.1.233 ◽

1986 ◽

Vol 86 (1) ◽

pp. 233-248

Author(s):

B.T. Atherton ◽

D.M. Meyer ◽

D.G. Simpson

Keyword(s):

Rat Heart ◽

Muscle Cells ◽

Neonatal Rat ◽

Cell Contact ◽

Heart Cells ◽

Cardiac Muscle Cells ◽

Rhodamine Phalloidin ◽

Intercalated Discs ◽

Neonatal Rat Heart ◽

Rat Heart Cells

The reorganization of myofibrils and the re-formation of intercalated discs was examined in neonatal rat cardiac muscle cells during the first 72 h of culture. Rhodamine phalloidin was used to monitor the organizational state of the myofibrils and antibodies to desmoplakin and vinculin were used as markers for the presence of desmosomes and fasciae adherentes, respectively. Tiny punctate desmosomes were observed between muscle cells after 24 h and apparently increased in number and/or size between 24 and 48 h in culture. Fasciae adherentes were not detectable with antibodies to vinculin until after 48 h in culture. Well-defined sarcomeres were restored after 48 h in culture. Once formed the sarcomeric organization of the myofibrils was found to be stable provided they were attached to the sarcolemma via intercalated discs. However, if the myofibrils attached to regions of the membrane that lacked intercalated discs the sarcomeres appeared to break down gradually centripetally. When myofibrils attached to the membrane at the free edges of cells that were not in contact with other muscle cells, the striations stopped abruptly at a considerable distance before the myofibril attached to the membrane. These non-striated terminals elongated between 48 and 72 h and were associated with focal contacts that contained vinculin. Overall the results suggest that cell-cell contact may be critical for the stabilization of normal myofibrillar structure in the heart.

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