scholarly journals Immunoelectron microscopical localization of phospholamban in adult canine ventricular muscle.

1987 ◽  
Vol 104 (5) ◽  
pp. 1343-1352 ◽  
Author(s):  
A O Jorgensen ◽  
L R Jones
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Cardiac Muscle ◽  
Subcellular Distribution ◽  
Immunogold Labeling ◽  
Relative Distribution ◽  
Ventricular Muscle ◽  
Myocardial Cells ◽  
Microscopical Localization ◽  
Cytoplasmic Side ◽  
Junctional Sarcoplasmic Reticulum

The subcellular distribution of phospholamban in adult canine ventricular myocardial cells was determined by the indirect immunogold-labeling technique. The results presented suggest that phospholamban, like the Ca2+-ATPase, is uniformly distributed in the network sarcoplasmic reticulum but absent from the junctional portion of the junctional sarcoplasmic reticulum. Unlike the Ca2+-ATPase, but like cardiac calsequestrin, phospholamban also appears to be present in the corbular sarcoplasmic reticulum. Comparison of the relative distribution of phospholamban immunolabeling in the sarcoplasmic reticulum with that of the sarcolemma showed that the density of phospholamban in the network sarcoplasmic reticulum was approximately 35-fold higher than that of the cytoplasmic side of the sarcolemma, which in turn was found to be three- to fourfold higher than the density of the background labeling. However, a majority of the specific phospholamban labeling within 30 nm of the cytoplasmic side of the sarcolemma was clustered and present over the sarcoplasmic reticulum in the subsarcolemmal region of the myocardial cells, suggesting that phospholamban is confined to the junctional regions between the sarcolemma and the sarcoplasmic reticulum, but absent from the nonjunctional portion of the sarcolemma. Although the resolution of the immunogold-labeling technique used (60 nm) does not permit one to determine whether the specific labeling within 30 nm of the cytoplasmic side of the sarcolemma is associated with the sarcolemma and/or the junctional sarcoplasmic reticulum, it is likely that the low amount of labeling in this region represents phospholamban associated with sarcoplasmic reticulum. These results suggest that phospholamban is absent from the sarcolemma and confined to the sarcoplasmic reticulum in cardiac muscle.

The distribution of calsequestrin in rat myocardial sarcoplasmic reticulum

1984 ◽  
Vol 42 ◽  
pp. 712-713
Author(s):  
A. O. Jorgensen ◽  
A. C.-Y. Shen ◽  
K. P. Campbell ◽  
G. Denney
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Ventricular Muscle ◽  
Myocardial Cells ◽  
Transverse Tubules ◽  
Purkinje Fibers ◽  
Immunofluorescence Localization ◽  
Band Region ◽  
Junctional Sarcoplasmic Reticulum

We have previously identified and purified calsequestrin from canine ventricular muscle. Immunofluorescence localization suggested that cardiac calsequestrin is confined to the lumen of the interior and peripheral junctional sarcoplasmic reticulum (SR) in adult mammalian ventricular myocardial cells. Immunolocalization of calsequestrin in myocardial cells without transverse tubules (chicken ventricular myocardial cells and sheep Purkinje fibers) clearly showed that calsequestrin as predicted was present in the lumen of peripheral junctional SR but absent from the lumen of the network SR. However, in addition calsequestrin was also present in the lumen of the corbular SR, bulbous ends on the network SR mostly confined to the I band region of myocardial cells.

Anchorfibers and the Topography of Junctional SR

1977 ◽  
Vol 35 ◽  
pp. 582-583
Author(s):  
James Junker ◽  
Joachim R. Sommer
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Cardiac Muscle ◽  
Single Filament ◽  
Relaxation Cycle ◽  
10 Nm Filaments ◽  
Junctional Sarcoplasmic Reticulum

Junctional sarcoplasmic reticulum (JSR) in all its forms (extended JSR, JSR of couplings, corbular SR) in both skeletal and cardiac muscle is always located at the Z - I regions of the sarcomeres. The Z tubule is a tubule of the free SR (non-specialized SR) which is consistently located at the Z lines in cardiac muscle (1). Short connections between JSR and Z lines have been described (2), and bundles of filaments at Z lines have been seen in skeletal (3) and cardiac (4) muscle. In opossum cardiac muscle, we have seen bundles of 10 nm filaments stretching across interfibrillary spaces and adjacent myofibrils with extensions to the plasma- lemma in longitudinal (Fig. 1) and transverse (Fig. 2) sections. Only an occasional single filament is seen elsewhere along a sarcomere. We propose that these filaments represent anchor fibers that maintain the observed invariant topography of the free SR and JSR throughout the contraction-relaxation cycle.

Calcium is released from the junctional sarcoplasmic reticulum during cardiac muscle contraction

1991 ◽  
Vol 260 (3) ◽  
pp. H989-H997 ◽  
Author(s):  
C. S. Moravec ◽  
M. Bond
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Muscle Contraction ◽  
Cardiac Muscle ◽  
Electron Probe Microanalysis ◽  
Electron Probe ◽  
Storage Site ◽  
Papillary Muscles ◽  
Cardiac Muscle Contraction ◽  
Intracellular Storage ◽  
Junctional Sarcoplasmic Reticulum

We have used electron-probe microanalysis (EPMA) to address the question of Ca2+ release by junctional sarcoplasmic reticulum (JSR) as well as Ca2+ regulation by mitochondria (MT) during cardiac muscle contraction. Hamster papillary muscles were rapidly frozen during relaxation or at the peak rate of tension rise (+dT/dt). Total Ca2+ content was measured by EPMA in the JSR, within a MT, over the A band, and in the whole cell, in nine cells per animal (five animals per group). JSR Ca2+ content was found to be significantly lower in muscles frozen at the peak of contraction [7.3 +/- 1.3 (mean +/- SE) mmol Ca2+/kg dry wt] than in those frozen during relaxation (12.5 +/- 1.9 mmol Ca2+/kg dry wt; P less than 0.01), suggesting that Ca2+ is released from this storage site during cardiac muscle contraction. In contrast, MT Ca2+ content did not change significantly during contraction (0.4 +/- 0.1 mmol/kg dry wt) compared with relaxation (0.1 +/- 0.2 mmol/kg dry wt). A third group of muscles was frozen during relaxation after pretreatment with 10(-7) M ryanodine. Ca2+ content of the JSR was significantly decreased (P less than 0.01) in this group of muscles, (6.4 +/- 1.8 mmol/kg dry wt) compared with those frozen during relaxation in the absence of the drug. This suggests that the intracellular storage site with a decreased Ca2+ content in muscles frozen at the peak of contraction is the ryanodine-releasable store. These results provide the first direct measurement of the Ca2+ content of both JSR and MT during a normal cardiac muscle contraction and demonstrate that Ca2+ is released from the JSR during muscle contraction.

scholarly journals SPHEROIDAL BODIES IN THE JUNCTIONAL SARCOPLASMIC RETICULUM OF LIZARD MYOCARDIAL CELLS

1974 ◽  
Vol 60 (3) ◽  
pp. 602-615 ◽  
Author(s):  
M. S. Forbes ◽  
N. Sperelakis
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Anolis Carolinensis ◽  
Myocardial Cells ◽  
En Face ◽  
Structural Details ◽  
Junctional Sarcoplasmic Reticulum

The sarcoplasmic reticulum (SR) of lizard (Anolis carolinensis) myocardial cells has been examined, with particular attention being paid to the structural details of the peripheral couplings (junctional SR). Spheroidal bodies are present within the opaque core of junctional SR; these can be seen both in sections made en face and in sections cut to show the apposition of the junctional SR with the sarcolemma. Opaque junctional processes extend between the sarcolemma and the peripheral junctional SR. The myocardial cells in addition contain some SR cisternae deep within the cells which also possess opaque cores composed of spheroids. Although the significance of the junctional SR spheroidal bodies is unknown, it is thought that they could act as a matrix on which enzymes such as calcium-specific ATPase may be located.

scholarly journals Localization of Ca2+ + Mg2+-ATPase of the sarcoplasmic reticulum in adult rat papillary muscle.

1982 ◽  
Vol 93 (3) ◽  
pp. 883-892 ◽  
Author(s):  
A O Jorgensen ◽  
A C Shen ◽  
P Daly ◽  
D H MacLennan
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Cardiac Muscle ◽  
Papillary Muscle ◽  
Functional Role ◽  
Transverse Tubules ◽  
Contraction Coupling ◽  
Adult Rat ◽  
Sarcoplasmic Reticulum Membrane ◽  
Rat Papillary Muscle ◽  
Junctional Sarcoplasmic Reticulum

Localization of the Ca2+ + Mg2+-ATPase of the sarcoplasmic reticulum in rat papillary muscle was determined by indirect immunofluorescence and immunoferritin labeling of cryostat and ultracryotomy sections, respectively. The Ca2+ + Mg2+-ATPase was found to be rather uniformly distributed in the free sarcoplasmic reticulum membrane but to be absent from both peripheral and interior junctional sarcoplasmic reticulum membrane, transverse tubules, sarcolemma, and mitochondria. This suggests that the Ca2+ + Mg2+-ATPase of the sarcoplasmic reticulum is antigenically unrelated to the Ca2+ + Mg2+-ATPase of the sarcolemma. These results are in agreement with the idea that the sites of interior and peripheral coupling between sarcoplasmic reticulum membrane and transverse tubules and between sarcoplasmic reticulum and sarcolemmal membranes play the same functional role in the excitation-contraction coupling in cardiac muscle.

scholarly journals Ultrastructural Localization of Glycolytic Enzymes on Sarcoplasmic Reticulum Vesicles

1998 ◽  
Vol 46 (4) ◽  
pp. 419-427 ◽  
Author(s):  
Kai Y. Xu ◽  
Lewis C. Becker
Keyword(s):  
Skeletal Muscle ◽  
Sarcoplasmic Reticulum ◽  
Pyruvate Kinase ◽  
Indirect Evidence ◽  
Ultrastructural Localization ◽  
Immunogold Labeling ◽  
Glycolytic Enzymes ◽  
Electron Microscopic ◽  
Gold Particles ◽  
Cytoplasmic Side

We have previously obtained indirect evidence that sarcoplasmic reticulum (SR) vesicles from cardiac and skeletal muscle contain the complete chain of glycolytic enzymes from aldolase to pyruvate kinase. To investigate directly whether pyruvate kinase and other glycolytic enzymes are anatomically associated with the SR, electron microscopic immunogold labeling studies were carried out in isolated SR vesicles using specific primary antibodies against selected glycolytic enzymes and Ca2+-ATPase, and appropriate secondary antibodies labeled with 6-nm or 12-nm gold particles. Pyruvate kinase was broadly dispersed on the cytoplasmic side of the SR membrane of both cardiac and skeletal muscle vesicles. With 6-nm gold particles, the density of binding to pyruvate kinase was 2522 ± 445 and 4171 ± 1379 particles/μm2 for cardiac and skeletal muscle SR, respectively. Binding densities to Ca2+-ATPase were similar (2550 ± 639 particles/μm2 for cardiac SR, 3877 ± 408 particles/μm2 for skeletal muscle SR). Immunogold labeling of ultrathin sections indicated that pyruvate kinase was attached to the SR membrane and located immediately adjacent to the Ca2+-ATPase. Aldolase and glyceraldehyde phosphate dehydrogenase were also found to be attached to the cytoplasmic side of SR vesicles and located in close proximity to Ca2+-ATPase. These results provide the first ultrastructural evidence that glycolytic enzymes are anatomically associated with SR membranes and located near the SR Ca2+-ATPase. The results further support the hypothesis that ATP generated by SR-associated glycolytic enzymes is coupled to SR Ca2+ active transport.

scholarly journals Evidence for the presence of calsequestrin in two structurally different regions of myocardial sarcoplasmic reticulum.

1984 ◽  
Vol 98 (4) ◽  
pp. 1597-1602 ◽  
Author(s):  
A O Jorgensen ◽  
K P Campbell
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Central Region ◽  
Protein A ◽  
Muscle Cells ◽  
Physiological Signals ◽  
Ventricular Muscle ◽  
Cardiac Muscle Cells ◽  
Band Region ◽  
Membrane Bound ◽  
Junctional Sarcoplasmic Reticulum

Localization of calsequestrin in chicken ventricular muscle cells was determined by indirect immunofluorescence and immuno-Protein A-colloidal gold labeling of cryostat and ultracryotomy sections, respectively. Calsequestrin was localized in the lumen of peripheral junctional sarcoplasmic reticulum, as well as in the lumen of membrane-bound structures present in the central region of the I-band, while being absent from the lumen of the sarcoplasmic reticulum in the A-band region of the cardiac muscle cells. Since chicken ventricular muscle cells lack transverse tubules, the presence of calsequestrin in membrane bound structures in the central region of the I-band suggests that these cells contain nonjunctional regions of sarcoplasmic reticulum that are involved in Ca2+ storage and possibly Ca2+ release. It is likely that the calsequestrin containing structures present throughout the I-band region of the muscle cells correspond to specialized regions of the free sarcoplasmic reticulum in the I-band called corbular sarcoplasmic reticulum. It will be of interest to determine whether Ca2+ storage and possibly Ca2+ release from junctional and nonjunctional regions of the sarcoplasmic reticulum in chicken ventricular muscle cells are regulated by the same or different physiological signals.

Effects of rest duration and ryanodine on changes of extracellular [Ca] in cardiac muscle from rabbits

1987 ◽  
Vol 253 (3) ◽  
pp. C398-C407 ◽  
Author(s):  
K. T. MacLeod ◽  
D. M. Bers
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Cardiac Muscle ◽  
Extracellular Space ◽  
Ventricular Muscle ◽  
Rest Interval ◽  
Continuous Stimulation ◽  
Rabbit Ventricular Muscle ◽  
Interpretive Framework ◽  
Rest Intervals

Cumulative depletions of extracellular Ca were measured using double-barreled Ca-sensitive microelectrodes in the extracellular space of rabbit ventricular muscle. Depletions were produced by 1-Hz stimulation after rest intervals of 10 s to 10 min. With longer rest intervals, depletion size increased while the first postrest contraction decreased in a reciprocal manner. The depletions may represent refilling of sarcoplasmic reticulum (SR) Ca stores that have become partially depleted of Ca during the rest. Within this interpretive framework, the longer the rest interval the lower the SR Ca content, so the SR is then capable of taking up larger amounts of Ca. This may be related to the rest decay of tension of the first postrest beat, since this is thought to be SR dependent. Ryanodine (1 microM) increased the size of the depletions after short rest intervals (less than 2 min) but not after longer (greater than or equal to 2 min) intervals. Ryanodine also increased the rate of Ca loss from the cell on cessation of stimulation. This increased rate of Ca loss with ryanodine may deplete the SR of Ca such that more Ca can be taken up during subsequent stimulation than in untreated muscles. Thus cumulative depletions after short rest intervals are enhanced by ryanodine. When a Ca load was produced during 1) quiescence [by removal of extracellular Na (Nao)] or 2) continuous stimulation (in the presence of 3 microM acetylstrophanthidin), addition of ryanodine (5-10 microM) did not produce any apparent Ca loss. Caffeine (10 mM), added after ryanodine, induced contractures accompanied by Ca efflux, implying there was Ca in the SR after ryanodine exposure. The results of other investigators have suggested that ryanodine may inhibit cardiac SR Ca release. The present study suggests that ryanodine also enhances the loss of cellular (and probably SR) Ca on cessation of stimulation but not when applied during continuous stimulation or quiescence.

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