Novel Structure of Skeletal Muscle Myosin’s Intermediate and Its Role in Crossbridge-Cycle

Two month, eight month and two year old rats were treated with 10 or 20 mg/kg of E. Coli endotoxin I. P. The eight month old rats proved most resistant to the endotoxin. During fixation the aorta, carotid artery, basil arartery of the brain, coronary vessels of the heart, inner surfaces of the heart chambers, heart and skeletal muscle, lung, liver, kidney, spleen, brain, retina, trachae, intestine, salivary gland, adrenal gland and gingiva were treated with ruthenium red or alcian blue to preserve the mucopolysaccharide (MPS) coating. Five, 8 and 24 hrs of endotoxin treatment produced increasingly marked capillary damage, disappearance of the MPS coating, edema, destruction of endothelial cells and damage to the basement membrane in the liver, kidney and lung.

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Ultra-Rapid Freezing of Isolated Skeletal Muscle Fibers

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100080316 ◽

1977 ◽

Vol 35 ◽

pp. 576-577

Author(s):

Joachim R. Sommer ◽

Nancy R. Wallace

Keyword(s):

Skeletal Muscle ◽

Sarcoplasmic Reticulum ◽

Granular Material ◽

Nuclear Envelope ◽

Intracellular Calcium ◽

Ruthenium Red ◽

Threshold Concentration ◽

Rapid Freezing ◽

Frog Skeletal Muscle ◽

Electrical Isolation

After Howell (1) had shown that ruthenium red treatment of fixed frog skeletal muscle caused collapse of the intermediate cisternae of the sarcoplasmic reticulum (SR), forming a pentalaminate structure by obi iterating the SR lumen, we demonstrated that the phenomenon involves the entire SR including the nuclear envelope and that it also occurs after treatment with other cations, including calcium (2,3,4).From these observations we have formulated a hypothesis which states that intracellular calcium taken up by the SR at the end of contraction causes the M rete to collapse at a certain threshold concentration as the first step in a subsequent centrifugal zippering of the free SR toward the junctional SR (JSR). This would cause a) bulk transport of SR contents, such as calcium and granular material (4) into the JSR and, b) electrical isolation of the free SR from the JSR.

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Electron Probe Analysis of Muscle

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100054583 ◽

1982 ◽

Vol 40 ◽

pp. 398-401

Author(s):

A. V. Somlyo ◽

H. Shuman ◽

A. P. Somlyo

Keyword(s):

Skeletal Muscle ◽

Smooth Muscle ◽

Sarcoplasmic Reticulum ◽

Resting State ◽

Binding Sites ◽

Electron Probe ◽

Frog Skeletal Muscle ◽

Electron Probe Analysis ◽

Probe Analysis

Electron probe analysis of frozen dried cryosections of frog skeletal muscle, rabbit vascular smooth muscle and of isolated, hyperpermeab1 e rabbit cardiac myocytes has been used to determine the composition of the cytoplasm and organelles in the resting state as well as during contraction. The concentration of elements within the organelles reflects the permeabilities of the organelle membranes to the cytoplasmic ions as well as binding sites. The measurements of [Ca] in the sarcoplasmic reticulum (SR) and mitochondria at rest and during contraction, have direct bearing on their role as release and/or storage sites for Ca in situ.

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How to sample the entire length of a single muscle fiber quick-frozen after electrical point stimulation for high resolution EM

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100124288 ◽

1992 ◽

Vol 50 (1) ◽

pp. 776-777

Author(s):

Joachim R. Sommer ◽

Teresa High ◽

Betty Scherer ◽

Isaiah Taylor ◽

Rashid Nassar

Keyword(s):

Skeletal Muscle ◽

High Resolution ◽

Action Potential ◽

Muscle Fiber ◽

Freeze Fracture ◽

Freeze Substitution ◽

Electrical Field Stimulation ◽

Skeletal Muscle Fiber ◽

Freeze Dried ◽

Quick Freezing

We have developed a model that allows the quick-freezing at known time intervals following electrical field stimulation of a single, intact frog skeletal muscle fiber isolated by sharp dissection. The preparation is used for studying high resolution morphology by freeze-substitution and freeze-fracture and for electron probe x-ray microanlysis of sudden calcium displacement from intracellular stores in freeze-dried cryosections, all in the same fiber. We now show the feasibility and instrumentation of new methodology for stimulating a single, intact skeletal muscle fiber at a point resulting in the propagation of an action potential, followed by quick-freezing with sub-millisecond temporal resolution after electrical stimulation, followed by multiple sampling of the frozen muscle fiber for freeze-substitution, freeze-fracture (not shown) and cryosectionmg. This model, at once serving as its own control and obviating consideration of variances between different fibers, frogs etc., is useful to investigate structural and topochemical alterations occurring in the wake of an action potential.

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Alterations in Ultrastructure of Skeletal Muscle From Newborn Guinea Pigs Following in Utero Exposure to Ethanol

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100120126 ◽

1985 ◽

Vol 43 ◽

pp. 686-687

Author(s):

C. Uphoff ◽

C. Nyquist-Battie ◽

T.B. Cole

Keyword(s):

Skeletal Muscle ◽

Guinea Pigs ◽

Muscle Development ◽

In Utero ◽

Ethanol Exposure ◽

Prenatally Exposed ◽

Chronic Alcoholic ◽

Test Kit ◽

Food And Water Intake ◽

Post Intubation

Ultrastructural alterations of skeletal muscle have been observed in adult chronic alcoholic patients. However, no such study has been performed on individuals prenatally exposed to ethanol. In order to determine if ethanol exposure in utero in the latter stages of muscle development was deleterious, skeletal muscle was obtained from newborn guinea pigs treated in the following manner. Six Hartly strain pregnant guinea pigs were randomly assigned to either the ethanol or the pair-intubated groups. Twice daily the 3 ethanol-treated animals were intubated with Ensure (Ross Laboratories) liquid diet containing 30% ethanol (6g/Kg pre-pregnant body weight per day) from day 35 of gestation until parturition at day 70±1 day. Serum ethanol levels were determined at 1 hour post-intubation by the Sigma alcohol test kit. For pair-intubation the Ensure diet contained sucrose substituted isocalorically for ethanol. Both food and water intake were monitored.

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Ultrastructural autoradiography of L6 skeletal-muscle cells exposed to a tritiated macrolide antibiotic (LY237216) in vitro

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100123465 ◽

1992 ◽

Vol 50 (1) ◽

pp. 612-613

Author(s):

S.L. White ◽

C.B. Jensen ◽

D.D. Giera ◽

D.A. Laska ◽

M.N. Novilla ◽

...

Keyword(s):

Skeletal Muscle ◽

Quantitative Analysis ◽

Macrolide Antibiotic ◽

Circle Method ◽

Apical Surface ◽

Polycarbonate Membrane ◽

L6 Cells ◽

Low Viscosity ◽

Erythromycin A

In vitro exposure to LY237216 (9-Deoxo-11-deoxy-9,11-{imino[2-(2-methoxyethoxy)ethylidene]-oxy}-(9S)-erythromycin), a macrolide antibiotic, was found to induce cytoplasmic vacuolation in L6 skeletal muscle myoblast cultures (White, S.L., unpubl). The present study was done to determine, by autoradiographic quantitative analysis, the subcellular distribution of 3H-LY237216 in L6 cells.L6 cells (ATCC, CRL 1458) were cultured to confluency on polycarbonate membrane filters (Millipore Corp., Bedford, MA) in M-199 medium (GIBCO® Labs) with 10% fetal bovine serum. The cells were exposed from the apical surface for 1-hour to unlabelled-compound (0 μCi/ml) or 50 (μCi/ml of 3H-LY237216 at a compound concentration of 0.25 mg/ml. Following a rapid rinse in compound-free growth medium, the cells were slam-frozen against a liquid nitrogen cooled, polished copper block in a CF-100 cryofixation unit (LifeCell Corp., The Woodlands, TX). Specimens were dried in the MDD-C Molecular Distillation Drier (LifeCell Corp.), vapor osmicated and embedded in Spurrs low viscosity resin. Ultrathin sections collected on formvar coated stainless steel grids were counter-stained, then individually mounted on corks. A monolayer of Ilford L4 nuclear emulsion (Polysciences, Inc., Warrington, PA) was placed on the sections, utilizing a modified “loop method”. The emulsions were exposed for 7-weeks in a light-tight box at 4°C. Autoradiographs were developed in Microdol-X developer and examined on a Philips EM410LS transmission electron microscope. Quantitative analysis of compound localization employed the point and circle approach of Williams; incorporating the probability circle method of Salpeter and McHenry.

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The “KIMWIPE” Effect in Ultra-Thin Cryosections

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100119119 ◽

1985 ◽

Vol 43 ◽

pp. 458-459

Author(s):

I. Taylor ◽

P. Ingram ◽

J.R. Sommer

Keyword(s):

Skeletal Muscle ◽

Electrical Stimulation ◽

Muscle Fibers ◽

Ice Crystals ◽

Crystal Formation ◽

Ice Crystal ◽

Thin Sections ◽

Skeletal Muscle Fibers ◽

Freeze Dried ◽

Ice Crystal Formation

In studying quick-frozen single intact skeletal muscle fibers for structural and microchemical alterations that occur milliseconds, and fractions thereof, after electrical stimulation, we have developed a method to compare, directly, ice crystal formation in freeze-substituted thin sections adjacent to all, and beneath the last, freeze-dried cryosections. We have observed images in the cryosections that to our knowledge have not been published heretofore (Figs.1-4). The main features are that isolated, sometimes large regions of the sections appear hazy and have much less contrast than adjacent regions. Sometimes within the hazy regions there are smaller areas that appear crinkled and have much more contrast. We have also observed that while the hazy areas remain still, the regions of higher contrast visibly contract in the beam, often causing tears in the sections that are clearly not caused by ice crystals (Fig.3, arrows).

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Two structural states of the Z-band in cardiac and skeletal muscle

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100157097 ◽

1989 ◽

Vol 47 ◽

pp. 1022-1023

Author(s):

J.P. Schroeter ◽

M.A. Goldstein ◽

J.P. Bretaudiere ◽

L.H. Michael ◽

R.L. Sass

Keyword(s):

Skeletal Muscle ◽

Cross Section ◽

Real Space ◽

Contractile State ◽

Contour Maps ◽

False Color ◽

Grey Scale ◽

Fourier Filtering ◽

Cardiac Muscles ◽

Enhancement Algorithm

We have recently established the existence of two structural states of the Z band lattice in cross section in cardiac as well as in skeletal muscle. The two structural states are related to the contractile state of the muscle. In skeletal muscle at rest, the Z band is in the small square (ss) lattice form, but tetanized muscle exhibits the basket weave (bw) form. In contrast, unstimu- lated cardiac muscle exhibits the bw form, but cardiac muscles exposed to EGTA show the ss form.We have used two-dimensional computer enhancement techniques on digitized electron micrographs to compare each lattice form as it appears in both cardiac and skeletal muscle. Both real space averaging and fourier filtering methods were used. Enhanced images were displayed as grey-scale projections, as contour maps, and in false color.There is only a slight difference between the lattices produced by the two different enhancement techniques. Thus the information presented in these images is not likely to be an artifact of the enhancement algorithm.

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