scholarly journals Helicoids in the T system and striations of frog skeletal muscle fibers seen by high voltage electron microscopy

1978 ◽  
Vol 22 (2) ◽  
pp. 145-154 ◽  
Author(s):  
L.D. Peachey ◽  
B.R. Eisenberg
Author(s):  
Lee D. Peachey ◽  
Clara Franzini-Armstrong

The effective study of biological tissues in thick slices of embedded material by high voltage electron microscopy (HVEM) requires highly selective staining of those structures to be visualized so that they are not hidden or obscured by other structures in the image. A tilt pair of micrographs with subsequent stereoscopic viewing can be an important aid in three-dimensional visualization of these images, once an appropriate stain has been found. The peroxidase reaction has been used for this purpose in visualizing the T-system (transverse tubular system) of frog skeletal muscle by HVEM (1). We have found infiltration with lanthanum hydroxide to be particularly useful for three-dimensional visualization of certain aspects of the structure of the T- system in skeletal muscles of the frog. Specifically, lanthanum more completely fills the lumen of the tubules and is denser than the peroxidase reaction product.


Author(s):  
Brenda R. Eisenberg ◽  
Lee D. Peachey

Analysis of the electrical properties of the t-system requires knowledge of the geometry of the t-system network. It is now possible to determine the network parameters experimentally by use of high voltage electron microscopy. The t-system was marked with exogenous peroxidase. Conventional methods of electron microscopy were used to fix and embed the sartorius muscle from four frogs. Transverse slices 0.5-1.0 μm thick were viewed at an accelerating voltage of 1000 kV using the JEM-1000 high voltage electron microscope at Boulder, Colorado and prints at x5000 were used for analysis.The length of a t-branch (t) from node to node (Fig. 1a) was measured with a magnifier; at least 150 t-branches around 30 myofibrils were measured from each frog. The mean length of t is 0.90 ± 0.11 μm and the number of branches per myofibril is 5.4 ± 0.2 (mean ± SD, n = 4 frogs).


Author(s):  
Craig H. Bailey ◽  
Lee D. Peachey

Our present understanding of the distribution and morphology of the sarcoplasmic reticulum (SR) in frog slow and twitch muscle fibers has been derived largely from the examination of thin sections by electron microscopy. This conventional approach to the study of an organelle as complex as the SR is limited to a degree by section thickness, and the extraction of three-dimensional information must usually be gathered from an extensive collection of two-dimensional images. The present study represents an alternative approach to the problem of investigating the three-dimensional organization of the SR by utilizing high voltage electron microscopy (HVEM) and examining stereoscopic images of selectively stained 1.0 /μm thick slices of muscle tissue.Slow and twitch fibers from the distal fiber bundles of the frog (Rana pipiens) cruralis muscle were processed for electron microscopy according to the selective SR staining technique (DAB-H2O2 and Os-ferrocyanide) developed by Waugh. Tissue slices from 0.25 to 1.0 μm in thickness were cut on a diamond knife, mounted on grids either with or without plastic support films, and examined using the JEM-1000 microscope at the University of Colorado operating at an accelerating voltage of 1000 kV.


Author(s):  
L. D. Ackerman ◽  
S. H. Y. Wei

Mature human dental enamel has presented investigators with several difficulties in ultramicrotomy of specimens for electron microscopy due to its high degree of mineralization. This study explores the possibility of combining ion-milling and high voltage electron microscopy as a means of circumventing the problems of ultramicrotomy.A longitudinal section of an extracted human third molar was ground to a thickness of about 30 um and polarized light micrographs were taken. The specimen was attached to a single hole grid and thinned by argon-ion bombardment at 15° incidence while rotating at 15 rpm. The beam current in each of two guns was 50 μA with an accelerating voltage of 4 kV. A 20 nm carbon coating was evaporated onto the specimen to prevent an electron charge from building up during electron microscopy.


Author(s):  
T. Mukai ◽  
T. E. Mitchell

Radiation-induced homogeneous precipitation in Ni-Be alloys was recently observed by high voltage electron microscopy. A coupling of interstitial flux with solute Be atoms is responsible for the precipitation. The present investigation further shows that precipitation is also induced at thin foil surfaces by electron irradiation under a high vacuum.


Author(s):  
G. E. Tyson ◽  
M. J. Song

Natural populations of the brine shrimp, Artemia, may possess spirochete- infected animals in low numbers. The ultrastructure of Artemia's spirochete has been described by conventional transmission electron microscopy. In infected shrimp, spirochetal cells were abundant in the blood and also occurred intra- and extracellularly in the three organs examined, i.e. the maxillary gland (segmental excretory organ), the integument, and certain muscles The efferent-tubule region of the maxillary gland possessed a distinctive lesion comprised of a group of spirochetes, together with numerous small vesicles, situated in a cave-like indentation of the base of the tubule epithelium. in some instances the basal lamina at a lesion site was clearly discontinuous. High-voltage electron microscopy has now been used to study lesions of the efferent tubule, with the aim of understanding better their three-dimensional structure.Tissue from one maxillary gland of an infected, adult, female brine shrimp was used for HVEM study.


Author(s):  
William H. Massover

Stereoscopic examination of thick sections of fixed and embedded biological tissues by high voltage electron microscopy has been shown to allow direct visualization of three-dimensional fine structure. The present report will consider the occurrence of some new technical problems in specimen preparation and Image interpretation that are not common during lower voltage studies of thin sections.Thick Sectioning and Tissue Coloration - Epon sections of 0.5 μm or more that are cut with glass knives do not have a uniform thickness as Judged by their interference colors; these colors change with time during their flotation on the knife bath, and again when drying onto the specimen support. Quoted thicknesses thus must be considered only as rough estimates unless measured in specific regions by other methods. Chloroform vapors do not always result in good spreading of thick sections; however, they will spread spontaneously to large degrees after resting on the flotation bath for several minutes. Ribbons of thick sections have been almost impossible to obtain.


Author(s):  
Benjamin M. Siegel

The potential advantages of high voltage electron microscopy for extending the limits of resolution and contrast in imaging low contrast objects, such as biomolecular specimens, is very great. The results of computations will be presented showing that at accelerating voltages of 500-1000 kV it should be possible to achieve spacial resolutions of 1 to 1.5 Å and using phase contrast imaging achieve adequate image contrast to observe single atoms of low atomic number.The practical problems associated with the design and utilization of the high voltage instrument are, optimistically, within the range of competence of the state of the art. However, there are some extremely important and critical areas to be systematically investigated before we have achieved this competence. The basic electron optics of the column required is well understood, but before the full potential of an instrument capable of resolutions of better than 1.5 Å are realized some very careful development work will be required. Of great importance for the actual achievement of high resolution with a high voltage electron microscope is the fundamental limitation set by the characteristics of the high voltage electron beam that can be obtained from the accelerator column.


Author(s):  
J.N. Turner ◽  
W.G. Shain ◽  
V. Madelian ◽  
R.A. Grassucci ◽  
D.L. Forman

Homogeneous cultures of astroglial cells have proved useful for studying biochemical, pharmacological, and toxicological responses of astrocytes to effectors of central nervous system function. LRM 55 astroglial cells, which were derived from a rat glioma and maintained in continuous culture, exhibit a number of astrocyte properties (1-3). Stimulation of LRM 55s and astrocytes in primary cell cultures with the beta-adrenergic agonist isoproterenol results in rapid changes of morphology. Studies with time lapse video light microscopy (VLM) and high-voltage electron microscopy (HVEM) have been correlated to changes in intracellular levels of c-AMP. This report emphasizes the HVEM results.


Author(s):  
Conly L. Rieder ◽  
Frederick J. Miller ◽  
Edwin Davison ◽  
Samuel S. Bowser ◽  
Kirsten Lewis ◽  
...  

In this abstract we Illustrate how same-section correlative light and high voltage electron microscopy (HVEM) of serial 0.25-0.50-μm sections can answer questions which are difficult to approach by EM of 60-100 nm sections.Starfish (Pisaster and Asterlas) eggs are fertilized at meiosis I when the oocyte contains two maternal centrosomes (e.g., asters) which form the poles of the first meiotic spindle. Immediately after fertilization a sperm aster is assembled in the vicinity of the male pronucleus and persists throughout meiosis. At syngamy the sperm aster splits to form the poles of the first mitotic spindle. During this time the functional and replicative properties of the maternal centrosome, inherited from the last meiotic division, are lost. The basis for this differential stability, of male and female centrosomes in the same cytoplasm, is a mystery.


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