scholarly journals QUANTITATIVE ASSAY OF ESTERASES IN END PLATES OF MOUSE DIAPHRAGM BY ELECTRON MICROSCOPE AUTORADIOGRAPHY

1972 ◽  
Vol 20 (12) ◽  
pp. 1059-1068 ◽  
Author(s):  
MIRIAM M. SALPETER ◽  
HELMUT PLATTNER ◽  
ANDREW W. ROGERS
Keyword(s):  
Electron Microscope ◽  
Predominantly White ◽  
Synaptic Cleft ◽  
Postsynaptic Membrane ◽  
Electron Microscope Autoradiography ◽  
Diisopropyl Fluorophosphate ◽  
Mouse Diaphragm ◽  
Microscope Autoradiography ◽  
Alternative Hypotheses ◽  
Motor End Plates

The inhibitor diisopropyl fluorophosphate (DFP) reacts covalently with a number of enzymes including acetylcholinesterase (AChE). The compound pyridine-2-aldoxime methiodide reactivates phosphorylated AChE faster than the other DFP-sensitive sites. Mouse diaphragm was incubated with 3H-DFP, and the radioactivity in the motor end plates was assessed by electron microscope autoradiography. Both the sites which are rapidly reactivated and those which are slowly reactivated by pyridine-2-aldoxime methiodide were evaluated. The distribution of silver grains was compatible with several alternative hypotheses, i.e., that both AChE and other DFP-sensitive enzymes are located on or around the postsynaptic membrane alone, are distributed uniformly throughout the synaptic cleft and are associated equally with both pre- and postsynaptic membranes. This distribution of developed grains was identical with that found in the larger end plates of mouse sternomastoid. The values for the number of molecules of 3H-DFP, whether expressed in terms of square micra of junctional membrane or of cubic micra of synaptic cleft, were also practically identical with those found for sternomastoid. These similarities between the diaphragm, a red muscle, and the sternomastoid, which is predominantly white, suggest that there may be a constant relationship between the sites that bind DFP and the unit dimensions of the subneural compartments concerned.

Quantitation of Fluorophosphate - Reactive Esterase Sites in Rat Liver Using Electron Microscope Autoradiography

1975 ◽  
Vol 33 ◽  
pp. 466-467
Author(s):  
G. C. Budd
Keyword(s):  
Electron Microscope ◽  
Rat Liver ◽  
Smooth Endoplasmic Reticulum ◽  
Average Concentration ◽  
Electron Microscope Autoradiography ◽  
Cytoplasmic Granules ◽  
Diisopropyl Fluorophosphate ◽  
Microscope Autoradiography ◽  
Cytoplasmic Structures ◽  
Silver Grains

Following the application of 10-4 molar tritium labeled diisopropyl fluorophosphate (3H-DFP) to fresh or glutaraldehyde fixed rat liver, fluorophosphate-reactive (FPR) sites within the hepatocytes were measured using quantitative electron microscope autoradiography (EMARG). Based on the distribution of autoradiographic silver grains, most of the FPR sites were concentrated in the rough and smooth endoplasmic reticulum and associated ground cytoplasm. Cytoplasmic granules, comprising autophagic and residual granules also contained FPR sites. The average concentration of sites within each of these cytoplasmic structures was obtained from combined morphometric and grain density analyses of the EMARGs and light microscope sections of the same epoxy embedded liver specimens.

scholarly journals Acetylcholinesterase in the fast extraocular muscle of the mouse by light and electron microscope autoradiography.

1978 ◽  
Vol 78 (1) ◽  
pp. 274-285 ◽  
Author(s):  
M M Salpeter ◽  
A W Rogers ◽  
H Kasprzak ◽  
F A McHenry
Keyword(s):  
Electron Microscope ◽  
Surface Area ◽  
Binding Sites ◽  
Extraocular Muscle ◽  
Extraocular Muscles ◽  
Axonal Membrane ◽  
Electron Microscope Autoradiography ◽  
Diisopropyl Fluorophosphate ◽  
Speed Of Response ◽  
Microscope Autoradiography

The distribution of acetylcholinesterase (ACHe) in the twitch fibers of the extraocular muscles of the mouse was examined by light and electron microscope autoradiography after labeling with radioactive diisopropyl fluorophosphate (DFP) with, and without, 2-pyridine aldoxime methiodide (2-PAM) reactivation. The values obtained were compared with those previously reported for the diaphragm and sternomastoid muscles. The extraocular muscles were studied because they differ from the other two muscles in that they are among the fastest of the mammalian muscles, yet their endplates have sparse junctional folds. They could thus provide information on the extent to which ACHe concentration is an invariant feature of endplate morphology and what, if any aspects may be related to their fast speed of response. We found, using light microscope autoradiography, that in the twitch fibers of the extraocular muscle, there is n average of 6.4 +/- 2.1 X 10(7) DFP-binding sites per endplate, of which 29% (1.8 X 10(7)) are reactivated by 2-PAM and are thus AChe. The morphology of the extraocular endplates allowed us to conclude, on statistical grounds, that the AChe site are probably localized not only along the surface area of the postjunctional membrane (PJM) but also along the surface of the presynaptic axonal membrane. Based on this localization, we calculate 7,800 DFP sites and 2,500 2-PAM-reactivated sites/micron 2 of surface area of pre-and postjunctional membrane. This stacking density of DFP-binding sites per surface area of membrane ( probably in the overlying sheets of basal lamina) is very similar to that in the diaphragm and sternomastoid muscles.

Incorporation of Cholesterol into Peripheral Nerve Myelin

1972 ◽  
Vol 30 ◽  
pp. 334-335
Author(s):  
Frank A. Rawlins
Keyword(s):  
Electron Microscope ◽  
Sciatic Nerve ◽  
Electron Microscope Autoradiography ◽  
Myelin Sheaths ◽  
Microscope Autoradiography ◽  
Grain Distribution ◽  
Sciatic Nerves ◽  
Autoradiographic Analysis ◽  
Old Mice ◽  
Short Times

Several speculations exist as to the site of incorporation of preformed molecules into myelin. The possibility that an autoradiographic analysis of cholesterol-1,2-H3 incorporation at very short times after injection might shed some light in the solution of that problem led to the present experiment.Cholesterol-1,2-H3 was injected intraperitoneally into 24 tenday old mice. The animals were then sacrificed at 10,20,30,40,60,90,120 and 180 min after the injection and the sciatic nerves were processed for electron microscope autoradiography. To analyze the grain distribution in the autoradiograms of cross and longitudinal sections from each sciatic nerve myelin sheaths were subdivided into three compartments named: outer 1/3, middle 1/3 and inner 1/3 compartments.It was found that twenty min. after the injection of cholesterol -1.2-H3 (Figs. 1 and 2), 55% of the total number of grains (t.n.g) found in myelin were within the outer 1/3 compartment, 9% were within the middle 1/3 and 36% within the inner 1/3 compartment

Synthesis and Localization of Sulphated Mucopolysaccharide in Megakaryocytes and Platelets of the Rat, An Analysis by Electron-Microscope Autoradiography

1972 ◽  
Vol 10 (3) ◽  
pp. 705-717
Author(s):  
G. G. MacPHERSON
Keyword(s):  
Electron Microscope ◽  
Golgi Apparatus ◽  
Blood Platelets ◽  
Total Activity ◽  
Electron Microscope Autoradiography ◽  
Dense Granules ◽  
Level Of Activity ◽  
Microscope Autoradiography ◽  
Almost All

Electron-microscope autoradiography has been used to investigate the synthesis and localization of sulphated mucopolysaccharide in megakaryocytes and blood platelets. Following 10-min incubation of bone marrow with 35S-sulpahte in vitro the majority of the activity in megakaryocytes was associated with the Golgi apparatus, but a substantial proportion was associated with other cytoplasmic organelles, suggesting either rapid transport or sulphation of mucopolysaccharide outside the Golgi apparatus. Three hours after the intravenous injection of 35SO4 only a small proportion of the total activity was associated with the Golgi apparatus, most being associated with demarcation membranes and dense granules, while 12 h after injection almost all the activity was associated with demarcation membranes and granules. A rising proportion of activity localized solely on the demarcation membranes suggested that they may possess some activity of their own. Autoradiographs of blood platelets prepared 72 h after the injection of 35SO4 were analysed. It was shown that most of the activity was associated with the α-granules, but there was strong evidence that the platelet membrane possessed a low level of activity.

Timing of nucleolar DNA replication in Amoeba proteus

1976 ◽  
Vol 22 (3) ◽  
pp. 521-530
Author(s):  
I. Minassian ◽  
L.G. Bell
Keyword(s):  
Electron Microscope ◽  
Dna Replication ◽  
Dna Synthesis ◽  
Central Region ◽  
Thymidine Incorporation ◽  
Tritiated Thymidine ◽  
Amoeba Proteus ◽  
Electron Microscope Autoradiography ◽  
Microscope Autoradiography ◽  
G2 Phase

Light- and electron-microscope autoradiography have been used to follow the incorporation of [3H]thymidine at different stages during the interphase of synchronously growing populations of Amoeba proteus. Two main patterns were found for tritiated thymidine incorporation, i.e. DNA synthesis. The major incorporation was in the central region of the nucleus, but a lesser degree of incorporation occurred in the nucleolar region. The bulk of this nucleolar DNA was found to be late replicating, i.e. it replicated during the G2 phase.

Studies on the Synthesis and Composition of Extracellular Mucilage in the Unicellular Red Alga Rhodella

1974 ◽  
Vol 16 (1) ◽  
pp. 1-21
Author(s):  
L. V. EVANS ◽  
MAUREEN E. CALLOW ◽  
ELIZABETH PERCIVAL ◽  
V. FAREED
Keyword(s):  
Electron Microscope ◽  
Uronic Acid ◽  
Red Alga ◽  
Electron Microscope Autoradiography ◽  
Golgi Bodies ◽  
Microscope Autoradiography ◽  
Reducing Substances

35SO42- has been used to investigate the production of extracellular mucilage by log-phase cells. Uptake of isotope occurs most rapidly in the light, when cells are actively dividing. The mucilage comprises about 50% carbohydrate, 16% protein and 10% sulphate. The major sugar is xylose; uronic acid, a small amount of galactose, glucose (trace) and 2 reducing substances are also present. Methylation studies have established the major linkages. Electron-microscope autoradiography shows that the mucilage is packaged in the Golgi bodies, passing to the plasmalemma in large vesicles. Sulphation of the mucilage occurs in the Golgi cisternae.

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