scholarly journals Freeze-fracture study on the erythrocyte membrane during malarial parasite invasion.

1981 ◽  
Vol 91 (1) ◽  
pp. 55-62 ◽  
Author(s):  
M Aikawa ◽  
L H Miller ◽  
J R Rabbege ◽  
N Epstein
Keyword(s):  
Erythrocyte Membrane ◽  
Cytochalasin B ◽  
Freeze Fracture ◽  
Malarial Parasite ◽  
Invasion Process ◽  
Parasite Invasion ◽  
Thin Sections ◽  
Intramembrane Particles ◽  
Fracture Study ◽  
Attachment Process

Invasion of erythrocytes by malarial merozoites requires the formation of a junction between the merozoite and the erythrocyte. Migration of the junction parallel to the long axis of the merozoite occurs during the entry of the merozoite into an invagination of the erythrocyte. Freeze-fracture shows a narrow circumferential band of rhomboidally arrayed particles on the P face of the erythrocyte membrane at the neck of the erythrocyte invagination and matching rhomboidally arrayed pits on the E face. The band corresponds to the junction between the erythrocyte and merozoite membranes observed in thin sections and may represent the anchorage sites of the contractile proteins within the erythrocyte. Intramembrane particles (IMP) on the P face of the erythrocyte membrane disappear beyond this junction. When the erythrocytes and cytochalasin B-treated merozoites are incubated together, the merozoite attaches to the erythrocyte membrane and a junction is formed between the two, but the invasion process does not advance further and no movement of the junction occurs. Although there is no entry of the parasite, the erythrocyte membrane still invaginates. Freeze-fracture shows that the P face of the invaginated erythrocyte membrane is almost devoid of the IMP that are found elsewhere on the membrane, suggesting that the attachment process in and of itself is sufficient to create a relatively IMP-free bilayer.

Myocardial sarcolemma in ischemia: a quantitative freeze-fracture study

1988 ◽  
Vol 255 (3) ◽  
pp. H467-H475 ◽  
Author(s):  
J. S. Frank ◽  
S. Beydler ◽  
N. Wheeler ◽  
K. I. Shine
Keyword(s):  
Electron Microscopy ◽  
Analysis Of Variance ◽  
Freeze Fracture ◽  
Mixed Effects ◽  
Fracture Face ◽  
Intramembrane Particles ◽  
Fracture Study ◽  
K Concentration ◽  
Freeze Fracture Electron Microscopy ◽  
Fracture Electron

Freeze-fracture electron microscopy permits the visualization of the intramembrane particles (IMP). These IMPs are presumably proteins responsible for the main functions of the membrane. Quantitative techniques (Clark-Evan statistics) were applied to determine in a critical manner whether IMP pattern shifts (random, clustered, or ordered) occur under the ischemic conditions (5-45 min with and without reperfusion) and whether this change is related to the experimental condition. In each case three hearts, eight replicas/heart, one area of 0.25 micron 2 of membrane fracture face/replica was measured to give a total of 6 micron 2 of membrane counted for each condition (control vs. ischemic). A mixed effects nested model analysis of variance was performed in each variable. We found that IMP aggregation can be present in some control membranes, but the degree of aggregation was greater and more consistent in membranes made ischemic and followed by reperfusion. Most striking was the significant clustering of IMPs in membranes from hearts ischemic for only 5 min. Reperfusion after only 5 min of ischemia reversed IMP clustering. Functionally at this time there is an increase in K+ concentration in the interstitial space that reaches approximately 15 mM within 10 min and reverses on reperfusion. The structural alteration in IMPs appears to parallel the function in ischemic hearts.

scholarly journals A thin-section and freeze-fracture study of microfilament-membrane attachments in choroid plexus and intestinal microvilli.

1978 ◽  
Vol 79 (3) ◽  
pp. 774-787 ◽  
Author(s):  
N S McNutt
Keyword(s):  
Choroid Plexus ◽  
Thin Section ◽  
Brush Border ◽  
Freeze Fracture ◽  
Similar Type ◽  
Heavy Meromyosin ◽  
Thin Sections ◽  
Intestinal Brush Border ◽  
Fracture Study ◽  
20 Nm

Choroid plexus and intestinal microvilli in thin sections have microfilaments in the cytoplasm adjacent to the membranes, and in replicas have broken strands of filaments in both cytoplasm and on E faces of plasm membranes. The microfilaments contain actin as indicated by their binding of heavy meromyosin (HMM). In sections of choroid plexus, the microfilaments are 7-8 nm in diameter and form a loose meshwork which lies parallel to the membrane and which is connected to the membranes both by short, connecting filaments (8 times 30 nm) and dense globules (approximately 15-20 nm). The filamentous strands seen in replicas are approximately 8 nm in diameter. Because they are similar in diameter and are connected to the membrane, these filamentous strands seen in replicas apparently represent the connecting structures, portions of the microfilaments, or both. The filamentous strands attached to the membrane are usually associated with the E face and appear to be pulled through the P half-membrane. In replicas of intestinal brush border microvilli, the connecting strands attaching core microfilaments to the membrane are readily visualized. In contrast, regions of attachment of core microfilaments to dense material at the tips of microvilli are associated with few particles on P faces and with few filamentous strands on the E faces of the membranes. Freeze-fracture replicas suggest a morphologically similar type of connecting strand attachment for microfilament-membrane binding in both choroid plexus and intestinal microvilli, despite the lack of a prominent core bundle of microfilaments in choroid plexus microvilli.

scholarly journals Low resistance junctions in crayfish. Structural changes with functional uncoupling.

1976 ◽  
Vol 70 (2) ◽  
pp. 419-439 ◽  
Author(s):  
C Peracchia ◽  
A F Dulhunty
Keyword(s):  
Gap Junctions ◽  
Conformational Changes ◽  
Structural Changes ◽  
Freeze Fracture ◽  
Particle Diameter ◽  
Electrical Measurements ◽  
Thin Sections ◽  
Giant Axons ◽  
Intramembrane Particles ◽  
Center Distance

Electrical uncoupling of crayfish septate lateral giant axons is paralleled by structural changes in the gap junctions. The changes are characterized by a tighter aggregation of the intramembrane particles and a decrease in the overall width of the junction and the thickness of the gap. Preliminary measurements indicate also a decrease in particle diameter. The uncoupling is produced by in vitro treatment of crayfish abdominal cords either with a Ca++, Mg++-free solution containing EDTA, followed by return to normal saline (Van Harreveld's solution), or with VAn Harreveld's solution containing dinitrophenol (DNP). The uncoupling is monitored by the intracellular recording of the electrical resistance at a septum between lateral giant axons. The junctions of the same septum are examined in thin sections; those of other ganglia of the same chain used for the electrical measurements are studied by freeze-fracture. In controls, most junctions contain a more or less regular array of particles repeating at a center to center distance of approximately 200 A. The overall width of the junctions is approximately 200 A and the gap thickness is 40-50 A. Vesicles (400-700 A in diameter) are closely apposed to the junctional membranes. In uncoupled axons, most junctions contain a hexagonal array of particles repeating at a center to center distance of 150-155 A. The overall width of the junctions is approximately 180 A and the gap thickness is 20-30 A. These junctions are usually curved and are rarely associated with vesicles. Isolated, PTA-stained junctions, also believed to be uncoupled, display similar structural features. There are reasons to believe that the changes in structure and permeability are triggered by an increase in the intracellular free Ca++ concentration. Most likely, the changes in permeability are caused by conformational changes in some components of the intramembrane particles at the gap junctions.

scholarly journals A freeze-fracture study of membrane events during neurohypophysial secretion.

1978 ◽  
Vol 78 (2) ◽  
pp. 542-553 ◽  
Author(s):  
D T Theodosis ◽  
J J Dreifuss ◽  
L Orci
Keyword(s):  
Plasma Membrane ◽  
Unit Area ◽  
Freeze Fracture ◽  
Core Material ◽  
Number Of Clusters ◽  
Intramembrane Particles ◽  
En Face ◽  
Large Particles ◽  
Mean Diameter ◽  
Fracture Study

Freeze-fracture was used to study the membrane events taking place during neurosecretory granule discharge (exocytosis) and subsequent membrane internalization (endocytosis) in axons of neurohypophyses from control and water-deprived rats. En face views of the cytoplasmic leaflet (P face) of the split axolemma reveal circular depressions that represent the secretory granule membranes fused with the plasma membrane during exocytosis. These depressions often contain granule core material in the process of extrusion into the extracellular space. The membrane surrounding some of the exocytotic openings shows a decreased number of intramembrane particles (mean diameter, 8 nm) which are elsewhere more numerous and evenly distrubuted on the fracture face. Endocytotic sites appear as smaller plasma membrane invaginations, with associated intramembrane particles. Moreover, such invaginations often contain large particles (mean diameter, 12 nm) that appear as clusters on en face views of the membrane leaflet. Quantitative analysis indicates that the number of exocytotic images increases significantly in glands from water-deprived rats. Concomitantly, the number of endocytotic figures per unit area of membrane is raised as is the number of clusters of large particles. The observations demonstrate that, in the neurohypophysis, it is possible to distinguish exocytosis morphologically from endocytosis and that the two events can be assessed quantitatively.

scholarly journals Analysis of the structure of intramembrane particles of the mammalian urinary bladder.

1980 ◽  
Vol 86 (2) ◽  
pp. 514-528 ◽  
Author(s):  
J D Robertson ◽  
J Vergara
Keyword(s):  
Urinary Bladder ◽  
Glass Surface ◽  
Hexagonal Lattice ◽  
Freeze Fracture ◽  
Transitional Epithelium ◽  
Thin Sections ◽  
Intramembrane Particles ◽  
Thin Sectioning ◽  
Space Constant ◽  
Dominant Structure

The luminal and discoid vacuole membranes of the superficial cell layer of the transitional epithelium of the mammalian urinary bladder have been studied by thin-sectioning and freeze-fracture-etch (FFE) electron microscope methods. For the FFE studies membranes were deposited on a cationized glass surface, covered by a thin copper disc, and fractured under liquid N2. Specimens were etched at -100 degrees C and replicated at -190 degrees C. A model of the lattice membrane derived from thin sections was used to predict the heights of the fracture faces above the glass surface. A hexagonal pattern of globular intramembrane particles spaced 160 A apart was seen in the external fracture (EF) face plaques as previously described and regarded as the dominant structure. However, very extensive areas of another pattern, seen before in only limited areas, have beeen found in the EF faces. The pattern consists of a smooth hexagonal lattice with the same space constant as the globular one but a different structure. By image analysis it consists of overlapping domains bordered by shared but incomplete metal rims. Each domain has a central spot of metal encircled by a shadow. The surface of the smooth lattice is partly complementary to the corresponding protoplasmic fracture (PF) face which shows a similar hexagonal lattice with the same space constant. The height of the smooth EF lattice above the glass substrate is the same as the plane of the center of the lipid bilayer predicted by the model. The mean heights of the particles of the globular EF lattice are greater than the total thickness of the membrane as predicted by the model and confirmed by measurements. The globular EF lattice is not complementary and it is concluded that the globular particles do not exist in the native membrane but arise artifactually during the preparatory procedures.

Freeze-fracture study of phagocytosis in Dictyostelium discoideum

1981 ◽  
Vol 51 (1) ◽  
pp. 63-84
Author(s):  
C. Favard-Sereno ◽  
M.A. Ludosky ◽  
A. Ryter
Keyword(s):  
Plasma Membrane ◽  
Dictyostelium Discoideum ◽  
Freeze Fracture ◽  
Thin Sections ◽  
Latex Beads ◽  
Fracture Study ◽  
Lateral Phase Separation ◽  
Membrane Changes ◽  
Freeze Fracture Electron Microscopy ◽  
Fracture Electron

The plasma membrane and its derivative, the phagosome membrane, were studied during and after ingestion of yeast of latex beads in Dictyostelium discoideum. Freeze-fracture electron microscopy, which provides information on the internal architecture of the membranes, and observation of thin sections of cells treated by cytochemical methods were used in parallel. For visualization of membrane sterols in the replicas, the cells were fixed in the presence of digitonin or the antibiotic filipin. No lateral phase separation occurred during yeast engulfment: the intramembranous particles (IMPs), phospholipids and sterols remained distributed at random in the forming phagosome membrane. In contrast architectural modifications of the membrane were observed upon phagosome internalization. Compared to the plasma membrane, the phagosome membrane displayed 2–3 times more IMPs a shift in the IMP size distribution and a higher sterol content. These changes were completed soon after phagosome closure; they were not related either to the nature of the ingested particles (yeast, latex beads) or to the pH in the membrane environment. The membrane changes too place when the phagosomes began to fuse with pre-existing digestive or autophagic vacuoles and lysosomes. Some of the experimental evidence suggests that the restructuring of the membrane may be related to the presence of hydrolases.

scholarly journals Ultrastructure of Na,K-transport vesicles reconstituted with purified renal Na,K-ATPase.

1980 ◽  
Vol 86 (3) ◽  
pp. 746-754 ◽  
Author(s):  
E Skriver ◽  
A B Maunsbach ◽  
P L Jørgensen
Keyword(s):  
Electron Microscopy ◽  
Freeze Fracture ◽  
Cation Transport ◽  
Coupled Transport ◽  
Vesicle Membrane ◽  
Thin Sections ◽  
Intramembrane Particles ◽  
Freeze Fracture Electron Microscopy ◽  
Fracture Electron ◽  
K Transport

To study the size and structure of the Na,K-pump molecule, the ultrastructure of phospholipid vesicles was examined after incorporation of purified Na,K-ATPase which catalyzes active coupled transport of Na+ and K+ in a ratio close to 3Na/2K. The vesicles were analyzed by thin sectioning and freeze-fracture electron microscopy after reconstitution with different ratios of Na,K-ATPase protein to lipid, and the ultrastructural observations were correlated to the cation transport capacity. The purified Na,K-ATPase reconstituted with phospholipids to form a very uniform population of vesicles. Thin sections of preparations fixed with glutaraldehyde and osmium tetroxide showed vesicles limited by a single membrane which in samples stained with tannic acid appeared triple-layered with a thickness of 70 A. Also, freeze-fracture electron microscopy demonstrated uniform vesicles with diameters in the range of 700-1,100 A and an average value close to 900 A. The vesicle diameter was independent of the amount of protein used for reconstitution. Intramembrane particles appeared only in the vesicle membrane after introduction of Na,K-ATPase and the frequency of intramembrane particles was proportional to the amount of Na,K-ATPase protein used in the reconstitution. The particles were evenly distributed on the inner and the outer leaflet of the vesicle membrane. The diameter of the particles was 90 A and similar to our previous values for the diameter of intramembrane particles in the purified Na,K-ATPase. The capacity for active cation transport in the reconstituted vesicles was proportional to the frequency of intramembrane particles over a range of 0.2-16 particles per vesicle. The data therefore show that active coupled Na,K transport can be carried out by units of Na,K-ATPase which appear as single intramembrane particles with diameters close fo 90 A in the freeze-fracture micrographs.

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