scholarly journals MEMBRANE DIFFERENTIATIONS IN FREEZE-FRACTURED MAMMALIAN SPERM

1974 ◽  
Vol 63 (2) ◽  
pp. 641-664 ◽  
Author(s):  
Daniel S. Friend ◽  
Don W. Fawcett
Keyword(s):  
Guinea Pig ◽  
Nuclear Envelope ◽  
Central Area ◽  
Ruthenium Red ◽  
Double Row ◽  
Thin Sections ◽  
Freeze Fracturing ◽  
Mammalian Sperm ◽  
Acrosomal Membrane ◽  
Principal Piece

A correlated thin-sectioning and freeze-fracturing study has been made of guinea pig and rat spermatozoa. In sections, the cell membrane over the acrosome has a concanavalin A and ruthenium red reactive glycocalyx which exhibits an ordered pattern related to the lattice of crystalline domains within the plane of the membrane revealed by freeze-fracturing. The cleaved acrosomal membrane also shows a finer linear periodicity in some areas. The membrane over the equatorial segment of the guinea pig acrosome is marked by a palisade of oblique ridges not observed in the rat. The plasmalemma of the postacrosomal region is rich in membrane intercalated particles, many randomly dispersed, others clustered in rectilinear arrays. A particle-poor zone is found just anterior to the posterior ring. The fold of redundant nuclear envelope posterior to the ring has many nuclear pores in close hexagonal array. The nuclear envelope lining the implantation fossa is devoid of pores. When cleaved it has a particle-free central area surrounded by a broad zone of large, closely packed, hollow particles. The membrane of the mid-piece in the guinea pig (but not the rat) contains linear strands of 6–8-nm particles oriented circumferentially. The membrane investing the principal piece exhibits the usual randomly distributed particles but in addition, a double row of larger (9 nm) particles runs longitudinally within the membrane over outer dense fiber 1. In the corresponding position in thin sections a local thickening of the membrane is discernible. These observations form a basis for further studies on the functional correlates of these regional specializations of the sperm membrane.

scholarly journals ACROSOMAL DISRUPTION IN SPERM

1974 ◽  
Vol 63 (2) ◽  
pp. 466-479 ◽  
Author(s):  
Daniel S. Friend ◽  
Irene Rudolf
Keyword(s):  
Plasma Membrane ◽  
Guinea Pig ◽  
Plasma Membranes ◽  
Freeze Fracture ◽  
Direct Consequence ◽  
Thin Sections ◽  
Geometric Patterns ◽  
Acrosomal Membrane ◽  
Principal Piece

"Capacitation" is a physiological event which alters sperm to permit rapid penetration through oocyte investments and fusion between gametes. Acrosomal "reaction," the physiological release of acrosomal contents, occurs after this facilitating process. In this study, acrosomal "disruption" of guinea pig and rat sperm was achieved in vitro by incubating sperm together with the follicular contents of superovulated mice. The samples contained both "reacted" and "disrupted" sperm. Thin sections of affected sperm revealed rupture and vesiculation of the plasma membrane overlying the acrosome, as well as loss of both the outer acrosomal membrane and the acrosomal content. Freeze-fracture revealed disintegration of the characteristic geometric patterns in regions of the acrosomal and plasma membranes thus disrupted and major modifications in particle distribution in the sperm tail. In the guinea pig, strands of 6–8-nm particles, usually confined to the plasma membrane of the midpiece, which overlies mitochondria, also appeared in the principal piece. Likewise, in rat sperm, bands of similarly small particles formed acute angles throughout the membrane of the principal piece. Compared with the membranes of control preparations, these membrane alterations are apparently a direct consequence of incubation with ovarian follicular contents.

scholarly journals Lateral diffusion of the PH-20 protein on guinea pig sperm: evidence that barriers to diffusion maintain plasma membrane domains in mammalian sperm

1987 ◽  
Vol 104 (4) ◽  
pp. 917-923 ◽  
Author(s):  
AE Cowan ◽  
DG Myles ◽  
DE Koppel
Keyword(s):  
Plasma Membrane ◽  
Guinea Pig ◽  
Domain Boundary ◽  
Lateral Diffusion ◽  
Fold Increase ◽  
Membrane Domains ◽  
Mammalian Sperm ◽  
Acrosomal Membrane ◽  
Percent Recovery ◽  
Lipid Probe

PH-20 protein on the plasma membrane (PH-20PM) is restricted to the posterior head of acrosome-intact guinea pig sperm. During the exocytotic acrosome reaction the inner acrosomal membrane (IAM) becomes continuous with the posterior head plasma membrane, and PH-20PM migrates to the IAM. There it joins a second population of PH-20 protein localized to this region of the acrosomal membrane (PH-20AM) (Cowan, A.E., P. Primakoff, and D.G. Myles, 1986, J. Cell Biol. 103:1289-1297). To investigate how the localized distributions of PH-20 protein are maintained, the lateral mobility of PH-20 protein on these different membrane domains was determined using fluorescence redistribution after photobleaching. PH-20PM on the posterior head of acrosome-intact sperm was found to be mobile, with a diffusion coefficient and percent recovery typical of integral membrane proteins (D = 1.8 X 10(-10) cm2/s; %R = 73). This value of D was some 50-fold lower than that found for the lipid probe 1,1-ditetradecyl 3,3,3',3'-tetramethylindocarbocyanine perchlorate (C14diI) in the same region (D = 8.9 X 10(-9) cm2/s). After migration to the IAM of acrosome-reacted sperm, this same population of molecules (PH-20PM) exhibited a 30-fold increase in diffusion rate (D = 4.9 X 10(-9) cm2/s; %R = 78). This rate was similar to diffusion of the lipid probe C14diI in the IAM (D = 5.4 X 10(-9) cm2/s). The finding of free diffusion of PH-20PM in the IAM of acrosome-reacted sperm supports the proposal that PH-20 is maintained within the IAM by a barrier to diffusion at the domain boundary. The slower diffusion of PH-20PM on the posterior head of acrosome-intact sperm is also consistent with localization by barriers to diffusion, but does not rule out alternative mechanisms.

Ultra-Rapid Freezing of Isolated Skeletal Muscle Fibers

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.

Membranous alterations in the cytoplasm and nucleus in a primitive neuroectodermal tumor of the brain

1978 ◽  
Vol 36 (2) ◽  
pp. 628-629
Author(s):  
C. N. Sun ◽  
C. Araoz ◽  
H. J. White
Keyword(s):  
Nuclear Envelope ◽  
Tumor Cells ◽  
Osmium Tetroxide ◽  
Primitive Neuroectodermal Tumor ◽  
Uranyl Acetate ◽  
Neuroectodermal Tumor ◽  
Annulate Lamellae ◽  
Lead Citrate ◽  
Thin Sections ◽  
The Brain

The ultrastructure of a cerebral primitive neuroectodermal tumor has been reported previously. In the present case, we will present some unusual previously unreported membranous structures and alterations in the cytoplasm and nucleus of the tumor cells.Specimens were cut into small pieces about 1 mm3 and immediately fixed in 4% glutaraldehyde in phosphate buffer for two hours, then post-fixed in 1% buffered osmium tetroxide for one hour. After dehydration, tissues were embedded in Epon 812. Thin sections were stained with uranyl acetate and lead citrate.In the cytoplasm of the tumor cells, we found paired cisternae (Fig. 1) and annulate lamellae (Fig. 2) noting that the annulate lamellae were sometimes associated with the outer nuclear envelope (Fig. 3). These membranous structures have been reported in other tumor cells. In our case, mitochondrial to nuclear envelope fusions were often noted (Fig. 4). Although this phenomenon was reported in an oncocytoma, their frequency in the present study is quite striking.

Ultrastructural and chemical evidences of heterogeneity and hormone dependence of certain glycocalyces

1978 ◽  
Vol 36 (2) ◽  
pp. 322-323
Author(s):  
B. Monis ◽  
D. Lis ◽  
I. Parlanti ◽  
A. R. Eynard ◽  
M. A. Valentich ◽  
...  
Keyword(s):  
Guinea Pig ◽  
Concanavalin A ◽  
Ruthenium Red ◽  
Transitional Epithelium ◽  
Membrane Material ◽  
Hormone Dependence ◽  
Cellulose Acetate Electrophoresis ◽  
Electrophoretic Data ◽  
Fat Globule ◽  
Collecting Tubules

We are gathering evidences which indicate ultrastructural variations and chemical heterogeneity of certain glycocalyces as well as hormone dependence of some of them. Thus, in the lumenal glycocalyx of renal collecting tubules of the guinea-pig granular and filamentous structures were seen (1, fig. 1). By isolation, chemical analysis and cellulose acetate electrophoresis in various buffers of tubular membrane material, glycopeptides and glycosaminoglycans were identified (fig. 2).Guinea-pig and rat transitional epithelium of urinary tract showed a filamentous lumenal glycocalyx demonstrable with ruthenium red (fig. 3) but which only in part stained with concanavalin A. Chemical and electrophoretic data indicated that urothelium contains glycoproteins, glycosaminoglycans and glycolipids.The glycocalyx of the fat globule membrane of milk of several species has a granular appearance as shown by cationic dyes and by concanavalin A (2, 3, fig. 4 and 5). Also, several glycoproteins were isolated and identified on polyacrilamide gel electrophoresis (fig. 6). Glycosaminoglycans and certain glycolipids such as sulfatides were chemically identified in this glycocalyx.

Ultrastructure of the lesion induced by toxin T-514 isolated from K. humboldtiana in the alveolar region of the lung

1992 ◽  
Vol 50 (1) ◽  
pp. 640-641
Author(s):  
Julio Sepúlveda-Saavedra ◽  
Beatriz González-Corona ◽  
Víctor A. Tamez Rodríguez ◽  
Ma. Victoria Bermúdez de Rocha ◽  
Alfredo Piñeyro López
Keyword(s):  
Electron Microscope ◽  
Guinea Pig ◽  
Macaca Fascicularis ◽  
Osmium Tetroxide ◽  
Lead Citrate ◽  
Severe Damage ◽  
Thin Sections ◽  
Alveolar Region ◽  
Histopathological Findings

It has been shown in previous studies that the toxin T-514 isolated from K. humboldtiana induces severe damage to the lung in treated rodents. Histopathological findings include edema, and alveolar hemorrage. However, the ultraestructure of the lesion has not been investigated. In this study we used two species of rodents: Hamster and guinea pig, and a primate: Macaca fascicularis. Animals received different single dosis of the toxin via intraperitoneal. Control animals received only the vehicle (propylen glycol). Inmediately after spontaneous death, lung samples were fixed in Karnovsky-Ito fixative, post fixed in osmium tetroxide and embedded in epon. Thin sections were prepared with an Ultratome V LKB, stained with uranly acetate and lead citrate, and studied in an electron microscope Zeiss-EM109.

An unusual cell surface modification: a double plasma membrane

1979 ◽  
Vol 39 (1) ◽  
pp. 355-372
Author(s):  
N.J. Lane ◽  
J.B. Harrison
Keyword(s):  
Plasma Membrane ◽  
Membrane Structure ◽  
Cell Layer ◽  
Peritrophic Membrane ◽  
Thin Sections ◽  
Double Membrane ◽  
Freeze Fracturing ◽  
Blood Sucking ◽  
Great Pressure ◽  
Insertion Sites

The occurrence of an unusual double plasma membrane structure is reported; it has been studied in conventional thin sections, after lanthanum-impregnation and with freeze-fracturing. This modification of the plasmalemma is found where the luminal cell membrane (I membrane) of gut microvilli in the haematophagous insect, Rhodnius prolixus, is surrounded by a second, outer membrane (O membrane), the 2 separated from one another by a highly regular I-O space of about 10 nm. Lanthanum impregnation reveals the presence of columns inclined at an angle, within this I-O space; as in the continuous junctions which link the lateral borders of these cells, these columns may maintain the very precise I-O distance. From the outer microvillar membranes radiate short spoke-like fibrils or sheets which encounter another more extensive system of myelin-like sheets. Freeze-fracturing reveals that the spoke-like sheets and the other ones which lie like a tube, around and parallel to the microvilli, contain linear ridges composed of particles, lying at random within layers of the myelin-like material which also extends into the lumen of the gut. The microvillar membanes, both O and I, fracture into faces containing rows of either PF particles or EF pits arranged as spiral ridges or grooves around the sides and across the tip of each microbillus. These could be the insertion sites of one or both of the I-O columns and spoke-like sheets while the sheets could represent a variant of peritrophic membrane. The double membrane may be a cellular device to increase the strength of the microvillar layer in these blood-sucking animals, since the cell layer must withstand great pressure owing to a sudden massive extension of the gut during a blood meal.

scholarly journals Immunocytochemical localization of histamine in secretory granules of rat peritoneal mast cells with conventional or rapid microwave fixation and an ultrastructural post-embedding immunogold technique.

1992 ◽  
Vol 40 (9) ◽  
pp. 1247-1256 ◽  
Author(s):  
G R Login ◽  
S J Galli ◽  
A M Dvorak
Keyword(s):  
Electron Microscopy ◽  
Mast Cells ◽  
Guinea Pig ◽  
Secretory Granules ◽  
Fixation Method ◽  
Structural Localization ◽  
Thin Sections ◽  
Aldehyde Fixation ◽  
Peritoneal Mast Cells ◽  
Rat Peritoneal Mast Cells

We used a post-embedding immunogold labeling approach to define the fine-structural localization of histamine in rat peritoneal mast cells that were fixed using either standard aldehyde fixation or a fast microwave-aldehyde fixation method. Specimens were processed routinely for electron microscopy. Thin sections were exposed first to guinea pig antihistamine antiserum and then to gold-conjugated goat IgG directed against guinea pig IgG. By transmission electron microscopy, gold particles were localized to the matrix of cytoplasmic granules. Control sections treated with non-immune sera did not show labeling of mast cells. Adsorption of antihistamine antiserum with purified histamine or histamine bound to agarose showed a significant reduction (p less than 0.005) in granule staining. We also confirmed that our isolation procedures yielded functionally competent mast cells which released histamine when stimulated with sheep anti-rat IgE antiserum or with compound 48/80. These studies define the conditions of fixation for electron microscopy that are appropriate for the localization of histamine in the granule matrix of rat peritoneal mast cells.

Pattern of Incorporation of [3H]Uridine into RNA of Amphibian Oocyte Nucleoli

1967 ◽  
Vol 2 (2) ◽  
pp. 145-150
Author(s):  
H. C. MACGREGOR
Keyword(s):  
Nuclear Envelope ◽  
Incubation Time ◽  
Core Material ◽  
Distinct Component ◽  
Thin Sections ◽  
The Core ◽  
Amphibian Oocyte ◽  
Nucleolar Rna ◽  
Silver Grains

Amphibian oocytes were incubated in vitro in the presence of [3H]uridine, and autoradiographs were made of nucleoli isolated from these oocytes and of sections of oocytes. After incubations of 2 h or less the nucleoli of oocytes larger than 0.6 mm diameter are asymmetrically labelled. With longer incubations nucleoli from oocytes of 0.6 to 1.1 mm diameter become more uniformly labelled. Those of oocytes larger than 1.2 mm diameter remain asymmetrically labelled whatever the incubation time. Autoradiographs of 1-µ sections through oocytes larger than 0.6 mm diameter show, after short incubations, asymmetrically labelled nucleoli. In these autoradiographs silver grains are concentrated over a distinct component of each nucleolus which is eccentrically placed towards the nuclear envelope. Thin sections of oocytes show nucleoli consisting of core and cortex. The core material is always concentrated into the half of the nucleolus which lies nearer the nuclear envelope. Autoradiographs of separated nucleolar cores and cortices from oocytes larger than 0.6 mm diameter show, after short incubations, silver grains over cores but not over cortices. Similar autoradiographs prepared from oocytes of 0.6 to 1.1 mm diameter, after longer incubations, show grains over cores and cortices. These results appear to indicate that nucleolar RNA is synthesized in the nucleolar core, in association with the nucleolar DNA, and is thence transferred to the cortex where it is built into ribonucleoprotein particles. Initial asymmetrical labelling is a consequence of the eccentric location of the nucleolar core. The nucleoli of oocytes smaller than 0.6 mm diameter always label symmetrically; such nucleoli consist entirely of core material. It is suggested that the nucleoli of oocytes larger than 1.2 mm diameter always label asymmetrically because transfer of RNA from core to cortex proceeds more slowly than in smaller oocytes.

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