Surface views of spermatozoa as revealed by fracture-flip

1989 ◽  
Vol 92 (3) ◽  
pp. 415-426
Author(s):  
C.A. Forsman ◽  
P. Pinto da Silva
Keyword(s):  
Plasma Membrane ◽  
Random Distribution ◽  
Freeze Fracture ◽  
Fracture Morphology ◽  
High Density ◽  
Surface Textures ◽  
Principal Piece ◽  
Boar Spermatozoa ◽  
Posterior Ring

We have used fracture-flip to produce new, macromolecular-resolution images of the surface of boar spermatozoa. Over the head, acrosomal and postacrosomal regions display sharply demarcated, subtly different surface textures. The rim is particle-poor, as well as a region above the oblique cords over the posterior ring. The tail shows two morphologically distinct domains: (1) the principal piece is covered by a high density of parallel-helical strands and a high density of large globular particles; (2) the midpiece and the neck are covered by smaller particles with apparent random distribution. Rectangular surface specializations frequently seen near the annulus display a waffle-like texture. With the notable exception of the parallel-helical strands of the principal piece the fracture-flip images of the boar spermatozoon can be directly related to the freeze-fracture morphology of its plasma membrane.

Freeze-fracture observations on changes in the distribution of Filipin-sterol complexes during epididymal maturation and capacitation of boar spermatozoa

1990 ◽  
Vol 48 (3) ◽  
pp. 90-91
Author(s):  
N. Seki ◽  
Y. Toyama ◽  
T. Nagano
Keyword(s):  
Plasma Membrane ◽  
Essential Role ◽  
Freeze Fracture ◽  
Final Concentration ◽  
Freeze Fracturing ◽  
Physiological Conditions ◽  
Epididymal Maturation ◽  
Cacodylate Buffer ◽  
Sperm Membrane ◽  
Boar Spermatozoa

It is believed that i ntramembra.nous sterols play an essential role in membrane stability and permeability. To investigate the distribution changes of sterols in sperm membrane during epididymal maturation and capacitation, filipin has been used as a cytochemical probe for the detection for membrane sterols. Using this technique in combination with freeze fracturing, we examined the boar spermatozoa under various physiological conditions.The spermatozoa were collected from: 1) caput, corpus and cauda epididymides, 2) sperm rich fraction of ejaculates, and 3)the uterus 2hr after natural coition. They were fixed with 2.5% glutaraldehyde in 0.05M cacodylate buffer (pH 7.4), and treated with the filipin solution (final concentration : 0.02.0.05%) for 24hr at 4°C with constant agitation. After the filipin treatment, replicas were made by conventional freeze-fracture technique. The density of filipin-sterol complexes (FSCs) was determined in the E face of the plasma membrane of head regions.

Bimodal redistribution of surface transmembrane glycoproteins during Ca2+-dependent secretion (acrosome reaction) in boar spermatozoa

1989 ◽  
Vol 93 (3) ◽  
pp. 467-479
Author(s):  
A.P. Aguas ◽  
P.P. da Silva
Keyword(s):  
Plasma Membrane ◽  
Acrosome Reaction ◽  
Freeze Fracture ◽  
Secretory Process ◽  
Membrane Domains ◽  
Cytoplasmic Vesicle ◽  
Acrosomal Membrane ◽  
Freeze Fracture Electron Microscopy ◽  
Boar Spermatozoa

We used the acrosome reaction of boar sperm cells to study the dynamics of surface transmembrane glycoproteins (TMG) during a secretory process. The acrosome reaction is the Ca2+-dependent fusion of a large cytoplasmic vesicle (the acrosome) with the overlying segment of the plasma membrane (acrosomal cap) that leads to the release of the acrosomal enzymes. After triggering the acrosome reaction in vitro (2 mM-CaCl2 in the presence of 10 microM-A23187), we used freeze-fracture electron microscopy to follow the topographical rearrangement of a population of acrosomal-cap large intramembrane particles that correspond to transmembrane proteins that bind wheat germ agglutinin. We found that these TMG move in the direction of either one of two opposite poles, proximal and distal, of the acrosomal cap. This bimodal movement of the TMG reorganizes the acrosomal cap into three extensive domains. The first two, on the apical rim and on the equator, are membrane domains to which the TMG are directed and where they accumulate. The third, a large in-between area of protein clearing, corresponds to the region from which TMG were preferentially located before displacement induced by the Ca2+ effect. The topography of these new membrane domains of the acrosomal cap becomes coincident with that of the structural domains of the subjacent acrosomal membrane. Mirroring of the acrosomal membrane by the plasma membrane is followed by fusion between the two membranes, formation of an exquisite labyrinth of hybrid-membrane tubules, followed by fission and release of the acrosomal contents through intertubular fenestrae.

Topographical stability of lateral regions of the Schizosaccharomyces pombe plasma membrane as revealed by invagination characteristics

1988 ◽  
Vol 34 (9) ◽  
pp. 1063-1068 ◽  
Author(s):  
Kanji Takeo ◽  
Kazuko Nishimura ◽  
Makoto Miyaji
Keyword(s):  
Plasma Membrane ◽  
Stationary Phase ◽  
Schizosaccharomyces Pombe ◽  
Freeze Fracture ◽  
Exponential Phase ◽  
High Density ◽  
The Stability

The characteristics, especially the stability, of plasma-membrane invaginations in Schizosaccharomyces pombe were studied using freeze-fracture techniques. The plasma membrane of exponential-phase S. pombe was characterized by the uninvaginated growing pole(s) and by the existence of particularly long invaginations, reaching up to 5 μm, in the nongrowing lateral regions. The stationary-phase plasma membrane was characterized by densely invaginated cell poles and by the high density of invaginations. After reinoculation of stationary-phase cells in fresh media, preexisting invaginations in the cell poles disappeared when the latter became the growing pole. In lateral regions of the plasma membrane, however, deep invaginations, characteristic of the stationary phase, were well conserved. The results obtained suggest the topographical stability of nongrowing lateral regions of the S. pombe plasma membrane.

Topology of morphologically detectable protein and cholesterol in membranes of polypeptide-secreting cells

1981 ◽  
Vol 296 (1080) ◽  
pp. 47-54 ◽  
Keyword(s):  
Plasma Membrane ◽  
Secretory Granule ◽  
Plasma Membranes ◽  
Secretory Pathway ◽  
Freeze Fracture ◽  
Fracture Morphology ◽  
Cholesterol Content ◽  
Intramembrane Particles ◽  
Exocrine Cells ◽  
Luminal Membrane

The freeze-fracture morphology of intracellular and plasma membranes in endocrine and exocrine polypeptide-secreting cells has been studied to detect changes while these membranes interact during secretion. A qualitative and quantitative evaluation of intramembrane particles and filipin binding as indicators of protein and cholesterol content of the membranes, respectively, reveals the following changes. From the forming of the maturing pole of the Golgi complex, membranes lose morphologically detectable protein and gain morphologically detectable cholesterol. The protein-poor, cholesterol-rich secretory granule membrane then interacts with a richly particulate plasma membrane in endocrine cells and with a moderately particulate luminal membrane in exocrine cells. The site of interaction between secretory granule and plasma membrane is characterized by a local clearing of intramembrane particles; by contrast, filipin-binding sites revealing cholesterol are present in this area. In exocrine cells, the fused secretory granule, which is initially rich in filipin-cholesterol complexes and poor in particles, appears to lose progressively its filipin labelling to resemble the poorly labelled luminal membrane. These findings, although they cannot be interpreted definitely at present, clearly show impressive changes of membrane structure along the secretory pathway and suggest that a corresponding degree of functional specialization is needed for proper interaction to occur.

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.

Ultrastructure of yeast plasma membrane revealed by a new high-resolution freeze-replica method

1987 ◽  
Vol 45 ◽  
pp. 964-965
Author(s):  
Tadashi Hirano ◽  
Akira Tanaka
Keyword(s):  
Plasma Membrane ◽  
High Resolution ◽  
Single Unit ◽  
Freeze Fracture ◽  
Fracture Morphology ◽  
Replica Method ◽  
Yeast Cells ◽  
Thin Sections ◽  
Yeast Protoplasts ◽  
Fracture Theory

The Freeze-fracture morphology of the plasma membrane and surface of yeast protoplasts has been investigated by a new high resolution freeze-replica method (Tanaka et al.1978). According to freeze-fracture theory, it is generally argued that the plane of cleavage breaks down into the bilayer and then follows the plane of the membrane between the two halves of the lipid. However, when we observed thin sections of replica film of the surface of the freeze-fractured face of intact yeast cells, the single unit membrane was clearly visible between the replica film and the cytoplasm (Fig. 1). Accordingly, in the case of yeast cells, we assume that the plane of cleavage breaks down between the plasma membrane and cell wall.On the other hand, Walzthöng et al. (1982) have shown that surface granules are an artifact or form of contamination produced under the conditions used for the ordinary freeze-replica method employing metal shadowing film.

Plasma membrane ultrastructure during plant protoplast plasmolysis, isolation and wall regeneration: a freeze-fracture study

1980 ◽  
Vol 42 (1) ◽  
pp. 401-415
Author(s):  
M.J. Wilkinson ◽  
D.H. Northcote
Keyword(s):  
Plasma Membrane ◽  
Time Course ◽  
Freeze Fracture ◽  
Fracture Morphology ◽  
Plant Protoplast ◽  
Suspension Cells ◽  
Isolated Protoplasts ◽  
Fracture Study ◽  
Membrane Ultrastructure ◽  
Hexagonal Arrays

The freeze-fracture morphology of the plasma membrane of cells and isolated protoplasts of plant callus suspensions has been investigated. Plasmolysis of suspension cells leads to the formation of 2 types of hexagonal arrays of intramembrane particles situated on the inner fracture face (PF). These arrays are interpreted as proteins that have ‘crystallized’ in the plane of the membrane as the area of surrounding lipid bilayer is reduced during protoplast retraction from the cell wall. Time-course studies have revealed no positive relationship between the distribution of hexagonal arrays and the occurrence of microfibrils regenerated around isolated protoplasts during periods of culture. No evidence for the specialized transport functions attributed to hexagonal arrays of plant cells by previous workers has been found.

scholarly journals The acrosomal membrane of boar sperm: a Golgi-derived membrane poor in glycoconjugates.

1985 ◽  
Vol 100 (2) ◽  
pp. 528-534 ◽  
Author(s):  
A P Aguas ◽  
P Pinto da Silva
Keyword(s):  
Plasma Membrane ◽  
Plasma Membranes ◽  
Biochemical Characterization ◽  
Freeze Fracture ◽  
Secretory Vesicle ◽  
High Density ◽  
Con A ◽  
Cytoplasmic Surface ◽  
Acrosomal Membrane ◽  
Boar Sperm

The acrosome is a large secretory vesicle of the sperm head that carries enzymes responsible for the digestion of the oocyte's investments. The event leads to sperm penetration and allows fertilization to occur. Release of the acrosomal enzymes is mediated by the interaction between sperm acrosomal and plasma membranes (acrosome reaction). Biochemical characterization of the acrosomal membrane has been restrained by a lack of methods to isolate uncontaminated fractions of the membrane. Here, we use new methods to expose the membrane to in situ cytochemical labeling by lectin-gold complexes. We study the topology and relative density of glycoconjugates both across and along the plane of the acrosomal membrane of boar sperm. Detachment of the plasma membrane from glutaraldehyde-fixed cells exposed the cytoplasmic surface of the acrosome to the lectin markers; freeze-fractured halves of the acrosomal membrane were marked by "fracture-label" (Aguas, A. P., and P. Pinto da Silva, 1983, J. Cell Biol. 97:1356-1364). We show that the cytoplasmic surface of the intact acrosome is devoid of binding sites for both concanavalin A (Con A) and wheat germ agglutinin (WGA). By contrast, it contains a high density of neuraminidase-resistant anionic sites detected by cationic ferritin. On freeze-fractured sperm, the receptors for Con A partitioned with the exoplasmic membrane half of the acrosomal membrane. The Con A-binding glycoconjugates were accumulated on the equatorial segment of the membrane. A low density of WGA receptors, as well as of intramembrane particles, was found on the freeze-fracture halves of the acrosomal membrane. The plasma membrane displayed, in the same preparations, a high density of receptors for both Con A and WGA. We conclude that the acrosome is limited by a membrane poor in glycoconjugates, which are exclusively exposed on the exoplasmic side of the bilayer. Regionalization of Con A receptors on the acrosome shows that sperm intracellular membranes, like the sperm surface, express domain distribution of glycocomponents.

Heterogeneity of Size and Distribution of Intramembrane Particles in the Plasma Membrane of Yeast

1977 ◽  
Vol 35 ◽  
pp. 596-597
Author(s):  
E. Keyhani
Keyword(s):  
Plasma Membrane ◽  
Candida Utilis ◽  
Synthetic Medium ◽  
Freeze Fracture ◽  
Translational Diffusion ◽  
Yeast Cells ◽  
Distilled Water ◽  
Intramembrane Particles ◽  
The Matrix ◽  
Water Cell

The matrix of biological membranes consists of a lipid bilayer into which proteins or protein aggregates are intercalated. Freeze-fracture techni- ques permit these proteins, perhaps in association with lipids, to be visualized in the hydrophobic regions of the membrane. Thus, numerous intramembrane particles (IMP) have been found on the fracture faces of membranes from a wide variety of cells (1-3). A recognized property of IMP is their tendency to form aggregates in response to changes in experi- mental conditions (4,5), perhaps as a result of translational diffusion through the viscous plane of the membrane. The purpose of this communica- tion is to describe the distribution and size of IMP in the plasma membrane of yeast (Candida utilis).Yeast cells (ATCC 8205) were grown in synthetic medium (6), and then harvested after 16 hours of culture, and washed twice in distilled water. Cell pellets were suspended in growth medium supplemented with 30% glycerol and incubated for 30 minutes at 0°C, centrifuged, and prepared for freeze-fracture, as described earlier (2,3).

Deformation of Yeast Plasma Membrane by Ethidium Bromide

1988 ◽  
Vol 46 ◽  
pp. 254-255
Author(s):  
E. Keyhani
Keyword(s):  
Plasma Membrane ◽  
Thin Section ◽  
Ethidium Bromide ◽  
Freeze Fracture ◽  
Mutagenic Effect ◽  
Yeast Cells ◽  
Hydrophobic Region ◽  
Thin Section Electron Microscopy ◽  
Section Electron ◽  
Deep Pits

The mutagenic effect of ethidium bromide on the mitochondrial DNA is well established. Using thin section electron microscopy, it was shown that when yeast cells were grown in the presence of ethidium bromide, besides alterations in the mitochondria, the plasma membrane also showed alterations consisting of 75 to 110 nm-deep pits. Furthermore, ethidium bromide induced an increase in the length and number of endoplasmic reticulum and in the number of intracytoplasmic vesicles.Freeze-fracture, by splitting the hydrophobic region of the membrane, allows the visualization of the surface view of the membrane, and consequently, any alteration induced by ethidium bromide on the membrane can be better examined by this method than by the thin section method.Yeast cells, Candida utilis. were grown in the presence of 35 μM ethidium bromide. Cells were harvested and freeze-fractured according to the procedure previously described.

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