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.

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.

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.

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.

A freeze-fracture-etched study of the physarum polycephalum plasma membrane during early formation of the macroplasmodial stage

1993 ◽  
Vol 51 ◽  
pp. 346-347
Author(s):  
Randolph W. Taylor ◽  
Henrie Treadwell
Keyword(s):  
Plasma Membrane ◽  
Life Cycle ◽  
Growth Phase ◽  
Filter Paper ◽  
Physarum Polycephalum ◽  
Freeze Fracture ◽  
Petri Dish ◽  
Wire Screen ◽  
Wide Mouth ◽  
Sterile Filter

The plasma membrane of the Slime Mold, Physarum polycephalum, process unique morphological distinctions at different stages of the life cycle. Investigations of the plasma membrane of P. polycephalum, particularly, the arrangements of the intramembranous particles has provided useful information concerning possible changes occurring in higher organisms. In this report Freeze-fracture-etched techniques were used to investigate 3 hours post-fusion of the macroplasmodia stage of the P. polycephalum plasma membrane.Microplasmodia of Physarum polycephalum (M3C), axenically maintained, were collected in mid-expotential growth phase by centrifugation. Aliquots of microplasmodia were spread in 3 cm circles with a wide mouth pipette onto sterile filter paper which was supported on a wire screen contained in a petri dish. The cells were starved for 2 hrs at 24°C. After starvation, the cells were feed semidefined medium supplemented with hemin and incubated at 24°C. Three hours after incubation, samples were collected randomly from the petri plates, placed in plancettes and frozen with a propane-nitrogen jet freezer.

scholarly journals Progesterone treatment of boar spermatozoa improves male pronuclear formation after intracytoplasmic sperm injection into porcine oocytes

Zygote ◽  
2002 ◽  
Vol 10 (2) ◽  
pp. 95-104 ◽  
Author(s):  
Mike Katayama ◽  
Takashi Miyano ◽  
Masashi Miyake ◽  
Seishiro Kato
Keyword(s):  
Plasma Membrane ◽  
Intracytoplasmic Sperm Injection ◽  
Acrosome Reaction ◽  
Sperm Chromatin ◽  
Oocyte Activation ◽  
Freezing And Thawing ◽  
Chromatin Decondensation ◽  
Progesterone Treatment ◽  
Pronuclear Formation ◽  
Boar Spermatozoa

Boar spermatozoa were prepared for intracytoplasmic sperm injection (ICSI) by two different treatments to facilitate sperm chromatin decondensation and improve fertilisation rates after ICSI in pigs: spermatozoa were either frozen and thawed without cryoprotectants, or treated with progesterone. Morphological changes of the sperm heads after the treatments were examined and then the activation of oocytes and the transformation of the sperm nucleus following ICSI were assessed. After freezing and thawing, the plasma membrane and acrosomal contents over the apical region of sperm head were lost in all the spermatozoa. Following treatment with 1 mg/ml progesterone, the acrosome reaction was induced in 61% of spermatozoa. After injection of three types of spermatozoa, non-treated spermatozoa and progesterone-treated (i.e. acrosome-reacted) spermatozoa induced oocyte activation, but frozen-thawed spermatozoa induced oocyte activation at a significantly lower rate. Sixty-two per cent of sperm heads remained orcein-negative for 6 h, however, resulting in delayed sperm chromatin decondensation and low male pronuclear formation in the oocytes injected with a non-treated spermatazoon. Since the treatments of freezing and thawing and progesterone for spermatozoa accelerated the initial change in sperm chromatin and the latter treatment induced oocyte activation earlier, it is considered that the delay in oocyte activation and decondensation of sperm chromatin after injection of non-treated spermatozoa is caused by the existence of the sperm plasma membrane. These results show that progesterone treatment efficiently induces the acrosome reaction in boar spermatozoa without destroying their potency for oocyte activation, and the induction of the acrosome reaction results in the promotion of male pronuclear formation after ICSI.

Lipid changes of goat sperm plasma membrane during epididymal maturation

1991 ◽  
Vol 1061 (2) ◽  
pp. 185-196 ◽  
Author(s):  
Ajay P.S. Rana ◽  
Gopal C. Majumder ◽  
Suniti Misra ◽  
Amitabha Ghosh
Keyword(s):  
Plasma Membrane ◽  
Epididymal Maturation ◽  
Sperm Plasma Membrane
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