Showing papers on "Sperm plasma membrane published in 1991"

Human sperm capacitation and the acrosome reaction.

Abstract: A model is presented that describes the mechanism of human sperm capacitation and the acrosome reaction. The processes of capacitation and the acrosome reaction are proposed to function in control of the activation/release of acrosomal enzyme(s) involved in sperm penetration through the zona pellucida. During capacitation, the sperm head membranes are biochemically modified, allowing the acrosome reaction to take place when the spermatozoon approaches or reaches the zona pellucida, resulting in the localized activation and release of the appropriate enzyme(s). Further, capacitation is presented as a continuing process that occurs during sperm transport through the female genital tract and is physiologically not completed until the spermatozoon reaches the oocyte (unless the spermatozoa are kept at a particular genital tract site for prolonged periods). The biochemical alterations that occur during capacitation are discussed. It is suggested that extensive modifications in the lipid bilayer structure, e.g. in the cholesterol or phospholipid content, are not part of capacitation because such changes would prematurely destabilize the membranes. Rather, such changes occur during the acrosome reaction. It is also proposed that the human sperm acrosome reaction has many similarities to the somatic cell exocytotic events which occur during the regulated pathway of secretion. One or more oocyte stimuli result in the activation of protein kinases, likely (but not necessarily) via activation of G-protein coupled receptors on the sperm plasma membrane and the formation of second messengers. The kinases phosphorylate and activate proteins, continuing the biochemical cascade that ultimately results in the acrosome reaction. The role of other enzyme systems such as those involved in ion transport, proteolysis, phospholipid metabolism (including that of arachidonic acid) and other metabolic events, is discussed. Calcium ion influx as initiator of the acrosome reaction is reconsidered. The proposed model also takes into consideration the structural events of membrane fusion.

How the egg regulates sperm function during gamete interaction: facts and fantasies.

Abstract: Although details of the molecular mechanism are not yet clear, considerable evidence suggests that the egg-specific extracellular matrix component ZP3 regulates an essential event of sperm function, the acrosome reaction. Spatial control of this exocytotic event appears to be exerted by immobilization of the triggering ligand, ZP3, in the zona pellucida matrix surrounding the egg. Our data suggest that the signal transduction pathway in sperm activated by this ligand involves highly conserved components that are involved in many other eukaryotic signalling events. Recent experiments indicate that the murine zona pellucida glycoprotein ZP3 regulates acrosomal exocytosis by aggregating its corresponding receptors (ZP3-Rs) located in the mouse sperm plasma membrane. In other experiments, we have identified a putative ZP3-R of mouse sperm with Mr 95,000. Indirect immunofluorescence localizes this ZP3-R, termed p95, to the acrosomal region of the mouse sperm head, which is the anticipated location for ZP3-Rs. Membrane fractionation studies indicate that p95 cofractionates with a plasma membrane-enriched preparation from sperm that contains zona pellucida-receptor activity. In addition to its role as a ZP3-R, p95 also serves as a substrate for a tyrosine kinase in response to zona pellucida binding. On the basis of the data presented here, and borrowing heavily from findings for other signalling systems, we have formulated two testable hypotheses that are compatible with the available data: either p95 is itself a protein tyrosine kinase receptor, or p95 serves as a ZP3 receptor and is separate from a protein tyrosine kinase that is activated during gamete interaction.(ABSTRACT TRUNCATED AT 250 WORDS)

HIV particles detected in spermatozoa of patients with AIDS

Abstract: In this paper the Authors describe the presence of HIV particles on and in mature spermatozoa either ejaculated by AIDS patients or incubated in vitro with HIV. Both kinds of spermatozoa have particles localized around the sperm organelles. In the first case, the nucleoid of the virus can be enveloped by a membrane-like coat or be devoid of it and form buddings in the plasma membrane. In the in vitro infected spermatozoa, only membrane enveloped nucleoids are present, and no process of budding can be found. The Authors conclude considering that the spermatozoa of the AIDS patients can be penetrated by the virus particles in different moments of their life, and show the HIV particles in different stages of their cycle: some of them have freshly penetrated the sperm, and are still contained in a membrane-like coat, others are replicated and are budding through the sperm plasma membrane. On the contrary, in vitro infected spermatozoa have only freshly penetrated virus particles, and lack buddings and membrane-free nucleoids. The presence of the HIV virus in spermatozoa is substantiated by labelling with monoclonal or polyclonal anti-HIV antibodies.

Fertilization in the Mouse

Abstract: To the murine spermatozoon, its conspecific egg in vivo must seem as remote and screened by obstacles as was the Holy Grail to the Knights of Chivalry. In order that the egg be fertilized, the sperm must overcome obstacles found in the female reproductive tract and surrounding the egg itself as well as prepare itself for the membrane fusion events that lead to its entry into the egg’s cytoplasm. In the laboratory, the technique of in vitro fertilization removes most of the naturally occurring impediments posed by the female reproductive tract and provides almost unobstructed access for the sperm to the egg and an unobstructed view for the investigator. The process by which the sperm and egg interact from time of initial contact to formation of male and female pronuclei may, as a result, be studied in detail. In the mouse, successful in vitro fertilization of isolated eggs was accomplished over 20 years ago (Whittingham, 1968). In the intervening period, clarification of the sequence of the reactions involved in mouse sperm—egg interaction leading to fertilization has begun, and the number and complexity of those reactions are now more fully appreciated. However, too much emphasis on in vitro experiments may serve to obscure the biology of fertilization. Section 2 of this chapter is, therefore, concerned with the natural route taken by the sperm to reach the egg and the ensuing events as fertilization occurs in vivo. Section 3 then describes in vitro studies aimed at understanding the reactions involved in direct sperm—egg interaction that culminates in fertilization of the egg. It is hoped that information gained from both in vivo and in vitro studies will permit the construction of a unifying description of the process of fertilization in the mammal.

Cryopreservation, actin localization and thermotropic phase transitions in ram spermatozoa.

Abstract: The effects of controlled stress, i.e. cooling, upon the distribution of actin in ram spermatozoa were examined to investigate the hypothesis that cytoskeletal proteins are involved in the maintenance of sperm plasma membrane integrity. The normal distribution of actin on the spermatozoon was initially determined. A monoclonal antibody (IgM) interacted exclusively with the post-acrosomal region and the principal piece of the flagellum. By the use of a polyclonal antibody, actin was detected on the acrosome (excluding the equatorial segment), the post-acrosomal region and the whole of the flagellum. The actin was present in its non-filamentous form. Spermatozoa fixed at 39 degrees C and then treated for the immunofluorescent detection of actin with the monoclonal antibody were mostly unstained (proportion stained = 4.4% (+/- 1.6; s.e.m.); n = 8 ejaculates). Provided spermatozoa were permeabilized by greater than 0.025% Triton X-100 before immunofluorescence, actin was localized in the postacrosomal region of all sperm heads, and to a minor extent on the principal piece of the flagellum. Use of the polyclonal antibody confirmed that the post-acrosomal antigen was unmasked by detergent treatment. Slow cooling, over 2-h periods to various temperatures between 5 and 15 degrees C, also induced an increase in the proportion of cells showing post-acrosomal actin immunoreactivity. Cooling through the temperature range 15 to 10 degrees C markedly increased the proportion of immunoreactive cells (mean +/- s.e.m.; 12 +/- 4.5% at 15 degrees C; 27 +/- 4.5% at 10 degrees C; n = 4 ejaculates). Further cooling to 5 degrees C failed to elicit increased staining. Ultrastructural examination of cooled spermatozoa confirmed that a subpopulation of spermatozoa exhibited post-acrosomal actin immunoreactivity after cooling. These results are compatible with the suggestion that actin fulfills a stabilizing function in spermatozoa.

Glycoprotein changes in the rat sperm plasma membrane during maturation in the epididymis.

Abstract: To investigate surface glycoprotein changes during post-testicular maturation, plasma membranes were isolated from proximal caput, distal caput, and cauda epididymal rat spermatozoa. Membrane glycoproteins were identified on Western blots of SDS-PAGE fractionated samples using biotinylated lectins and Vecta-stain reagents; these were compared to glycoproteins present in cauda epididymal luminal fluid. Lens culinaris agglutinin, Pisum sativum agglutinin, peanut agglutinin, wheat germ agglutinin, Ricinus communis agglutinin, Ulaex europaeus agglutinin, and Dolichol biflorus agglutinin each bound a specific subset of the polypeptides present. Several types of glycoprotein changes were noted including their appearance, loss, alteration of staining intensity, and alteration of electrophoretic mobility. Some maturation-dependent sperm surface glycoproteins co-migrated with glycoproteins present in epididymal fluid. This approach of direct analysis of the glycoproteins in purified plasma membranes identifies a broader spectrum of maturation-related surface changes occurring within the epididymis than are noted with surface labeling procedures.

Fertilization in the Mouse

Abstract: The mouse has been used extensively in studies of mammalian fertilization, especially during the last 20 years or so. Among the many reasons for choosing mice for such studies are (1) the tremendous wealth of knowledge available about mouse developmental and reproductive biology, (2) the relatively low cost and ease with which mice can be obtained, housed, and handled, (3) the well-established protocols available for reliably obtaining and culturing relatively large numbers of mouse gametes, (4) the well-established protocols available for carrying out fertilization with mice, both in vivo and in vitro, and (5) the firm belief that fertilization in mice is an appropriate model for understanding many aspects of human fertilization. These and other factors have made the mouse a principal player in mammalian fertilization research and, consequently, a major contributor to our understanding of the mammalian fertilization process. Several manuals are available that describe mouse husbandry and experimental manipulation of mouse gametes and embryos (Rafferty, 1970; Daniel, 1971, 1978; Hogan et al., 1986; Burki, 1986; Monk, 1987).

Cross-linking a maturation-dependent ram sperm plasma membrane antigen induces the acrosome reaction.

Abstract: ESA152 is a highly hydrophobic 18 kDa sialoglycoprotein, which becomes expressed on ram sperm in the proximal cauda epididymis. ESA 152 is expressed on all regions of the sperm surface, most strongly on the posterior region of the head, most weakly on the anterior region of the head. In this paper, we show that induction of the acrosome reaction with Ca2+ ionophore causes ESA152 to be redistributed from the posterior to the anterior region of the head plasma membrane. Cross-linking ESA152 with bivalent antibody causes similar redistribution and induces the acrosome reaction, Induction of the acrosome reaction with ESA152 antibody requires Ca2+ but is insensitive to (10 ng/ml) pertussis toxin.

In-vitro effects of anti-sperm antibodies on human sperm movement

Abstract: The present study was designed to investigate whether autoantibodies to external domains of the sperm plasma membrane affect the movement of normal motile spermatozoa. Eight sera and 20 seminal plasma samples containing high levels of anti-sperm antibodies as well as antibodies eluted from the sperm fraction of 19 autoimmune ejaculates were incubated with donor's motile spermatozoa, obtained by swim-up migration in Tyrode's solution. Sperm movement was analysed using 1 s exposure microphotography when greater than 70% of the spermatozoa were coated with antibodies (after 30-90 min of incubation). At least 50 tracks of progressively motile spermatozoa were analysed in order to obtain the mean values of the amplitude of lateral head displacement (ALH) and the velocity of progression (VSL). Serum antibodies and sperm eluted antibodies had quite consistent but opposite effects on sperm movement; serum antibodies increased ALH and decreased VSL whereas eluted antibodies decreased ALH and increased VSL. Seminal antibodies did not affect these two parameters significantly. Furthermore, seminal antibodies and sperm eluted antibodies obtained from the same ejaculates had distinct effects on ALH and/or VSL. This diversity was apparently not linked to antibody isotype or localization on the sperm membrane; it might be due to differences in the composition of the extracellular media. These results suggest a dynamic effect of anti-sperm antibodies on sperm movement, a possibility that merits further investigation.

Inhibition of sperm-zona pellucida tight binding by sperm immobilizing antibodies as assessed by the hemizona assay (HZA).

Abstract: Researchers collected serum samples from 23 infertile patients with sperm immobilizing antibodies (SI-Ab) and 1 pregnant patient from the Department of Obstetrics and Gynecology at the Hyogo Medical College in Japan to screen sera to determine whether they contained factors to inhibit sperm-zona pellucida tight binding. They used the recently developed hemizona assay (HZA) to test for this binding. The HZA assay showed that all 23 serum samples inhibited sperm-zona pellucida tight binding. The hemizona index (HZI) ranged from 3-53 with a mean of 18.1 (standard deviation of = or - 12) compared to a normal (HZI) of 100. Serum samples with titers >10 of SI50 inhibited sperm-zona binding as well as those with titers -or= 10 of SI50 (HZIs=17.3 vs. 18.9; p>.1). All 23 serum samples bound to the surface of sperm plasma membrane after 1 hour coincubation as evidenced by the fact that they all demonstrated >50% IgG beads bound. Further the results of the indirect immunobead test (I-IBT) showed that positive sera (+or= 20% IgG beads) significantly inhibited binding more than negative sera ( .1). No association existed between HZI and site of IB binding. The researchers interpreted theses results to mean that sera with both SI-Ab and antibodies recognized I-IBT for IgG and IgA may play a significant role to inhibit the sperm-zona pellucida tight binding. In conclusion physicians should expect patients with low HZI to have more problems conceiving than those with normal HZI. In vitro fertilization using heat inactivated human cord serum or donor serum may help them to conceive.

Reduction of the fertilizing capacity of sea urchin sperm by cannabinoids derived from marihuana. II. Ultrastructural changes associated with inhibition of the acrosome reaction.

Abstract: Pretreatment of Strongylocentrotus purpuratus sperm with delta 9-tetrahydrocannabinol (THC) prevents the triggering of the acrosome reaction by egg jelly. Examination of THC-treated sperm by transmission electron microscopy reveals that the membrane fusion reaction between the sperm plasma membrane and the acrosomal membrane is completely blocked. Electron-dense deposits are present in the subacrosomal fossa and in the centriolar fossa. The nuclear envelope is fragmented in close proximity to the electron-dense deposits. The electron-dense deposits are not bound by a limiting membrane, stain positively for lipid with thymol and farnesol, and disappear from THC-treated sperm that are extracted with chloroform:methanol (2:1) after glutaraldehyde fixation. The electron-dense deposits are lipid in nature and may be a hydrolytic product of the nuclear envelope. Electron-dense deposits are seen in sperm after 1-10 min treatment with 5-100 microM THC. The electron-dense deposits disappear after removal of THC from the sperm by washing, but the fragmented nuclear envelope in the subacrosomal fossa persists. Cannabidiol (CBD) and cannabinol (CBN) also inhibit the triggering of the acrosome reaction by egg jelly and produce ultrastructural changes in the sperm identical to those elicited by THC. Enhanced phospholipase activity stimulated by THC, CBD, and CBN may be the cause of the accumulation of lipid deposits in the sperm. Metabolites derived from this modification of membrane phospholipids may prevent triggering of the acrosome reaction by egg jelly and thereby inhibit fertilization.

Sperm cells of the pollen tubes of Brassica: Ultrastructure and isolation.

Abstract: Sperm cells of pollen tubes grown both in vivo and in vitro form a male germ unit. Extensions from both sperm cells of each pollen tube are closely associated with the tube nucleus. A high yield (2.7 × 104. 20 mg−1 pollen grains germinated) of intact sperm cells was obtained following release by osmotic shock from pollen tubes grown in vitro. Structural integrity of isolated sperm was maintained by isolation at low temperature in an osmotically balanced medium. At 4° C many isolated sperm pairs were still enclosed within the pollentube inner plasma membrane. Sperm cells not enclosed within this membrane no longer remained connected as a pair. During isolation vesicles formed on the sperm cell surface from disruption of the fibrillar components bridging the periplasmic space. Both in the pollen tube and after isolation the sperm nucleus is in close association with at least one region of the sperm plasma membrane. Sperm isolated at room temperature showed the presence of nucleopores, and nuclei were euchromatic, instead of heterochromatic as in intact sperm in the pollen tube.

Use of Neonatal Tolerization and Chemical Immunosuppression for the Production of Monoclonal Antibodies to Maturation‐Specific Sperm Surface Molecules

Abstract: Mammalian sperm acquire functional maturity as they move from the caput to the cauda epididymidis. Changes occur in the protein/glycoprotein composition of the sperm plasma membrane during this time, and may be essential to the maturation process. The production of monoclonal antibody (Mab) probes to the maturation-specific molecules has been difficult since new proteins comprise a minor portion of total membrane proteins. This report describes a protocol for enhancing the production of Mabs to maturation specific molecules. By injecting neonatal mice with caput epididymal sperm plasma membranes, in combination with chemical immunosuppression at adulthood, the mice were made tolerant to the antigens expressed on the caput sperm membranes. Subsequent immunization with cauda epididymal sperm plasma membranes allowed the production of Mabs to the maturation-specific moieties without the necessity for extensive antigen purification procedures. The majority of the resulting Mabs recognize cauda, not caput, epididymal sperm plasma membranes as determined by enzyme-linked immunosorbent assay (ELISA), immunocytochemistry on unfixed cells, and Western blot analyses, even though the protein profile from caput epididymal sperm plasma membranes is very similar to that from cauda membranes. The five Mabs described also recognize cauda fluid antigens, suggesting that the maturational changes on the sperm plasma membranes arise from interactions with the epididymal fluid. Use of the tolerization/immunosuppression protocol has provided Mab tools to assist in the study of sperm maturation during epididymal transit.

Gamete interactions and the fate of sperm organelles in fertilized echinoderm eggs.

Abstract: Investigations of gamete fusion, sperm entry and the fate of the sperm nucleus, plasma membrane, mitochondrion, and axonemal complex in fertilized echinoderm eggs are reviewed. The timing of gamete fusion with respect to the onset of electrical activity characteristic of the activated egg and the affects of fixation conditions on the stability of fusing membranes are discussed. Observations from investigations using cationized ferritin labeled gametes and immunogold cytochemistry to demonstrate the mixing of sperm plasma membrane components within the egg plasma membrane, in particular along the surface of the fertilization cone, are compared with results from studies in somatic cells. Transformations of the sperm nucleus into a male pronucleus, consisting of sperm nuclear envelope breakdown, chromatin dispersion, and formation of a pronuclear envelope, are correlated with recent biochemical observation of similar processes in other cellular systems. Fates of the sperm mitochondrion and axonemal complex are examined.

Water insoluble fraction of egg yolk maintains porcine sperm motility by activating adenylate cyclase

Abstract: The decrease in motility of porcine cauda epididymal sperm was less than that of caput epididymal sperm in the medium containing bicarbonate. This may be due to the difference of sensitivity of adenylate cyclase to bicarbonate between mature and immature sperm; activation of mature sperm enzyme by bicarbonate was higher than that of immature sperm. Nondialysable fraction of egg yolk prevented the decrease in motility of immature sperm in the presence of bicarbonate, but it was not effective for the motility of mature sperm under the same condition, because only bicarbonate is sufficient for the maintenance of its motility. In the absence of bicarbonate, both mature and immature sperm required egg yolk to maintain motility. The favorable effect of egg yolk on the motility is ascribed to the enhancement of intracellular cAMP level. Partial fractionation of egg yolk showed that water-insoluble lipoprotein fraction contains factor(s) which activates adenylate cyclase in sperm plasma membrane. This is the first report in which high molecular weight activator of the sperm enzyme was demonstrated.

A Comparison of Mammalian Sperm Membranes

Abstract: Mammalian spermatozoa exhibit considerable species differences in their size and shape, yet they all possess the same set of cellular organelles assembled on a common architectural theme. The polarized spermatozoon is partitioned into distinct segments or domains, distinguished by specific subsets of the cellular organelles (Eddy, 1988; Fawcett, 1975). These include the acrosomal and postacrosomal segments of the head, followed by the connecting piece, midpiece, principal piece, and end piece segments of the flagellum. During fertilization, different segments perform specific functions in generating motility, in binding the zona pellucida, in penetrating the egg coats, and in fusing with the egg plasma membrane (Wassarman, 1987; Yanagimachi, 1988). The sperm plasma membrane plays a central role in regulating these functions, and it varies in structure and molecular composition in the different domains. In this chapter we discuss domain-specific properties of the plasma membrane and address mechanisms that may maintain their unique properties.

Fertilization: A model for cell-cell interaction

Abstract: Many interactions between mammalian cells are known to involve carbohydrate moieties of plasma membrane glycoconjugates and corresponding carbohydrate-binding proteins. Examples are found in systems as diverse as embryogenesis, lymphocyte homing (Lasky, 1991), tumor invasion (Gabius and Nagel, 1988) and fertilization (Miller and Ax, 1990). Although the overall process of fertilization is unquestionably unique, it is made up of a series of steps that are not unique and are commonly used by a variety of both normal and tumor cells.

Facts and Fantasies1

Abstract: Although details of the molecular mechanism are not yet clear, considerable evidence suggests thatthe egg-specific extracellularmatrix component ZP3 regulates an essential event of sperm function,the acrosome reaction,Spatialcontrol of this exocytotic event appears to be exerted by immobilization of the triggering ligand, ZP3, in the zona pellucidamatrix surrounding the egg. Our data suggest that the signal transduction pathway in sperm activated by this ligand involves highly conserved components that are involved in many other eukaryotic signalling events, Recent experiments indicate that the murine zona pellucida glycoprotein ZP3 regulates acrosomal exocytosis by aggregating its corresponding receptors (ZP3-Rs) located in the mouse sperm plasma membrane. In other experiments, we have identified a putative ZP3-R of mouse sperm with M, 95000. Indirectimmunofluorescence localizes this ZP3-R, termed p95, to the acrosomal region of the mouse sperm head, which is the anticipated location for ZP3-Rs. Membrane fractionation studies indicate that p95 cofractionates with a plasma membrane-enriched preparation from sperm that contains zona pellucida-receptor activity. In addition to its role as a ZP3-R, p95 also serves as a substrate for a tyrosine kinase in response to zona pellucida binding. On the basis of the data presented here, and borrowing heavily from fmdings for other signalling systems, we have formulated two testable hypotheses that are compatible with the available data: either p95 is itself a protein tyrosine kinase receptor, or p95 serves as a ZP3 receptor and is separate from a protein tyrosine kinase that is activated during gamete interaction, Examination of these hypotheses will allow further exploration of the relationshipbetween sperm binding to the zona pellucida and triggering of the acrosome reaction,