Showing papers on "Sperm published in 1988"
TL;DR: These studies demonstrate capacitation of bovine sperm by heparin requires at least a 4-h exposure of sperm toHeparin and suggest that plasma membrane changes prior to an AR can be detected by exposure of bovo sperm to LC.
Abstract: Capacitation of bovine sperm was evaluated by determining the ability of sperm to fertilize bovine oocytes in vitro and to undergo an acrosome reaction upon exposure to lysophosphatidylcholine (LC). Incubation of sperm with heparin (10 micrograms/ml) increased the percentage of oocytes fertilized, but this required exposing sperm to heparin for at least 4 h before adding them to oocytes. There was no effect on the percentage of motile or acrosome-reacted sperm after exposure of noncapacitated sperm to 100 micrograms/ml LC for 15 min. When sperm were incubated for 4 h with heparin, exposure to 100 micrograms/ml LC for 15 min had no effect on the percentage of sperm that were motile, but the percentage of acrosome-reacted sperm increased from less than 10% to over 70%. The acrosome reactions (ARs) induced by LC were synchronous, reached maximal levels within 15 min, and differed (p less than 0.001) between sperm incubated under capacitating (with heparin) and noncapacitating conditions (without heparin). The time course required for heparin to capacitate sperm as judged by in vitro fertilization and to render sperm sensitive to LC induction of the AR were found to be similar. The percentage of ARs induced by LC and percentage of oocytes fertilized by sperm were found to be heparin-dose-dependent, with the maximum responses occurring at 5-10 micrograms/ml heparin. The correlation between the mean fertilization and LC-induced AR percentages was 0.997 (p less than 0.01). These studies demonstrate capacitation of bovine sperm by heparin requires at least a 4-h exposure of sperm to heparin and suggest that plasma membrane changes prior to an AR can be detected by exposure of bovine sperm to LC.
1,580 citations
TL;DR: By evaluating sperm morphology with the proposed strict criteria, its predictive value in in vitro fertilization is enhanced.
1,095 citations
TL;DR: Assessment of the ability of the antioxidants, butylated hydroxytoluene, and vitamin E, to curtail the peroxidative damage inflicted by cells in response to centrifugation revealed a significant improvement of sperm function in the presence of vitamin E.
Abstract: The mechanisms responsible for mediating the influence of sperm preparation protocols on human sperm function have been investigated. Techniques that involved the separation of motile spermatozoa prior to centrifugation were found to yield sperm suspensions of highest quality. If the spermatozoa were centrifuged prior to isolation of the motile cells, sperm function was impaired. The detrimental effects of centrifugation were associated with a sudden burst of reactive oxygen species production by a discrete subpopulation of cells (characterized by significantly diminished motility and fertilizing capacity) that could be separated from normal functional spermatozoa on Percoll gradients. If unfractionated sperm suspensions were subjected to centrifugation, the reactive oxygen species generated by this subpopulation impaired the functional competence of normal spermatozoa in the same suspension. Assessment of the ability of the antioxidants, butylated hydroxytoluene, and vitamin E, to curtail the peroxidative damage inflicted by such cells in response to centrifugation revealed a significant improvement of sperm function in the presence of vitamin E.
732 citations
TL;DR: The use of the polymerase chain reaction for analysing DNA sequences in individual diploid cells and human sperm shows that two genetic loci can be co-amplified from a single sperm, which may allow the analysis of previously inaccessible genetic phenomena.
Abstract: The use of the polymerase chain reaction for analysing DNA sequences in individual diploid cells and human sperm shows that two genetic loci can be co-amplified from a single sperm, which may allow the analysis of previously inaccessible genetic phenomena.
642 citations
TL;DR: Zona pellucida glycoproteins are responsible for species-restricted binding of sperm to unfertilized eggs, inducing sperm to undergo acrosomal exocytosis, and preventing sperm from binding to fertilized eggs.
623 citations
TL;DR: ZP2 serves as a secondary receptor for sperm during the fertilization process in mice and that maintenance of binding of acrosome-reacted sperm to eggs may involve a sperm, trypsin-like proteinase.
348 citations
TL;DR: It is concluded that galactose, located in alpha-linkage at the nonreducing terminus of O-linked oligosaccharides, is at least one of the sugar determinants on ZP3 responsible for binding of sperm to the zona pellucida.
Abstract: During fertilization in mice, zona pellucida glycoprotein ZP3 mediates initial sperm-egg interactions by serving as receptor for sperm. Purified egg ZP3, as well as ZP3-derived O-linked oligosaccharides, exhibit sperm receptor activity in vitro. We report that treatment of purified egg ZP3 and ZP3-derived O-linked oligosaccharides with either alpha-galactosidase or galactose oxidase results in loss of sperm receptor activity. In the latter case, sperm receptor activity can be restored to the oxidized glycoprotein and O-linked oligosaccharides by treatment with sodium borohydride. We conclude that galactose, located in alpha-linkage at the nonreducing terminus of O-linked oligosaccharides, is at least one of the sugar determinants on ZP3 responsible for binding of sperm to the zona pellucida.
346 citations
TL;DR: The results of this study show that mature hamster oocytes contain significantly more glutathione (GSH), about 8 mM, and hence more disulfide reducing power, as compared with GV or pronuclear oocytes, which is the first evidence that modulation of GSH levels during oocyte maturation and fertilization may be a mechanism by which sperm nuclear decondensing activity is regulated.
342 citations
TL;DR: 100% effective contraception was obtained in male and female guinea pigs immunized with PH-20, and the contraceptive effect was long-lasting and reversible: immunized females, mated at intervals of six to fifteen months after immunization, progressively regained fertility.
Abstract: Immunization of male and female animals with extracts of whole sperm cells is known to cause infertility. Also, men and women who spontaneously produce antisperm antibodies are infertile but otherwise healthy. Although the critical sperm antigens are unknown, these observations have led to the proposal that sperm proteins might be useful in the development of a contraceptive vaccine. The guinea pig sperm surface protein PH-20 is essential in sperm adhesion to the extracellular coat (zona pellucida) of the egg, a necessary initial step in fertilization. Here, we report that 100% effective contraception was obtained in male and female guinea pigs immunized with PH-20. Antisera from immunized females had high titres, specifically recognized PH-20 in sperm extracts, and blocked sperm adhesion to the egg zona pellucida in vitro. The contraceptive effect was long-lasting and reversible: immunized females, mated at intervals of six to fifteen months after immunization, progressively regained fertility.
282 citations
TL;DR: The distribution of protamines in sperm obtained from a select group of infertile males producing an elevated level of large sperm heads, in contrast, was different from that of the fertile males.
Abstract: Protamines were extracted from the sperm of fertile and infertile human males and the relative proportion of protamines 1, 2, and 3 were determined by scanning microdensitometry following electrophoresis of total protamine in polyacrylamide gels. The proportion of the three protamines was found to be similar in sperm obtained from different normal males. The distribution of protamines in sperm obtained from a select group of infertile males producing an elevated level of large sperm heads, in contrast, was different from that of the fertile males.
279 citations
TL;DR: It is concluded that the human zona pellucida, or material intimately associated with it, can induce acrosome reactions in human sperm.
Abstract: We have used two approaches to test the ability of the human zona pellucida to induce acrosome reactions in human sperm. First, nonviable human oocytes were incubated for 1 min in a suspension of capacitated sperm (of which fewer than 5% were acrosome-reacted) to allow binding of about 200 sperm per oocyte. Some of the oocytes were fixed immediately, and the remainder were fixed after a further 1-h incubation without free-swimming sperm. As determined by light microscopy, sperm on the zona were only 3 +/- 2% (avg. +/- SD) acrosome-reacted at 1 min, and the incidence increased to 46 +/- 15% during the next hour. Electron microscopy confirmed that most sperm on the zona at 1 min were acrosome-intact. A few sperm were in an early stage of the acrosome reaction. Acrosome reactions occurring on the zona during the subsequent hour appeared to be morphologically normal. Second, treatment of sperm in suspension with acid-disaggregated zonae (2 to 4 zonae/microliter) increased the incidence of acrosome-reacted sperm from 3 +/- 1% to 24 +/- 4%. We conclude that the human zona pellucida, or material intimately associated with it, can induce acrosome reactions in human sperm.
TL;DR: Primate species with multi-male breeding systems produced ejaculates with relatively higher sperm motility and with relatively more motile sperm than did primates with single-male systems.
TL;DR: Multivariate analysis showed that the best prognostic indicator of fertility was given by the percentage of motile sperm and the Multiple Anomalies Index, particularly in patients with primary infertility.
Abstract: A prospective study of 394 infertile men was conducted over 3 years following a primary semen analysis. The cumulative pregnancy rate was 43 and 64% after 1 and 3 years, respectively. The pregnancy rate was significantly higher in the secondary infertile group. The study of various sperm factors and the occurrence of pregnancy showed that they were not of equal significance in predicting male fertility potential. The percentage of pregnancies decreased significantly only when the sperm concentration was less than 5 x 10(6)/ml. The pregnancy rate increased significantly with the percentage of motile sperm. The percentage of sperm with normal morphology was also found to be significantly higher when a pregnancy occurred than when the couple remained infertile (43.6% vs 37.7%). In a detailed morphological analysis of the sperm, six abnormalities (microcephaly, double head, amorphous head, cytoplasmic droplet, bent tail and coiled tail) were found to be significantly more frequent when a pregnancy did not occur. The most predictive value was given by the Multiple Anomalies Index (MAI), which is the mean number of abnormalities observed per abnormal sperm. The pregnancy rate was significantly lower after both 1 and 3 years when the MAI was greater than 1.6. Multivariate analysis showed that the best prognostic indicator of fertility was given by the percentage of motile sperm and the MAI, particularly in patients with primary infertility.
TL;DR: The primary finding is that the assay of chromatin structure stability performed on killed spermatozoa was as highly correlated with the heterospermic performance of semen as the best of the classical tests for semen quality.
Abstract: Data obtained by the sperm chromatin structure assay (SCSA) on spermatozoa from nine bulls were correlated with fertility, measured by heterospermic performance (-0.94, P less than 0.01) and by alternate tests of sperm quality, including motility, acrosome integrity, Sephadex filtration and morphology of spermatozoa (all significant at P less than 0.05 to P less than 0.01). The SCSA uses flow cytometry to determine the susceptibility of nuclear DNA to low pH-induced denaturation in situ as measured by the ratio of acridine orange binding to double- or single-stranded DNA. The error associated with multiple SCSA measurements was relatively low. The primary finding is that the assay of chromatin structure stability performed on killed spermatozoa was as highly correlated with the heterospermic performance of semen as the best of the classical tests for semen quality. The SCSA may therefore be a highly useful technique for evaluation of sperm quality.
TL;DR: It is shown that an influx of extracellular Ca2+ is responsible for initiating the acrosome reaction in human sperm treated with human follicular fluid, and this is the first demonstration in mammalian sperm that a potentially physiological stimulus can cause an increase in intracellularCa2+ concomitant with the acosome reaction.
Abstract: The role of Ca2+ in the human sperm acrosome reaction was investigated using the fluorescent calcium indicator fura-2. Previous experiments have shown that a Sephadex G-75 column fraction of human follicular fluid can stimulate the human sperm acrosome reaction [Suarez SS, Wolf DP, Meizel S (1986): Gamete Res 14:107-121]. Using fura-2, we demonstrated that this Sephadex G-75 fraction also stimulates a rapid, transient increase in intracellular free Ca2+. This Ca2+ transient is blocked either by chelation of extracellular calcium or by addition of the Ca2+ antagonist La3+. We have also been able to stimulate the acrosome reaction in human sperm without significant loss of motility, using the divalent cation ionophore ionomycin. Acrosome reactions stimulated by whole follicular fluid, the G-75 fraction, or ionomycin are all blocked by removal of extracellular Ca2+. These results strongly suggest that an influx of extracellular Ca2+ is responsible for initiating the acrosome reaction in human sperm treated with human follicular fluid. This is the first demonstration in mammalian sperm that a potentially physiological stimulus can cause an increase in intracellular Ca2+ concomitant with the acrosome reaction.
TL;DR: Oocytes from 5 of 11 patients microinjected with human spermatozoa demonstrated successful pronuclear formation and correlated with the incidence of pregnancy in these patients transferred with same-source oocytes inseminated by standard protocols, promoting the supposition of clinical feasibility of assisted fertilization by sperm microinjection.
TL;DR: It is suggested that calmodulin-dependent protein phosphatase plays a major role in the calcium-dependent regulation of flagellar motility and stimulated curvolinear velocity and lateral head amplitude and inhibited beat cross frequency suggesting a role for this enzyme in axonemal function.
Abstract: Preliminary data demonstrated that the inhibition of reactivated sperm motility by calcium was correlated with inhibited protein phosphorylation. The inhibition of phosphorylation by Ca2+ was found to be catalyzed by the calmodulin-dependent protein phosphatase (calcineurin). Sperm from dog, pig, and sea urchin contain both the Ca2+-binding B subunit of the enzyme (Mr 15,000) and the calmodulin-binding A subunit with an Mr of 63,000. The sperm A subunit is slightly higher in Mr than reported for other tissues. Inhibition of endogenous calmodulin-dependent protein phosphatase activity with a monospecific antibody revealed the presence of 14 phosphoprotein substrates in sperm for this enzyme. The enzyme was localized to both the flagellum and the postacrosomal region of the sperm head. The flagellar phosphatase activity was quantitatively extracted with 0.6 M KCl from isolated flagella from dog, pig, and sea urchin sperm. All salt-extractable phosphatase activity was inhibited with antibodies against the authentic enzyme. Preincubation of sperm models with the purified phosphatase stimulated curvolinear velocity and lateral head amplitude (important components of hyperactivated swimming patterns) and inhibited beat cross frequency suggesting a role for this enzyme in axonemal function. Our results suggest that calmodulin-dependent protein phosphatase plays a major role in the calcium-dependent regulation of flagellar motility.
TL;DR: The relationship between testes size, ejaculate quality, and sperm competition in birds was analysed using data collected in artificial insemination studies to hypothesized that ejaculatequality should be superior in species with intense sperm competition compared with other species.
Abstract: The relationship between testes size, ejaculate quality (volume, sperm concentration, number of sperm per ejaculate) and sperm competition in birds was analysed using data collected in artificial insemination studies. I hypothesized that ejaculate quality, because of natural selection, should be superior in species with intense sperm competition compared with other species. In regression analyses, testes weight increased with body weight, with an exponent less than one, and ejaculate volume increased with testes weight with an exponent not significantly different from one, whereas sperm number per ejaculate increased with testes weight with an exponent larger than one. Species with relatively large testes also produced ejaculates with a high sperm concentration. Monogamous species with a relatively low intensity of sperm competition copulate rarely, but deliver ejaculates with a relatively large number of sperm. Monogamous species with a high intensity of sperm competition copulate frequently, but produce ejaculates with a relatively small number of sperm. Males of polygynous species, which also experience intense sperm competition, copulate rarely with specific females, but produce many ejaculates per male each with a relatively small number of sperm.
TL;DR: It is concluded that patients with severe sperm head abnormalities have a lower ability to establish successful pregnancies, even though fertilization may be achieved.
TL;DR: The results suggest that spermatozoa of salmonid fish acquire the potential for motility as a result of the increase in seminal bicarbonate concentration and pH that occurs as spermutozoa pass from the testis to the sperm duct.
Abstract: Spermatozoa of rainbow trout and chum salmon, which have no potential for motility in the testis, acquire that potential in the sperm duct. This paper demonstrates that there is little difference between the levels of sodium, potassium, calcium, magnesium, chloride and osmolality of the seminal plasma in the testis and in the sperm duct. However, the bicarbonate concentration of the seminal plasma and the pH value of semen were higher in the sperm duct than in the testis. When immotile spermatozoa obtained from the testis were incubated in artificial seminal plasma with a high pH and containing HCO3-, spermatozoa became motile within 1 h. These results suggest that spermatozoa of salmonid fish acquire the potential for motility as a result of the increase in seminal bicarbonate concentration and pH that occurs as spermatozoa pass from the testis to the sperm duct.
TL;DR: In this article, the same six species of sperm nuclei were induced to undergo decondensation in vitro by treatment with DTT and detergent, and the resulting changes in nuclear size were monitored by phase-contrast microscopy and flow cytometry.
Abstract: To assess the structural stability of mammalian sperm nuclei and make interspecies comparisons, we microinjected sperm nuclei from six different species into hamster oocytes and monitored the occurrence of sperm nuclear decondensation and male pronucleus formation. The time course of sperm decondensation varied considerably by species: human and mouse sperm nuclei decondensed within 15 to 30 min of injection, and chinchilla and hamster sperm nuclei did so within 45 to 60 min, but bull and rat sperm nuclei remained intact over this same period of time. Male pronuclei formed in oocytes injected with human, mouse, chinchilla, and hamster sperm nuclei, but rarely in oocytes injected with bull or rat sperm nuclei. However, when bull sperm nuclei were pretreated with dithiothreitol (DTT) in vitro to reduce protamine disulfide bonds prior to microinjection, they subsequently decondensed and formed pronuclei in the hamster ooplasm. Condensed rat spermatid nuclei, which lack disulfide bonds, behaved similarly. The same six species of sperm nuclei were induced to undergo decondensation in vitro by treatment with DTT and detergent, and the resulting changes in nuclear size were monitored by phase-contrast microscopy and flow cytometry. As occurred in the oocyte, human sperm nuclei decondensed the fastest in vitro, followed shortly by chinchilla, mouse, and hamster and, after a lag, by rat and bull sperm nuclei. Thus species differences in sperm nuclear stability exist and appear to be related to the extent and/or efficiency of disulfide bonding in the sperm nuclei, a feature that may, in turn, be determined by the type(s) of sperm nuclear protamine(s) present.
TL;DR: It is demonstrated that hCG, in the absence of exogenous FSH, can complete spermiogenesis in men with partial gonadotropin deficiency, and the response to hCG inMen with IHH is predicted by the initial testicular volume.
Abstract: This study was designed to determine whether exogenous hCG alone can complete spermiogenesis in men with isolated hypogonadotropic hypogonadism (IHH). hCG was administered to 22 men with IHH until maximal testicular growth was achieved. Their mean testicular volume increased from 5.5 ± 1.1 (±se) mL (pretreatment) to 10.8 ± 1.6 mL (maximum) during treatment (P < 10−6). The maximum mean testicular volume was highly positively correlated with initial volume (r = 0.84; P < 10−6). All men attained normal serum testosterone levels, and 7 of 22 men achieved supraphysiological serum estradiol levels. During hCG treatment, 14 of the 22 men had sperm appear in their semen. Six of 11 men with complete gonadotropin deficiency, defined as an initial mean testicular volume less than 4 mL, became sperm positive during hCG treatment. In contrast, 9 of 11 men with partial gonadotropin deficiency (initial mean testicular volume of 4 mL or more) produced sperm during treatment (P < 0.001). Sperm concentration was highly pos...
TL;DR: Significantly more spermatozoa were recorded in the ooplasm of follicle oocytes than in that of cumulus oocytes, thus suggesting that the effect of the follicle on fertilizability may partly depend on an action on the plasma membrane of the oocyte.
Abstract: In order to investigate the effect of the somatic cells of the follicle on the preparation of the oocyte for fertilization three experiments were carried out. In the first, pig oocytes, cultured for 46 h in the presence of extroverted follicles (follicle oocytes) or surrounded by the cumulus (cumulus oocytes), were exposed to sperm in an in vitro fertilization system. Follicle oocytes showed a higher rate of fertilization than that recorded in cumulus oocytes (80% vs. 47%). In addition, significantly more sperm penetrated into the ooplasm of follicular oocytes (3.77/egg) than into that of cumulus oocytes (1.42/egg). To investigate the reason for the observed fertilization difference zona-free oocytes were studied in the second experiment. Significantly more spermatozoa were recorded in the ooplasm of follicle oocytes than in that of cumulus oocytes, thus suggesting that the effect of the follicle on fertilizability may partly depend on an action on the plasma membrane of the oocyte. A further effect of the follicular tissue was on cytoplasmic maturation: only follicular oocytes were capable of consistently promoting male pronucleus formation. In cumulus oocytes, sperm that entered the cytoplasm remained in a condensed form near the female pronucleus. In the third experiment cumulus oocytes and denuded oocytes were matured in medium that had been previously used to mature follicle oocytes. This conditioned medium was alone able to affect sperm penetration and male pronucleus formation in cumulus oocytes, but it did not exert any influence on denuded oocytes. This suggests that the observed effect of the follicle is mediated by soluble factors that, however, cannot influence the oocyte without some direct cell-oocyte contract.
TL;DR: The kinetics of change in the extent of hyperactivation and in acrosomal loss, although measured in different cell populations, are consistent with an association between these two events.
Abstract: The occurrence and time course of capacitation, acrosomal loss, and hyperactivated motility require quantitative definition in order to characterize fertile human sperm. In this study, video microscopy and digital image analysis were used to measure curvilinear (VCL) and straight line (VSL) velocity, average linearity of progression (UN [100 x VSL/VCUJ), maximum and mean amplitude of lateral head displacement (ALH), beat-cross-frequency (BCF), DANCE (VCL x meanALH) and DANCEMEAN (meanALH/(UIN/100)). These parameters were measured for sperm in semen and in the swim-up fraction of washed cells during incubation for up to 24 h under in vitro fertilization (IVF) conditions. Acrosomal loss was monitored in the same population of washed cells by an immunofluorescence end-point assay. The greatest increase in mean values of motility parameters was observed when seminal sperm were washed free of seminal plasma. Increases continued for up to 6 h of incubation. Two subpopulations of hyperactivated sperm were identified; one type, not found in semen, showed star-spin trajectories, and constituted 3.0, 3.8, 4.5, and 4.1% of the swim-up population after 0, 3, 6 and 24 h of incubation. The second type, termed transitional showed a more progressive trajectory and constituted less than 1% in semen. In total, hyperactivated cells constituted 0.8% of cells in semen, 14.5% of the swim-up population with no incubation, and 23.1, 22.7, and 19.4% after 3, 6, and 24 h of incubation, respectively. Acrosomal loss in the swim-up population was delayed during the first 3 h of incubation, then increased from near 5% at 3 h to 7 and 12% at 6 and 24 h, respectively. The kinetics of change in the extent of hyperactivation and in acrosomal loss, although measured in different cell populations, are consistent with an association between these two events.
TL;DR: The main emphasis in this chapter is on monogamy, extrapair copulations (EPCs), and associated behaviors, although sperm competition within other avian mating systems is also discussed.
Abstract: Publisher Summary The chapter discusses a review of sperm competition and behavioral adaptations to sperm competition in birds. Sperm competition is the competition between spermatozoa of different males to fertilize the eggs of a single female. The main emphasis in this chapter is on monogamy, extrapair copulations (EPCs), and associated behaviors, although sperm competition within other avian mating systems is also discussed. Four factors are known to influence the probability of extrapair paternity: (1) the timing and success of copulation— that is, sperm transfer by different males, (2) the relative numbers of copulations by different males, (3) the duration of sperm storage, and (4) sperm precedence. Extrapair paternity can be detected using genetic markers of two main types: those that are continuous variables (e.g., morphological characters) and those that are discontinuous variables (e.g., plumage color, enzyme polymorphisms). The chapter also discusses case studies on magpie ( pica pica ), common guillemot ( utria aalge ), swallow ( Hirundo rustica ), ringdove ( Barbary dove, Streptopelia risoria ), and indigo bunting ( Passerina cyanea ).
TL;DR: A test for human sperm binding to the zona pellucida (ZP) was developed using oocytes which failed to fertilize in vitro and should be useful in studies of human gamete interaction.
TL;DR: It is concluded that capacitation is an essential prerequisite for the induction of acrosomal exocytosis in bovine sperm by the zona pellucida.
TL;DR: The morphology of the human sperm AR, as initiated by human follicular fluid, is not unique, but is similar to that described for other mammalian sperm.
Abstract: It has been suggested that the morphology of the human sperm acrosome reaction (AR) is markedly different from that of other mammalian sperm. The present study examines the fine structural events of the early stages of the human sperm AR as initiated by preovulatory human follicular fluid. Human sperm, capacitated in vitro for 6 hr at 40 degrees C (90% motility) were diluted with equal volumes of follicular fluid before fixing in cacodylate-buffered glutaraldehyde at 5, 10, 15, 20, and 180 sec. Fixed sperm were treated with either tannic acid or thiocarbohydrazine and OsO4. Over 2,000 sperm were viewed. By 5 sec, 2% of the sperm had initiated the AR. By 15 sec, 8% of the sperm were in some stage of the reaction, and after 180 sec 40% of the sperm had completed the acrosome reaction. The initial stages of the human AR are characterized by a swelling or decondensation of the acrosomal matrix. The fusion between the plasma membrane and outer acrosomal membrane begins at the most anterior tip of the head and progresses toward the equatorial segment. Fusion and fenestration ended at the equatorial segment. With thiocarbohydrazine + OsO4 fixation the fused membranes are distinct hybrid vesicles with the outer acrosomal membrane being far more electron dense. The acrosomal matrix is retained by 20 sec, but by 180 sec the matrix is completely dispersed, even when viewed after tannic acid fixation. Also by 180 sec, vesicles were being progressively lost. It is therefore concluded that the morphology of the human sperm AR, as initiated by human follicular fluid, is not unique, but is similar to that described for other mammalian sperm.
TL;DR: The data suggest that avoiding glycerol toxicity either by reducing the concentration used or by adding Glycerol at a lower temperature, or both, may improve human sperm cryosurvival rates.