bluuebunny/crossref_metadata_2025_split huggingface dataset:
Ultrastructural analysis of abnormalities in the morphology of the second meiotic spindle in ethanol‐induced parthenogenones
Authors: O'Neill G. T. , McDougall R. D. , Kaufman M. H. 
Published: February 2005

AbstractA high frequency of parthenogenetic activation occurs when ovulated mouse oocytes are briefly exposed to a dilute solution of ethanol in vitro. Cytogenetic analyses of parthenogenones at metaphase of the first cleavage division have confirmed that parthenogenetic activation, per se, does not increase the incidence of chromosome segregation errors during the completion of the second meiotic division. Ethanol‐induced activation, however, significantly increases the incidence of aneuploidy. The ultrastructural changes that occur in the morphology and organization of the second meiotic spindle apparatus in ethanol‐ and hyaluronidase‐activated oocytes is reported here. Abnormalities in the arrangement of microtubule arrays and chromosome position were principally observed in ethanol‐activated oocytes at anaphase and telophase of the second meiotic division, but were only rarely observed in hyaluronidase‐activated oocytes. It is proposed that the abnormalities in spindle morphology and chromosome displacement observed in ethanol‐activated oocytes represent the initial events that lead to chromosome segregation errors following exposure to this agent.
Cytochemical study of intracellular calcium in hamster spermatozoa during the acrosome reaction
Authors: Ruknudin A. 
Published: February 2005

AbstractCalcium was localized by a pyroantimonate technique in hamster spermatozoa during the acrosome reaction and pyroantimonate precipitates were observed in the anterior region of the acrosome. The calcium was also localized in the postacrosomal lamina of spermatozoa undergoing the acrosome reaction. Spermatozoa, incubated in capacitating medium containing verapamil, showed denser precipitates with an increase in concentration of this drug. Ionophore A23187 enhanced binding of calcium to the acrosomal region. The sodium channel inhibitor amiloride inhibited the acrosome reaction and the pyroantimonate precipitates were absent in these spermatozoa, whereas ionophore monensin enhanced the acrosome reaction. This suggests that the Na+/Ca++ antiporter may be responsible for intracellular Ca++ regulation during the acrosome reaction in hamster spermatozoa.
Proteins of the acrosomal region in mouse sperm: Immunological probes reveal post‐testicular modifications
Authors: Lakoski Katherine , Williams Carmen , Saling Patricia 
Published: February 2005

AbstractDue to the central role the acrosomal region plays in sperm‐egg interactions, monoclonal antibodies (mAbs) were used to identify components of this domain in mouse sperm. Sev eral sperm proteins that localize specifically to the anterior acrosomal region are described here in terms of electrophoretic mobility, susceptibility to proteolytic degradation, and post‐translational modification during epididymal transit. Six different mAbs were used, each recognizing a distinctive antigen (Ag) or set of Ags in cauda epididymal mouse sperm: a doublet of 185/200 Kd (M42 mAb); 150‐160 Kd (M5 mAb); 105 Kd (W71 mAb); 21, 35, and 60 Kd (M41 mAb); 27 and 33 Kd (W33 mAb); and 57 and 86 Kd (W108 mAb). Previously reported work implicates two of these, M42 Ag and M5 Ag, as participants in sperm‐zona interaction (Saling and Lakoski: Biol Reprod 33:527‐536, 1985; Saling: Dev Biol 117:511‐519, 1986; and Lakoski et al.: Biol Reprod 38:221‐233, 1988).Recognition of some (M42, M5, W108), but not all (W33), of the Ags by their corre sponding mAbs was affected by sperm incubation with proteases (trypsin or collagenase). Evidence of post‐translational modification during epididymal maturation was suggested by altered electrophoretic mobility of several of the Ags (M42, M5, W33, and W108) accompanying sperm transit from proximal to distal epididymis. Retention of sperm within the caput epididymis prevented structural alterations for the four proteins exam ined, indicating that spatial rather than temporal factors are critical for Ag modification in maturing mouse sperm.
Identification, isolation, and properties of a plasma membrane protein involved in the adhesion of boar sperm to the porcine zona pellucida
Authors: Peterson R. N. , Hunt W. P. 
Published: February 2005

AbstractBoar sperm plasma membranes contain an integral protein (Mr 55 kDa) that apparently functions in the adhesion of sperm to the zona pellucida (Peterson and Hunt: J Cell Biol 105:170a, 1987.) In experiments described in this report, the protein is identified after additional steps of purification involving lectin affinity chromatography and preparative PAGE. An active form of the adhesion protein (APz) develops or becomes first exposed in the corpus epididymis and is fully active in the cauda epididymis; a significant portion of this conformationally labile protein, while integral to the plasma membrane, cannot be solubilized by nonionic detergents and may be associated with the membrane skeleton. APz does not exhibit enzymatic properties thought possibly to be involved in sperm‐zona interaction in this and other species. Galactosyltransferase substrates and inhibitors and anliproteases including soybean trypsin inhibitor, pepstatin, leupeptin, and p‐aminoben‐zamidine failed to block sperm from binding to porcine eggs. Boar sperm proacrosin and antiproacrosin antibody failed to inhibit sperm‐egg binding. When plasma membranes or fractions containing APz that bind to dextran sulfate agarose were chromatographed on L‐fucose agarose, a sugar which binds proacrosin, plasma membrane proteins that bound to the column failed to absorb anti‐APz antibody, Anti‐APz was absorbed by fractions that did not contain proacrosin. These data indicate that APz is not proacrosin. Since anti‐APz monovalcnt antibody raised from whole cauda or corpus sperm plasma membranes or from chromatographic fractions containing APz completely block capacitated sperm from binding to eggs, and since the ability of this antibody to be absorbed develops as sperm become capable of binding to eggs, we view AP, to be the major and perhaps only plasma membrane protein involved in the adhesion of capacitated boar sperm to eggs prior to the acrosome reaction.
A consistently successful procedure for in vitro fertilization of golden hamster eggs
Authors: Bavister Barry D. 
Published: February 2005

AbstractComplete details are described for the first time of the procedures used in the author's laboratory for obtaining in vitro fertilization (IVF) of golden hamster eggs leading to the first cleavage division. These IVF procedures have been developed during the past 20 years and are very reproducible: IVF of at least 75% of eggs is routinely achieved, and on average 65% of inseminated eggs undergo the first cleavage division in vitro. These results can easily be obtained by inexperienced investigators. The ease and reproducibility of the hamster IVF procedures make them very suitable for studies of sperm:egg interaction and associated events. Studies in the author's laboratory have included analysis of sperm fertilizing ability under chemically defined conditions, the presence of sperm acrosome reaction stimulating factors in the egg investments, maturation of oocytes in vitro, the block to polyspermy, and the contribution of egg aging to fertilization anomalies. In addition, the motility of hamster sperm under chemically defined conditions is used in a routine screening protocol for detecting contaminants in the culture milieu. Golden hamster gametes of Ter several distinct advantages for IVF studies, including the large size of the sperm acrosome, the persistence of the very large sperm tail in the ooplasm for many hours following fertilization, and the translucence of the ooplasm, which facilitates observation of the sperm tail and pronuclei. The female golden hamster exhibits a regular 4 day estrous cycle, with distinctive indications of estrus and postestrus phases. Because of the advantages of using the golden hamster, the procedures described in this report may be useful to other investigators wishing to conduct research using IVF. Essentially the same IVF procedures can be used with monkey and bovine gametes.
Capacitation of hamster spermatozoa with the divalent cation chelators D‐penicillamine, L‐histidine, and L‐cysteine in a protein‐free culture medium
Authors: Andrews Jane C. , Bavister Barry D. 
Published: February 2005

AbstractBy using a chemically defined (protein‐free) culture medium that supports sperm viability but not capacitation or the acrosome reaction, we have determined that hamster spermatozoa can be chemically capacitated in vitro by the divalent cation chelators D‐penicilla‐mine, L‐histidine, and L‐cysteine in the absence of bovine serum albumin (BSA). Washed cauda epididymal spermatozoa were preincubated (1–2 × 106 sperm/ml) for 3, 4, or 6 hr at 37°C in 5% CO2 in air. The basic culture medium used for sperm preincubation and for sperm:egg coincubation was a modified Tyrode's solution (protein‐free) containing 10 mM sodium lactate, 100 μM sodium pyruvate, and 1.0 mg/ml polyvinylalcohol (TLP‐PVA). Sperm viability was maintained in all preincubation and coincubation media with PHE (20 μM D‐penicillamine, 100 μM hypotaurine, and 1.0 μM epinephrine). The low control sperm preincubation medium consisted of TLP‐PVA. In some cases the high control preincubation medium also contained 3 mg/ml BSA (TALP‐PVA). The experimental preincubation medium was TLP‐PVA with additional D‐penicillamine (125 or 500 μM), or L‐histidine (10, 100, or 1,000 μM) or L‐cysteine (25, 75, or 125 μM). After preincubation, sperm were coincubated (2 × 104 sperm/ml) with cumulus‐free hamster eggs in TALP‐PVA ± additional D‐penicillamine, L‐histidine, or L‐cysteine for 1.5 hr, fixed, and evaluated for percent egg penetration as an index of sperm capacitation. The results demonstrate that hamster spermatozoa can be chemically capacitated in vitro with D‐penicillamine (500 μM: range of mean penetration values, 53.6%–84.3%), L‐histidine (100 μM: range of mean values, 24.8%–56.3%) or L‐cysteine (75 μM: 51.3%) in the absence of exogenous protein.
Morphogenesis of the decapitated and decaudated sperm defect in two brothers
Authors: Baccetti B. , Burrini A. G. , Collodel G. , Magnano A. R. , Piomboni P. , Renieri T. , Sensini C. 
Published: February 2005

AbstractThe “decapitated sperm” defect, found in both of two sterile brothers, may be assumed to have a genetic origin. The present material suggests that the term “decapitated spermatozoa” is not exact, because detached heads and tails were found in the brothers' ejaculate that could be regarded as “decapitated tails” and “decaudated heads.” The present report describes frequent, more or less advanced stages of detachment. Both heads and tails showed a normal structure in which only the postnuclear region was deficient, lacking basal plate and implantation fossa. A break at a different level of the midpiece, and therefore three kinds of separation, were observed. The defect, according to the present research, must originate in the testicular region, whereas the detachment occurs in the epididymis.
Fertilization and development of mouse eggs injected under the zona pellucida with single spermatozoa treated to induce the acrosome reaction
Authors: Lacham Orly , Trounson Alan , Holden Carol , Mann Jeff , Sathananthan Henry 
Published: February 2005

AbstractMouse spermatozoa were capacitated and acrosome reacted by incubation for 30 min to 6 h in modified Tyrode's medium (T6) and in cGMP. The fertilization rates of eggs microin‐jected with a spermatozoon from samples incubated for 30 min, 2 h, and 6 h in T6 and in cGMP were 36%, 34%, 29%, and 43%, respectively. These rates were not correlated to the percentage of acrosome‐reacted spermatozoa identified after these treatments. The lack of association could be explained by the selection of an increased proportion of acrosomc‐reacted spermatozoa actually used for microinjection, particularly in samples incubated for 30 min and 2 h in T6. Both acrosome‐intact and acrosome‐reacted spermatozoa were identified under the zona in the eggs which failed to fertilize. Fertilized eggs developed at very high rates to the blastocyst stage in vitro.
Maintenance of meiotic arrest in mouse oocytes by purines: Modulation of cAMP levels and cAMP phosphodiesterase activity
Authors: Downs Stephen M. , Daniel Susan A. J. , Bornslaeger Elayne A. , Hoppe Peter C. , Eppig John J. 
Published: February 2005

AbstractHypoxanthine and adcnosine are present in preparations of mouse ovarian follicular fluid, and these purines maintain mouse oocytes in meiotic arrest in vitro (Eppig et al.: Biology of Reproduction 33:1041–1049, 1985). The first hypothesis tested in this study is that purines which maintain meiotic arrest act by maintaining meiosis‐arresting levels of cyclic adenosine monophosphatc (cAMP) in the oocyte. Oocyte‐cumulus cell complexes were incubated in control medium (no added purines), or medium containing 0.75 mM adenosine, 4 mM hypoxanthine, or both for 3 hr and the percentage of the oocytes that underwent germinal vesicle breakdown (GVB) and the cAMP content of the intact complexes and the oocytes were determined. Adenosine alone had little inhibitory effect on GVB at this time point but sustained higher levels of cAMP in the oocytes. Hypoxanthine maintained 80% of cumulus cell‐enclosed oocytes in meiotic arrest and also sustained higher cAMP levels in the oocytes. The additon of adenosine to hypoxanthine‐containing medium increased the percentage of oocytes maintained in meiotic arrest, and increased the amount of cAMP in the oocytes above that maintained by either hypoxanthine or adenosine alone. Neither hypoxanthine, adenosine, nor hypoxanthine plus adenosine altered the cAMP content of intact complexes when assayed after 3 hr culture. Microinjection of an inhibitor of the catalytic subunit of cAMP‐dcpendent protein kinase induced GVB in denuded oocytes cultured in medium containing hypoxanthine. This purne, therefore, maintained meiotic arrest by sustaining elevated cAMP levels within the oocytes.The second hypothesis tested in this study is that purines maintain meiosis‐arresting levels of cAMP, at least in part, by inhibiting cAMP phosphodiestcrase activity. In descending order of potency, 3‐isobutyl‐l‐methylxanthine (IBMX), guanosine, hypoxanthine, adenosine, and xanthosine inhibited cAMP phosphodiesterase in oocyte lysates. Moreover, like the potent phosphodiesterase inhibitor IBMX, hypoxanthine augmented the cAMP meiotic arrest and cAMP accumulation mediated by follicle‐stimulating hormone (FSH) in intact complexes. Therefore, inhibition of oocyte phosphodiesterase appears to be one mechanism by which the purines could maintain meiosis‐arresting levels of cAMP.
Effects of semen preservation on boar spermatozoa head membranes
Authors: Buhr M. M. , Canvin A. T. , Bailey J. L. 
Published: February 2005

AbstractHead plasma membranes were isolated from the sperm‐rich fraction of boar semen and from sperm‐rich semen that had been subjected to three commercial preservation processes: Ex tended for fresh insemination (extended), prepared for freezing but not frozen (cooled), and stored frozen for 3‐5 weeks (frozen‐thawed). Fluorescence polarization was used to determine fluidity of the membranes of all samples for 160 min at 25°C and also for membranes from the sperm‐rich and extended semen during cooling and reheating (25 to 5 to 40°C, 0.4°C/min). Head plasma membranes from extended semen were initially more fluid than from other sources (P < 0.05). Fluidity of head membranes from all sources decreased at 25°C, but the rate of decrease was significantly lower for membranes from cooled and lower again for membranes from frozen‐thawed semen. Cooling to 5°C reduced the rate of fluidity change for plasma membranes from the spernvrich fraction, while heating over 30°C caused a signifi cantly greater decrease. The presence of Ca++ (10 mM) lowered the fluidity of the head plasma membranes from sperm‐rich and extended semen over time at 25°C but did not affect the membranes from the cooled or frozen‐thawed semen. The change in head plasma membrane fluidity at 25°C may reflect the dynamic nature of spermatozoa membranes prior to fertilization. Extenders, preservation processes and temperature changes have a strong influence on head plasma membrane fluidity and therefore the molecular organization of this membrane.
Ultrastructural and histochemical changes in the murine zona pellucida during the final stages of oocyte maturation prior to ovulation
Authors: Kaufman M. H. , Fowler R. E. , Barratt E. , McDougall R. D. 
Published: February 2005

AbstractThe ultrastructural morphology of the mouse zona pellucida was studied in preovulatory follicles from the ovaries of immature mice treated with exogenous gonadotrophins. The ovaries were fixed in the presence of cetylpyridinium chloride, which precipitates carbohydrates, so that their loss during fixation and processing is substantially reduced. The semi‐thin araldite sections obtained from osmicated material were viewed by conventional light microscopy, while the ultra‐thin sections were examined by transmission electron microscopy. A parallel series of semi‐thin sections of non‐osmicated ovaries was deresined and subsequently stained with periodic acid Schiff (PAS). The morphological appearance of the zona pellucida in preovulatory oocytes changed during the final stages of oocyte maturation. A close correlation was observed between the ultrastructural appearance of the zona pellucida and that observed following PAS staining during the period studied. Real differences were observed in the location, density, and distribution of glycoconjugates within the zona pellucida during the final stages of oocyte maturation prior to and immediately following germinal vesicle breakdown. Similar changes in the zona were observed in adult females autopsied during proestrus and oestrus. The changes in the density and distribution of glycoconjugates are likely to have important consequences for fertilization by affecting sperm‐zona binding and sperm‐egg interactions.
Catalase activity in human spermatozoa and seminal plasma
Authors: Jeulin C. , Soufir J. C. , Weber P. , Laval‐Martin D. , Calvayrac R. 
Published: February 2005

AbstractCatalase activity was determined in human semen by measuring the oxygen burst with a Clark electrode, after H2O2 addition. Significant catalase activities (mean ± SD) were found in migrated, motile spermatozoa (44 ± 17 nmoles O2/min/108 cells) and in seminal plasma of normozoospermic men (129 ± 59 nmoles O2/min/ml). It has been demonstrated that seminal catalase originated from prostate; however, its activity was not correlated with the usual prostatic markers (such as citric acid and zinc). Our data suggest a multiglandular function secreted by this organ. The catalase activities measured in seminal samples from asthenozo‐ospermic, infertile men were found lower than those from normozoospermic subjects. The understanding of the relative contribution of the different enzyme systems against O2 toxicity (superoxide dismutase, catalase, glutathione peroxidase) seem to be a priority area of research to understand disturbances of sperm function.
Morphology of immature bovine oocytes
Authors: de Loos F. , van Vliet C. , van Maurik P. , Kruip Th. A. M. 
Published: February 2005

AbstractBovine cumulus oocyte complexes (COCs) as used for in vitro maturation and fertilization can be classified into different categories by light microscopical inspection. We have distinguished four categories based on compactness and transparency of the cumulus investment and homogeneity and transparency of the ooplasm. The four categories were studied for their morphological characteristics at the ultrastructural level and for their developing capacity in an in vitro maturation system. In categories 1 and 2 oocytes, organelles were evenly distributed. In categories 3 and 4, oocytes organelles were clustered and the distribution of the organelles mimicked the characteristics of oocytes during final maturation. Cumulus cell process endings penetrated the cortex of the oocyte or were located superficial to the cortex of the oocyte. In category 1 oocytes, most of the process endings penetrated the cortex. In category 4 oocytes, most of the process endings did not penetrate. In categories 2 and 3 oocytes, both forms of process endings did occur.After in vitro maturation, only category 4 oocytes showed a decreased developing capacity. Categories 1–3 oocytes showed equal developing capacity in an in vitro maturation system.
Parthenogenetic activation of unfertilized mouse oocytes by exposure to 1,2‐propanediol is influenced by temperature, oocyte age, and cumulus removal
Authors: Shaw J. M. , Trounson A. O. 
Published: February 2005

AbstractCumulus‐intact and ‐denuded unfertilized oocytes from two mouse strains were exposed to 1.5 m ethanol (EtOH) or two cryoproteclant solutions, 1.5 M propanediol (PROH) or 1.5 M dimethylsulfoxide (DMSO), for 4.5 min at 27°C, and the proportion of activating or degenerating oocytes studied. Exposure to DMSO did not significantly increase activation above that of oocytes not exposed to DMSO. Treatment of oocytes in PROH resulted in the activation of up to 87% of viable oocytes. This was significantly higher (P <01) than in control oocytes and comparable to the rate of activation after treatment with EtOH (59–96% activation). In solutions at 1°C, 47% of control oocytes were activated, which was not significantly different from the rate of activation in EtOH (36%) or PROH (50%) at 1°C. Following treatment with PROH, up to 87% of oocytes degenerated within a period of 6 h in vitro. The age of the oocytes (h post hCG) and the time of cumulus removal with the enzyme hyaluronidase, relative to the time of exposure to the chemicals, influenced the level of degeneration in most groups. Significantly fewer oocytes degenerated when cumulus cells were removed before treatment (0–31%) than when the cumulus was left intact throughout the treatment and 6 h culture period (10–87%). Exposure to PROH at 1°C reduced oocyte degeneration to 5%. We conclude that PROH causes significantly greater losses of oocytes as a result of parthenogenetic activation and degeneration than of exposure to DMSO.
In Vitro maturation and fertilization of domestic cat follicular oocytes
Authors: Johnston L. A. , O'Brien S. J. , Wildt D. E. 
Published: February 2005

AbstractThe time course and conditions necessary for oocyte maturation and subsequent fertilization in vitro were studied in the domestic cat. Darkly pigmented oocytes surrounded by cumulus cells and a tight corona radiata were collected from ovaries removed at ovariohysterectomy. After culture in Eagle's minimum essential medium, oocytes were evaluated for nuclear maturation by analyzing chromosomal spreads. Oocytes achieved metaphase II after intervals of 40–48 hr of in vitro incubation. The incidence of maturation was enhanced (P<0.05) when oocytes were recovered from inactive (54%) or follicular (56%) stage donors compared to those recovered from luteal phase (29%) or pregnant (35%) cats. The proportion of oocytes successfully maturing in vitro in medium containing no hormone supplementation (37%) was less (P<0.01) than counterparts cultured in follicle‐stimulating hormone (FSH) only (48%) or FSH and luteinizing hormone (LH) (54%). The efficiency of maturation was not influenced (P >0.05) by either maintenance/transport temperature (4°C vs. 22°C) or delaying recovery of oocytes from antral follicles (2–8 hr vs. 24–32 hr). Approximately 36% of the in vitro matured oocytes cocultured with spermatozoa demonstrated evidence of fertilization; however, there appeared to be a critical development period for maximizing the incidence of fertilization. These results demonstrate that domestic cat antral oocytes are capable of maturing in vitro, and maturation is influenced by the reproductive status of the donor and the presence of gonadotropins in the culture medium. These oocytes are capable of forming embryos and developing to at least the 16‐cell stage in vitro.