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  • 1
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 35 (1993), S. 197-208 
    ISSN: 1040-452X
    Keywords: Capacitation ; Cell death ; Destabilization ; Fluo-3 ; Spermatozoa ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Boar spermatozoa loaded with the Ca2+ probe fluo-3 were incubated in various Tyrode's-based media similar to those used for in vitro fertilization (IVF), and samples were then analysed by two-colour flow cytometry; propidium iodide was included in the media to detect membrane-damaged (“dead”) cells.If media contained bicarbonate/CO2 (a component thought to promote capacitation), part of the live sperm population experienced a considerable influx of Ca2+ into both head and tail compartments. The percentage of responding cells reached a maximum after about 30 min, but both during and after this period there was also a steady increase in the number of dead cells. This bicarbonatemediated increase in cell death took place in the absence of external Ca2+. Evidence was obtained that the entry of propidium iodide was preceded by a change in permeability of the plasma membrane, detectable by leakage of carboxydichlorofluorescein, and it was therefore deduced that the Ca2+ influx detected by fluo-3 was due to destabilization of the plasma membrane. A similar response could be produced by both caffeine and papaverine (best known as phosphodiesterase inhibitors), but neither cyclic AMP nor activators of adenylate cyclase had any effect. There was no influence of substrate on the process, but, in comparison to poly(vinyl alcohol), serum albumin enhanced it.The precise relevance of this destabilization to capacitation is not yet clear, but it seems significant that the process is mediated or enhanced by components often specifically included in IVF media, and that different individual cells respond after different times. © 1993 Wiley-Liss, Inc.
    Additional Material: 10 Ill.
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  • 2
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Gamete Research 4 (1981), S. 407-432 
    ISSN: 0148-7280
    Keywords: acrosin ; acrosome reaction ; calcium ; hyaluronidase ; ionophore ; spermatozoa ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A system has been developed for inducing a calcium-dependent acrosome reaction in ram spermatozoa in vitro using the calcium ionophore A23187. The resultant reaction is accompanied by release of the acrosomal enzymes hyaluronidase and acrosin, but there is no release of the cytoplasmic enzyme glucose 6-phosphate isomerase. In any given cell, the visible acrosome reaction apparently takes place rapidly, but there is a variable delay before the reaction occurs. Under optimum conditions, about 90% of treated spermatozoa show an acrosome reaction within one hour.Preincubation of the spermatozoa with the proteinase inhibitors p-amino-benzamidine or p-nitrophenylguanidinobenzoate allows two stages of the reaction to be distinguished ultrastructurally, a membrane fusion stage followed by a dispersal of the acrosomal matrix. In the presence of the inhibitors, the first stage is delayed but is completed within 1 hour, whereas the second remains largely incomplete.In the presence of calcium, ionophore concentrations which induce an acrosome reaction abolish sperm motility rapidly and completely. However, by adding serum albumin shortly after addition of ionophore, motility can be preserved while the acrosome reaction occurs as usual; the motility pattern observed under these conditions is of the “whip-lash” or “activated” type.Although the motile ionophore-treated spermatozoa were unsuccessful at penetrating normal mature sheep oocytes in vitro, they were able to penetrate zona-free oocytes, after which swelling and decondensation of the sperm head took place.
    Additional Material: 15 Ill.
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  • 3
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Gamete Research 9 (1984), S. 425-440 
    ISSN: 0148-7280
    Keywords: acrosin ; acrosome reaction ; enzyme localization ; immunocytochemistry ; proacrosin ; spermatozoa ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Proacrosin and acrosin were localized immunocytochemically at the electron microscope level in ram spermatozoa undergoing an ionophore-induced acrosome reaction. Antigenicity was preserved after fixation with 0.5% w/v ethyl-(dimethylaminopropyl)-carbodimide, and an antibody preparation was used that reacted with all major forms of ram acrosin. All stages of the acrosome reaction could be observed in a single preparation. At the earliest stage, labeling was observed throughout the acrosomal contents, which were just beginning to disperse. As dispersal proceeded, labeling diminished, being associated only with visible remnants of the acrosomal matrix. By the time the acrosome had emptied, almost no labeling could be detected on the inner acrosomal membrane.The relationship between matrix dispersal and proacrosin activation was studied in isolated ram sperm heads. While proacrosin was prevented from activating, the acrosomal matrix remained compact; but as activation proceeded, the matrix decondensed and dispersed in close parallel. By the time proacrosin activation was complete, the acrosomal contents had almost entirely disappeared.We conclude that proacrosin is distributed throughout the acrosomal contents as an intrinsic constituent of the acrosomal matrix. During the acrosome reaction, proacrosin activation occurs, resulting directly in decondensation of the matrix. All the contents of the acrosome including acrosin disperse and, by the time the acrosome is empty and the acrosomal cap is lost, only occasional traces of acrosin remain on the inner acrosomal membrane. Since the acrosomal cap is normally lost during the earliest stages of zona penetration, acrosin's role in fertilization is unclear: it does not appear to be a zona lysin bound to the inner acrosomal membrane.
    Additional Material: 10 Ill.
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  • 4
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Gamete Research 2 (1979), S. 75-87 
    ISSN: 0148-7280
    Keywords: acrosin ; activation ; inhibitor ; proacrosin ; spermatozoa ; zymogen ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The sperm-specific proteinase acrosin (EC 3.4.21.10) is found in spermatozoa as a zymogen. We have looked for different forms of this zymogen in testicular, epididymal, and ejaculated spermatozoa from ram and have compared total sperm extracts made immediately after cell disruption with extracts made later from isolated sperm heads. We have concluded that the autoactivatable zymogen form, known generally as proacrosin, is the only form of acrosin within intact mature ram spermatozoa; no other zymogen form was detected, although lower levels of proacrosin were found in some samples of testicular spermatozoa. From studies of the activation process, it appears that ram proacrosin is truly autoactivatable; no evidence could be found for the involvement of any auxiliary enzyme. Estimations of the molecular weight of proacrosin using gel chromatography (60,000) and SDS-polyacrylamide gel electrophoresis (51,300) indicated that the zymogen is monomeric. Comparison with the molecular weight of ram acrosin (44,000 or 40,000, using the two respective methods) indicated that a single acrosin molecule is derived from each zymogen molecule. The sperm acrosin inhibitor (molecular weight 11,000 or 8,000) was present in testicular spermatozoa as well as in ejaculated spermatozoa; there was no evidence that it was produced as a result of zymogen activation.
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