摘要:

AbstractIn mammalian fertilization, sperm bind to thezona pellucida, a glycoprotein matrix forming a shell surrounding the oocyte. Subsequently, one of the bound sperm penetrates thezonaand fertilizes the egg. The adhesion between sperm andzonais mediated by complementary receptor‐ligand pairs. Recent biochemical evidence has identified likely candidates for these molecules in the mouse. Biophysical studies have predicted that very few (possibly as few as one) bonds are needed to tether a motile sperm to thezona. We have used the data characterizing the putative receptors of the mouse sperm to predict the number of bonds they can form with thezonaligands. Our calculations indicate that few bonds probably form between the sperm andzonaduring the initial contact when the sperm is captured, supporting the hypothesis that fertilization depends on the action of a very few sperm‐zonabo

ISSN:0148-7280    

DOI:10.1002/mrd.1120240103

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractTesticular spermatozoa and sperm development in the archaeogastropodCalliotropis glyptusWatson (Trochoidae: Trochidae) are examined using transmission electron microscopy and formalin‐fixed tissues. During spermiogenesis, the acrosome, formed evidently through fusion of Golgi‐derived proacrosomal vesicles, becomes deeply embedded in the condensing spermatid nucleus. Two centrioles (proximal and distal), both showing triplet microtubular substructure, are present in spermatids—the distal centriole giving rise to the sperm tail and its associated rootlet. During formation of the basal invagination in the spermatid nucleus, centrioles, and rootlet move towards the nucleus and come to lie totally within the basal invagination. Mitochondria are initially positioned near the base of the nucleus but subsequently become laterally displaced. Morphology of the mature spermatozoon is modified from that of the classic primitive or ect‐aquasperm type by having 1) the acrosome embedded in the nucleus (the only known example within the Mollusca), 2) a deep basai invagination in the nucleus containing proximal and distal centrioles and an enveloping matrix (derived from the rootlet), 3) laterally displaced periaxonemal mitochondria, and 4) a tail extending from the basal invagination of the nucleus. Implantation of the acrosomal complex and centrioles within imaginations of the nucleus and lateral displacement of mitochondria effectively minimize the length of the sperm head and midpiece. Such modifications may be associated with motility demands, but this remains to be established. The unusual features ofC. glyptus spermatozoa, though easily derivable from ‘typical’ trochoid sperm architecture, may prove useful in delineating the genusCalliotropisor tracing its relationship to other genera within the trochid subfamily M

ISSN:0148-7280    

DOI:10.1002/mrd.1120240104

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractEffects of reduced glutathione (GSH), oxidized glutathione (GSSG), or glutathione reductasc (GR) supply were studied on the ability of hamster oocytes to be fertilized by human sperm. Zona‐free oocytes were pretreated with these compounds prior to sperm insemination. Oocyte pretreatment with high concentrations of GSH or GSSG (50 or 100 mM. 30 min) significantly increased the penetrated oocyte rate (PR). Polyspermy was not increased except when high concentrations of GSH (100 mM) were used. Incubation of oocytes with GR (1 or 10IU/ml) prior to sperm insemination induced increasing dose‐dependent PR. Polyspermy increased significantly with 10 mM GR in oocyte incubation medium. Oocyte incubation for 30 min with the sulfhydryl blocking agent iodoacetamide (1 mM) led to a drastic decrease in oocyte penetration and in polyspermy.Our results demonstrate an original way to increase the efficacy of human‐hamster heterospecific fertilization. Various hypotheses are discussed explaining these observations which open new investigations for heterospecific and homospecific in vitro fertiliz

ISSN:0148-7280    

DOI:10.1002/mrd.1120240105

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractHamster oocytes were frozen using a 1,2‐propanediol‐sucrose procedure, which resulted in over 90% survival. After thawing and zona removal the oocytes were compared with non‐frozen oocytes in a zona‐free hamster egg test employing spermatozoa from human semen donors and suspected infertility patients. Similar data were obtained, indicating that propancdiol‐sucrose frozen hamster eggs may be used in place of fresh eggs for convenience and to a void scheduling

ISSN:0148-7280    

DOI:10.1002/mrd.1120240106

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

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 i

ISSN:0148-7280    

DOI:10.1002/mrd.1120240107

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractRemoval of epididymal fluids from epididymal sperm suspension is an important step for the study of sperm motility, capacitation, and the acrosome reaction. The technique of washing should minimize damage to viable spermatozoa but at the same time efficiently remove debris, non‐sperm cells, and biological fluids. We examined sperm motility and fertilizability in vitro of rat epididymal spermatozoa after washing with Percoll continuous gradient. Nine milliliters (ml) of 50%N‐2‐hydroxyethylpiperazine‐N1‐2‐ethanesulfonicacid (HEPES) buffered Percoll solution was centrifuged at 20,000g for 45 minutes to form a continuous gradient. One hundred to 300μ of sperm suspension was loaded onto the surface of the gradient and centrifuged at 150gand 1,500gfor 10 minutes. Two main layers of spermatozoa were formed, one of high (lower layer) and one of low (upper layer) motility. At centrifugation 1,500g, the sperm density and motility in the lower layer were greater than at 150g.Spermatozoa from both layers at 150gand l,500gwere diluted with modified Krebs‐Ringer's bicarbonate solution (mKRB) and preincubated for 5 hours. Superovulated eggs collected from 21–25‐day‐old Wistar strain immature rats were introduced into the preincubated sperm suspension for insemination and fixed 5‐5.5 hours later for observation of fertilization. Spermatozoa from both layers, 150g and 1,500 g, showed the same fertilizability in vitro as control spermatozoa. From these results we conclude that Percoll gradients can be used for washing rat epididymal sperm for the study of sperm physiology i

ISSN:0148-7280    

DOI:10.1002/mrd.1120240108

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractIn previous studies we have shown differences in the function of morphologically normal and abnormal sperm by evaluating their flagellar movements and swimming trajectories. In this study we have compared the capability of morphologically normal and abnormal human sperm to undergo an acrosome reaction after incubation with human follicular fluid. Semen samples were studied from 6 research donors and 21 semen evaluation patients. All men had normal semen by clinical criteria. Semen was prepared either by a two‐step Percoll gradient centrifugation or the sperm werediluted, washed, and centrifuged three times. Sperm suspensions were incubated for 24 hours in a modified Tyrode's medium, containing 2.6% bovine serum albumin, prior to dilution with human follicuiar fluid. The percentage of acrosome reactions among viable sperm was assessed after 15 minutes using the supra vital Hoescht stain and fluoresceinated pea lectin. Sperm head size was measured with an ocular micrometer and normal values were defined as length 3–5 μm and width 2–3 μm. At least 25 viable normal sperm, and 25 viable abnormal sperm were analyzed for acrosome reactions on each slide. With Percoll separation the percentage of acrosome reactions (mean ± sem) for normal sperm was 38i ± 3% vs. 22 ± 2% for abnormal sperm (P<0.005). After washing, the comparable values were 12 ± 1 % vs. 5 ± 1 % (P<0.005). The incidence of spontaneous acrosome reactions (24 hours of incubation, no follicular fluid) was also higher for normal sperm than abnormal sperm (9 ± 1 % vs. 4 ± 1 %,P<0.01). These data demonstrate an association between normal sperm morphology and acro

ISSN:0148-7280    

DOI:10.1002/mrd.1120240109

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractHatching in vitro was studied following zona drilling of 507 two‐cell mouse embryos using three methods: 1) acidic Tyrode's (AT), 2) partial zona dissection (PZD) using a sharp micronecdle, and 3) zona chiseling (CH), using a large beveled needle. PZD and CH were performed while the embryos were kept in a sucrose/PBS solution. Hatching was compared to 191 umnicromanipulated controls. The incidences of cavitation and completion of hatching did not differ between groups, however more micromanipulated embryos (20–25%) hatched partially than controls (9%). The zona pellucida thinned in 59/59 (100%) control blastocysts during expansion, but in only 3/205 (2%) micromanipulated blastocysts. The hatching gap was wide in all control embryos, but smaller in 96/129 (75%) micromanipulated embryos. Partially hatched blastocysts with a ‘figure‐8’ shape were found in 59/129 (46%) micromanipulated embryos and in none of the 39 hatching controls. Hatching usually occurred a day earlier in micromanipulated embryos as 214/218 (98%) had started extruding on day 5 as compared to 20/59 (27%) control blastocysts. Fifty percent of 1‐day‐old human oocytes were fertilized following PZD and reinsemination and 15/31 (48%) were monospermic. Thirteen monospermic embryos cleaved, six compacted and four cavitatcd—of these, three extruded through the PZD incision upon expansion. The zonae did not thin and one blastocyst twinned spontaneously as it was caught between the thick ridges of the PZD hole. Results indicate that the hatching process is abnormal following zona drilling; more embryos start hatching, extrusion occurs earlier, and many become trapped which may lead to artificial twinning or the formation of trophob

ISSN:0148-7280    

DOI:10.1002/mrd.1120240110

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractIn a previous study, it was shown that cumulus cell‐enclosed germinal vesicle (GV)‐stage oocytes, isolated from pregnant mares' serum gonadotropin (PMSG)‐primed immature (22–24 day old) mice and that underwent spontaneous maturation in vitro, exhibited frequencies of embryonic development similar to oocytes stimulated to mature and ovulate in vivo by administration of gonadotropins [Schroeder AC, Eppig JJ, (1984) Dev Biol 102:493–497]. In the present study, the effect of the hormonal state of the oocyte donor on the capacity of in vitro matured oocytes to be fertilized and undergo pre‐ and post‐implantation development was explored further. Oocytes were isolated at the GV‐stage from the following groups of mice: 1) unprimed immature mice; 2) adult cycling mice; 3) unprimed Snell dwarf (dw) mice that have undetectable levels of growth hormone (GH), prolactin, and thyroid‐stimulating hormone (TSH); and 4) primed and unprimed hypogonadal (hpg) mice that have undetectable levels of circulating gonadotropins. Oocytes maturing in vitro after isolation from normal unprimed immature or adult mice at all stages of the estrous cycle acquired full developmental capacity. GV‐stage oocytes isolated from dwarf mice showed embryonic development equivalent to normal ( + /?) littermate controls. Therefore, GH, TSH, or prolactin are not required during oogencsis in vivo to promote the acquisition of competence to complete embryogenesis after maturation in vitro. Oocytes from hypogonadal mice had a much reduced capacity for preimplantation development when compared with normal littermates. Administration of PMSG to the hypogonadal mice significantly increased the developmental capacity of oocytes that underwent maturation in vitro. Gonadotropins, therefore, have a beneficial effect on the oocytc's capacity for em

ISSN:0148-7280    

DOI:10.1002/mrd.1120240111

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractWe created enlarged octaploid mouse blastomeres by subjecting four‐cell embryos to a large (>2.000 V/cm) dc field of brief duration (10 μsec). This electrofusion pulse caused three to four of the blastomcres to fuse in 60% of the embryos tested. Modifications of fusion chamber and medium enabled fusion of up to 20 embryos per pulse, greatly increasing the yield for this fusion method. The effectiveness of the electrofusion pulse depended upon such parameters as embryonic cell cycle time and the pH and temperature of the electrofusion medium. There was no discernable lag in the onset of the third cleavage division or the time of cavitation in fused blastomeres. These fused blastomeres also underwent polarization of their apical surfaces at the same time as controls in spite of their increased cell size. These results suggest that octaploid mouse blastomeres created via electrofusion divide normally through the blastocyst stage and polarize at the same time and in the same sequence as smaller control blastomeres. This suggests that the mechanisms underlying cell division, cavitation, and cortical polarization are not affected by changes in cellular size or ploi

ISSN:0148-7280    

DOI:10.1002/mrd.1120240112

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY