bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genome organisation in the murine sperm nucleus
Authors: Jennings Carol , Powell Don 
Published: September 2008

SummaryThe organisation of DNA sequences in the murine sperm nucleus was studied usingin situhybridisation of biotinylated DNA probes. The efficiency of this reaction was assessed using a dispersed repetitive DNA probe. Telomeric DNA was distributed around the nucleus. Centromeric and ribosomal DNA sequences occupied restricted domains in the sperm nucleus. DNA sequences for a transgene and a cluster of homeogenes occupied different, and rather less defined, domains. Together these results imply that both repetitive and protein-coding sequences are arranged in the nucleus in an ordered fashion.
Long-lasting exocytosis and massive structural reorganisation in the egg periphery during cortical reaction inPlatynereis dumerilii(Annelida, Polychaeta)
Authors: Kluge B. , Lehmann-Greif M. , Fischer A. 
Published: September 2008

SummaryThe course of the cortical reaction in thePlatynereis dumeriliiegg is described from live observation and from sectioned fixed material and is found to differ in several aspects from the course of cortical reactions in better-known systems. Cortical granules are unusually numerous. They are discharged by exocytosis during a period of about 25 min following fertilisation (18°C). Most of the surplus membrane material brought to the egg surface by exocytosis is set free into the perivitelline space. Swelling of egg jelly precursor secreted by cortical granule exocytosis may be causal for the detachment of the vitelline envelope from the egg cell surface which, however, remains attached punctately to the vitelline envelope by about 30000 microvilli. Under the strain of the distending vitelline envelope, the bases of the microvilli move and line up, pulling the cell surface into a network of ridges. The grooves in between the ridges are the sites of exocytoses. Cytochalasin B, generally destabilising actin filaments, induces rupture of the microvilli and exaggerated distension of the vitelline envelope during the cortical reaction. In a final phase of the cortical reaction the vitelline envelope wrinkles and falls back onto the egg cell surface, the microvilli shorten and the egg cell transiently becomes deformed by local contractions. The cortical reaction in the nereid egg is discussed as a process of distortion and reorganisation of the egg cortex and plasmalemma. The abundance of cortical granules accommodating egg jelly precursor in thePlatynereisoocyte is attributed to the mode of so-called diffuse oogenesis characteristic of nereids, i.e. of differentiation of oocytes freely suspended in the coelomic fluid. In nereids, egg jelly therefore forms after fertilisation as opposed to ovulation.
The double spermatogenesis in scutigera coleptratrata (Myriapoda, Chilopoda): macro-and Microspermaltocyte growth
Authors: Pruescu Carol C. , Descamps Michel , Fabre Marie-Chantal , Serra Antoni 
Published: September 2008

SummaryIn Scutigera coleoptrata, double spermatogenesis (macro and micro germinal cells) was reinvesti
gted by means of both image analysis and electron microscopy. Image analysis of the cell-lines showed no differences in DNA content between the two types of spermatogesis.The only difference found, with the exception of the size and the related number of organelles, was that in the macro-cell line the chromatin was well dispersed and no condensed, which is the opposite of what was observed in the macro-cell line. It is asumed that the level of activity related to this condensation process is at the origin of the two pathways of development.
Calcium influx, fertilisation potential and egg activation inFucus serratus
Authors: Roberts Stephen , Brownlee Colin 
Published: September 2008

SummaryFertilisation in the marine algaFucus serratusis accompanied by increased influx of Ca2+from the external medium. The onset of this increase, monitored with the Mn2+fluorescence quench technique, corresponded with the depolarisation phase of the fertilisation potential. External Ca2+was necessary for the onset of the fertilisation potential and the early activation events, including cell wall exocytosis. Removal of Ca2+from, or addition of Sr2+to, the external medium during the fertilisation potential reduced the magnitude of the depolarisation and prolonged its duration. While fertilisation potentials could not be elicited in the presence of 0.1 mM Ca2+, addition of Ba2+in the presence of 0.1 mM Ca2+allowed normal fertilisation potential and egg activation. Microinjection of ryanodine or cyclic guanosine 5'-monophosphate (cGMP) did not induce cytoplasmic Ca2+elevation or egg activation. Inositol 1,4,5-triphosphate [Ins(l,4,5)P3] produced a transient elevation of cytoplasmic Ca2+, monitored using ratio photometry, but did not cause cell wall exocytosis except at the site of microinjection. The results demonstrate an essential role for Ca2+influx duringFucusegg activation. The relative importance of influx and intracellular Ca2+release inFucusegg activation is discussed.
Localisation of two classes of acetylcholine receptor-like molecules in sperms of different animal species
Authors: Baccetti B. , Burrini A.G. , Collodel G. , Falugi C. , Moretti E. , Piomboni P. 
Published: September 2008

SummaryThe distribution of different classes of acetylcholine (ACh) receptor-like molecules in sperms of different invertebrate and vertebrate species is described. ACh receptor molecules belong to one of two classes: muscarinic receptors (mAChRs), associated with signal transduction mechanisms in the inner domain of the cell, and nicotinic receptors (nAChRs), capable of opening Na+channels when activated by the ligand. Molecules immunologically related to mAChRs and to ACh can be identified by specific antibodies, and revealed by immunofluorescent or immunogold staining; the nicotinic receptor-like molecules are localised as curare-sensitive affinity sites for α-bungarotoxin. In all species studied, both classes of receptors were found, with a similar distribution. Muscarinic-like molecules were found mainly in the sperm head regions of most species; such a localisation may be correlated to a function in sperm–egg interaction, for instance in the regulation of the block to polyspermy. Nicotinic-like molecules are present mainly in the tail and in the post-acrosomal region of most animals, thus confirming their function in the regulation of sperm propulsion, but are also present at the acrosomal region of most species. The distribution patterns of the different classes of molecules indicate that both may be in sperm–egg interactions, in addition to their known function in the regulation of sperm propulsion.
Different calcium-dependent pathways control fertilisation-triggered glycoside release and the cortical contraction in ascidian eggs
Authors: McDougall Alex , Sardet Christian , Lambert Charles. C 
Published: September 2008

SummaryFertilisation of ascidian eggs induces the rapid release of a cell surfaceN-acetylglycosaminidase that blocks sperm binding to vitelline coat sperm receptors resulting in a block to polyspermy. Fertilisation also triggers a large contraction of the egg (thus stimulating ooplasmic segregation) that is completed within 5 min of insemination. In eggs of the ascidianPhallusia mammillata, glycosidase release and cortical contractions are blocked by BAPTA-AM [bis-(o-aminophenoxy-ethane-N,N,N',N' -tetraacetic acid, tetra(acetoxymethyl)-ester], a cell-permeant calcium chelator, indicating that both processes are probaly dependent on a rise in intracellular calcium levels. Both glycosidase release and the cortical contraction are induced by treatment of the egg with the protein synthesis inhibitor emetine, while only the glycosidase release is induced by isoproterenol, carbachol or acetylcholine. Previous work with ryanodine demonstrated that ryanodine also caused glycosidase release but not the cortical contraction Inversely, activation by ionomycin in calcium-free sea water causes cortical contractions but not glycosidase release. Thus the two processes can be activated independently. Dextran-coupled (10kDa) calcium green-1 injected eggs show an increase in intracellular calcium 30–40s before the cortical contraction is triggered by fertilisation or ionomycin- induced activation. This confirms previous findings that the cortical contraction is a consequence of the activation calcium the triggered by te sperm. The extracellular calcium requirement for the glycosidase release suggests that calcium influx may be more important for this phase of egg activation. Thus activation eggs appears to involve two independent pathways involving calcium.
Pronuclear formation and intracellular glutathione content ofin vitro-matured porcine oocytes followingin vitrofertilisation and/or electrical activation
Authors: Funahashi Hiroaki , Stumpf Todd T. , Cantley Thomas C. , Kim Nam-Hyung , Day Billy N. 
Published: September 2008

SummaryPronuclear formation and intracellular content of glutathione, containing reduced and oxidised forms, in porcine oocytes maturedin vitrowere determined following insemination and/or electrical stimulation. After insemination, sperm penetration had occurred as early as 3 h and female pronuclei had formed by 6 h with complete development by 12 h. Male pronuclear formation occurred, primarily, between 9 and 12 h after insemination. Glutathione content of the oocytes decreased following sperm penetration and remained at a depressed level until 12 h. After electrical stimulation, oocyte activation had occurred and female pronuclei had formed by 3 and 6 h, respectively. Oocyte glutathione content did not change as a result of oocyte activation. When oocytes were exposed to an electrical pulse and then spermatozoa, female pronuclear formation was observed by 3 h after stimulation/insemination. Sperm penetration was observed between 3 and 9 h. However, the incidence of male pronuclear formation observed at 12 h was extremely low, although sperm decondensation had occurred in some oocytes. Oocyte glutathione content had not decreased by 6 h following electrical activation. These results demonstrate that the changes in glutathione content in porcine oocytes following fertilisationin vitrodiffer from those due to electrical activation. Further, the decreased intracellular glutathione content in oocytes activated by sperm penetration appears to be due to the presence of a sperm factor.
Intracytoplasmic sperm injection in dysmorphic human oocytes
Authors: Alikani Mina , Palermo Gianpiero , Adler Alexis , Bertoil Massimo , Blake Marlena , Cohen Jacques 
Published: September 2008

SummaryFertilisation and development of dysmorphic human oocytes recovered from hyperstimulated ovaries have been evaluated following intracytoplasmic sperm injection (ICSI) for treatment of male infertility. A total of 2968 oocytes at metaphase II of meiosis were injected, of which 806 (27.2%) were dysmorphic at the light microscopic level. Cytoplasmic abnormalities included granularity, areas of necrosis, organelle clustering, vacuoles, and accumulating saccules of smooth endoplasmic reticulum. Anomalies of the first polar body and zona pellucida, as well as non-spherical shapes of oocytes, were also noted. Contrary to previous findings linking some dysmorphisms to non-assisted fertilisation failure, in this study no single abnormality led to a reduction in the fertilisation rate, nor was fertilisation compromised in oocytes with multiple abnormalities. The incidence of normal fertilisation (two pronuclei and two polar bodies) was 69% in both the dysmorphic and non-dysmorphic oocytes. While overall pregnancy and implantation results were not altered in the group of patients (n = 242) in whom at least one dysmorphic oocyte was injected, exclusive replacement of embryos which originated from dysmorphic oocytes led to a higher incidence of early pregnancy loss. It is concluded that aberrations in the morphology of human oocytes – most probably a product of controlled ovarian stimulation – are of little or no consequence to fertilisation or early cleavage after ICSI. It is possible, however, that these embryos have a reduced potential for implantation and further development.
Comparison of the acrosome reaction-inducing ability of the outer and inner surfaces of the zona pellucida and oolemma: a study using the golden hamster
Authors: Yazawa Hiroyuki , Radaelli Giuseppe , Yanagimachi Ryuzo 
Published: September 2008

SummaryThe outer surface of hamster zona pellucida has a strong ability to induce the sperm acrosome reaction, whereas neither the inner surface of the zona nor the oolemma has this ability. Therefore, the spermatozoa that fertilise oocytes after subzonal insemination of spermatozoa (SUZI) must have been either acrosome-reacted prior to injection or acrosome-reacted spontaneously while swimming within the perivitelline space. The outer surface of fresh mouse zona is not capable of inducing the acrosome reaction of hamster spermatozoa. When fixed with glutaraldehyde, however, it became capable of inducing the hamster sperm acrosome reaction rather efficiently. The reason for this is not clear.
Estimation by radiation inactivation of the minimum functional size of acrosome-reaction-including substance (ARIS) in the starfish,Asterias amurensis
Authors: Ushiyama Akira , Chiba Kazuyoshi , Shima Akihiro , Hoshi Motonori 
Published: September 2008

SummaryIn the starfishAsterias amurensis, the jelly coat of the eggs contains a glycoprotein essential for the induction of the acrosome reaction in homologous spermatozoa that is termed the acrosome-reaction-inducing substance (ARIS).ARIS is a highly sulphated and fucose-rich glycoprotein of extremely high molecular mass(>104kDa). ARIS was irradiated with high-energy eletrons in order to estimate the minimum size required for its biological activity. The minimum functional unit or target size of ARIS was estimated to bec. 14 kDa by target size analysis. ARIS was significantly disintegrated by the irradiation, yet the total sugar content was not apparently reduced. The binding of125I-labelled ARIS to spermatozoa competed with that of irradiated ARIS, although the affinity of ARIS was much reduced after irradiation.
The effect of 6-dimethylaminopurine (6-DMAP) on DNA synthesis in activated mammalian oocytes
Authors: Ledda S. , Loi P. , Bogliolo L. , Moor R.M. , Fulka J. 
Published: September 2008

SummaryMouse, sheep and bovine metaphase II oocytes were briefly preincubated in medium with 6-dimethyl-aminopurine (6-DMAP) and thereafter activated with ethanol before further culture in 6-DMAP supplemented medium. The presence of 6-DMAP enhances the efficiency of activation and the speed of pronuclear formation but has no effect on DNA synthesis. These results are discussed in the context of recently published data showing that the preincubation of condensed chromosomes with 6-DMAP blocks DNA synthesis when nuclei become reformed.
Protein synthesis is not required for male pronuclear formation in bovine zygotes
Authors: Chian R.C. , Sirard M.A. 
Published: September 2008

SummaryFollowing fertilisation, the sperm triggers a series of intracellular changes which initiate oocyte activation and pronuclear formation. Oocyte activation can also be induced artificially by several chemicals, such as the calcium ionophore A23187. The sperm nucleus is transformed into the male pronucleus through the interaction of oocyte cytoplasmic factors. The profile of protein synthesis is different in bovine oocytes following fertilisation and parthenogenetic activation. The formation of male and female pronuclei was not blocked by the presence of the protein synthesis inhibitor cycloheximide. These results suggest that bovine oocyte activation by sperm and parthenogenetic activation induce different cytoplasmic responses for protein synthesis and that new protein synthesis is not required for male pronuclear formation in bovine zygotes.
Bovine oocyte plasma membrane binding sites for sperm plasma membrane duringin vitrooocyte maturation and fertilisation
Authors: Wheeler Emily , Berger Trish , Behboodi Esmail 
Published: September 2008

SummaryThe experimental objective was to determine whether the capability of bovine oocyte plasma membrane to bind sperm changes duringin vitrooocyte maturation and fertilisation. Binding was quantified by the intensity of tetramethylrhodamine isothiocyanate (TRITC) fluorescence at the periphery of oocytes following incubation with biotinylated sperm plasma membrane proteins and subsequent incubation with TRITC-avidin. Bovine oocytes were maturedin vitro. Sample groups were removed after 0,6 and 22 h, or inseminated and further cultured for 24 or 48 h. Oocytes were denuded of cumulus cells and zona pellucida and co-incubated with 56 μg biotinylated bovine sperm plasma membrane protein for 45 min in 150 μl drops of saline-BSA. Controls were incubated for the same time period in the absence of sperm plasma membrane proteins. All oocytes were rinsed, incubated with TRITC-avidin and subsequently fixed and transferred to mounting medium. Oocytes were scanned with a confocal microscope and analysed using ImageQuant software. The binding of sperm plasma membrane was quantified by integrated fluorescent intensity in standardised ellipses spaced around the plasma membrane of the oocyte. Values are expressed as mean intensity units per 320 pixel ellipse. Binding of sperm plasma membrane continued to increase throughoutin vitrooocyte maturation and fertilisation (9051, 24318 and 49953 for 0 and 22 hin vitromatured oocytes and fertilised oocytes, respectively;p= 0.0001). A dramatic decrease in sperm plasma membrane binding to the oocyte plasma membrane was observed in 2-cell embryos (mean intensity = 24477,p= 0.0001). The observed binding was primarily due to the binding of sperm plasma membrane proteins, as control oocytes incubated with TRITC- avidin only were barely visible (integrated fluorescence intensity values ranged from 8 to 3757).
In vitrofertilisation of Chinese hamster oocytes by spermatozoa that have undergone ionophore A23187-induced acrosome reaction, and their subsequent development into blastocysts
Authors: Tateno Hiroyuki , Kamiguchi Yujiroh 
Published: September 2008

SummaryTo enhance potential use of the Chinese hamster,Cricetulus griseus, in developmental and cytogenetic studies of mammalian gametes and embryos, techniques forin vitrofertilisation and embryo culture were developed in the species. Spermatozoa were recovered from the vasa deferentia of mature males, and incubated in modified TYH medium for 1 h at 37°C under 5% CO2in air. They were then treated with ionophore A23187 (20¼M) for 10min to induce the acrosome reaction. Following ionophore treatment, superovulated oocytes were collected from hormonally stimulated females and incubated with the acrosome-reacted spermatozoa for 2 h at 37°C under 5% CO2in air. In this study, 245 oocytes ova (98.0%) were determined to be monospermic. The monospermic ova were then cultured in TYH supplemented with 1mM hypotaurine under the same gas phase. Within 30h of fertilisation, 182 ova (93.8%) cleaved to the 2-cell stage, and subsequently 163 ova (84.0%) developed beyond the 2-cell stage. Thus, obstinate developmental arrest at the 2-cell stage(‘2-cell block’) was not observed in this species. Ultimately, 65.5% of monospermic ova reached morula to blastocyst stages.
Cyclic AMP reversal of hypoxanthine-arrested preimplantation mouse embryos is EDTA-dependent
Authors: Dienhart Mary K. , Downs Stephen M. 
Published: September 2008

SummaryHypoxanthine can block preimplantation mouse embryo developmentin vitroat the 2- to 4-cell stages, and this has recently been shown to be reversed by cAMP-elevating agents. However, the extent of this hypoxanthine-induced arrest is determined by the culture conditions and strain of mouse. Whitten's and KSOM/AA are two embryo culture media that support preimplantation development to the blastocyst stage. This study was undertaken to examine the influence of several components in these media on hypoxanthine-arrested preimplantation mouse embryos and to test the hypothesis that reversal of the hypoxanthine block by cAMP-elevating agents requires cooperative interaction with the chelator, EDTA. Initial experiments demonstrated that embryo development was blocked in the presence of hypoxanthine in Whitten's medium but not in KSOM/AA; furthermore, removal of EDTA from KSOM/AA rendered this medium incapable of supporting high levels of development to blastocyst (9%), whereas high numbers of blastocysts (80%) formed in Whitten's medium, which does not contain the chelator. Consequently, Whitten's medium was used to test our hypothesis. It has previously been demonstrated that the phosphodiesterase inhibitor, IBMX, can reverse the developmental arrest imposed by hypoxanthine in EDTA-supplemented Earle's basic salt solution, but in the present study the addition of IBMX to Whitten's medium resulted in a block to development and failed to reverse the hypoxanthine arrest. These disparate effects can be explained by the presence or absence of EDTA. Supplementing Whitten's medium with EDTA reverses the IBMX effect, but not the hypoxanthine-induced block. While IBMX alone is unable to reverse the hypoxanthine block in Whitten's medium, development is greatly enhanced by the simultaneous addition of EDTA and IBMX. Similar results were obtained with the cAMP analogue, 8-AHA-cAMP. The data therefore support our hypothesis that the reversal of the hypoxanthine-induced arrest by cAMP-elevating agents is critically dependent on the presence of EDTA. We contrast this with the situation in mouse oocytes, where the hypoxanthine-induced meiotic arrest is not reversed by the addition of EDTA and/or cAMP-elevating agents.
DNA synthesis, microtubule and nuclear dynamics in porcine parthenotes
Authors: Liu Lin , Lee Caroline , Moor Robert M. 
Published: September 2008

SummaryParthenogenetically activated mammalian oocytes have been used in the past decade as cytoplasts, in an attempt to support the development of nuclear transplant embryos. The present experiments were undertaken to study the DNA synthesis and the organisation of microtubules, nuclear envelope and chromatin during the first cell cycle of electrically activated porcine oocytes (parthenotes) maturedin vitroby using immunocytochemistry and laser scanning confocal microscopy. The results showed that pronuclear-like (PN) formation began 4–5 h post-activation (hpa), whilst DNA synthesis as revealed by bromodeoxyuridine incorporation was initiated 5–6 hpa, with a maximum number of labelled oocytes (73%) around 11 hpa, and persisted in some parthenotes until 15–16 hpa. In the metaphase II (MII) oocytes, microtubules were detected only in the metaphase II spindle; no lamin A/C antigen was observed. Electrical DC pulses resulted in 91% of MII oocytes being activated and confocal microscopy Indicated that microtubules were assembled in the spindle first for the extrusion of a second polar body, and for the second time for division from one to two cells. Nuclear envelope, indicated by anti-lamin A/C stain, was formed around the time of PN formation and surrounded the nuclear chromatin of 1- and 2-cell parthenogenotea. These results demonstrate that the apparent normality in both DNA synthesis and dynamics of microtubules and nuclear envelope is involved with chromosomal organisation in the parthenotes. In addition, the use of electrically activated IVM oocytes for both nuclear transfer and parthenogenetic studies in pigs is discussed.