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R M Moor

Publications and source records attributed to R M Moor.

At least 109 records · Page 6Linked to original sources
PubMed1985

cAMP in ovine oocytes: localization of synthesis and its action on protein synthesis, phosphorylation, and meiosis.

In the first series of experiments, the source of cAMP in the sheep oocyte was studied. Cholera toxin was shown to be a potent stimulator of cAMP in isolated sheep oocytes, demonstrating the presence of adenyl cyclase. There was no evidence for transmission of cAMP from stimulated myocardial cell monolayers to cumulus-enclosed oocytes even though the existence of a concentration gradient of cAMP and of intercellular communication were demonstrated. However, gonadotrophin-stimulated follicle shells were able to induce a rise in the cAMP content of denuded or cumulus-enclosed oocytes in the same dish, independently of cell contact. Further experiments were designed to study the effects of a cholera toxin-stimulated rise in cAMP on the maturation of oocytes. When applied to cumulus-oocyte complexes, cholera toxin did not block germinal vesicle breakdown (GVBD), nor the accompanying changes in protein synthesis and phosphorylation, although there was evidence for a delaying effect. There were, however, indications that the toxin was inducing abnormalities that became gross when the concentration was raised to 1 microgram/ml. This high concentration of cholera toxin was able to block the maturation of oocytes in intact, gonadotrophin-treated follicles, although once again abnormalities were evident. We conclude that the role of cAMP in the maturation of the sheep oocyte is different from that proposed in the mouse.

Animals↗
PubMed1985

Gonadotrophin-induced abnormalities in sheep oocytes after superovulation.

The development of preovulatory follicles involves an initial phase of somatic cell differentiation and a final phase, initiated by the LH surge, when both the somatic and germinal compartments alter. Abnormalities in this pattern of compartmental development after superovulation have been identified by examining follicles from control, PMSG- and FSH-treated sheep. The pattern of proteins synthesized by oocytes from untreated sheep did not differ after culture of follicles in hormone-free medium from that of germinal vesicle oocytes in vivo. Similarly, 93.5% of oocytes from sheep injected with a pituitary gonadotrophin (FSH-P) synthesized the unchanged germinal vesicle pattern of proteins during culture in an hormonally neutral culture environment. By contrast, the administration of the placental gonadotrophin, PMSG, induced in 28% of oocytes changes in the pattern of synthesis which are normally associated with maturation. An examination of follicular steroidogenesis showed that both total output and particularly oestrogen secretion was over twice as high in follicles from PMSG-treated as compared with FSH-treated animals (P less than 0.01). We conclude that the compartmental pattern of development and steroidogenesis is grossly perturbed in many follicles from PMSG-treated animals. Premature activation of the germinal compartment results in aged or abnormal oocytes and a hostile reproductive tract.

Animals↗
PubMed1985

Temperature-induced abnormalities in sheep oocytes during maturation.

The susceptibility of sheep oocytes to temperature changes during maturation in vitro was tested by reducing the incubation temperature to 20 degrees C at various stages of meiosis. Cooling induced chromosomal abnormalities including disorganized metaphase plates and multipolar spindles in 28-54% of oocytes cooled at all stages of meiosis from germinal vesicle breakdown (GVBD) to metaphase II. The time of GVBD (8-11 h after the start of culture) was the most sensitive to cooling, whereas fewest abnormalities were found in oocytes cooled in late metaphase I (16-19 h). In addition to the chromosomal abnormalities, unusual vesicles appeared in the cytoplasm of oocytes cooled at 8-11 h and 12-15 h. No abnormalities in protein synthesis were detected by one-dimensional SDS gel electrophoresis. The consequences of the abnormalities for the developmental potential of the cooled oocytes were tested by transfer to recipient ewes and fertilization in vivo. After 12 days of development only 6% and 11% oocytes cooled at 12-15 h and 20-23 h respectively had developed to expanded blastocysts, compared with 44% of control oocytes. The results demonstrated that maturing sheep oocytes are very sensitive to a drop in temperature.

Animals↗
PubMed1984

Changes in protein phosphorylation during the maturation of mammalian oocytes in vitro.

Cumulus-enclosed sheep oocytes were cultured in gonadotrophin-containing medium for up to 9 hr and were then incubated for 3 hr in the presence of [32P]phosphate. The incorporation of 32P into TCA-insoluble material was measured, and oocyte proteins were separated by one- and two-dimensional gel electrophoresis. Incorporation of [32P]phosphate into protein increased after 3 hr culture and again after 9 hr, the time of germinal vesicle breakdown (GVBD). Qualitative and quantitative changes in the phosphorylation of proteins occurred over the 12-hr period studied. One of the most prominent changes was the appearance of a band of Mr 33,000, which was absent at 0-3 hr but appeared with increasing intensity with longer periods of culture. Two-dimensional electrophoresis revealed that the bulk of material in this band was a neutral polypeptide. No significant incorporation of [32P]phosphate was found in ribosomal extracts of oocytes.

Animals↗
PubMed1984

Protein synthesis and secretion by hamster, rat and mouse jejunum.

The synthesis and secretion of [35S]methionine-labelled proteins by hamster, rat and mouse jejunum has been measured in vitro after 60 min incubation in culture medium. Twenty-three major bands of radioactively labelled proteins were detected by PAGE analysis of enterocyte extracts. Three of these proteins having mol. wts of 28,000, 43,000 and 60,000 appeared to be synthesised preferentially by older enterocytes. A small number of radioactively labelled proteins also appeared in culture medium at the end of incubation. Three of these proteins, accounting for most of the recovered radioactivity, had mol. wts of 28,000, 43,000 and 60,000. Secretion of these proteins was inhibited by monensin. Further experiments showed these secreted proteins to be acidic and possibly glycoprotein in nature. Their rapid turnover, selective secretion and changing abundance in enterocytes of different ages all suggest that they may have important functions to perform in the intestine.

Age Factors↗
PubMed1983

Somatic control of protein synthesis in mammalian oocytes during maturation.

The extent to which signals from the follicular compartment regulate protein synthesis in sheep oocytes is considered. Evidence is derived from experiments on the control of actin synthesis and also from a quantitative analysis of protein synthesis in oocytes matured in follicles in which specific somatic signals have been altered. Actin represents about 10% of the total protein synthesized by oocytes enclosed by follicle cells but under 2% in denuded oocytes. Although follicle cells synthesize and release actin, no evidence of its uptake into oocytes has been obtained. Instead, the results suggest that regulatory signals of somatic origin are transmitted directly into the oocyte by intercellular transport. A variety of cell types in addition to those of follicular origin support actin synthesis in oocytes, provided only that functional communication between the somatic and germinal compartment is maintained. Finally, the results on precursor uptake, actin turnover and RNA inhibition, and those of Dawn Giebelhaus on actin mRNA measurements, indicate that the somatic signals act on post-transcriptional events, possibly by influencing the mobilization of stored actin message. The selective inhibition of 17 alpha-hydroxylase in follicles depresses oestrogen and androgen and enhances progesterone biosynthesis. These steroid alterations, if induced during the first 8 h of maturation, lead to developmental abnormalities at fertilization. In addition, these steroidogenic changes alter the synthesis of a small number of acidic proteins in the maturing oocyte. It is postulated that steroid signals regulate the synthesis of certain proteins associated specifically with maturation.

Actins↗
PubMed1983

Time-dependent effects of alpha-amanitin on nuclear maturation and protein synthesis in mammalian oocytes.

The addition of alpha-amanitin to extrafollicular, cumulus-enclosed ovine oocytes at explantation inhibits meiotic maturation and prevents many of the changes in protein synthesis that normally accompany maturation. By contrast, these inhibitory effects are considerably reduced by either delaying the addition of the drug for 1-4 h or by denuding the oocytes of all associated cumulus cells at the onset of culture. The observations that the inhibitory effect of cordycepin on nuclear maturation is also time-dependent and cumulus-cell-dependent and that the oocyte is susceptible to cordycepin for longer than its sensitivity to alpha-amanitin are consistent with the differential effects of these drugs on RNA synthesis. It is concluded that a transcriptional event at the onset of maturation is essential for the initiation of those changes in protein synthesis required for the regulation of nuclear and cytoplasmic maturation. It is uncertain, however, whether this transcriptional event occurs within the cumulus cells or within the oocyte.

Amanitins↗
PubMed1983

The role of steroid signals in the maturation of mammalian oocytes.

The resumption of meiosis in mammalian oocytes is associated with sequential changes in follicular steroidogenesis, the principal features of which are an initial stimulation and then suppression of the secretion of oestrogens and androgens, followed by a steady increase in the secretion of progesterone. The addition of steroid enzyme inhibitors to isolated follicles in vitro alters the normal profile of steroids secreted during maturation and induces intracellular changes in the oocytes which are expressed as nuclear abnormalities at fertilization. Different abnormalities are induced by selectively modifying steroid biosynthesis during maturation, the maturing oocyte being more sensitive to an imbalance in the steroid profile rather than the total inhibition of steroid secretion. The cause of the developmental aberrations may either result from or be associated with the abnormal patterns of proteins synthesized by oocytes denied the support of the correct balance or sequence of steroids during maturation.

Androgens↗
PubMed1982

Cell interactions and actin synthesis in mammalian oocytes.

In this study, we have examined the profiles of proteins synthesized by mammalian oocytes in the presence and absence of cumulus cells. The results show that the patterns of protein synthesis are broadly similar in cumulus-enclosed and denuded oocytes, but that as a consequence of removing the cumulus cells, the presence of a 45,000-dalton band is greatly reduced. This band, identified by comigration studies on two-dimensional polyacrylamide gels as actin, is strongly synthesized in both isolated cumulus cells and in oocytes labeled in the presence of cumulus cells, but is only weakly synthesized by oocytes incubated in the absence of cumulus cells. We suggest that the presence of newly synthesized actin in the oocyte is dependent upon cellular cooperation between the cumulus cells and oocytes.

Actins↗
PubMed1981

Cyclic AMP in mammalian follicle cells and oocytes during maturation.

Intact ovarian follicles and isolated oocytes were cultured for 30 sec to 18 hr in the presence of gonadotrophins, and the cyclic AMP content in the follicle and oocyte was measured. The basal content of cyclic AMP in ovine oocytes before gonadotrophic stimulation was 6.3 +/- 0.7 fmol/oocyte or 8 microM. There was no fall in oocyte cyclic AMP concentration as an immediate response to the gonadotrophins, but at 1, and 12-18 hr after stimulation, the concentration in both the oocytes and follicle cells was considerably elevated. There was no comparable increase in intracellular cyclic AMP in oocytes denuded of follicle cells before culture, even when both gonadotrophins and phosphodiesterase inhibitors were included in the medium. We conclude that the signal which initiates oocyte maturation in mammals differs from that of amphibia, where an early fall in intracellular cyclic AMP is essential for the resumption of meiosis. Moreover, cellular interactions within the mammalian follicle are necessary for the characteristic periods of increased cyclic AMP in oocytes during maturation.

Animals↗
PubMed1981

Follicle cell regulation of protein synthesis and developmental competence in sheep oocytes.

The developmental capacity of sheep oocytes cultured outside the follicle was greatly increased by the presence of high concentrations of gonadotrophins (10 micrograms/ml) in the medium. However, even under these conditions, the developmental capacity of the oocytes was only half that of oocytes cultured within the intact follicle. The presence of the cumulus was essential for development; nearly all denuded oocytes failed to undergo cleavage. Maturational changes in the oocyte involving increased amino acid uptake increased incorporation and specific changes in protein synthesis were inhibited by the follicle cells; this suppression was alleviated by gonadotrophic hormones. The cumulus cells suppressed amino acid incorporation and, to some extent, the changes in protein synthesis. However, the suppression of amino acid uptake required the presence of the whole follicle. Patterns of protein synthesis by oocytes cultured outside the follicle differed from those in oocytes cultured within the follicle, irrespective of the presence of the cumulus or gonadotrophins. Analysis of single oocytes cultured outside the follicle showed that the protein profiles varied markedly even under identical culture conditions.

Amino Acids↗
PubMed1981

Selective effect of gonadotrophins on cell coupling, nuclear maturation and protein synthesis in mammalian oocytes.

Individual gonadotrophic hormones were used to examine the degree to which changes in intercellular coupling between somatic and germ cells initiate meiotic maturation, regulate protein synthesis or alter the ultrastructure of the ovine oocyte. Follicle Stimulating Hormone (FSH; 50 ng ml-1) suppressed intercellular coupling to the same extent as that observed during oocyte maturation in vivo. At low concentrations FSH did not, however, initiate resumption of meiosis. By contrast, luteinising hormone (LH; 100-500 ng ml-1) invariably initiated meiosis in oocytes cultured within the follicle but did not disrupt intercellular coupling. We conclude that nuclear maturation is not dependent upon the disruption of cell contact between the oocyte and the surrounding follicle cells. The profile of proteins synthesized by untreated oocytes differed greatly from that of oocytes matured for 18 h in follicles treated with a combination of FSH and LH. Pretreatment of follicles with either FSH or LH at low concentrations resulted in the synthesis of an intermediate and more variable pattern of proteins. No correlation was found between changes in protein synthesis and the extent of junctional communication between the cumulus cells and oocyte. Membrane vesiculation and lysosomal change in the transzonal processes are early structural changes associated with the suppression of intercellular coupling in oocytes. These changes in coupling probably result in the relocation of intracellular organelles in the final stages of oocyte maturation.

Animals↗
PubMed1980

Fine structure of the sheep oocyte during antral follicle development.

The structure and distribution of organelles within oocytes of developing antral follicles were studied qualitatively and quantitatively. In the smallest category of follicle (0.2-0.4 mm diam.) the Golgi was present in a peripheral position, the endoplasmic reticulum was distended and mitochondria were intimately associated with it. Processes from surrounding cumulus cells were in contact with the oolemma which was thrown up into slender villi, increasing the oocyte surface area 5-fold. In these and all subsequent follicles the oocyte cytoplasm contained numerous 30 nm particles or vesicles. As the follicles grew to about 2.0 mm the mitochondria became located in a peripheral band, interior to which were numerous large vesicles, and the villi became shorter and thicker. In follicles > 2.0 mm the mitochondrial band was largely dispersed and cortical granules rested close to the oolemma. Little structural change was then observed until late oestrus. In oocytes from preoovulatory follicles at this stage an internal dense zone was formed in the zona pellucida. Most of the cumulus cell processes degenerated and most mitochondria assumed a hooded appearance.

Animals↗
PubMed1980

Effect of follicular steroids on the maturation and fertilization of mammalian oocytes.

Pronuclear development was used to measure the effects on ovine oocytes of altering follicular steroidogenesis during maturation in vitro. Follicular steroid secretion was altered using enzyme inhibitors and exogenous steroid supplementation. Abnormalities induced during maturation were measured 24 h after tranfer of oocytes to the oviducts of inseminated hosts. The presence throughout maturation of aminoglutethimide, an inhibitor of the conversion of cholesterol to pregnenolone, reduced steroid secretion to 7% of that in controls and decreased from 77% to 33% the number of normal oocytes. Abnormalities were substantially reduced by the addition of aminoglutethimide during the final 8 h of maturation only. The inhibition of 17 alpha-hydroxylase enzymes with SU10603 reduced oestrogen and testosterone secretion to about 10% of control levels but had no effect on progestin secretion. Only 13% of oocytes matured in the continual presence of SU10603 underwent normal fertilization. The number of oocytes undergoing normal fertilization was increased to about 50% by (i) delaying the addition of SU10603 until the last 8 h of the maturation period or (ii) adding exogenous steroids to follicles cultured with inhibitor from explantation. It is concluded that oocytes require a specific intra-follicular steroid environment for the completion of the full maturation process. Alterations to the steroid profile during maturation induce changes in the oocyte which are expressed as gross abnormalities at fertilization.

Aminoglutethimide↗