Georgiana Jagiello

Alterations of mammalian oocyte meiosis I with divalent cations and calmodulin

Experiments using a Ca2+/Mg2+, serum free media were carried out aimed at clarifying proposed effects of these divalent cations on in vitro meiotic maturation of mouse and cow oocytes. Agents known to perturb intracellular Ca2+ or calmodulin were also studied. Total absence of both cations restricts both oocyte species from completing meiosis I. Media containing Mg2+ and no Ca2+ permitted some maturation in both species. Absence or small amounts of Mg2+ in the media containing control amounts of Ca2+ was much more inhibitory for the cow than the mouse oocyte. Studies of mouse oocyte maturation with Verapamil, Epinephrine and A23187 demonstrated an inhibition of maturation perhaps by the intracellular Ca2+ changes these agents are alleged to induce. A dependency of mouse oocyte maturation on active Ca-Calmodulin complexes was suggested by the calmodulin inhibitor studies.

Further Studies Of Inhibitors Of In Vitro Mammalian Oocyte Maturation

Studies of luteinizing hormone (LH)-mediated serum inhibition and of follicular fluid and granulosa cell inhibition of in vitro porcine, ewe, and cow oocyte meiotic maturation have been carried out. A porcine serum inhibitor or inhibitors with a molecular weight above 15,000, similar to the ewe and cow serum inhibitor(s), have been demonstrated. An LH-inactivated inhibitor or inhibitors in fluid from medium-sized ewe and cow follicles have been shown and a minimally effective porcine follicular fluid inhibitor has been confirmed. The estimated molecular weight of these inhibitors is below 1,000. Granulosa cells from the three species were not found to be dramatically inhibitory. It is postulated that the follicular fluid inhibitor(s) may be derived from the larger serum molecule(s) by an action of LH within an intermediate cell type.

Quantitation of ribosomal RNA genes in fetal human oocyte nuclei using rRNA: DNA hybridization in situ: Evidence for increased multiplicity

Abstract Several experimental observations provide evidence for the existence of an amplification-type phenomenon involving the genes for ribosomal RNA (rRNA) during meiotic prophase in the human oocyte. In previous investigations multiple micronucleoli, in addition to primary nucleoli, were shown to be present in late pachytene and in diplotene human oocyte nuclei. In the present study, detailed quantitative analysis of grain counts from more than 1 000 cells following hybridization in situ indicated the presence of greater than the expected 4C number of ribosomal genes in the oocyte nuclei, most notably in these same stages. The extent of this increase over the expected 4C amount of DNA complementary to rRNA was approx. 2-fold in oocytes early in meiotic prophase and rose to approx. 4-fold in late pachytene and early diplotene oocytes. These results constitute the first evidence for the presence of extra rDNA in mammalian oocyte nuclei, the occurrence of which is clearly consistent with earlier cytological and ultrastructural observations.

SO2 and its metabolite: effects on mammalian egg chromosomes.

Abstract The ubiquity of sulfur dioxide as an air pollutant combined with reports of effects of the gas and its blood metabolite on DNA and chromosomes, prompted a study of possible mutagen action on mammalian oocytes. Utilizing in vitro and in vivo systems, mouse, ewe, and cow oocytes were examined for cytologic evidence of structural or numerical damage in meiosis. Fragmentation with rearrangement seen at M 1 and M 2 and anaphase lagging seen in vitro in each species are classically known to be etiologic in chromosome abnormalities and must be considered, especially in contaminated areas with foci of fetal loss involving ewe and cow.

The localization of rDNA in small, nucleolus-like structures in human diplotene oocyte nuclei

SummarySmall, nucleolus-like structures were demonstrated in the nuclei of human diplotene oocytes. At least some of these bodies were shown to be true micronucleoli by virtue of their ability to bind rRNA during RNA-DNA hybridization in situ.

Cytological observations of deletions in pachytene stages of oogenesis and spermatogenesis in the mouse

Cytological examination of pachytene stages of oogenesis and spermatogenesis revealed the presence of diagnostic loops in cells heterozygous for each one of six radiation induced lethal albino alleles in the mouse. Such loops were never found in any of the normal sibs or other controls. Such evidence supports the contention that all six of these mutations are deletions as indicated previously for five of them by complementation tests. An incidental finding was that of a pleiomorphic region on a medium sized pachytene bivalent in both sexes. This was not restricted to mutant cells but was also found in control cells. Whereas the pachytene deletion loops characterize lethal albino allele carriers, the observation of the pleiomorphic region reveals an apparently normal Cytological phenomenon in the mouse, the significance of which remains to be determined.

A Pachytene map of the mouse oocyte

A map of the mouse oocyte has been constructed utilizing early and mid/late pachytene stages. Each bivalent was clearly identified. At the mid/late stage 195 chromomeres corresponded directly to precursor chromomeres in the early stage, whilst 84 chromomeres were derived from two chromomeres from the early stage. The X and 13 bivalents were found to have a single chromomere in the early stage which later yielded two sites. There were approximately twice as many total chromomeres as mitotic bands, with good correspondence between pachytene chromomeres and major mitotic bands. Application of the use of mapping to a previously reported bivalent bearing a pleiomorphic region is noted.

Meiosis suppression by caffeine in female mice.

SummaryThe effects of caffeine on meiosis of mouse ova in vitro and in vivo have been studied. Concentrations of 200 μg/cc or more suppressed entry of ova into in vitro meiosis without production of any numerical or structural abnormalities at dose levels above or below that. Acute superovulation by the standard technique was prevented at a dose of 0.25 mg/g/body weight and examination of the intrafollicular ova revealed presistence of the germinal vesicle stage. Chronic administration of the same dose (0.25 mg/g/bw) throughout the period from weaning to puberty did not induce any cytologically detectable meiotic abnormalities in ova studied in vitro at the end of that period. It is suggested that the reduced fertility seen by some previous authors may have been due to the suppression of entry into meiosis of the ova with diminished numbers of ova being ovulated.

Colcemid effects on homologue pairing and crossing over during fetal mouse oogenesis.

Colcemid was administered to gestational day 13 female mice to test effects on homologue pairing, synapsis and recombination of fetal oogenesis. Pairing abnormalities were detected in pachytene oocytes by light and electron microscopy examination of bivalents and synaptonemal complexes. Reduction of total chiasmata per treated diplotene oocyte (22.74) compared to controls (31.07) was found.

Baboon late diplotene oocytes contain micronucleoli and a low level of extra rDNA templates

Abstract Oocytes from ovaries of near-term baboon ( Papio cynocephalus ) fetuses were predominantly in late diplotene of meiosis. The oocytes contained one or two large primary nucleoli and many additional small, nucleolus-like structures, some of which hybridized 125 I-18 S and -28 S rRNA in situ . These structures were morphologically identical to micronucleoli previously demonstrated in human oocytes during earlier stages of meiotic prophase. Quantitative grain count analysis following rRNA:DNA hybridization in situ suggested the presence of a low-level (approximately fourfold) excess of rDNA templates over the predicted 4C number.