Oogenesis of Tilapia mossambica. I. Oogonia and meiotic prophase oocytes

Using light and electron microscopy and autoradiography, the morphology and synthesis of DNA, RNA and proteins in oogonia and early meiotic prophase oocytes in Tilaria mossabique were studied. According to dimensions and morphological features observed it is possible to distinguish between two groups of oogonia: large oogonia corresponding to type A spermatogonia of mammals, and small actively dividing oogonia, located in groups and identical to type B spermatogonia. The morphology of oogonia and of the early meiotic prophase oocytes well compares with the pattern described for other species of bony fishes. In the cytoplasm of these cells dense bodies, nuage-material, free ribosomes, large mitochondria with lamellar cristae and Golgi cisterns are available. In the oocyte nuclei at zygotene and pahytene stages 3H-thymidine incorporation was seen mainly into the nucleolus-associated chromatin. Besides, the formation of a heterochromatin cape and the synaptonemal complex was observed. Incorporation of 3H-uridine and 3H-leucine in the nuclei of these cells was very poor.     

Continuous loss of oocytes throughout meiotic prophase in the normal mouse ovary

The number of germ cells reaches the maximum just prior to entry into meiosis, yet decreases dramatically by a few days after birth in the female mouse, rat, and human. Previous studies have reported a major loss at the pachytene stage of meiotic prophase during fetal development, leading to the hypothesis that chromosomal pairing abnormalities may be a signal for oocyte death. However, the identification as well as the quantification of germ cells in these studies have been questioned. A recent study using Mouse Vasa Homologue (MVH) as a germ cell marker reached a contradictory conclusion claiming that oocyte loss occurs in the mouse only after birth. In the present study, we established a new method to quantify murine germ cells by using Germ Cell Nuclear Antigen-1 (GCNA-1) as a germ cell marker. Comparison of GCNA-1 and MVH immunolabeling revealed that the two markers identify the same population of germ cells. However, nuclear labeling of GCNA-1 was better suited for counting germ cells in histological sections as well as for double labeling with the antibody against synaptonemal complex (SC) proteins in chromosome spreading preparations. The latter experiment demonstrated that the majority of GCNA-1-labeled cells entered and progressed through meiotic prophase during fetal development. The number of GCNA-1-positive cells in the ovary was estimated by counting the labeled cells retained in chromosome spreading preparations and also in histological sections by using the ratio estimation method. Both methods demonstrated a continuous decline in the number of GCNA-1-labeled cells during fetal development when the oocytes progress through meiotic prophase. These observations suggest that multiple causes are responsible for oocyte elimination.     

Ultrastructural changes in female germ cells during the meiotic prophase in the rat: a special study of membranes after cryofracture

Ultrastructural changes in the nuclear and cytoplasmic elements in the germ cells of female rats were followed before meiotic prophase (15.50 days post-co?tum and 17.25 days post-co?tum) and during it (17.75 days post-co?tum to birth). We observed: modifications in the nuclear envelope which was thick during the oogonial stage, becoming thinner when the chromosomes entered preleptotene stage. The thinning of the envelope was due to the disappearance of the chromatin material lining it; variations in the number and distribution of germ cell nuclear pores according to stage; the pores were first scattered in small clusters of 6 to 8 over the entire nuclear membrane. From the preleptotene to zygotene stage, these clusters enriched in pores to form large areas. Finally, in the pachytene and diplotene stages, clusters of more than 100 pores were seen; nucleolar fragmentation from the preleptotene stage, followed by the formation of a new active nucleole in the diplotene; polarization of the mitochondria in the oldest oogonia just before the beginning of meiotic prophase. This polarization disappeared after the onset of the meiotic processes, then appeared again near the developing Golgi apparatus at the end of the pachytene stage; the formation of large gap junctions and numerous bands of tight junctions between the somatic cells; these formations contrasted with small gap junctions, and the tight junctions became scarce just before the meiotic process began. These observations, as well as those concerning nuclear pore distribution were made using the cryofracture technique.     

Apoptosis in mouse fetal and neonatal oocytes during meiotic prophase one

Background  

The vast majority of oocytes formed in the fetal ovary do not survive beyond birth. Possible reasons for their loss include the elimination of non-viable genetic constitutions arising through meiosis, however, the precise relationship between meiotic stages and prenatal apoptosis of oocytes remains elusive. We studied oocytes in mouse fetal and neonatal ovaries, 14.5–21 days post coitum, to examine the relationship between oocyte development and programmed cell death during meiotic prophase I.     

Heterochromatin and nucleolar organizers during first meiotic prophase in quail oocytes

The study of chromosomes in oocytes of the quail shows, at the pachytene stage, that microchromosomes are made of a euchromatic segment and a heterochromatic juxtacentromeric region. The heterochromatic regions of the microchromosomes amalgamate between themselves so as to constitute bulky chromocentres from which radiate the euchromatic segments which remain free. At late pachytene, nucleoli appear at the contact of these chromocentres. When the oocytes reach the diplotene stage, the nucleoli become quite large. They are stuck against chromocentres and establish a very close relationship with the euchromatic segments of the microchromosomes which surround or penetrate them. These observations lead one to think that the euchromatic segments of microchromosomes could be bearing nucleolar organizers. The close relations that the nucleolar organizers develop with the bulk of the nucleolus could explain its Feulgen-positive character in the quail.     

Ultrastructural studies of late meiotic prophase nuclei of spermatocytes in Ascaris suum

Post pachytene stages of meiotic prophase in males of Ascaris suum have been analyzed with the electron microscope. No synaptonemal-like polycomplexes (PCs) have been observed in the nucleoplasm or cytoplasm during the period from pachytene to diakinesis. From Serially sectioned diplotene nuclei it was found that the bivalents are located near the periphery of the nuclei, the central part of the nuclei being vacant. Each nucleus contains one nucleolus. Up to 1 m long stretches of unpaired lateral elements (LEs) are found in some of the diplotene bivalents. These LEs are morphologically similar to unpaired LEs in early zygotene nuclei. Partial 3-dimensional reconstruction of two nuclei shows that the bivalents contain some small stretches of synaptonemal complex (SC) up to 1.9 m long. Some bivalents at diakinesis show remnants of SCs. At this stage chromosomes are fibrous, condensed, attached to the nuclear envelope and mostly with a rounded profile in cross section. The synchronous development of the spermatocytes and small bivalents at diplotene in A. suum make this system a good object for the study of localization of SC remnants.     

Structural organization of the meiotic prophase chromatin in the rat testis

Pachytene nuclei were isolated from rat testes by the unit gravity sedimentation technique and contained histone variants H1a, H1t, TH2A, TH2B, and X2 in addition to the somatic histones H1bde, H1c, H2A, H2B, H3, and H4. The basic organization of the pachytene chromatin namely the nucleosome repeat length and the accessibility to micrococcal nuclease, was similar to that of rat liver interphase chromatin. However, when digested by DNase I, the susceptibility of pachytene chromatin was 25% more than liver chromatin under identical conditions. Nucleosome core particles were isolated from both liver and pachytene nuclei and were characterized for their DNA length and integrity of the nucleoprotein on low ionic strength nucleoprotein gels. While liver core particles contained all the somatic histones H2A, H2B, H3, and H4, in the pachytene core particles, histone variants TH2A, X2, and TH2B had replaced nearly 60% of the respective somatic histones. A comparison of the circular dichroism spectra obtained for pachytene and liver core particles indicated that the pachytene core particles were less compact than the liver core particles. Studies on the thermal denaturation properties of the two types of core particles revealed that the fraction of the pachytene core DNA melting at the premelting temperature region of 55-60 degrees C was significantly higher than that of the liver core DNA.     

Maintenance of meiotic prophase arrest in vertebrate oocytes by a Gs protein-mediated pathway

Maintenance of meiotic prophase arrest in fully grown vertebrate oocytes depends on an elevated level of cAMP in the oocyte. To investigate how the cAMP level is regulated, we examined whether the activity of an oocyte G protein of the family that stimulates adenylyl cyclase, Gs, is required to maintain meiotic arrest. Microinjection of a dominant negative form of Gs into Xenopus and mouse oocytes, or microinjection of an antibody that inhibits the Gs G protein into zebrafish oocytes, caused meiosis to resume. Together with previous studies, these results support the conclusion that Gs-regulated generation of cAMP by the oocyte is a common mechanism for maintaining meiotic prophase arrest in vertebrate oocytes.     

Patterns of RNA synthesis in early meiotic prophase oocytes from fetal mouse ovaries

In a study of the early meiotic prophase stages of mouse oogenesis from d12 of gestation to 10d post-partum the patterns of RNA synthesis during these stages of oogenesis using H³-uridine incorporation as visualized by light microscope autoradiography are reported. We find that chromosomal RNA synthesis occurs in all stages except early to mid-pachytene, the time of maximum chromosome condensation. Diplotene and dictyate nuclei are the most heavily labelled stages. Nucleolar labelling ceases before leptotene and reappears in late pachytene or early diplotene, even though nucleoli can be identified in all stages except early to mid-pachytene.     

Onset and progress of meiotic prophase in the oocytes in the B6.YTIR sex-reversed mouse ovary

When the Y chromosome of a Mus musculus domesticus male mouse (caught in Tirano, Italy) is placed on a C57BL/6J genetic background, approximately half of the XY (B6.YTIR) progeny develop into normal-appearing but infertile females. We have previously reported that the primary cause of infertility can be attributed to their oocytes. To identify the primary defect in the XY oocyte, we examined the onset and progress of meiotic prophase in the B6.YTIR fetal ovary. Using bromo-deoxyuridine incorporation and culture, we determined that the germ cells began to enter meiosis at the developmental ages and in numbers comparable to those in the control XX ovary. Furthermore, the meiotic prophase appeared to progress normally until the late zygotene stage. However, the oocytes that entered meiosis early in the XY ovary failed to complete the meiotic prophase. On the other hand, a considerable number of oocytes entered meiosis at late developmental stages and completed the meiotic prophase in the XY ovary. We propose that the timing of entry into meiosis and the XY chromosomal composition influence the survival of oocytes during meiotic prophase in the fetal ovary.     

Ultrastructural changes in the cell wall,nucleus and cytoplasm of pollen mother cells during meiotic prophase I inLycopersicon esculentum (Mill.)

Summary Throughout the premeiotic to late prophase I stages of meiosis in the anthers of tomato (Lycopersicon esculentum) extensive changes occurred in the ultrastructure of pollen mother cells (PMCs). During early prophase, the wall of each PMC developed a layered appearance and was broadened both by the widening of the middle lamella as well as by intensive deposition of microfibrils in the wall. By late prophase, however, the microfibrils adjacent to the plasmalemma dissipated. At the same time, callose was deposited between the wall and the plasmalemma. The nucleus of the PMCs also underwent changes. During early prophase, the nucleolus consisted of a linear series of three segments, with a separation of the granular and fibrillar portions. By late prophase, the nucleoli were less distinct as the nucleus was highly vacuolate. Mitochondria were initially simple with lightly stained matrix and few cristae but, during the course of prophase, they acquired a more densely-stained matrix with dilated cristae. Plastids remained relatively undifferentiated and, at late prophase, many were convoluted in appearance and constricted at intervals indicating their division. Cytoplasmic connections between adjacent PMCs were broad enough to permit the passage of organelles and were retained through to metaphase I. These cytological and wall changes appear to be a prerequisite for the subsequent development of microspores.Abbreviations PMC pollen mother cell - NOR nucleolus organizing region     

DNA reduplication during meiotic prophase in the oocytes of Carausius morosus Br. (Insecta,Cheleutoptera)

Cytological investigations combined with cytophotometric DNA determinations on Feulgen stained squash preparations of ovarioles from third and fourth instar nymphs of Carausius morosus revealed that during meiotic prophase a largely despiralized stage follows pachytene and that in this stage an extra reduplication of the nuclear DNA takes place (pachyreduplication phase). Meiotic prophase then proceeds with tetrapachytene, a pachytene-like stage with twice the amount of DNA as compared to oocyte nuclei in earlier meiotic stages, and with more than half the somatic number of elements, being probably autobivalents.Supported by Deutsche Forschungsgemeinschaft.     

Coordinating the events of the meiotic prophase

Meiosis is a specialized type of cell division leading to the production of gametes. During meiotic prophase I, homologous chromosomes interact with each other and form bivalents (pairs of homologous chromosomes). Three major meiotic processes--chromosome pairing, synapsis and recombination--are involved in the formation of bivalents. Many recent reports have uncovered complex networks of interactions between these processes. Chromosome pairing is largely dependent on the initiation and progression of recombination in fungi, mammals and plants, but not in Caenorhabditis elegans or Drosophila. Synapsis and recombination are also tightly linked. Understanding the coordination between chromosome pairing, synapsis and recombination lends insight into many poorly explained aspects of meiosis, such as the nature of chromosome homology recognition.     

Nucleolar structures in chromosome and SC preparations from human oocytes at first meiotic prophase

Summary We describe a comparative study of the behavior of nucleolar structures and their relationship with nucleolar chromosomes and synaptonemal complexes at first meiotic prophase of human oocytes in an attempt to elucidate the nature of this cellular organization and to learn more about maternal nondisjunction. The number of main nucleoli varies along the different stages of prophase I and is usually low. It shows an increase from leptotene to pachytene and a decrease from pachytene to diplotene related to a decrease and an increase of main nucleoli volume, respectively. The methodology employed has enabled us to analyze in detail dark bodies, round bodies, dense bodies, and main nucleoli in chromosome or synaptonemal complex spreads. The relationship between nucleolar chromosomes or synaptonemal complexes and the nucleoli implies the existence, in a very reduced space, of chromosomal regions that contain homologous sequences and that are often unpaired. This situation may facilitate the production of heterologous pairing and chromosomal exchanges between nonhomologous chromosomes and finally result in aneuploidy. THus, the situation explained above together with the differences between the oocyte and spermatocyte NOR cycles could be one of the reasons for the higher incidence of aneuploidies of maternal origin at meiosis I.     

Incorporation of H3-thymidine into the oocytes of a sterlet in the early meiotic prophase]

DNA synthesis in meiotic oocytes of the sterlet (A. ruthenus) has been studied during early prophase stages using H3-thymidine. The pattern of H3-thymidine incorporation is similar to that in oocytes of Amphibia and Osteichthyes. In the oogonia as well as in the leptotene and zygotene oocytes, the label is predominantly localized over chromosomes. An intensive incorporation of H3-thymidine into the material of the heterochromatic "cap" has been observed during pachytene. Thus, the main synthesis of extra DNA in the sterlet oocytes occurs during pachytene. No DNA in synthesized by the diplotene oocytes.     

Chromatin dynamics in the male meiotic prophase

During the male meiotic prophase in mouse and man, pairing and recombination of homologous chromosomes is accompanied by changes in chromatin structure. In this review, the dynamics of assembly and disassembly of the chromatin-associated complexes that mediate sister chromatid cohesion (cohesin) and maintain chromosome pairing (the synaptonemal complex) are described. Special features of the meiotic S phase are discussed, and also the dynamics of several key players that act together after the S phase at sites of meiotic double-strand break DNA repair. Current knowledge on histone modifications that occur during the male meiotic prophase is discussed, with special attention for the inactive chromatin of the X and Y chromosomes that constitutes the sex body. Finally, it is discussed that in the future, it will be possible to view the true chromatin dynamics during male meiosis in time, in living cells, through analysis of fluorescent-tagged proteins expressed in transgenic mice, using advanced fluorescent microscopy techniques.     

Autoradiographic study of RNA synthesis during meiotic prophase in the human oocyte

The incorporation of 3H-uridine in oogonia and oocytes during meiotic prophase I was studied in three human fetuses 13, 18, and 19 weeks old. Following a 40- or 60-min pulse, intense nuclear and nucleolar labeling was observed in oogonia. During the preleptotene chromosome condensation stage, the heteropycnotic masses were unlabeled, while numerous silver grains were seen on the filaments persisting around these masses. During leptotene, chromosomal and nucleolar RNA synthesis was significant, but less than that in the oogonia. The rate of incorporation declined rapidly during zygotene and fell to a very low level at early pachytene. Throughout pachytene no nucleolar RNA synthesis was observed. Chromosomal RNA synthesis progressively recovered during middle pachytene, was of moderate intensity at late pachytene, and increased again at early diplotene. Nucleolar RNA synthesis was very intense at early diplotene, at the same time as nucleolar size and basophilia increased.