Differentiation of somatic cells in the ovariuteri of the apoikogenic scorpion Euscorpius italicus (Chelicerata,Scorpiones, Euscorpiidae)

In apoikogenic scorpions, growing oocytes protrude from the gonad (ovariuterus) and develop in follicles exposed to the mesosomal (i.e. hemocoelic) cavity. During subsequent stages of oogenesis (previtellogenesis and vitellogenesis), the follicles are connected to the gonad surface by prominent somatic stalks. The aim of our study was to analyze the origin, structure and functioning of somatic cells accompanying protruding oocytes. We show that these cells differentiate into two morphologically distinct subpopulations: the follicular cells and stalk cells. The follicular cells gather on the hemocoelic (i.e. facing the hemocoel) surface of the oocyte, where they constitute a cuboidal epithelium. The arrangement of the follicular cells on the oocyte surface is not uniform; moreover, the actin cytoskeleton of these cells undergoes significant modifications during oocyte growth. During initial stages of the stalk formation the stalk cells elongate and form F-actin rich cytoplasmic processes by which the stalk cells are tightly connected to each other. Additionally, the stalk cells develop microvilli directed towards the growing oocyte. Our findings indicate that the follicular cells covering hemocoelic surfaces of the oocyte and the stalk cells represent two distinct subpopulations of epithelial cells, which differ in morphology, behavior and function.     

Surface glycoproteins bearing alpha-GalNAc terminated chains accompany pyriform cell differentiation in lizards.

The present investigation demonstrates that in squamate reptiles, as already reported for Podarcis sicula (Andreuccetti et al., 2001), the differentiation of pyriform cells from small, stem follicle cells is characterized by the progressive appearance on the cell surface of glycoproteins bearing alpha-GalNAc terminated O-linked side chains. Using a lectin panel (WGA, GSI-A4, GSI-B4, PSA UEA-I, PNA, Con-A, DBA, LCA, BPA, SBA), we demonstrated that, during previtellogenesis, the pattern of distribution of DBA binding sites over the follicular epithelium dramatically changes. In fact, binding sites first appear in follicular epithelium at the time that small cells begin to differentiate; in such follicles, labeling is evident on the cell surfaces of small and intermediate cells. Later on, as the differentiation progresses, the binding sites also become evident on the cell surface of pyriform cells. Once differentiated, the pattern of the distribution of DBA binding sites over the follicular epithelium does not change. By contrast, during the phase of intermediate and pyriform cell regression, DBA binding sites gradually decrease, so that the monolayered follicular epithelium of vitellogenic follicles, constituted only by small cells, shows no binding sites for DBA. It is noteworthy that binding sites for DBA are present on small cells located in contact with the oocyte membrane, but not on those located under the basal lamina or among pyriform cells, and therefore not engaged in the differentiation into pyriform cells. This finding demonstrates that, in squamates, the pattern of distribution of alpha-N-GalNAc containing glycoproteins significantly changes during previtellogenesis, and that these modifications are probably related to the differentiation of small stem cells into highly specialized pyriforms.     

Structural and functional dynamics of oogenesis in Glossina austeni: general features, previtellogenesis and nurse cells.

Glossina austeni oogenesis throughout its nine-day pregnancy cycle is described with the focus on previtellogenic stages. The ultrastructural details of the oocyte-nurse cell relationship and cyst formation is presented. The oocyte develops in a syncytial association with 15 nurse cells with the entire unit surrounded by a follicular epithelium. The nurse cells have large elaborate nucleoli. Evidence of nuclear emissions and the presence of an unusual cytoplasmic membrane association were found. A variety of nuclear inclusions are seen in the oocyte. Glycogen, lipid, ribosomes and membrane organelles accumulate in the oocyte during previtellogenesis.     

Dynamics of the oocyte cortical cytoskeleton during oogenesis in Rhodnius prolixus

The oocyte cortex undergoes dramatic changes during oogenesis in Rhodnius prolixus. Despite numerous studies examining oogenesis in the telotrophic ovariole, none has investigated the ultrastructural details of the oocyte cortex, in particular, the lateral cortical cytoskeleton. Indirect immunofluorescent staining of sections, rhodamine phalloidin staining of whole mounts and scanning and transmission EM of permeabilized and unpermeabilized preparations revealed the dynamic changes of the oocyte cortex from early previtellogenesis through to late vitellogenesis. During early previtellogenesis, oocytes 50-150 mum in length have a smooth oolemma, with no discernible cortical cytoskeleton. During mid to late previtellogenesis (oocytes 150-350 mum in length) a tightly woven network of microfilaments and microtubules forms, excluding mitochondria and Golgi complexes from the lateral cortex. At the onset of vitellogenesis, the follicuiar epithelium becomes patent, and there is an increase in microvilli covering the lateral oocyte surface. The microfilament cores form a discrete pattern that corresponds to the imprint of the follicle cells on the oocyte surface. While the lateral microfilament cytoskeleton becomes more elaborate, the lateral microtubule cytoskeleton diminishes, remaining sparse throughout vitellogenesis. The oocyte cortical cytoskeleton undergoes dramatic changes during oogenesis. These cortical dynamics are intricately related to the cellular and molecular processes that occur during oogenesis.     

Early oogenesis in the ascidian Ciona savignyi

Summary

Morphology of the germinal epithelium and the early follicular oocyte in the ascidian Ciona savignyi as examined by electron microscopy. The oogenetic part of the germinal epithelium contains oocytes at two different stages and the dark and clear cells. The smaller oocyte contains synaptonemal complexes. The larger oocyte in the initial phase of growth has a conspicuous nucleolus, electron-dense materials and some mitochondria close to the nuclear envelope. The nucleus of the larger oocyte is round and has the smooth contour. The dark cell contains a relatively large nucleus and is sometimes connected to each other by an intercellular bridge. Therefore, the dark cell, which has been suggested to be the progenitor cell of two kinds of accessory cells, may be also the oogonium. The early follicular oocyte just after migration from the germinal epithelium retains most of cytological features similar to those of the larger oocyte. However, the nuclear contour of the early follicular oocyte is uneven. This difference in the nuclear contour probably indicates that such a follicular oocyte is in the second phase of growth.     

Structure of ovaries and oogenesis in the snow scorpionfly boreus hyemalis (LINNE)(MECOPTERA : BOREIDAE)

The ovaries of the snow scorpionfly, Boreus hyemalis (Mecoptera : Boreidae) are panoistic and comprise 7–8 ovarioles. Each ovariole consists of a terminal filament, elongated vitellarium, and ovariole stalk (=pedicel) only ; in adult specimens, functional germaria are absent. Five consecutive stages of oogenesis i.e., early, mid- and late previtellogenesis, vitellogenesis, and choriogenesis have been distinguished in imagines. Oocyte nuclei (=germinal vesicles) of previtellogenic oocytes contain numerous polymorphic multiple nucleoli (or nucleolar masses), endobodies, and chromatin aggregations. Next to the nuclear envelope, large accumulations of nuage material are localized. The ooplasm of late previtellogenic oocytes is differentiated into transparent (perinuclear) and opaque (peripheral) regions. Ultrastructural investigations have revealed that within the latter, abundant ribosomes as well as mitochondria, elements of endoplasmic reticulum, Golgi complexes, annulate lamellae, symbiotic bacteroids, lipid droplets and distinctive accumulations of membrane-free clathrin-like cages are present. Early- and mid previtellogenic oocytes are invested with flat somatic cells that gradually transform into a follicular epithelium. In the vicinity of 3-cell junctions, neighbouring follicular cells are joined by narrow intercellular bridges. During late previtellogenesis, numerous microvilli develop on the oocyte surface. They interdigitate with morphologically similar but less frequent microvilli of the follicular cells. Concurrently, first endocytotic vesicles appear in the cortical ooplasm. In the context of presented results, the phylogenetic relationships between mecopterans (boreids) and fleas are discussed.     

Ultrastructural and quantitative dynamics of the granulosa of ovarian follicles of the lizard Gerrhonotus coeruleus (family Anguidae)

The progression of ovarian follicular development in the Northern Alligator Lizard has been documented ultrastructurally and by enumeration of cells, with a focus on changes in the granulosa component of the follicle. The pattern of cellular differentiation of the granulosa entails, as in other lizards, the transformation of a simple, cuboidal epithelium in small follicles into a complex layer consisting of three types of cells. Marked differences in size and ultrastructure of the cell types indicate different functional states: the smallest cells are little differentiated and serve primarily as stem cells to other granulosa cells throughout follicular growth, whereas the larger "intermediate" and "pyriform" cells do not divide and show ultrastructural features indicative of synthetic activity. Contrary to some views that this latter cell type is the final step in cellular differentiation and provides organelles and cytoplasm to the oocyte through an intercellular bridge, the results of this study suggest that only relatively small molecules such as ribosomal RNA might pass between cells. Further, these observations support the interpretation that a heterogeneous granulosa results from the fusion in early follicular stages of some cells that are in surface contact with the oocyte. Several of the cytological features of the larger granulosa cell types are seen in the oocyte and in germ-line cells generally, such as highly dispersed chromatin, large nucleoli, abundant nuclear pores, mitochondrial "rosettes," annulate lamellae, "ribosome bodies," and surface microvilli. This strongly suggests that the cytology of large granulosa cells is induced by the oocyte. The heterogeneous granulosa persists only through previtellogenesis and at the onset of exogenous yolk uptake by the oocyte it becomes a secondarily homogeneous layer. The appearance of the granulosa at this stage is similar to that of reptiles whose granulosa remains a single-cell layer throughout folliculogenesis (e.g., turtles and crocodilians). Thus, although follicular development has been scrutinized in only a few representative genera of reptiles to date, the course of follicular development among lizards is similar in detail and involves the transitory development of a heterogeneous population of cells. This feature appears to be exclusive to the squamate reptiles.     

Subcortical microtubule network separates the periplasm from the endoplasm and is responsible for maintaining the position of accessory nuclei in hymenopteran oocytes

Oocytes of hymenopterans are equipped with peculiar organelles termed accessory nuclei. These organelles originate from the germinal vesicle (oocyte nucleus) and gather preferentially at the anterior pole. To gain insight into the mechanism of uneven (asymmetrical) distribution of accessory nuclei, the organization of the microtubule cytoskeleton in the oocytes of two hymenopterans Chrysis ignita and Cosmoconus meridionator has been studied. It is shown that during late previtellogenesis two networks of microtubules are present along the contact zone between the oocyte and enveloping follicular epithelium. The external one is associated with belt desmosomes connecting neighbouring follicular cells. The internal network is composed of randomly orientated microtubules and separates transparent, organelle-free periplasm from the endoplasm. All cellular organelles and the germinal vesicle are localized in the endoplasm. Accessory nuclei are accumulated in the anterior endoplasm; they always lie in direct contact with the subcortical network. Treatment with colchicine results in the disappearance of the periplasm as well as in the redistribution of cellular organelles including accessory nuclei. Presented findings suggest that subcortical microtubules play an important role in the positioning of accessory nuclei throughout the ooplasm.     

Oogenesis: From Oogonia to Ovulation in the Flagfish,Jordanella floridae Goode and Bean, 1879 (Teleostei: Cyprinodontidae)

We provide histological details of the development of oocytes in the cyprinodontid flagfish, Jordanella floridae. There are six stages of oogenesis: Oogonial proliferation, chromatin nucleolus, primary growth (previtellogenesis [PG]), secondary growth (vitellogenesis), oocyte maturation and ovulation. The ovarian lamellae are lined by a germinal epithelium composed of epithelial cells and scattered oogonia. During primary growth, the development of cortical alveoli and oil droplets, are initiated simultaneously. During secondary growth, yolk globules coalesce into a fluid mass. The full‐grown oocyte contains a large globule of fluid yolk. The germinal vesicle is at the animal pole, and the cortical alveoli and oil droplets are located at the periphery. The disposition of oil droplets at the vegetal pole of the germinal vesicle during late secondary growth stage is a unique characteristic. The follicular cell layer is composed initially of a single layer of squamous cells during early PG which become columnar during early vitellogenesis. During primary and secondary growth stages, filaments develop among the follicular cells and also around the micropyle. The filaments are seen extending from the zona pellucida after ovulation. During ovulation, a space is evident between the oocyte and the zona pellucida. Asynchronous spawning activity is confirmed by the observation that, after ovulation, the ovarian lamellae contain follicles in both primary and secondary growth stages; in contrast, when the seasonal activity of oogenesis and spawning ends, after ovulation, the ovarian lamellae contain only follicles in the primary growth stage. J. Morphol. 277:1339–1354, 2016. © 2016 Wiley Periodicals, Inc.     

Oogenesis and ovarian histology in two populations of the viviparous lizard Sceloporus grammicus (Squamata: Phrynosomatidae) from the central Mexican Plateau

The annual histological changes in ovarian morphology (oogenesis, follicular atresia, and corpus luteum) are described for the Mexican lizard Sceloporus grammicus, in two populations that inhabit contrasting environments (vegetation categories, climate, precipitation, and temperature) from Hidalgo State, Mexico. Two germinal beds were situated on the dorsal surface of each ovary of this species. In both the populations, oogenesis involves two major processes: previtellogenesis and vitellogenesis. The histological changes during previtellogenesis are similar to those for other reptilian sauropsids, whereas vitellogenesis differs and the features of this last process are described for the first time. In early previtellogenesis, primary oocytes have fibrillar chromosomes and the ooplasm stains slightly. The primordial follicles are surrounded by a granulosa composed of cuboidal follicular cells. During late previtellogenesis, the oocyte had an eccentric nucleus with lamp‐brush chromosomes and multiple nucleoli. The granulosa becomes multilayered and polymorphic, containing three cell types: small, intermediate, and pyriform. The zona pellucida was homogeneous and clearly observed. In early vitellogenesis, the oocyte showed several small acidophilic granules distributed in the center and the periphery of the oocyte. As vitellogenesis progresses, the yolk platelets move toward the central area of the oocyte and they fuse to form acidophilic and homogeneous yolk. Lipid droplets were distributed irregularly in the ooplasm of the oocyte. In Zacualtipán, the results revealed a strong seasonal reproductive activity. Females had vitellogenic follicles from July to September, and pregnant females were founded from September to March. In Tizayuca, the results showed an unusual pattern of reproductive activity. Females with vitellogenic follicles and pregnant females were found throughout the year, indicating continuous reproduction. We suggest that the observed differences in reproductive activity from these populations indicate adaptative fine tuning in response to local environmental conditions. These results contribute to the knowledge of variation in vitellogenesis and reproductive strategies of this species and among spiny lizards overall. J. Morphol. 275:949–960, 2014. © 2014 Wiley Periodicals, Inc.     

Structure of ovaries and formation of egg envelopes in the stonefly,Leuctra autumnalis Aubert, 1948 (Plecoptera: Leuctridae). Ultrastructural studies

The investigation of ovaries and the formation of egg envelopes of the stonefly Leuctra autumnalis was carried out with light and transmission electron microscopes. The ovary of the studied species is paired and consists of several dozen panoistic ovarioles opening individually to the oviduct. The process of egg capsule formation already begins in previtellogenesis. At this time the follicular cells secrete precursors of the vitelline envelope. Analysis of the presented data suggests that the oocyte itself also takes part in the formation of the vitelline envelope during late vitellogenesis. Simultaneously, the follicular cells produce precursors of further layers of the egg capsule, i.e. two-layered chorion and extrachorion, consisting of two gelatinous layers and a flocculent one. The completely developed capsule contains channels, probably micropylar ones.     

Oogenesis in the viviparous matrotrophic lizard Mabuya brachypoda

Oogenesis in the lizard Mabuya brachypoda is seasonal, with oogenesis initiated during May-June and ovulation occurring during July-August. This species ovulates an egg that is microlecithal, having very small yolk stores. The preovulatory oocyte attains a maximum diameter of 0.9-1.3 mm. Two elongated germinal beds, formed by germinal epithelia containing oogonia, early oocytes, and somatic cells, are found on the dorsal surface of each ovary. Although microlecithal eggs are ovulated in this species, oogenesis is characterized by both previtellogenic and vitellogenic stages. During early previtellogenesis, the nucleus of the oocyte contains lampbrush chromosomes, whereas the ooplasm stains lightly with a perinuclear yolk nucleus. During late previtellogenesis the ooplasm displays basophilic staining with fine granular material composed of irregularly distributed bundles of thin fibers. A well-defined zona pellucida is also observed. The granulosa, initially composed of a single layer of squamous cells during early previtellogenesis, becomes multilayered and polymorphic. As with other squamate reptiles, the granulosa at this stage is formed by three cell types: small, intermediate, and large or pyriform cells. As vitellogenesis progresses the oocyte displays abundant vacuoles and small, but scarce, yolk platelets at the periphery of the oocyte. The zona pellucida attains its maximum thickness during late oogenesis, a period when the granulosa is again reduced to a single layer of squamous cells. The vitellogenic process observed in M. brachypoda corresponds with the earliest vitellogenic stages seen in other viviparous lizard species with larger oocytes. The various species of the genus Mabuya provided us with important models to understand a major transition in the evolution of viviparity, the development of a microlecithal egg.     

Confocal scanning laser microscopy of the follicular epithelium in ovarioles of the stick insect Carausius morosus

Ovarian follicles of the stick insect Carausius morosus were analyzed by confocal laser microscopy and immunocytochemistry with a view to studying cell polarity in the follicular epithelium. Such probes as anti-α-tubulin antibodies and Rh-phalloidin were employed to establish how the follicle cell cytoskeleton changes during ovarian development. Data show that α-tubulin prevails over the basal end, while F-actin appears more abundant along the apical end of the follicle cells. This finding was further corroborated by immunogold cytochemistry, showing that label along the basal end is primarily associated with microtubules, while that along the apical end is due to follicle cell microvilli interdigitating with the oocyte plasma membrane. A monoclonal antibody specifically raised against a vitellin polypeptide was used to investigate the role the follicular epithelium might play in relation to vitellogenin (Vg) uptake by the oocyte. Data show that under these conditions label is restricted to the intercellular channels of the follicular epithelium, thus providing further support to the notion that Vg enters the oocyte through the extracellular pathway leading from the basement lamina to the oocyte surface. By contrast, the use of a monoclonal antibody raised against a fat-body-derived protein of 85 kDa that is specifically sulfated within the follicle cells provides evidence for the existence of an alternative way of gaining access to the oocyte surface, that is by transcytosis through the follicular cell epithelium. These findings confirm our earlier observations on stick insect ovarioles whereby polarization in the follicular epithelium is primarily addressed to sustain a transcytotic vesicular traffic between opposite poles of the follicle cell of Vg toward the oocyte surface.     

cGnRH II involvement in pyriform cell apoptosis

We have investigated whether gonadotrophin-releasing hormone (GnRH) is involved in triggering the apoptotic death of pyriforms, the nurse cells that cooperate in oocyte growth during mid- to late previtellogenesis in the lizard Podarcis sicula. Our immunocytochemical analyses demonstrate that pyriforms express GnRH receptors and that, in late previtellogenesis, they are up-regulated by cGnRH II. The hormone however does not trigger receptor synthesis and activation, events that therefore must be under the control of other regulatory factors. Our results also indicate that in vitro treatment of pyriforms with cGnRH II induces DNAse I activation and DNA laddering, clear cytological evidence of apoptosis, but not Fas/Fas-L synthesis or caspase activation. We conclude that cGnRH II is pro-apoptotic to pyriform cells and that it exerts its effects by activating an alternative cell death pathway, probably involving calcium as first messenger and DNase I as first executioner. This work was financed by a Progetto Giovani Ricercatori entitled “Le vie della morte nelle cellule follicolari nutrici di Podarcis sicula ” to S.T. and by a PRIN grant (2007) to Prof. Piero Andreuccetti.     

Oogenesis of microlecithal oocytes in the viviparous teleost Heterandria formosa

Viviparous teleosts exhibit two patterns of embryonic nutrition: lecithotrophy (when nutrients are derived from yolk that is deposited in the oocyte during oogenesis) and matrotrophy (when nutrients are derived from the maternal blood stream during gestation). Nutrients contained in oocytes of matrotrophic species are not sufficient to support embryonic development until term. The smallest oocytes formed among the viviparous poeciliid fish occur in the least killifish, Heterandria formosa, these having diameters of only 400 μm. Accordingly, H. formosa presents the highest level of matrotrophy among poeciliids. This study provides histological details occurring during development of its microlecithal oocytes. Five stages occur during oogenesis: oogonial proliferation, chromatin nucleolus, primary growth (previtellogenesis), secondary growth (vitellogenesis), and oocyte maturation. H. formosa, as in all viviparous poeciliids, has intrafollicular fertilization and gestation. Therefore, there is no ovulation stage. The full‐grown oocyte of H. formosa contains a large oil globule, which occupies most of the cell volume. The oocyte periphery contains the germinal vesicle, and ooplasm that includes cortical alveoli, small oil droplets and only a few yolk globules. The follicular cell layer is initially composed of a single layer of squamous cells during early previtellogenesis, but these become columnar during early vitellogenesis. They are pseudostratified during late vitellogenesis and reduce their height becoming almost squamous in full‐grown oocytes. The microlecithal oocytes of H. formosa represent an extreme in fish oogenesis typified by scarce yolk deposition, a characteristic directly related to matrotrophy. J. Morphol., 2011. © 2010 Wiley‐Liss, Inc.     

Morphological changes in the follicular epithelium in oogenesis of the Peking duck (Anas boschas domestica) and other poultry]

The morphology of the follicular epithelium during the course of oogenesis in poultry (duck goose, hen, turkey) and at the first stages of oocyte growth in some wild birds (finch, totmit, wood-pecker, pigeon) was studied. The general patterns of the follicular epithelium changings are similar in the both groups of birds. In the process of the oocyte growth the flat follicular epithelial monolayer changes to cubic, prismatic, pseudostratified epithelium. It leads sometimes to the impression of multi-layerness owing to its irregular structure. During the oocyte rapid growth the surface of the oocyte increases causing tension on the epithelium layer. This process again turns the pseudostratified epithelium into primatic, cubic and flat epithelium. Pseudostratified structure of the follilicular epithelium is regarded as adaptation to the necessity of the rapid tension during rapid oocyte growth. Correlations of the follicular epithelium morphology with the oocyte diameter are established. Existance of a temporary multilayer stage is discussed with arguments against the interpretation of this stage as a real multilayer.     

Gap junctions between the follicle cells and the oocyte during oogenesis in an insect,Tribolium destructor/Coleoptera

Summary Oocyte-follicle cell gap junctions inTribolium occur in all oogenetic stages studied. During early previtellogenesis the junctions are found exclusively between lateral membranes of oocyte microvilli and the membrane of prefollicle cells. In late previtellogenesis and vitellogenesis the junctions are located between the tips of oocyte microvilli and the flat membranes of the follicle cells. During previtellogenesis gap junctions are infrequent, whereas in the phase of yolk accumulation their number increases considerably, exceeding 17 junctions/m² of the follicle cell membrane. It could be shown by microinjection of a fluorescent dye that gap junctions are in a functional state during vitellogenesis. Possible roles of heterologous gap junctions in oogenesis are discussed.     

A comparative histochemical study of fish (Channa maruleus) and amphibian (Bufo stomaticus) oogenesis

Summary Comparative histochemical studies on the fish (Channa maruleus) and amphibian (Bufo stomaticus) oogenesis demonstrate a great similarity in the growth and differentiation of their egg follicle. The ooplasm, germinal vesicle and egg-membranes show distinct morphological and cytochemical changes during previtellogenesis and vitellogenesis.During previtellogenesis the various components of the follicle are engaged in the synthesis of protoplasm as shown by the proliferation of yolk nucleus substance, mitochondria and some lipid bodies in the ooplasm and of nucleoli in the germinal vesicle. The substance of the yolk nucleus consisting of proteins, lipoproteins and RNA first appears adjacent to the nuclear membrane. Numerous mitochondria of lipoprotein composition, and some lipid bodies consisting of unsaturated phospholipids lie in association with the yolk nucleus which forms substratum for the former. The lipid bodies, present inside the germinal vesicle, follicular epithelium, and adjacent to the plasma membrane in association with some pinocytotic vacuoles, have been considered to play a significant role in the active transport of some substances from the environment into the ooplasm and from the latter into the germinal vesicle. The follicular epithelium itself is very poorly developed, negating its appreciable role in the contribution of specific substances into the oocyte, which seem to be contributed by the germinal vesicle showing a considerable development of nuclear sap, basophilic granules and nucleoli consisting of RNA and proteins; many large nucleoli bodily pass into the cytoplasm during the previtellogenesis of Channa, where their substance is gradually dissolved. The intense, diffuse, basophilic substance of the cytoplasm is believed due to free ribosomes described in many previous ultrastructural studies.During vitellogenesis, the various deutoplasmic inclusions, namely carbohydrate yolk, proteid yolk and fatty yolk, are deposited in the ooplasm. The carbohydrate yolk bodies rich in carbohydrates originate in association with the plasma membrane and correspond to vesicles and cortical granules of previous studies. The proteid yolk consisting of proteins and some lipoproteins, and fatty yolk containing first phospholipids and some triglycerides and then triglycerides only are deposited under the influence of yolk nucleus substance, mitochondria and cytoplasm. The mitochondria and yolk nucleus substance foreshadow in some way the pattern of these two deutoplasmic inclusions and persist at the animal pole of mature egg while the other inclusions of previtellogenesis disappear from view. The pigment granules, which also show a gradient from the animal to vegetal pole in Bufo, are also formed in association with yolk nucleus substance and mitochondria. Some glycogen also appears in both the species. The nuclear membrane becomes irregular due to the formation of lobes. The lipid bodies of the germinal vesicle come to lie outside the nuclear membrane, suggesting active transport of some substances into the ooplasm; many nucleoli bodily pass into the ooplasm of Bufo, where they are gradually absorbed. The amount of basophilic granules is considerably increased in the germinal vesicle during vitellogenesis. Various egg-membranes such as outer epithelium, thin theca, single-layered follicular epithelium, zona pellucida or vitelline membrane surround the vitellogenic oocytes. The zona pellucida formed between the oocyte and follicle cells consists of a carbohydrate-protein complex. The follicle cells show lipid droplets, mitochondria and basophilic substance in their cytoplasm. The various changes that occur in the components of the follicle during vitellogenesis seem to be initiated by gonadrotrophins formed under the influence of specific environmental conditions.The author wishes to express sincere appreciation and gratitude to Dr. Gilbert S. Greenwald, who has made the completion of this investigation possible.Ph. D. Population Council Post-doctoral Fellow.     

Block of mitochondrial apoptotic pathways in lizard ovarian follicle cells as an adaptation to their nurse function

Pyriforms are ovarian follicle nurse cells that undergo apoptosis at the end of previtellogenesis and are completely eliminated by the epithelium. This event is accompanied by the active transfer of organelles and macromolecules to the oocyte via an intercellular bridge. Since it would be a nonsense for damaged mitochondria to reach the oocyte, we have postulated that pyriform cells have adapted their apoptotic machinery to prevent mitochondrial degradation. To verify this hypothesis, we have studied mitochondrial morphology and functionality during follicle cell regression. Cytological and biochemical evidence indicates that mitochondria in pyriforms maintain their size, organization and membrane potential. This clearly indicates that they are not involved in apoptosis signalling/progression. This block would favour both the oocyte, by increasing the pool of organelles available from follicle cells, and also the regressing pyriforms, by maintaining the energy resources required for completion of their nurse function. The block is probably attributable to an over-expression of Bcl-2 and might be carried out by sequestering cytochrome c inside the organelles. As demonstrated by in vitro experiments, the mitochondrial apoptosis pathway can be activated by stress induction, such as serum deprivation, but not following physiological pro-apoptotic signalling, such as treatment with gonadotrophin-releasing hormone. These studies were supported by a grant from the MIUR (PRIN project: Molecular responses of embryonic, differentiated and tumoral cells exposed to cadmium intoxication).     

Structure of the ovary and the differentiation of follicular epithelium in the pig louse, Haematopinus suis (Insecta: Phthiraptera)

The female reproductive system of the pig louse, Haematopinus suis (Insecta: Phthiraptera) is composed of paired ovaries, lateral oviducts, and a common oviduct that leads into a vagina. Clusters of mycetocytes (= cells filled with symbiotic organisms) are associated with lateral oviducts. Each ovary is composed of five loosely arranged ovarioles of the polytrophic-meroistic type. An individual ovariole is covered by a basal lamina and is composed of a terminal filament, germarium, and vitellarium. The terminal filament is composed of large, disc-shaped cells that are orientated perpendicularly to the long axis ofthe ovariole. The basal part of the terminal filament is separated from the germarium by a well-developed transverse septum. The germarium is short and filled with clusters of oogonial cells. In each cluster the cells arejoined by intercellular bridges, filled with fusomal material. Within the cluster, only one cell, the future oocyte, enters the prophase of the first meiotic division; the other cells differentiate into nurse cells. The basal part ofthe germarium is filled with the somatic prefollicular cells. The boundary between the germarium and the vitellarium is not distinct. The vitellarium contains linearly arranged ovarian follicles in subsequent stages of oogenesis (previtellogenesis, vitellogenesis and choriogenesis). Each follicle consists of an oocyte and 7 nurse cells and is surrounded by follicular cells. During oogenesis the follicular cells diversify, so that ultimately, five morphologically distinct subpopulations of these cells can be distinguished: (1) cells in contact with the nurse cells, (2) anterior cells, (3) mainbody cells, (4) posterior cells, and (5) interfollicular cells. Interestingly, the follicular cells associated with the anterior part of the oocyte, i.e. located in space at the oocyte/nurse cell border (fold cells) are mitotically active throughout previtellogenesis. It might be suggested, in this context, that the separation of the oocyte from the nurse cell compartment is brought about by mitotic divisions, consequent multiplication and centripetal migration of these cells.