An electron microscopic study of early gonadogenesis in the hermaphroditic appendicularian Oikopleura gracilis (Tunicata,Oikopleuridae)

The postembryonic development of the gonad in the hermaphroditic appendicularian O. gracilis was studied using transmission electron microscopy. The primordial germ cells were detected first in 10-h-old larvae and represent migrating primordial germ syncytium (mPGS) localized in the hemocoel of the tail/trunk junction and several haemocoel areas of the digestive compartment. The mPGS consisted of primordial germ nuclei (PGN) 2 μm in diameter, and elongate somatic-line nuclei 1.8 μm in diameter. In 12.5-h-old juveniles the gonad primordium 40 × 90 μm in size, was separated by a narrow space of haemocoel between the gut and the epidermis of the reproductive compartment. The gonad primordium consisted of the central syncytial part of primordial germ nuclei (PGN), enclosing a single layer of somatic epithelium. In 3-day-old juveniles, the gonad was differentiated into testis and ovary. The testis, 400 × 550 μm in size, is a syncytium of spermatogonial nuclei, covered by a single layer of somatic epithelium. The ovaries, 350 × 850 μm in size, consist of a syncytium with nurse nuclei and meiotic nuclei. The hermaphroditic gonad originates from extragonadal mPGS. Early gonadogenesis in appendicularians has ultrastructural features in common with early gonadogenesis in ascidians.     

Gonads in Histiostoma mites (Acariformes: Astigmata): Structure and development

The development of male and female gonads in arrhenotokous and thelytokous species of Histiostoma was studied using transmission electron microscopy (TEM). All instars were examined: larvae, protonymphs, facultative heteromorphic deutonymphs (=hypopi), tritonymphs, and adults. In testis primordium, spermatogonia surrounding a testicular central cell (TCC) with a gradually enlarging, branched nucleus are present already at the larval stage. Spermatogonia and the TCC are connected via narrow, tubular intercellular bridges revealing that the TCC is a germline cell. Spermatocytes appear at the protonymphal stage. At the heteromorphic deutonymph stage, the testis primordium is similar to that of the protonymph, but in the tritonymph it is much larger and composed as in the adult: spermatids as well as sperm cells are present. The latter are congregated ventrally in the testis at the entrance of the deferent duct.In the larval ovary, an eccentrically located ovarian nutritive cell (ONC) is surrounded by oogonia which are connected with the ONC via tubular intercellular bridges. In later stages, the ovary grows and oocytes appear in the protonymph. Meiotic synaptonemal complexes in oocytes occur from the tritonymph stage. At about the time of the final molting, tubular intercellular bridges transform into peculiar diaphragm-crossed bridges known only in Histiostoma mites. In the adult female, growing oocytes at the end of previtellogenesis lose intercellular bridges and move ventro-laterally to the ovarian periphery towards the oviduct entrance. Vitellogenesis occurs in oviducts.Germinal cells in both the testis and ovary are embedded in a few somatic stroma cells which may be well discernible already in the larval ovary; in the testis, somatic stroma cells are evident not earlier than the end of the tritonymphal stage. The ovary has a thin wall of flat somatic cells, whereas the testis is covered by a basal lamina only.The obtained results suggest that gonads in Histiostoma and other Astigmata originate from two primordial cells only.     

Ultrastructure of the body cavities in juveniles and adults of the appendicularian Oikopleura gracilis (Tunicata,Chordata) suggests how the heart may have evolved in tunicates

The organization of the body cavities is an important morphological trait that can be used for establishing the phylogenetic relationships between different groups of animals. In the present study, the hemocoel and coelomic systems of 10‐hr‐old juveniles and adults of the hermaphroditic oikopleurid Oikopleura gracilis were examined using light and transmission electron microscopy. The trunk hemocoel in 10‐hr‐old juveniles was represented by small clefts containing layers of extracellular matrix of adjacent tissues or interstices with migrating primordial germ syncytium. The wide hemocoel in the tail contained extracellular strands, subdividing the hemocoel into hemal sinuses. In adults, a large hemocoel appeared in the trunk and tail, and also contained extracellular strands. The hermaphroditic gonad was surrounded by its own lining, separating it from the hemocoel. The gamete‐filled cavity in the ovary and testis appeared only at late‐stage gonadogenesis, when the pre‐spawning reduction of syncytium occurred in the gonads. The true coelom in 10‐hr‐old juveniles and adults was represented by the pericardium. The lining of the pericardium consisted of myoepithelial and peritoneal cells. In the myoepithelial cells of 10‐hr‐old juveniles, myofibrils had been formed. The myoepithelial cells of adults had several parallel rows of completely differentiated myofibrils. The substantial reduction of the coelomic and circulatory systems in O. gracilis evidently results from the extreme shortening of ontogeny in appendicularians. Development in O. gracilis from early juvenile to adult involves the following steps, which also suggest how the tunicate heart may have evolved: a single‐layered coelomic sac gives rise to a grooved pericardium with an open hemal sinus (simple heart). In ascidians, this simple heart in turn gives rise to a closed tubular, double‐layered heart–pericardial complex, with a separate pericardial cavity and a closed heart, whose wall is formed by specialized myocardium.     

The effect of inhibitors upon intracellular cyclic AMP levels and chick myoblast differentiation.

The small free-living nematode Caenorhabditis elegans is usually found as a hermaphrodite, but occasionally true males appear in the population. This study provides an account of gonadogenesis in the normal male and in a mutant that is a temperature-sensitive sex transformer.Male and hermaphrodite gonads develop from morphologically identical primordia. The small primordial gonad lies on the ventral side of the worm in the coelomic cavity. The gonadial primordium contains four nuclei at parturition. As this primordium develops in a hermaphrodite, it produces a double-armed, mirror symmetrical gonad that produces first sperm and then eggs. In the male, however, this primordium develops into an asymmetrical structure composed of a ventrally located testis, a loop region, a seminal vesicle, and a vas deferens. The male gonad presents a linear sequence of nuclei in successive stages of spermatogenesis beginning with a mitotic region in the testis, followed by clearly distinguishable stages of meiosis throughout the loop region to the seminal vesicle.A temperature-sensitive sex transformer mutant, tsB202, has been isolated. tsB202 carries an autosomal recessive mutation in linkage group II that at restrictive temperature transforms an XX hermaphrodite into a phenotypic male, complete with a normal male gonad and vestigial external genitalia. These transformed males are classified as pseudomales because they do not exhibit mating behavior. Temperature shift experiments have determined the specific temporal sequences of gonadogenesis, oogenesis, and spermatogenesis. Proper manipulation of the temperature regimen causes the production of intersexes. In one intersex, a male gonad complete with sperm, seminal vesicle, and vas deferens also contains oocytes. In another intersex produced by the complementary temperature shift, a hermaphrodite-shaped gonad develops that produces only sperm and no oocytes.     

Ultrastructure of somatic tissues in the ovaries of a chaetognath (Ferosagitta hispida)

Transmission electron microscopy reveals that the ovaries of Ferosagitta hispida contain four somatic tissues. A myoepithelial ovary wall, continuous with a thin layer of peritoneocytes lining the coelomic cavity, encloses a fluid-filled ovarian space in which oocytes develop. Lamellar extensions of a “follicular reticulum” branch throughout the ovarian space and ensheath developing oocytes. This tissue has been overlooked in most previous studies of chaetognath ovaries. A bipartite oviductal complex extends the length of each ovary just within the lateral ovary wall. It consists of a flattened, blindly ending cellular tube, herein referred to as the cellular sheath, and an enclosed syncytium. Sheath cells secrete an electron-dense product into the ovarian space. Those sheath cells directly bordering the syncytium are contractile and are joined to the to the syncytium by gap junctions and microvillar interdigitations. The syncytium contains a complex of membrane-bounded lumina. The latter sometimes enclose sperm received during mating or ovulated eggs. Thus the syncytium serves both as a seminal receptacle and as a duct for passage of eggs to the outside. Contrary to several classical reports, the cellular sheath and syncytium of the oviductal complex do not separate at ovulation to form a temporary oviductal lumen.     

Trichinella spiralis: Morphological characteristics of male and female intestine-infecting larvae

The sex of encysted and excysted intestine-infecting T. spiralis larvae can be distinguished by the following morphological characteristics: the male larva has a long (approx 50 μm) rectum, and the anterior part of the testis is curved posteriorly. The female larva has a shorter rectum (approx 25 μm), a telogonic ovary, coiled uterine and seminal receptacle primordia, and a vaginal primordium. In paraffin sections males can be recognized by the spermatocytes which are of the same size. The oocytes vary in size: the smallest are located in the ventral portion, the largest on the dorsal portion of the ovary. Sex of the larvae can be differentiated by the length of the rectum as early as the tenth day, by the curvature of the anterior part of the testis and by the uterine primordium by the eleventh day, and by the presence of the vaginal primordium by the thirteenth day of intramuscular development. Farre's Organ is believed to be the primordium of the seminal receptacle.     

The Oikopleura coenocyst, a unique chordate germ cell permitting rapid, extensive modulation of oocyte production

The ability to adjust reproductive output to environmental conditions is important to the fitness of a species. The semelparous, chordate, Oikopleura dioica, is particularly adept in producing a highly variable number of oocytes in its short life cycle. Here we show that this entails an original reproductive strategy in which the entire female germline is contained in a single multinucleate cell, the "coenocyst". After an initial phase of syncytial nuclear proliferation half of the nuclei entered meiosis whereas the other half became highly polyploid. The inner F-actin network, with associated plasma membranes, formed a highly ramified infrastructure in which each meiotic nucleus was contained in a pseudo-compartmentalized pro-oocyte linked to the common cytoplasm via ring canals. At a set developmental time, a subset of the pro-oocytes was selected for synchronous growth and the common coenocyst cytoplasm was equally partitioned by transfer through the ring canals. Examination of related species indicated that the coenocyst arrangement is a conserved feature of Appendicularian oogenesis allowing efficient numerical adjustment of oocyte production. As Appendicularia are the second most abundant class of zooplankton, with a world-wide distribution, the coenocyst is clearly a common and successful reproductive strategy on a global scale.     

Morphological and histochemical examination of male and female gonads in Homarus gammarus (L. 1758)

The reproductive organs of both male and female European lobsters (Homarus gammarus) are H-shaped gonads that lie dorsal to the gut on the large hepatopancreas. The ovary consists of a pair of tubular, parallel lobules with a connecting bridge. The germarium of the ovary containing oogonia is concentrated in the center of the ovarian lobe. As oogonesis proceeds, the oocytes move to the peripheral regions of the ovary. The follicle cells begin to surround the oocytes in the previtellogenic stage, and the mature oocytes are completely surrounded by the follicle cells. Carbohydrates exist in both early and late vitellogenic oocytes that give PAS positive reaction. However, their rising protein content in late vitellogenic oocytes makes them stain with Bromophenol blue. Testes show convoluted lobules with a germinal epithelium and a central collecting duct, and the paired vasa deferentia have three distinct parts. Spermatophores are nonpedunculate and tubular, which extrude as a continuous column and consist of a sperm mass covered with primary and secondary layers. The primary layer stains with Bromophenol Blue and gives a PAS positive reaction. But the secondary layer only weakly stains with Bromophenol Blue. The histochemical results may indicate that the function of the two layers is different.     

The cytoskeleton organizes germ nuclei with divergent fates and asynchronous cycles in a common cytoplasm during oogenesis in the chordate Oikopleura

Germline cysts are conserved structures in which cells initiating meiosis are interconnected by ring canals. In many species, the cyst phase is of limited duration, but the chordate, Oikopleura, maintains it throughout prophase I as a unique cell, the coenocyst. We show that despite sharing one common cytoplasm with meiotic and nurse nuclei evenly distributed in a 1:1 ratio, both entry into meiosis and subsequent endocycles of nurse nuclei were asynchronous. Coenocyst cytoskeletal elements played central roles as oogenesis progressed from a syncytial state of indistinguishable germ nuclei, to a final arrangement where the common cytoplasm had been equally partitioned into resolved, mature oocytes. During chromosomal bouquet formation in zygotene, nuclear pore complexes clustered and anchored meiotic nuclei to the coenocyst F-actin network opposite ring canals, polarizing oocytes early in prophase I. F-actin synthesis was required for oocyte growth but movement of cytoplasmic organelles into oocytes did not require cargo transport along colchicine-sensitive microtubules. Instead, microtubules maintained nurse nuclei on the F-actin scaffold and prevented their entry into growing oocytes. Finally, it was possible to both decouple meiotic progression from cellular mechanisms governing oocyte growth, and to advance the timing of oocyte growth in response to external cues.     

Renewal of the gonads in Styela clava (Urochordata: Ascidiacea) as revealed by autoradiography with tritiated thymidine

DNA-synthesizing cells in the gonads of the ascidian Styela clava were labeled with tritiated thymidine and detected with autoradiography. In the testis, spermatogonia and primary spermatocytes are labeled after 1 hr. Labeled spermatozoa occur in the lumen of the testis follicles after 10 days and in the sperm ducts after 20 days. In the ovary, only germ cells (oogonia and pre-leptotene primary oocytes) and follicle cells are labeled after 1 hr. By 60 days, oocytes with basophilic cytoplasm (15–65 μ in diameter) are labeled; test cells embedded in larger eosinophilic oocytes (150 μ in diameter) are also labeled. Germ cells give rise to both oocytes and follicle cells. Through continued cell division, follicle cells give rise to test cells.     

Sex Reversal in Zebrafish fancl Mutants Is Caused by Tp53-Mediated Germ Cell Apoptosis

The molecular genetic mechanisms of sex determination are not known for most vertebrates, including zebrafish. We identified a mutation in the zebrafish fancl gene that causes homozygous mutants to develop as fertile males due to female-to-male sex reversal. Fancl is a member of the Fanconi Anemia/BRCA DNA repair pathway. Experiments showed that zebrafish fancl was expressed in developing germ cells in bipotential gonads at the critical time of sexual fate determination. Caspase-3 immunoassays revealed increased germ cell apoptosis in fancl mutants that compromised oocyte survival. In the absence of oocytes surviving through meiosis, somatic cells of mutant gonads did not maintain expression of the ovary gene cyp19a1a and did not down-regulate expression of the early testis gene amh; consequently, gonads masculinized and became testes. Remarkably, results showed that the introduction of a tp53 (p53) mutation into fancl mutants rescued the sex-reversal phenotype by reducing germ cell apoptosis and, thus, allowed fancl mutants to become fertile females. Our results show that Fancl function is not essential for spermatogonia and oogonia to become sperm or mature oocytes, but instead suggest that Fancl function is involved in the survival of developing oocytes through meiosis. This work reveals that Tp53-mediated germ cell apoptosis induces sex reversal after the mutation of a DNA–repair pathway gene by compromising the survival of oocytes and suggests the existence of an oocyte-derived signal that biases gonad fate towards the female developmental pathway and thereby controls zebrafish sex determination.     

Ultra- and microstructure of the female reproductive system of Matsucoccus matsumurae

The ultra- and microstructure of the female reproductive system of Matsucoccus matsumurae was studied using light microscopy, scanning and transmission electron microscopy. The results revealed that the female reproductive system of M. matsumurae is composed of a pair of ovaries, a common oviduct, a pair of lateral oviducts, a spermatheca and two pairs of accessory glands. Each ovary is composed of approximately 50 telotrophic ovarioles that are devoid of terminal filaments. Each ovariole is subdivided into an apical tropharium, a vitellarium and a short pedicel connected to a lateral oviduct. The tropharium contains 8–10 trophocytes and two early previtellogenic oocytes termed arrested oocytes. The trophocytes degenerate after egg maturation, and the arrested oocytes are capable of further development. The vitellarium contains 3–6 oocytes of different developmental stages: previtellogenesis, vitellogenesis and choriogenesis. The surface of the vitellarium is rough and composed of a pattern of polygonal reticular formations with a center protuberance. The oocyte possesses numerous yolk spheres and lipid droplets, and is surrounded by a mono-layered follicular epithelium that becomes binucleate at the beginning of vitellogenesis. Accessory nuclei are observed in the peripheral ooplasm during vitellogenesis.     

Reproduction and sexual development in a freshwater gastrotrich

Summary Six small cells are present in each of the bilateral gonads of parthenogenically reproductive Lepidodermella squammata. Early in the extended postparthenogenic phase of the life history, these cells undergo limited proliferation followed by differentiation. Primary oocytes of three types are present 0.3 days after deposition of the final parthenogenic egg: small oocytes with presynaptic nuclei; intermediate oocytes with nuclei containing synaptonemal complexes; and larger oocytes with a germinal vesicle. Oocytes persist without further development at least until day four of the postparthenogenic phase. Older isolated animals may contain and even deposit an enlarged egg, but successful progeny does not result. Oocytes are located at the anterior pole of each of the bilateral gonads, adjacent to developing male tissues producing sperm. More posterior cells in the gonad are initially undifferentated in the postparthenogenic phase. Dorsal and central cells first show specialization for secretory activity, and by day four contain peripheral layers of RER and central accumulations of polymorphic secretion droplets. The posterior and ventral cells produce secretion droplets that aggregate into an enlarging bilobed structure called the X-body. Two or three cells in each gonad contribute secretions to the X-body, which is intracellular in a secondary syncytium formed by the contributing cells. Functions for the postparthenogenic gametes and for the X-body are not yet demonstrated.     

Organization of Interna in Sacculina polygenea (Crustacea: Rhizocephala)

We performed histological studies on trophic and reproductive systems of colonial interna in Sacculina polygenea, a parasite of the coastal crab Hemigrapsus sanguineus. The trophic system that performs functions of absorption, accumulation, and transportation of nutrients from the hemolymph of the host comprises the trophic epithelium of distal canals and transporting trunks. The reproductive system of interna consist of nuclei (early stages of development of the primordia of externae) and the primordia of externae in later stages of development. It has been shown that during morphogenesis of the nucleus two primordia arise, a primordium of the externa itself and a primordium of its trophic system. In the primordium of the ovary, we found oogonia; early oocytes and vitellogenic oocytes were found in the ovaries of the late primordia of the externae. The damaging effects of the interna on the ovaries and testes of the crab host are discussed. Thus, we have found numerous elements of reproductive and trophic systems in the colonial interna of S. polygenea. The term individual is proposed to be used for the externa in rhizocephalan barnacles with its trophic system.     

A New Model of Development of the Mammalian Ovary and Follicles

Ovarian follicular granulosa cells surround and nurture oocytes, and produce sex steroid hormones. It is believed that during development the ovarian surface epithelial cells penetrate into the ovary and develop into granulosa cells when associating with oogonia to form follicles. Using bovine fetal ovaries (n = 80) we identified a novel cell type, termed GREL for Gonadal Ridge Epithelial-Like. Using 26 markers for GREL and other cells and extracellular matrix we conducted immunohistochemistry and electron microscopy and chronologically tracked all somatic cell types during development. Before 70 days of gestation the gonadal ridge/ovarian primordium is formed by proliferation of GREL cells at the surface epithelium of the mesonephros. Primordial germ cells (PGCs) migrate into the ovarian primordium. After 70 days, stroma from the underlying mesonephros begins to penetrate the primordium, partitioning the developing ovary into irregularly-shaped ovigerous cords composed of GREL cells and PGCs/oogonia. Importantly we identified that the cords are always separated from the stroma by a basal lamina. Around 130 days of gestation the stroma expands laterally below the outermost layers of GREL cells forming a sub-epithelial basal lamina and establishing an epithelial-stromal interface. It is at this stage that a mature surface epithelium develops from the GREL cells on the surface of the ovary primordium. Expansion of the stroma continues to partition the ovigerous cords into smaller groups of cells eventually forming follicles containing an oogonium/oocyte surrounded by GREL cells, which become granulosa cells, all enclosed by a basal lamina. Thus in contrast to the prevailing theory, the ovarian surface epithelial cells do not penetrate into the ovary to form the granulosa cells of follicles, instead ovarian surface epithelial cells and granulosa cells have a common precursor, the GREL cell.     

Cloning and Characterization of a Rice Field Eel vasa-Like Gene cDNA and Its Expression in Gonads During Natural Sex Transformation

The vasa (vas)-related gene encodes an RNA helicase protein member of the DEAD-box family and plays key roles in germ-cell formation in higher metazoans. Using degenerate PCR and RACE, we cloned the vasa gene of the rice field eel (Monopterus albus), which is homologous to the Drosophila vasa gene. We named it ma-vas (Monopterus albus vas). Ma-vas encodes a protein of 618 amino acids, which contains all of the known characteristics of vasa homologs. RT-PCR analysis revealed that ma-vas was exclusively expressed in the gonads of the female, intersex, and male. During gonadal natural sex reversal, ma-vas is expressed in oocytes at all stages of oogenesis, in degenerating oocytes of ovotestis, and in spermatogonia and spermatocytes at early stages. The vasa positive signal was also observed in the peripheral layer of late ovary. It was not found, however, in that layer of the testis. Alkaline phosphatase (AKP) staining on the ovary and testis also indicated that some cells had differentiational potential in the peripheral layer of the ovary, suggesting that spermatogonia might arise from cells with AKP and vasa-positive staining in the peripheral layer of the female gonad.     

Morphological and histochemical studies on oogenesis in Callosobruchus analis Fabr. (Bruchidae-Coleoptera)

The ovary in Callosobruchus analis consists of telotrophic ovarioles with the so called nurse cells confined to one chamber at the anterior end of the ovariole. There are three types of lipids in the ovary: (1) L₁ bodies that are present in the early oocytes, in the posterior prefollicular tissue and in the follicular epithelium and contain unsaturated phospholipids; (2) L₂ bodies that have a complete or incomplete sheath of phospholipids and a triglyceride core; (3) L₃ bodies that are formed of highly saturated triglycerides. Lipids are absent from the trophic tissue. In a mature oocyte the L₁ and L₂ bodies are cortical in distribution while the L₃ bodies are centrally located. The mitochondria contain lipoproteins with RNA. The yolk spheres are acid mucopolysaccharides and protein in nature. The precursors of the yolk spheres appear first in the cortical coplasm and are absent from the follicular epithelium or the trophic tissue. The nucleolus of the oocyte shows evidence of extrusions that are believed to pass into the ooplasm. There are no nutritive cords connecting the trophic tissue to the oocytes; nor is there any evidence of any histochemically demonstrable nutritive material being contributed to the oocyte by the trophic tissue. The circumstantial evidence points towards a contribution of the raw materials to the oocyte by the haemolymph either through or in between the follicular epithelium in some soluble form or as submicroscopic particles.