Origin of the gonad in the juvenile of a solitary ascidian, Ciona intestinalis

In the just-metamorphosed juveniles of Ciona intestinalis, a round mass of tissue debris derived from the resorbed tadpole tail is situated in the broad space enclosed by the peritoneal membrane and the epidermis around the ventral side of the esophagus. In living juveni es, the origin of the gonad rudiment was traced back to the mass of tissue debris. Electron microscopically, the round mass was a clump of irregular-shaped phagocytotic cells engulfing degenerated cell fragments. On the surface of the cell clump, a small number of singly occurring round cells were found and identified as primordial germ cells on the basis of morphological continuity to obvious germ cells in later stages. Presence of nuage around the nucleus characterized the germ cells. In a few days the germ cells assembled to form a solid slender body (gonad rudiment) together with smaller somatic cells. The gonad rudiment left the space around the esophagus, moving into the narrow mesenteric space connecting the stomach and intestine on the fourth day after metamorphosis. It gradually increased in size by proliferation of the germ cells and somatic cells. The solid gonad rudiment changed into an oval vesicle with an eccentrically located cavity on about the seventh day after metamorphosis. The vesicle comprised a thinner wall made of a simple epithelium without germ cells and a thicker wall containing germ cells and somatic cells.     

Primordial germ cells originate from the endodermal strand cells in the ascidian Ciona intestinalis

The origin of germ cells in the ascidian is still unknown. Previously, we cloned a vasa homologue (CiVH) of Ciona intestinalis from the cDNA library of ovarian tissue by polymerase chain reaction and showed that its expression was specific to germ cells in adult and juvenile gonads. In the present study, we prepared a monoclonal antibody against CiVH protein and traced the staining for this antibody from the middle tailbud stage to young adulthood. Results showed that positive cells are present in the endodermal strand in middle tailbud embryos and larvae. When the larval tail was absorbed into the trunk during metamorphosis, the CiVH-positive cells migrated from the debris of the tail into the developing gonad rudiment, and appeared to give rise to a primordial germ cell (PGC) in the young juvenile. The testis rudiment separated from the gonad rudiment, the remainder of which differentiated into the ovary. PGCs of the testis rudiment and the ovary rudiment differentiated into spermatogenic and oogenic cells, respectively. When the larval tail containing the antibody-positive cells was removed, the juveniles did not contain any CiVH-positive cells after metamorphosis, indicating that the PGCs in the juvenile originated from part of the larval tail. However, even in such juveniles, positive cells newly appeared in the gonad rudiment at a later stage. This observation suggests that a compensatory mechanism regulates germline formation in C. intestinalis.     

Germ Cells Are Not Required to Establish the Female Pathway in Mouse Fetal Gonads

The fetal gonad is composed of a mixture of somatic cell lineages and germ cells. The fate of the gonad, male or female, is determined by a population of somatic cells that differentiate into Sertoli or granulosa cells and direct testis or ovary development. It is well established that germ cells are not required for the establishment or maintenance of Sertoli cells or testis cords in the male gonad. However, in the agametic ovary, follicles do not form suggesting that germ cells may influence granulosa cell development. Prior investigations of ovaries in which pre-meiotic germ cells were ablated during fetal life reported no histological changes during stages prior to birth. However, whether granulosa cells underwent normal molecular differentiation was not investigated. In cases where germ cell loss occurred secondary to other mutations, transdifferentiation of granulosa cells towards a Sertoli cell fate was observed, raising questions about whether germ cells play an active role in establishing or maintaining the fate of granulosa cells. We developed a group of molecular markers associated with ovarian development, and show here that the loss of pre-meiotic germ cells does not disrupt the somatic ovarian differentiation program during fetal life, or cause transdifferentiation as defined by expression of Sertoli markers. Since we do not find defects in the ovarian somatic program, the subsequent failure to form follicles at perinatal stages is likely attributable to the absence of germ cells rather than to defects in the somatic cells.     

Intact fetal ovarian cord formation promotes mouse oocyte survival and development

Background  

Female reproductive potential, or the ability to propagate life, is limited in mammals with the majority of oocytes lost before birth. In mice, surviving perinatal oocytes are enclosed in ovarian follicles for subsequent oocyte development and function in the adult. Before birth, fetal germ cells of both sexes develop in clusters, or germline cysts, in the undifferentiated gonad. Upon sex determination of the fetal gonad, germ cell cysts become organized into testicular or ovarian cord-like structures and begin to interact with gonadal somatic cells. Although germline cysts and testicular cords are required for spermatogenesis, the role of cyst and ovarian cord formation in mammalian oocyte development and female fertility has not been determined.     

Development of ovaries in bovine fetuses

The growth of ovaries, development of germ cells, formation of sex cords, folliculogenesis and dependence of these processes on the gonad morphogenesis stages were studied on 68 embryos and foetuses at the age of 1.5 to 9 months. Sex differentiation of ovaries was shown to take place in 1.5 month old embryo. The cords of connective tissue's cortical stroma appear also in 1.5 month old embryo, they develop in the dorsoventral direction and reach the gonad's covering epithelium in 6 month old foetuses. The formation of the medulla rudiment starts in 1.5 month old embryo when the gonad is separated from mesonephros and connected with it via the ovary gate. In 4 month old foetuses the ovary net transforms into a stellate structure. Important morphogenetic processes, such as the development of the ovary somatic elements, entry of the oocytes into meiotic prophase, formation of the sex cords and folliculogenesis, develop in the dorsoventral direction Germ cells in 9 month old foetuses are enclosed into primordial or, growing follicles.     

Gonadal morphogenesis and sex differentiation in the viviparous fish Chapalichthys encaustus (Teleostei, Cyprinodontiformes, Goodeidae)

This study describes the structural and ultrastructural characteristics of gonadal sex differentiation and expression of Vasa, a germline marker, in different developmental stages of embryos and newborn fry of the barred splitfin Chapalichthys encaustus, a viviparous freshwater teleost endemic to Mexico. In stage 2 embryos, the gonadal crest was established; gonadal primordia were located on the coelomic epithelium, formed by scarce germ and somatic cells. At stage 3, the undifferentiated gonad appeared suspended from the mesentery of the developing swimbladder and contained a larger number of germ and somatic cells. At stages 4 and 5, the gonads had groups of meiotic and non-meiotic germ cells surrounded by somatic cells; meiosis was evident from the presence of synaptonemal complexes. These stages constituted a transition towards differentiation. At stage 6 and at birth, the gonad was morphologically differentiated into an ovary or a testis. Ovarian differentiation was revealed by the presence of follicles containing meiotic oocytes, and testicular differentiation by the development of testicular lobules containing spermatogonia in mitotic arrest, surrounded by Sertoli cells. Nuage, electron-dense material associated with mitochondria, was observed in germ cells at all gonadal stages. The Vasa protein was detected in all of the previously described stages within the germ-cell cytoplasm. This is the first report on morphological characteristics and expression of the Vasa gene during sexual differentiation in viviparous species of the Goodeidae family. Chapalichthys encaustus may serve as a model to study processes of sexual differentiation in viviparous fishes and teleosts.     

Posthatching development of Alligator mississippiensis ovary and testis

We investigated ovary and testis development of Alligator mississippiensis during the first 5 months posthatch. To better describe follicle assembly and seminiferous cord development, we used histochemical techniques to detect carbohydrate‐rich extracellular matrix components in 1‐week, 1‐month, 3‐month, and 5‐month‐old gonads. We found profound morphological changes in both ovary and testis. During this time, oogenesis progressed up to diplotene arrest and meiotic germ cells increasingly interacted with follicular cells. Concomitant with follicles becoming invested with full complements of granulosa cells, a periodic acid Schiff's (PAS)‐positive basement membrane formed. As follicles enlarged and thecal layers were observed, basement membranes and thecal compartments gained periodic acid‐methionine silver (PAMS)‐reactive fibers. The ovarian medulla increased first PAS‐ and then PAMS reactivity as it fragmented into wide lacunae lined with low cuboidal to squamous epithelia. During this same period, testicular germ cells found along the tubule margins were observed progressing from spermatogonia to round spermatids located within the center of tubules. Accompanying this meiotic development, interstitial Leydig cell clusters become more visible and testicular capsules thickened. During the observed testis development, the thickening tunica albuginea and widening interstitial tissues showed increasing PAS‐ and PAMS reactivity. We observed putative intersex structures in both ovary and testis. On the coelomic aspect of testes were cell clusters with germ cell morphology and at the posterior end of ovaries, we observed “medullary rests” resembling immature testis cords. We hypothesize laboratory conditions accelerated gonad maturation due to optimum conditions, including nutrients and temperature. Laboratory alligators grew more rapidly and with increased body conditions compared with previous measured, field‐caught animals. Additionally, we predict the morphological maturation observed in these gonads is concomitant with increased endocrine activities. J. Morphol. 2010. © 2009 Wiley‐Liss, Inc.     

Gonadal development and sex differentiation in the cichlid fish Cichlasoma dimerus (Teleostei, Perciformes): a light- and electron-microscopic study

Although the overall pattern and timing of gonadal sex differentiation have been established in a considerable number of teleosts, the ultrastructure of early stages of gonadal development is not well documented. In this study, gonads from larval and juvenile stages of laboratory-reared Cichlasoma dimerus were examined at the light-microscopic and ultrastructural levels. This freshwater species adapts easily to captivity and spawns with high frequency during 8 months of the year, providing an appropriate model for developmental studies. Larvae and juveniles were kept at a water temperature of 26.5 +/- 1 degrees C and a 12:12 hour photoperiod. Gonadal development was documented from 14-100 days postfertilization, covering the period of histologically discernible sex differentiation. Gonadal tissue was processed according to standard techniques for light and electron microscopy. C. dimerus, a perciform teleost, is classified as a differentiated gonochorist, in which an indifferent gonad develops directly into a testis or ovary. On day 14, the gonadal primordium consists of a few germ cells surrounded by enveloping somatic cells. Ovarian differentiation precedes testicular differentiation, as usual in teleost fishes. The earliest signs of differentiation, detected from day 42 onward, include the onset of meiotic activity in newly formed oocytes, which is soon accompanied by increased oogonial mitotic proliferation and the somatic reorganization of the presumptive ovary. The ovarian cavity is completely formed by day 65. Numerous follicles containing perinucleolar oocytes are observed by day 100. In contrast, signs of morphological differentiation in the presumptive testis are not observed until day 72. By day 100, the unrestricted lobular organization of the testis is evident. The latest stage of spermatogenesis observed by this time of testicular development is spermatocyte II.     

Ovarian development in 46,XY gonadal dysgenesis

Summary In human the XY ovary is degenerative, there being scant evidence of persistence of that organ beyond the perinatal period. Here we describe indications of functional ovarian tissue in a 17-year-old female with male karyotype, H-Y⁺ cellular phenotype, and some signs of the Turner syndrome. Her gonads were removed after the onset of secondary amenorrhea. Histological examination revealed a degenerative right ovary devoid of germ cells and follicles, and a left streak gonad. There was no trace of testicular development in either side.     

Gonadal morphogenesis and sex differentiation in intraovarian embryos of the viviparous fish Zoarces viviparus (Teleostei, Perciformes, Zoarcidae): a histological and ultrastructural study

It is essential to know the timing and process of normal gonadal differentiation and development in the specific species being investigated in order to evaluate the effect of exposure to endocrine-disrupting chemicals on these processes. In the present study gonadal sex differentiation and development were investigated in embryos of a viviparous species of marine fish, the eelpout, Zoarces viviparus, during their intraovarian development (early September to January) using light and electron microscopy. In both sexes of the embryos at the time of hatching (September 20) the initially undifferentiated paired bilobed gonad contains primordial germ cells. In the female embryos, ovarian differentiation, initiated 14 days posthatch (dph), is characterized by the initial formation of the endoovarian cavity of the single ovary as well as by the presence of some early meiotic oocytes in a chromatin-nucleolus stage. By 30 dph, the endoovarian cavity has formed. By 44 dph and onward, the ovary and the oocytes grow in size and at 134 dph, just prior to birth, the majority of the oocytes are at the perinucleolar stage of primary growth and definitive follicles have formed. In the presumptive bilobed testis of the male embryos, the germ cells (spermatogonia), in contrast to the germ cells of the ovary, remain quiescent and do not enter meiosis during intraovarian development. However, other structural (somatic) changes, such as the initial formation of the sperm duct (30 dph), the presence of blood vessels in the stromal areas of the testis (30 dph), and the appearance of developing testicular lobules (102 dph), indicate testicular differentiation. Ultrastructually, the features of the primordial germ cells, oogonia, and spermatogonia are similar, including nuage, mitochondria, endoplasmic reticulum, and Golgi complexes.     

Stem cell factor and c-kit gene expression and protein localization in the sheep ovary during fetal development

The aim of this study was to investigate stem cell factor and c-kit gene expression and protein localization in the mesonephros and ovary of sheep fetuses at different days of gestation, using RNA in situ hybridization and immunohistochemical procedures. At days 24 and 26 of gestation, stem cell factor mRNA and protein were present in cells throughout the developing gonad and mesonephros. From day 28 to day 40 of gestation, stem cell factor mRNA and protein became increasingly localized to the cortical region of the ovary, where most germ cells were present as actively proliferating oogonia. From day 40 to day 90 of gestation, stem cell factor mRNA and protein localization were confined mainly to the ovarian cortex. Moreover, within the cortical region, stem cell factor mRNA was low or absent where follicles were first forming and highest in the outer ovarian cortex, where germ cells were undergoing mitosis or the early stages of meiosis. In contrast, stem cell factor protein was present in newly forming follicles, as well as in mitotic and meiotic germ cells, which is consistent with the presence of both membrane-bound and soluble forms of this ligand. However, by day 90 of gestation, both stem cell factor mRNA and protein were observed in the granulosa cells of most (> 90%) primordial follicles. C-kit mRNA and protein were observed from day 24 of gestation in both germ cells and somatic cells but, with increasing gestational age, preferentially in germ cells (for example, pre-meiotic germ cells and both isolated oocytes and follicle-enclosed oocytes). C-kit protein, but not mRNA, was also observed in germ cells that were undergoing meiosis. The results indicate that the cells containing stem cell factor mRNA within the ovary up to day 90 of gestation originated from the gonadal blastema and from cells that migrated from the mesonephros before day 28 of gestation. Since stem cell factor mRNA was absent in both mesonephric cells migrating after day 28 of gestation and in regions where follicles were first forming, it is suggested that these later migrating mesonephric cells are the progenitors of the granulosa cells in the first forming follicles. In conclusion, during follicle formation, c-kit mRNA is localized to germ cells whereas c-kit, together with stem cell factor protein, is localized to both germ cells and somatic cells, consistent with the hypothesis that the presence of this receptor-ligand pair is essential to prevent apoptosis.     

Phylogenetic significance of the structure of adult ovary and oogenesis in a primitive chilognathan diplopod,Hyleoglomeris japonica Verhoeff (Glomerida,Diplopoda)

Some histological details of the adult ovary of Hyleoglomeris japonica are described for the first time in the glomerid diplopods. The ovary is a single, long sac-like organ extending from the 4th to the 12th body segment along the median body axis, lying between the alimentary canal and the ventral nerve cord. The ovarian wall consists of a layer of thin ovarian epithelium which surrounds a wide ovarian lumen. A pair of longitudinal “germ zones,” including female germ cells, runs in the lateral ovarian wall. Each germ zone consists of two types of oogenetic areas: 1) 8–12 narrow patch-shaped areas for oogonial proliferation, arranged metamerically in a row along each of the dorsal and ventral peripheries, and 2) the remaining wide area for oocyte growth. Oogonial proliferation areas include oogonia, very early previtellogenic oocytes, and young somatic interstitial cells, among the ovarian epithelial cells. The larger early previtellogenic oocytes in the oogonial proliferation areas are located nearer to the oocyte growth area, and migrate to the oocyte growth area. They are surrounded by a layer of follicle cells and are connected with the ovarian epithelium of the oocyte growth area by a portion of their follicles. They grow into the ovarian lumen, but their follicles are still connected with the oocyte growth area. Various sizes of the previtellogenic and vitellogenic oocytes in the ovarian lumen are connected with the oocyte growth area; the smaller oocytes are connected nearer to the dorsal and ventral oogonial proliferation areas, while the larger ones are connected nearer to the longitudinal middle line of the oocyte growth area. Following the completion of vitellogenesis and egg membrane formation in the largest primary oocytes, the germinal vesicles break down. Ripe oocytes are released from their follicles directly into the ovarian lumen to be transported into the oviducts. Ovarian structure and oogenesis of H. japonica are very similar to those of other chilognathan diplopods. At the same time, however, some characteristic features of the ovary of H. japonica are helpful for understanding the structure and evolution of the diplopod ovaries. Some aspects of the phylogenetic significance in the paired germ zones of H. japonica are discussed. J. Morphol 231:277–285, 1997. © 1997 Wiley-Liss, Inc.     

Degeneration of germ line cells in amphibian ovary

Ogielska, M., Rozenblut, B., Augustyńska, R., Kotusz, A. 2010. Degeneration of germ line cells in amphibian ovary. —Acta Zoologica (Stockholm) 91 : 319–327 We studied the morphology of degenerating ovarian follicles in juvenile and adult frogs Rana temporaria, Rana lessonae and Rana ridibunda. Degeneration of primordial germ cells was never observed and was extremely rare in oogonia and early oocytes in a cyst phase in juveniles. Previtellogenic oocytes were rarely affected. Three main types of atresia were identified. In type I (subdivided into stages A–D), vitellogenic oocytes are digested by proliferating follicle cells that hypertrophy and become phagocytic. A – germinal vesicle shrinks, nucleoli fuse, oocyte envelope interrupts, and follicular cells hypertrophy; B – follicular cells multiply and invade the oocyte; C – entire vesicle is filled by phagocytic cells; D – degenerating phagocytes accumulate black pigment. Type II is rare and resembles breakdown of follicles and release of ooplasm. In type III, observed in previtellogenic and early vitellogenic oocytes, ooplasm and germinal vesicle shrink, follicle cells do not invade the vesicle, and condensed ooplasm becomes fragmented. The residual oogonia in adult ovaries (germ patches) multiply, but soon degenerate.     

埃塞俄比亚塔纳湖鲤科鱼类体内亚美尼亚许氏绦虫Khawia armeniaca (Cholodkovsky, 1915) Shulman, 1958 (绦虫纲: 鲤蠢目) 的形态鉴定

在埃塞俄比亚境内塔纳湖中发现了1种寄生于鲤科鱼类: 间魮(Labeobarbus intermedius)和Labeobarbus tsanensis体内的许氏绦虫, 经形态学鉴定其为亚美尼亚许氏绦虫Khawia armeniaca (Cholodkovsky, 1915)(绦虫纲: 鲤蠢目)。该绦虫鉴别特征为头节呈半球状, 边缘光滑, 无皱褶; 睾丸分布区域从虫体中部至阴茎囊之前; 卵巢前卵黄腺分布于虫体中后部至阴茎囊前部区域, 少数排列在阴茎囊之后; 子宫起始于阴茎囊后方弯曲环绕直至卵巢后部区域, 有少许卵黄腺排列在子宫和卵巢的两侧。此外, K. armeniaca卵巢呈滤泡状或蝴蝶状, 雌雄生殖孔分离但彼此距离很近, 开口于体表并形成同一生殖腔, 且雄性生殖孔位于雌性生殖孔前方。卵巢后卵黄腺数目少于100个, 分布于虫体末端, 卵黄腺距离卵巢后翼较远, 部分样品内卵黄腺接近卵巢后翼。     

New testicular mechanisms involved in the prevention of fetal meiotic initiation in mice

In mammals, early fetal germ cells are unique in their ability to initiate the spermatogenesis or oogenesis programs dependent of their somatic environment. In mice, female germ cells enter into meiosis at 13.5 dpc whereas in the male, germ cells undergo mitotic arrest. Recent findings indicate that Cyp26b1, a RA-degrading enzyme, is a key factor preventing initiation of meiosis in the fetal testis. Here, we report evidence for additional testicular pathways involved in the prevention of fetal meiosis. Using a co-culture model in which an undifferentiated XX gonad is cultured with a fetal or neonatal testis, we demonstrated that the testis prevented the initiation of meiosis and induced male germ cell differentiation in the XX gonad. This testicular effect disappeared when male meiosis starts in the neonatal testis and was not directly due to Cyp26b1 expression. Moreover, neither RA nor ketoconazole, an inhibitor of Cyp26b1, completely prevented testicular inhibition of meiosis in co-cultured ovary. We found that secreted factor(s), with molecular weight greater than 10 kDa contained in conditioned media from cultured fetal testes, inhibited meiosis in the XX gonad. Lastly, although both Sertoli and interstitial cells inhibited meiosis in XX germ cells, only interstitial cells induced mitotic arrest in germ cell. In conclusion, our results demonstrate that male germ cell determination is supported by additional non-retinoid secreted factors inhibiting both meiosis and mitosis and produced by the testicular somatic cells during fetal and neonatal life.     

Gametogenesis in Fundulus heteroclitus

Our present understanding of the structure of the gonads andof gametogenesis in Fundulus helerodilus is briefly reviewed. The testes contain spermatogenic tubules which distally arecomposed of linearly arranged germinal cysts with clones ofsynchronously developing germ cells, and proximally of efferentducts with mature sperm. Within the tubules, those cysts containingspermatogonia are found close to the testicular periphery, whereascysts with mature sperm are contiguous with the efferent ducts.Cytological details of the three principal stages of spermatogenesis,namely spermatocytogenesis, meiosis, and spermiogenesis, arepresented along with examples of the various somatic cells ofthe testis. The ovary consists of numerous ovigerous lamellae which containrandomly arranged follicles in various stages of development.Since follicular growth is asynchronous, follicles of all sizesare present in the ovary during the breeding season and eggsare continuously ovulated into the ovarian lumen. Oocyte growthis divided into five distinct stages: Primary growth (substagesA and B), cortical alveolus formation, vitellogenesis, oocytematuration (substages A and B), and the ovulated egg. This seriesof stages is based both on cytological observations and physiologicalevents.     

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.     

Ultrastructural aspects of gonadal morphogenesis in Bufo bufo (Amphibia Anura) 1. Sex differentiation

The morphogenesis of gonads in Bufo bufo tadpoles was studied, and ultrastructural differences between sexes were identified. All specimens analyzed initially developed gonads made up of a peripheral fertile layer (cortex) surrounding a small primary cavity. Subsequently a central layer of somatic cells (medulla) developed. Both layers were separated by two uninterrupted basal laminae between which a vestige of the primary cavity persisted. During female differentiation, the peripheral layer continued to be the fertile layer. In males, the central layer blended into the peripheral layer and the basal laminae disappeared. The somatic cells of the central layer came into direct contact with the germ cells; this did not occur in females. Testicular differentiation continued with the migration of germ cells towards the center of the gonad. The somatic elements surrounding the germ cells appeared to play an active role in their transfer to the center of the gonad. The peripheral layer shrank and became sterile. Two basal laminae then re-formed to separate the fertile central layer from the peripheral sterile one. Germ cells have always been thought to perform a passive role in sex differentiation in amphibians. Following the generally accepted "symmetric model", the mechanism of gonad development is symmetrical, with cortical somatic cells determining ovarian differentiation and medullary somatic cells determining testicular differentiation. In contrast, we found that sex differentiation follows an "asymmetric" pattern in which germ cells tend primarily toward a female differentiation and male differentiation depends on a secondary interaction between germ cells and medullary somatic cells.     

Germ cells are not the primary factor for sexual fate determination in goldfish

The presence of germ cells in the early gonad is important for sexual fate determination and gonadal development in vertebrates. Recent studies in zebrafish and medaka have shown that a lack of germ cells in the early gonad induces sex reversal in favor of a male phenotype. However, it is uncertain whether the gonadal somatic cells or the germ cells are predominant in determining gonadal fate in other vertebrate. Here, we investigated the role of germ cells in gonadal differentiation in goldfish, a gonochoristic species that possesses an XX-XY genetic sex determination system. The primordial germ cells (PGCs) of the fish were eliminated during embryogenesis by injection of a morpholino oligonucleotide against the dead end gene. Fish without germ cells showed two types of gonadal morphology: one with an ovarian cavity; the other with seminiferous tubules. Next, we tested whether function could be restored to these empty gonads by transplantation of a single PGC into each embryo, and also determined the gonadal sex of the resulting germline chimeras. Transplantation of a single GFP-labeled PGC successfully produced a germline chimera in 42.7% of the embryos. Some of the adult germline chimeras had a developed gonad on one side that contained donor derived germ cells, while the contralateral gonad lacked any early germ cell stages. Female germline chimeras possessed a normal ovary and a germ-cell free ovary-like structure on the contralateral side; this structure was similar to those seen in female morphants. Male germline chimeras possessed a testis and a contralateral empty testis that contained some sperm in the tubular lumens. Analysis of aromatase, foxl2 and amh expression in gonads of morphants and germline chimeras suggested that somatic transdifferentiation did not occur. The offspring of fertile germline chimeras all had the donor-derived phenotype, indicating that germline replacement had occurred and that the transplanted PGC had rescued both female and male gonadal function. These findings suggest that the absence of germ cells did not affect the pathway for ovary or testis development and that phenotypic sex in goldfish is determined by somatic cells under genetic sex control rather than an interaction between the germ cells and somatic cells.     

Histological and ultrastructural investigation of early gonad development and sex differentiation in Adriatic sturgeon (Acipenser naccarii,Acipenseriformes, Chondrostei)

Gonad development and sex differentiation from embryos to 594‐day‐old individuals were investigated in farmed Acipenser naccarii using light and transmission electron microscopy. The migrating primordial germ cells first appear along the dorsal wall of the body cavity in embryos 1.5 days before hatching. The gonadal ridge, containing a few primary primordial germ cells (PGC‐1) surrounded by enveloping cells, appears in 16‐day‐old larvae. At 60 days, the undifferentiated gonad is lamellar and PGC‐1 multiply, producing PGC‐2. In 105‐day‐old juveniles, a distinct germinal area with advanced PGC‐2 appears on the lateral side near the mesogonium and the first blood vessels are visible. At 180 days, putative ovaries with a notched gonadal epithelium and putative testes with a smooth one appear, together with adipose tissue on the distal side. In 210‐day‐old juveniles, active proliferation of germ cells begins in the putative ovaries, whereas putative testes still contain only a few germ cells. The onset of meiosis and reorganization of stromal tissue occurs in ovaries of 292‐day‐old individuals. Ovaries with developed lamellae enclosing early oocyte clusters and follicles with perinucleolar oocytes occur at 594 days. Meiotic stages are never found, even in anastomozing tubular testes of 594‐day‐old individuals. Steroid producing cells are detected in the undifferentiated gonad and in the differentiated ones of both sexes. Anatomical differentiation of the gonad precedes cytological differentiation and female differentiation largely precedes that of the male. Gonad development and differentiation are also associated with structural changes of connective tissue, viz. collagen‐rich areas are massive in developing testes and reduced in ovaries. J. Morphol., 2008. © 2008 Wiley‐Liss, Inc.