Functions of the testicular gland in two blenniid fishes, Salaria (=Blennius) pavo and Lipophrys (= Blennius) dalmatinus (Blenniidae, Teleostei) as revealed by electron microscopy and enzyme histochemistry

The functions of the testicular gland in two different blenniid fishes, Salaria pavo and Lipophrys dulmutinus , are described by fine structural and enzyme histochemical methods. In the testes of the two fishes no mature spermatozoa are found. Sperrniogenesis occurs only until the spermatidal stage. Spermatids are released into the testicular gland. During the spawning period the testicular gland functions in differentiation of spermatids, nutrition of spermatids, and secretion of sialomucins. After spawning, the testicular gland has phagocytotic functions in resorbtion of remaining spermatids, which are transformed into lipids in the gland cells. During the interspawning period the testicular gland is a storage reservoir for lipids and phospholipids which are re-transformed into testicular gland secretion in the next reproductive season. Testicular gland cells themselves do not have steroidogenetic functions, but steroids are synthesized by interstitial cells homologous to Leydig cells in other fish. Possible explanations for the reduction of the testis in L. dalmatinus and implications of the testicular gland in taking over testicular functions are discussed.     

Breeding ecology and nest aggregations in a population of Salaria pavo (Pisces: Blenniidae) in an area where nest sites are very scarce

In this paper we present data on the breeding ecology and patterns of nest aggregation in a population of Salaria pavo (Pisces: Blenniidae), in an area where hard substrates are very scarce. The study site was at Ria Formosa, an extensive littoral lagoon on the south coast of Portugal. The only hard materials available are bricks, tiles, stones and debris that clam culturists use to delimit their fields. The males of S. pavo establish nests intertidally in the holes of the bricks where they guard the eggs. Data were collected at low tide inspections of these artificial ridges and underwater behavioural observations during high tide. The main results are: (i) the same male can establish sequentially more than one nest in the same breeding season; the number of males that stay within the bricks increases before the start of the breeding season and declines towards its end; (ii) there is a large excess of mature males that do not establish nests and they are significantly smaller than the nesting males, suggesting very strong competition for nest sites; (iii) the nests of several males occur in adjacent holes of the same brick, sometimes being entirely surrounded by other nests. Thus, in this population, there is virtually no defended territory around the nest, a situation not known for other blenniid species. This pattern of nest aggregation implies both strong competition among males and reduction of aggression between neighbour parental fishes. This can be explained by the extreme scarcity and spatial distribution of available nest sites.     

Histological, fine-structural and histochemical differences in the testicular glands of gobiid and blenniid fishes

The testicular gland (t.g.) is a glandular tissue situated adjacent to the testis of blenniid and several gobiid species. In the present study the t.g. of Blennius pavo Risso and Gobius niger L. were compared by histological and histochemical methods. In B. pavo the spermatozoa have to cross the t.g. to reach the vas deferens and thus they come into contact with the gland cells, whereas in G. niger the vas deferens is situated between the testis and the t.g. The fine structure and histo-chemistry of the t.g. cells reveal that in B.pavo the cells of the t.g. have exocrine as well as endocrine functions. The t.g. cells of B. pavo contain large amounts of lipids, form a secretion containing acid mucopolysaccharides, show positive reaction for acid phosphatase, and some cells stain for 3β-HSD and G6PD. The function of the t.g. of G. niger is exclusively endocrine. Characteristics of the gland cells of this species are well developed smooth ER and tubulovesicular or paracristal-line mitochondria. The stainings for 3β-HSD, G6PD and UDPGD give strong positive results in the whole t.g., indicating the presence of steroids and steroid glucuronides.     

Gonadal sex differentiation in Atlantic halibut

The process of gonadal sex differentiation in 338 Atlantic halibut Hippoglossus hippoglossus larvae, ranging in size from 10 mm L_s to 230 mm L _F, is described histologically. Gonadal sex differentiation occurred by 38.0 mm L _F, which coincided with the weaned, post-metamorphic, settled stage in the life cycle. This was a gradual process that coincided with other organogenesis in the developing larvae.     

Germ cell-less expression in medaka (Oryzias latipes)

The gene germ cell-less (gcl) plays an important role in the early differentiation of germ cells in Drosophila. We isolated the gcl homolog of the model teleost medaka (Oryzias latipes) using degenerated primers and an ovary cDNA bank. The predicted amino acid sequence of medaka gcl showed 92, 68 and 31% overall identity to mouse, human and Drosophila gcl respectively. RT-PCR revealed stronger expression in the ovary and weaker expression in testis, brain, heart, liver and muscle tissue. Expression in early embryos indicates the presence of maternal mRNA. By in situ hybridisation (ISH), gcl could not be detected in embryos. In contrast to vasa, ISH revealed expression of gcl in the ovary but not in the testis. Mol. Reprod. Dev. 67: 15-18, 2004.     

中国林蛙性腺的发育及温度对其性别分化的影响

为探讨幼蛙性别分化与温度的关系,在恒温和变温条件下培养中国林蛙（Rana chensinensis)受精卵至变态完成,结果表明：（1）胚胎发育到24期时生殖嵴开始出现,25期个别原始生殖细胞（PGCs)已迁移到生殖嵴中,生殖细胞与生殖嵴共同发育成生殖腺;（2）胚胎发育到31期生殖腺出现性别分化,卵巢分化初期较易识别,而精巢分化不明显;…（3）卵巢分化完成于37期,精巢分化完成于变态之后,两侧生殖腺等大;（4）胚胎发育从30期开始,性别分化对温度较为敏感,低温利于雌性化,高温利于雄性化;（5）15－25℃为变温培养时性比发生变化的敏感温度区,缓慢升温雄性比较显著增加,缓慢降温雌性比例显著增加。     

Genetic diversity and differentiation between Palearctic and Nearctic populations of Aedimorphus (=Aedes) vexans (Meigen, 1830) (Diptera,Culicidae)

Genetic diversity was studied at allozyme loci in two Palearctic and one Nearctic population of Aedimorphus (=Aedes) vexans, a species of public health and veterinary importance. The population from Serbia was the most polymorphic (P= 35%) with the highest observed heterozygosity (H_o= 0.027). The lowest observed heterozygosity (H_o= 0.010) was obtained for the Nearctic population. All analyses based on individual (STRUCTURE analysis) and population level (pairwise F_ST,Nm values, AMOVA, Nei's D value) revealed significant structuring between Nearctic and Palearctic populations, indicating a lack of gene flow and thus, the presence of independent gene pools. Taxon‐specific alleles at the diagnostic Ao, Hk‐2, Hk‐3, Hk‐4, Idh‐1, and Idh‐2 loci were used for identification and separation of Nearctic and Palearctic populations. Population genetics study provided valuable information on the correct distinction of Am. vexans populations and their adaptive potential that could find a future use in the studies of vector competence and development of vector‐control strategies.     

Sexual differentiation and juvenile intersexuality in the European sea bass (Dicentrarchus labrax)

Sexual differentiation was studied at the histological level using a mixture of 30 families of sea bass Dicentrarchus labrax . Most of the fish (93%) differentiated into males as usually observed in farmed populations. All testes were differentiated when the males reached 12 cm and no more undifferentiated fish were found from 419 days post-fertilization (p.f.). In 28% of the males, among the biggest, sexual differentiation had already begun at 168 days p.f. (8.3–9.5 cm) and these fish started spermatogenesis in their first year of life. The other males differentiated later and remained immature at the end of their first year of life. Ovaries could be identified at the histological level from the age of 168 days p.f. (7.9–9.0 cm) and the females became significantly longer than the males from the age of 191 days p.f., i.e. during the process of ovarian differentiation. In the studied group, 62% of the males developed intratesticular oocytes. Such intersexuality had no consequence on growth rate. Intratesticular oocytes were also recorded in testes of wild males originating from Atlantic (Britain and Gulf of Gascogne) and West Mediterranean showing that juvenile intersexuality is not restricted to farmed populations but is a widespread phenomenon in sea bass.     

Differentiation of female chicken primordial germ cells into spermatozoa in male gonads

In avian species, the developmental fate of different-sex germ cells in the gonads is unclear. The present study attempted to confirm whether genetically female germ cells can differentiate into spermatozoa in male gonads using male germline chimeric chickens produced by the transfer of primordial germ cells (PGC), and employing molecular biological methods. As a result of Southern hybridization, specific sequences of the W chromosome (the female specific sex chromosome in birds) were detected in the genomic DNA extracted from one out of four male germline chimeric chickens. When two-color in situ hybridization was conducted on the spermatozoa of this germline chimera, 0.33% (average) of the nuclei of each semen sample showed the fluorescent signal indicating the presence of the W chromosome. The present study shows that female PGC can differentiate into spermatozoa in male gonads in the chicken. However, the ratio of produced W chromosome-bearing (W-bearing) spermatozoa fell substantially below expectations. It is therefore concluded that most of the W-bearing PGC could not differentiate into spermatozoa because of restricted spermatogenesis.     

Differentiation of female primordial germ cells in the male testes of chicken (Gallus gallus domesticus)

In our previous studies, we demonstrated that female primordial germ cells (PGCs) have the ability to differentiate into W chromosome-bearing (W-bearing) spermatozoa in male gonads of germline chimeric chickens. In this study, to investigate the differentiation pattern of female PGCs in male gonads in chickens, three germline chimeric chickens were generated by injecting female PGCs into the male recipient embryos. After these male chimeras reached sexual maturity, the semen samples were analyzed for detecting W-bearing cells by PCR and in situ hybridization analyses. The results indicated that the female PGCs had settled and differentiated in their testes. A histological analysis of the seminiferous tubule in those chimeras demonstrated that the W-bearing spermatogonia, spermatocytes, and round spermatids accounted for 30.8%, 32.7%, and 28.4%, respectively. However, the W-bearing elongating spermatid was markedly lower (7.7%) as compared to the W-bearing round spermatid. The W-bearing spermatozoa were hardly ever observed (0.2%). We concluded that although female PGCs in male gonads are capable of passing through the first and second meiotic division in adapting themselves to a male environment, they are hardly complete spermiogenesis.     

Sexual differentiation of brain and behavior in the zebra finch: Critical periods for effects of early estrogen treatment

In order to determine the critical period(s) during which estrogen alters sexually dimorphic behavior and neuroanatomy in zebra finches (Poephila guttata), nestlings were injected daily 20 μg estradiol benzoate (EB) during posthatching week 1, week 2, week 3, or weeks 1, 2, and 3. At 7 months of age, birds were implanted with testosterone propionate and tested with female partners for singing, dancing, and copulatory mounting. Brains were subsequently processed for morphometry, and the volumes of the song system nuclei HVC, area X, and RA and the soma sizes and densities of neurons in RA were determined. Males given EB during week 1 failed to mount. Females given EB during week 1 were fully masculinized with respect to dancing and RA neuron soma size and density, and were partially masculinized with respect to song nuclei volumes and singing. Treatment beginning after week 1 was ineffective or less effective for all measures. Only for RA neuron measures was treatment for all three weeks more effective than week 1 treatment. Thus the first post-hatching week is the most influential period of those tested for effects of exogenous estrogen on sexual differentiation in this species, and is a period during which both masculinization of females and demasculinization of males is possible. 1994 John Wiley & Sons, Inc.     

多能干细胞向雄性生殖细胞定向分化的研究进展

生殖健康是生命科学领域关注的核心之一,各种原因所致男性不育亟待解决,然而由于伦理限制等原因,缺少合适的具有人类遗传背景的研究模型开展研究。胚胎干细胞（ESCs）和诱导多能干细胞（iPSCs）均属于多能干细胞,具有多向分化潜能。一方面,可利用ESCs?/?iPSCs向生殖细胞分化的模型研究人类生殖细胞的发育规律,另一方面,在此基础上,可建立带有人类疾病遗传背景的iPSCs模型,体外诱导其向雄性生殖细胞分化,利用该模型研究男性不育的发病机制。由于精子在体内的形成遵循一定规律,体外环境中不同发育阶段的生殖细胞在不同诱导因子作用下才可稳定地往下一阶段定向分化,因此,诱导ESCs?/?iPSCs向雄性生殖细胞方向分化时,诱导因子的种类和加入时间的选择应根据生殖细胞的体内发育特征而定,并且在诱导的不同阶段循序加入,以此模拟精子在体内的形成过程,进而更好地研究男性不育的发病机制。本文将对多能干细胞向雄性生殖细胞定向分化的常用诱导因子及存在问题和展望进行综述,为相关研究的开展提供借鉴。     

EED is required for mouse primordial germ cell differentiation in the embryonic gonad

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Cell cycle analysis of fetal germ cells during sex differentiation in mice

Background information. Primordial germ cells in developing male and female gonads are responsive to somatic cell cues that direct their sex‐specific differentiation into functional gametes. The first divergence of the male and female pathways is a change in cell cycle state observed from 12.5 dpc (days post coitum) in mice. At this time XY and XX germ cells cease mitotic division and enter G₁/G₀ arrest and meiosis prophase I respectively. Aberrant cell cycle regulation at this time can lead to disrupted ovarian development, germ cell apoptosis, reduced fertility and/or the formation of germ cell tumours. Results. In order to unravel the mechanisms utilized by germ cells to achieve and maintain the correct cell cycle states, we analysed the expression of a large number of cell cycle genes in purified germ cells across the crucial time of sex differentiation. Our results revealed common signalling for both XX and XY germ cell survival involving calcium signalling. A robust mechanism for apoptosis and checkpoint control was observed in XY germ cells, characterized by p53 and Atm (ataxia telangiectasia mutated) expression. Additionally, a member of the retinoblastoma family and p21 were identified, linking these factors to XY germ cell G₁/G₀ arrest. Lastly, in XX germ cells we observed a down‐regulation of genes involved in both G₁‐ and G₂‐phases of the cell cycle consistent with their entry into meiosis. Conclusion. The present study has provided a detailed analysis of cell cycle gene expression during fetal germ cell development and identified candidate factors warranting further investigation in order to understand cases of aberrant cell cycle control in these specialized cells.     

KIF2Abeta: A kinesin family member enriched in mouse male germ cells, interacts with translin associated factor-X (TRAX)

Translin associated factor X (TRAX) is a binding partner of TB-RBP/Translin. A cDNA encoding the 260 C-terminal amino acids of KIF2Abeta was isolated from mouse testis cDNAs in a yeast two-hybrid library screen for specific TRAX-interacting proteins. KIF2Abeta was expressed predominantly in the mouse testis and enriched in germ cells. The interaction of full-length KIF2Abeta or its C-terminus with TRAX was verified using in vitro synthesized fusion proteins. Deletion mapping of the TRAX-binding region of KIF2Abeta indicated that amino acids 514-659 were necessary and sufficient for the interaction in vivo. Confocal microscopy studies using GFP-fusion proteins demonstrated that KIF2Abeta colocalizes with TRAX in a perinuclear location. KIF2Abeta does not interact with TB-RBP, suggesting that either TRAX can function as an adaptor molecule for motor proteins and TB-RBP, or that this interaction reveals an undescribed role for TRAX in germ cells. The interaction with KIF2Abeta suggests a role for TRAX in microtubule-based functions during spermatogenesis.     

Sexual dimorphism and seasonal variation in submandibular gland histology of Bolomys lasiurus (Rodentia,Muridae)

Wild rodents (Bolomys lasiurus) of both sexes were caught in a cerrado grassland area during the dry (July-September) and rainy (January-March) seasons of Brazil. Fasted animals were perfused with Karnovsky fixative through the left ventricle, under ether anesthesia, and the submandibular gland was processed for embedding in historesin. Histological and histometric data show sexual dimorphism at both seasons. In the volume percentage of the granular convoluted tubules (GCT) and their secretory granules, the males exhibited higher values. The absolute volume occupied by these structures, however, was dimorphic only in the rainy season. The diameter of the GCT, the height of its epithelium, and its total length were also greater in males during the rainy season. The absolute volumes of the acini and of the ductal tree were identical in both sexes in the dry and rainy seasons but the acinar diameter increased in the males and females during the rainy season. The sexual dimorphism and the seasonal variations now described in the B. lasiurus submandibular glands could be explained by the augmented reproductive activity of the males in the rainy period.