Epididymal maturation and ejaculation are key events for further in vitro capacitation of boar spermatozoa

Mammalian spermatozoa acquire functionality during epididymal maturation, and the ability to penetrate and fertilize the oocyte during capacitation. The aim of this study was to assess the effects of epididymal maturation, ejaculation and in vitro capacitation on sperm viability, acrosome integrity, mitochondrial activity, membrane fluidity, and calcium influx, both as indicators of capacitation status and sperm motility. Results indicated that boar spermatozoa acquired the ability to move in the epididymal corpus; however, their motility was not linear until the ejaculation. Epididymal spermatozoa showed low membrane fluidity and intracellular calcium content; ejaculation led to an increased calcium content, while membrane fluidity showed no changes. Acrosome integrity remained constant throughout the epididymal duct and after ejaculation and in vitro capacitation. The frequency of viable spermatozoa with intact mitochondrial sheath was higher in caput and ejaculated samples than in corpus and cauda samples, whereas the frequency of spermatozoa with high membrane potential was significantly lower in cauda samples. In vitro capacitation resulted in a decreased frequency of viable spermatozoa with intact mitochondrial sheath and an increased frequency of spermatozoa with high membrane potential in ejaculated samples. These results indicated that both epididymal maturation and ejaculation are key events for further capacitation, because only ejaculated spermatozoa are capable of undergoing the set of changes leading to capacitation.     

Natural cyclic degeneration by a sequence of programmed cell death modes in <Emphasis Type=Italic>Semibalanus balanoides</Emphasis> (Linnaeus, 1767) (Crustacea,Cirripedia Thoracica)

The male reproductive system of the barnacle Semibalanus balanoides (Linnaeus, 1767) is a perfect model system for investigating naturally occurring cyclic nonpathological degeneration and programmed cell death (PCD). Every year after copulation, the barnacle degenerates its gonads, eggs, spermatids and penis initiated by severe environmental constraints. This is apparently a useful strategy for saving energy. By careful ultrastructural and immunohistochemical analysis of the male reproductive organs, we identified autophagic cell death in the penis and spermatids, both morphologically on ultrathin sections as well as histochemically for the first time on semithin tissue sections with the new antibody against microtubule-associated protein 1 light chain 3 (LC3). Apoptosis in the testis and vesicula seminalis was determined based on the morphological characters on the apoptosis-specific antibody against the apoptosis inducing factor (AIF), and on positive TUNEL (terminal in situ nick end labeling) nuclei. Secondary necrosis in late degrading stages occurred in all tissues investigated, based on morphological characters and weak TUNEL staining. Moreover, the ultrastructural screening showed transient forms of cell death in the ductus ejaculatorius epithelium, the epidermis, and the longitudinal muscles of the penis. According to the immunohistochemical tests, the cyclic degeneration of the male reproductive system follows a sequence of programmed cell death modes from autophagic cell death via apoptosis to secondary necrosis.     

草地螟雄蛾生殖系统的形态和结构

针对目前关于草地螟Loxostege sticticalis L.雄成虫生殖系统结构和功能的研究相对缺乏的状况,本文利用光镜和扫描电镜系统研究了草地螟雄蛾的内外生殖器官及相关结构。草地螟雄蛾内外生殖器官集中于腹部第5¹0腹节;内生殖器官位于510腹节;内生殖器官位于5⁸腹节内腔中,由睾丸(testes)、贮精囊(seminal vesicle)、输精管(vas deferens)、附腺(accessory gland)和射精管(ejaculatory ducts)组成;外生殖器官为腹部第9、10腹节的特化结构,包括抱握器(harpes)、背兜(tegumen)、基腹弧(vinculum)、钩形突(uncus)、颚形突(gnathos)、阳茎囊(aedeagal caecum)和阳茎(phallus)。外生殖器中抱握器的端刺(furcella)方向为雄蛾区别于其它种类的一个重要形态学特征。该研究首次明确了草地螟雄蛾的生殖系统结构,并为锥额野螟蛾属中种间分类提供更多的科学依据。     

山羊体外受精的研究

通过在山羊卵母细胞体外成熟培养液中添加不同的血清和不同浓度的卵泡液 ,在体外成熟培养液中培养不同的时间 ,以及采用不同的精子获能方法来摸索效率较高的体外受精方法体系。在山羊卵母细胞体外成熟培养液中添加FCS、EGS及不同浓度的卵泡液 ,成熟培养时间分别为 16、2 0、2 4和 2 7h ,山羊新鲜精液用钙离子载体法和肝素法进行获能处理后用于体外受精 ,比较其体外成熟率和体外受精率。添加FCS、EGS和 2 0 %卵泡液组的成熟率无显著差异 ,显著高于添加 10 %和 3 0 %卵泡液组的成熟率 ;但添加FCS和EGS组的受精率显著高于添加10 %、2 0 %、3 0 %卵泡液组的受精率。培养 2 4h组和 2 7h组的成熟率显著高于另外两组 ,而培养 2 7h组的受精率显著高于其余各组。用钙离子载体法处理的山羊精子的顶体反应率和体外受精率显著高于肝素法。EGS可以代替FCS添加于成熟培养液中 ,对COCs的成熟率和受精率没有明显的影响 ,但卵泡液不能完全代替FCS的作用。培养时间为 2 7h ,精子用钙离子载体法处理获能后能得到较高的体外受精率     

甲壳动物的生殖量与环境关系：Ⅰ.枝角类

生殖量是生殖潜力(reproductivepotential)的一个主要标志,也是生态系统(含生产力)和种群生态学(含数量变动)的一个主要内容。它在很大程度上是由遗传因子(基因)决定的,表现在各类动物的生殖量有很     

Characterization,expression, and functional analysis of testis-specific serine/threonine kinase 1 (Tssk1) in the pen shell Atrina pectinata

Abstract

Testis-specific serine/threonine kinase 1 (Tssk1) was named since its function was determined in mouse spermiogenesis. In our research, we cloned a homolog of the pen shell Atrina pectinata Tssk1, ApTssk1, and determined its expression characteristics at mRNA. The full length of ApTssk1 cDNA was 1517 bp, with an open reading frame of 1089 bp (58–1146), which encodes a peptide including 362 amino acids. The homologous analysis indicated that the deduced amino acid sequences of ApTssk1 shared a close identity with other reported Tssk1. In addition, the highly conserved fragment containing a serine/threonine protein kinase catalytic (S-TKc) domain was predicted to exist in ApTssk1. Results from the RT-qPCR display that ApTssk1 was only transcribed in the male gonad of A. pectinata, and was highest in the mature testis. Therefore, this study indicated that ApTssk1 might play a functional role during spermatogenesis in A. pectinata.     

X-Y chromosome univalency in the testes of hyperthermic mice: I. Concomitant formation of multinucleated giant cells

Exposure of mice to an environment of approximately 35°C and 65% relative humidity for 2–5 days caused dissociation of the X-Y chromosome bivalent in diakinesis-metaphase I (primary) spermatocytes. Dissociation of autosomal bivalents, chromosome fragmentation, aneuploidy, and polyploidy were not significantly affected by heat stress. Heat stress also caused formation of multinucleated giant cells in the mouse testis. Giant cell formation followed a distribution similar to X-Y dissociation, reaching highest levels 5 days after heat stress and returning to control levels 50–70 days after heat stress. Spermatogonial activity was not permanently damaged by these conditions, and the germinal epithelium was able to repopulate. We believe that cell aberration or cell death resulting from either or both of these anomalies causes much of the decreased fertility found after subjecting scrotal mammals to high temperatures.     

Semicystic spermatogenesis and reproductive strategy in Ophidion barbatum (Pisces, Ophidiidae)

In this paper the testicular structure and spermatogenesis of Ophidion barbatum are studied and the reproductive strategy of this species is analysed. The species has a rare type of spermatogenesis, called semicystic, in which the cyst ruptures in a stage prior to the spermatozoon stage. The most peculiar feature of the spermatozoon is its elongated shape, not typical of an oviparous species. Taking into account our results, the type of ovary and the spawning method, this species shows specialized characteristics which are fairly uncommon among oviparous species. The analysis of other biological aspects of the species, such as the capacity of the males to produce sounds, the low population density, the habit of individual members of burying themselves and the crepuscular habits, combined with our observations concerning their reproductive strategies, lead us to suggest a unique mating behaviour.     

Cloning,Characterization and Primary Function Study of a Novel Gene,Cymg1,Related to Family 2 Cystatins

Cystatins are cysteine proteinase inhibitors,We found two expression sequence tags (ESTs),CA463109 and AV042522,from a mouse testis library using Digital differential display (DDD).By electricalhybridization,a novel gene,Cymgl(GenBank accession No.AY600990),which has a full length of 0.78 kb,and contains four exons and three introns,was cloned from a mouse testis eDNA library.The gene is locatedin the 2G3 area of chromosome 2.The full eDNA encompasses the entire open reading frame,encoding 141amino acid residues.The protein has a cysteine protease inhibitor domain that is related to the family 2cystatins but lacks critical consensus sites important for cysteine protease inhibition.These characteristicsare seen in the CRES subfamily,which are related to the family 2 cystatins and are expressed specifically inthe male reproductive tract.CYMG1 has a 44%(48/108)identity with mouse CRES and 30%(42/140)identity with mouse cystatin C.Northern blot analysis showed that the Cymgl is specifically expressed inadult mouse testes.Cell location studies showed that the GFP-tagged CYMG 1 protein was localized in thecytoplasm of HeLa cells,lmmunohistochemistry revealed that the CYMG1 protein was expressed in mousetestes spermatogonium,spermatocytes,round spermatids,elongating spermatids and spermatozoa.RT-PCRresults also showed that Cymgl was expressed in mouse testes and spermatogonium.The Cymgl expressionlevel varied in different developmental stages:it was low 1 week postpartum,steadily increased 2 to 5 weekspostpartum,and was highest 7 weeks postpartum.The expression level at 5 weeks postpartum was main-tained during 13 to 57 weeks postpartum.The Cymgl expression level in the testes over different develop-mental stages correlates with the mouse spermatogenesis and sexual maturation process.All these indicatethat Cymgl might play an important role in mouse spermatogenesis and sexual maturation. Cystatins are cysteine proteinase inhibitors,We found two expression sequence tags(ESTs),CA463109 and AV042522,from a mouse testis library using Digital differential display (DDD).By electricalhybridization,a novel gene,Cymgl(GenBank accession No.AY600990),which has a full length of 0.78 kb,and contains four exons and three introns,was cloned from a mouse testis eDNA library.The gene is locatedin the 2G3 area of chromosome 2.The full eDNA encompasses the entire open reading frame,encoding 141amino acid residues.The protein has a cysteine protease inhibitor domain that is related to the family 2cystatins but lacks critical consensus sites important for cysteine protease inhibition.These characteristicsare seen in the CRES subfamily,which are related to the family 2 cystatins and are expressed specifically inthe male reproductive tract.CYMG1 has a 44%(48/108)identity with mouse CRES and 30%(42/140)identity with mouse cystatin C.Northern blot analysis showed that the Cymgl is specifically expressed inadult mouse testes.Cell location studies showed that the GFP-tagged CYMG 1 protein was localized in thecytoplasm of HeLa cells,lmmunohistochemistry revealed that the CYMG1 protein was expressed in mousetestes spermatogonium,spermatocytes,round spermatids,elongating spermatids and spermatozoa.RT-PCRresults also showed that Cymgl was expressed in mouse testes and spermatogonium.The Cymgl expressionlevel varied in different developmental stages:it was low 1 week postpartum,steadily increased 2 to 5 weekspostpartum,and was highest 7 weeks postpartum.The expression level at 5 weeks postpartum was main-tained during 13 to 57 weeks postpartum.The Cymgl expression level in the testes over different develop-mental stages correlates with the mouse spermatogenesis and sexual maturation process.All these indicatethat Cymgl might play an important role in mouse spermatogenesis and sexual maturation.     

Cloning of a novel gene,<Emphasis Type=Italic>Cymg1</Emphasis>, related to family 2 cystatins and expressed at specific stages of mouse testis development

We have cloned a novel gene,Cymg1 (GenBank accession number AY600990), from a mouse testis cDNA library.Cymg1 is located in 2G3 of mouse chromosome 2. The cDNA includes an open reading frame that encodes 141 amino acid residues. The encoded polypeptide has a cysteine protease inhibitor domain found in the family 2 cystatins but lacks critical consensus sites important for cysteine protease inhibition. These characteristics are seen in the proteins of the CRES subfamily of the family 2 cystatins which are expressed specifically in the reproductive tract. CYMG1 protein shows 44% identity with mouse CRES and 30% identity with mouse cystatin C. Northern blot analysis showed that theCymg1 gene was specifically expressed in adult mouse testis. RT-PCR also showed thatCymg1 was expressed in testis and spermatogonial cells.Cymg1 expression level varied in the different developmental stages of mouse testis, and were coincidental with spermatogenesis and sex maturation. These results indicate thatCymg1 may play important roles in mouse spermatogenesis and sex maturation     

Cloning, characterization and primary function study of a novel gene, Cymg1, related to family 2 cystatins

Cystatins are cysteine proteinase inhibitors. We found two expression sequence tags (ESTs), CA463109 and AV042522, from a mouse testis library using Digital differential display (DDD). By electrical hybridization, a novel gene, Cymg1 (GenBank accession No. AY600990), which has a full length of 0.78kb, and contains four exons and three introns, was cloned from a mouse testis cDNA library. The gene is located in the 2G3 area of chromosome 2. The full cDNA encompasses the entire open reading frame, encoding 141 amino acid residues. The protein has a cysteine protease inhibitor domain that is related to the family 2 cystatins but lacks critical consensus sites important for cysteine protease inhibition. These characteristics are seen in the CRES subfamily, which are related to the family 2 cystatins and are expressed specifically in the male reproductive tract. CYMG1 has a 44% (48/108) identity with mouse CRES and 30% (42/140) identity with mouse cystatin C. Northern blot analysis showed that the Cymg1 is specifically expressed in adult mouse testes. Cell location studies showed that the GFP-tagged CYMG1 protein was localized in the cytoplasm of HeLa cells. Immunohistochemistry revealed that the CYMG1 protein was expressed in mouse testes spermatogonium, spermatocytes, round spermatids, elongating spermatids and spermatozoa. RT-PCR results also showed that Cymg1 was expressed in mouse testes and spermatogonium. The Cymg1 expression level varied in different developmental stages: it was low 1 week postpartum, steadily increased 2 to 5 weeks postpartum, and was highest 7 weeks postpartum. The expression level at 5 weeks postpartum was maintained during 13 to 57 weeks postpartum. The Cymg1 expression level in the testes over different developmental stages correlates with the mouse spermatogenesis and sexual maturation process. All these indicate that Cymg1 might play an important role in mouse spermatogenesis and sexual maturation.     

Reproductive anatomy of three nulliparous female Asian elephants: The development of artificial breeding techniques

Detailed gross examinations of the reproductive tracts of three mature female nulliparous Asian elephants were conducted to develop artificial insemination (AI) techniques. Of primary concern was the determination of the length characteristics and the size and configuration of the foramina between segments of the tract. The elephants were 13, 28, and 40 years of age and had been maintained in captivity for most of their lives. One elephant died naturally and two were euthanized for health reasons. The reproductive tracts of two of the elephants were manually palpated in situ via the urogenital canal. A fibreoptoscope was used to visualize the internal structures of the terminal reproductive tract of one elephant and to deposit dye into the vagina. The reproductive organs were removed from the body cavity, dissected, measured, and photographed. The major anatomical obstacles to overcome for standard AI procedures (the passage of an AI pipette into the reproductive tract) were the length of the urogenital canal (85–97 cm), the constriction at the urogenital-vaginal junction, and the tight cervix. The reproductive anatomy was compared to that of previous dissections reported in the literature.     

The reproductive cycle of Norway lobster

The reproductive biology of female Norway lobster Nephrops norvegicus was studied throughout an annual cycle from January to December 2007 in Pagasitikos Gulf, a large semi-enclosed Gulf in the central west Aegean Sea, in Greece. Six ovarian maturation stages were described to follow gonadal development, based on the combined external observation and histological examination of the ovary. Reproduction showed clear seasonality both in terms of ovarian maturation and brooding period. The proportion of fully mature females in the catch increased from January to the summer months with a peak in June. The species has a protracted brooding period that peaks in November and December, while the release of eggs from females' pleopods occurs from January to March. The size at which 50% of females reached sexual maturity was estimated, using a logistic model, to be 28.1 mm of carapace length. The undiscovered reproductive dynamics will be valuable for optimizing population models and management strategies for this important fishery resource.     

Spermatogenesis in a monogenean, Diclidophora merlangi.

Halton D. W. &; Hardcastle A. 1976. Spermatogenesis in a monogenean, Diclidophora merlangi. International Journal for Parasitology6: 43–53. Development of the spermatozoa in the testis of a polyopisthocotylean fish-gill fluke, Diclidophora merlangi, has been examined by light and electron microscopy. Spermatogonial cells are typically undifferentiated and display numerous free ribosomes and relatively little cytoplasm. Successive mitotic divisions produce spermatocytes which are characterized by expansion of the ER and the development of Golgi complexes. Nuclear division is followed by incomplete cytokinesis so that spermatocytes and subsequent stages are joined and develop syncytially. Nuclear synaptonemal complexes mark the first division of the meiotic phase, the second giving rise to a rosette of 32 spermatids. During spermateleosis, the spermatid nucleus condenses and migrates into a conical-shaped projection of cytoplasm. A centriole-like structure and basal bodies, anchored by a pair of attached rootlets, produce axial filaments that grow out from the spermatid and eventually fuse with the nuclear projection. Spermatozoa are then released from the residual cytoplasm. Each spermatozoon is approximately 325 μm in length and 2 μm maximum diameter and in section shows a nucleus, mitochondrion, paired axial units which conform to the “9 +1” pattern described for other platylelminthes, particles of β-glycogen, and a line of micro-tubules around the inner aspect of the limiting membrane.