Assembly of the Inner Perivitelline Layer, a Homo log of the Mammalian Zona Pellucida: An Immunohistochemical and Ultrastructural Study

The avian inner perivitelline layer (IPVL), a homologous structure to the mammalian zona pellucida, is deposited between the granulosa cells and the oocyte cell membrane during folliculogenesis. The glycoprotein meshwork of the IPVL forms a 3-dimensional matrix and possesses important functions in the fertilization process: it contributes to the binding of avian spermatozoa to the oocyte and induces acrosomal exocytosis. In contrast to the zona pellucida of mammals, the IPVL does not prevent the physiological polyspermy found in birds. Previous studies have shown that in the Japanese quail (Cotumix japonica) at least 5 glycoproteins are constituents of the IPVL (ZP1, ZP2, ZP3, ZP4, and ZPD). In this study, we investigated the spatiotennporal assembly pattern of the IPVL during folliculogenesis using immunohistochemical and ultrastructural methods. The obtained results clearly show that these glycoproteins are incorporated into the IPVL at distinct points during follicular development, supporting the hypothesis that ZP2 and ZP4 form a type of prematrix into which ZP1, ZP3, and ZPD are integrated at a later stage of development. Copyright (C) 2011 S. Karger AG, Base

Cellular analysis of cleavage-stage chick embryos reveals hidden conservation in vertebrate early development

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributed.Birds and mammals, phylogenetically close amniotes with similar post-gastrula development, exhibit little conservation in their post-fertilization cleavage patterns. Data from the mouse suggest that cellular morphogenesis and molecular signaling at the cleavage stage play important roles in lineage specification at later (blastula and gastrula) stages. Very little is known, however, about cleavage-stage chick embryos, owing to their poor accessibility. This period of chick development takes place before egg-laying and encompasses several fundamental processes of avian embryology, including zygotic gene activation (ZGA) and blastoderm cell-layer increase. We have carried out morphological and cellular analyses of cleavage-stage chick embryos covering the first half of pre-ovipositional development, from Eyal-Giladi and Kochav stage (EGK-) I to EGK-V. Scanning electron microscopy revealed remarkable subcellular details of blastomere cellularization and subgerminal cavity formation. Phosphorylated RNA polymerase II immunostaining showed that ZGA in the chick starts at early EGK-III during the 7th to 8th nuclear division cycle, comparable with the time reported for other yolk-rich vertebrates (e.g. zebrafish and Xenopus). The increase in the number of cell layers after EGK-III is not a direct consequence of oriented cell division. Finally, we present evidence that, as in the zebrafish embryo, a yolk syncytial layer is formed in the avian embryo after EGK-V. Our data suggest that several fundamental features of cleavage-stage development in birds resemble those in yolk-rich anamniote species, revealing conservation in vertebrate early development. Whether this conservation lends morphogenetic support to the anamniote-to-amniote transition in evolution or reflects developmental plasticity in convergent evolution awaits further investigation

Nurturing a gender-responsive approach to climate-smart agriculture in Guinayangan, Quezon

Coconut-based farming systems in Guinayangan, Quezon offer special opportunities for achieving multiple objectives, including carbon sequestration, economic empowerment of women and reduction of risks from variable and extreme weather. This info note discusses the gender-based role inequalities within coconut-based farming systems that can be addressed through agroforestry-based, climate-smart agriculture that features small livestock, fruit trees and root and tuber crops as understory crops. Numerous Climate-Smart Villages, spread across the municipality of Guinayangan, now serve as proof of concept, providing evidence that climate-smart agriculture based on agroforestry interventions are gender sensitive

Sperm Motility Regulation in Male and Female Bird Genital Tracts

Sperm drastically change their flagellar movement in response to the surrounding physical and chemical environment. Testicular sperm are immotile; however, they gain the competence to initiate motility during passage through the male reproductive tract. Once ejaculated, the sperm are activated and promptly initiate motility. Unlike mammals, ejaculated sperm in birds are stored in specialized tubular invaginations referred to as sperm storage tubules (SSTs), located between the vagina and uterus, before fertilization. The resident sperm in the SSTs are in a quiescent state and then re-activated after release from the SSTs. It is thought that avian sperm can undergo motility change from quiescent to active state twice; however, the molecular mechanism underlying sperm motility regulation is poorly understood. In this short review, we summarize the current understanding of sperm motility regulation in male and female bird reproductive tracts. We also describe signal transduction, which regulates sperm motility, mainly derived from in vitro studies

Stimulation of ZPC Production by Follicle-Stimulating Hormone in the Granulosa Cells of Japanese Quail (Coturnix Japonica)

Avian perivitelline membrane, an investment homologous to the zona pellucida of mammalian oocytes, is composed of two glycoproteins. Previous studies have indicated that one of the components, a glycoprotein homologous to mammalian ZPC, is produced and secreted by the granulosa cells of maturing follicles in quail ovary. In the present study, to investigate the hormonal regulation of ZPC production, [3H] leucine incorporation into ZPC was compared in granulosa cells cultured in a medium containing follicle-stimulating hormone (FSH), luteinizing hormone, or prolactin, and a stimulatory effect of FSH was observed. By measuring ZPC protein in the medium and mRNA in the cells with Western blotting and Northern blotting analyses, respectively, we found that addition of FSH stimulated not only ZPC protein production but also ZPC mRNA expression in the granulosa cells. These results suggested that FSH stimulates ZPC protein production at the gene transcription level

Effects of Cadmium Administration on Reproductive Performance of Japanese Quail (Coturnix japonica)

The effects of a single dosage of cadmium (Cd) administration on the survival rate, body weight (BW), egg production, egg quality characteristics and hatching outputs in 0, 0.1, 0.3, 1, 3 and 10mg CdCl2/kg BW injected laying quail were studied. It was found that a 10mg Cd injection induced 41.7% mortality. A retainable significant BW loss in the 3mg dosed birds and a decreased egg production in the 1mg and 3mg dosed birds were observed during the first 3 days after injection, and the recovering of the egg production rate was delayed as the doses of Cd injection increased. Cd administration significantly increased the shell membrane weight in 0.1mg and 0.3mg injection, decreased the eggshell thickness in 0.3mg injection, and lowered the fertility rate in 0.3mg injection during the first 3 days of injection. The present study suggests that exposure to Cd, even by a single dosage, can hamper quail reproduction for a very short time, mainly by decreasing egg production and thinning the eggshell

Binding of Xenoestrogens and Phytoestrogens to Estrogen Receptor β of Japanese Quail (Coturnix japonica)

In order to examine the binding affinity of various estrogenic compounds to estrogen receptor β (ERβ), the cDNA encoding the hinge domain, the ligand-binding domain, and the C-terminal domain of quail ERβ was constructed and transfected into competent Escherichia coli. The binding assay performed using the supernatant of the cell lysates showed that bacterially-expressed ERβ has one single class of binding site for estradiol-17β with a dissociation constant of 4.90±0.16×10-9M. The competition studies indicated that the relative binding affinities for the synthetic estrogens, diethylstilbestrol and ethinyl estradiol, are very high, while those for the xenoestrogens, bisphenol A and nonylphenol, are very low. Coumestrol, known as one of phytoestrogens, can compete with estradiol-17β with higher binding affinity for ERβ than ERα

Cyclin D1 gene expression is essential for cell cycle progression from the maternal-to-zygotic transition during blastoderm development in Japanese quail

Embryogenesis proceeds by a highly regulated series of events. In animals, maternal factors that accumulate in the egg cytoplasm control cell cycle progression at the initial stage of cleavage. However, cell cycle regulation is switched to a system governed by the activated nuclear genome at a specific stage of development, referred to as maternal-to-zygotic transition (MZT). Detailed molecular analyses have been performed on maternal factors and activated zygotic genes in MZT in mammals, fishes and chicken; however, the underlying mechanisms remain unclear in quail. In the present study, we demonstrated that MZT occurred at blastoderm stage V in the Japanese quail using novel gene targeting technology in which the CRISPR/Cas9 and intracytoplasmic sperm injection (ICSI) systems were combined. At blastoderm stage V, we found that maternal retinoblastoma 1 (RB1) protein expression was down-regulated, whereas the gene expression of cyclin D1 (CCND1) was initiated. When a microinjection of sgRNA containing CCND1-targeted sequencing and Cas9 mRNA was administered at the pronuclear stage, blastoderm development stopped at stage V and the down-regulation of RB1 did not occur. This result indicates the most notable difference from mammals in which CCND-knockout embryos are capable of developing beyond MZT. We also showed that CCND1 induced the phosphorylation of the serine/threonine residues of the RB1 protein, which resulted in the degradation of this protein. These results suggest that CCND1 is one of the key factors for RB1 protein degradation at MZT, and the elimination of RB1 may contribute to cell cycle progression after MZT during blastoderm development in the Japanese quail. Our novel technology, which combined the CRISPR/Cas9 system and ICSI, has the potential to become a powerful tool for avian-targeted mutagenesis

Expression of Transferrin and Albumin in the Sperm-Storage Tubules of Japanese Quail and their Possible Involvement in Long-Term Sperm Storage

Because of the presence of sperm storage tubules (SSTs) in the utero-vaginal junction (UVJ) in the oviduct, once ejaculated sperm enter the female reproductive tract, they can survive for a prolonged period in domestic birds; however, the specific mechanisms involved in sperm maintenance within the SST remain to be elucidated. In this study, we showed that transferrin (TF) and albumin (ALB) are expressed in SSTs. When UVJ extracts were subjected to size-exclusion column chromatography, we obtained fractions that extend sperm longevity in vitro. LC-MS/MS analysis of the two major proteins in the fractions identified these proteins as TF and ALB. Immunohistochemical analysis using specific antisera against TF and ALB indicated that both proteins were localized not only in the SSTs, but also in the surface epithelium of the UVJ. When the ejaculated sperm were incubated with either purified TF or ALB, sperm viability increased after 24 h. These results indicated that oviductal TF and ALB are involved in the process of sperm storage in SSTs and may open a new approach for technological improvement to prolong sperm longevity in vitro

Prolactin and Growth Hormone in Birds: Protein Structure, Gene Structure and Genetic Variation

Prolactin (PRL) and Growth hormone (GH) are secreted from the anterior pituitary gland and affect a wide variety of physiological functions in birds. A causal relationship between plasma PRL concentration and broody behaviour is well known. Similarly, a relationship between plasma GH concentration and body growth is well documented. Following the cloning of cDNA of PRL and GH in the chicken and turkey, the levels of mRNA were measured. Levels of PRL or GH mRNA in the anterior pituitary gland were generally correlated with the plasma levels of PRL or GH. Using the PRL or GH cDNA clone as a probe, the genomic DNA clone of PRL and GH was obtained in several galliform species. The availability of cDNA and genomic DNA sequence also allowed the determination of sequence variation of PRL and GH genes in chicken and turkey. The genetic variation of the turkey and chicken PRL promoter and allelic variation of PRL have been shown to be associated with the plasma levels of PRL and expression of incubation behaviour. Likewise, allelic variants of the GH have been proposed as a mechanism to explain variation in egg production in the chicken. Recently, the duck PRL and GH gene was cloned and sequenced. Genetic variations were also detected in the duck GH gene. The presence of variation and/or polymorphic sequence may be important since a variety of body types and egg production traits are found among the duck breeds. Thus, genetic variation in the PRL and GH genes of domestic fowl may applicable to marker assisted selection. Cloning of cDNA and genomic DNA of PRL and GH was conducted mainly in the domestic species. The sequence information of PRL and GH in the domestic species makes it possible to clone and characterize the PRL and GH in altricial species. The sequence of PRL and GH in the various avian species will provide useful information for studying the physiological function and general and species specific mechanisms of gene expression in birds