Establishment of cell-lines stably expressing recombinant Japanese eel follicle-stimulating hormone and luteinizing hormone using CHO-DG44 cells: fully induced ovarian development at different modes.

The gonadotropins (Gth), follicle-stimulating hormone (Fsh) and luteinizing hormone (Lh), play central roles in gametogenesis in vertebrates. However, available information on their differential actions in teleost, especially , is insufficient. In this study, we established stable CHO-DG44 cell lines expressing long-lasting recombinant Japanese eel Fsh and Lh with extra O-glycosylation sites (Fsh-hCTP and Lh-hCTP), which were produced in abundance.

Effect of GnRHa on plasma levels of Fsh and Lh in the female greater amberjack Seriola dumerili.

The effects of gonadotropin-releasing hormone agonist (GnRHa) on plasma levels of follicle-stimulating hormone (Fsh) and luteinising hormone (Lh) are reported for female greater amberjack Seriola dumerili with post-vitellogenic ovarian oocytes. Five females were implanted with pellets containing GnRHa (600 μg kg body weight), while five other females were injected with saline. All females implanted with GnRHa-containing pellets ovulated 36-42 h post-implantation.

Photoperiodic regulation of plasma gonadotropin levels in previtellogenic greater amberjack (Seriola dumerili).

In Seriola species, exposure to a long photoperiod regime is known to induce ovarian development. This study examined photoperiodic effects on pituitary gene expression and plasma levels of follicle-stimulating hormone (Fsh) and luteinizing hormone (Lh) in previtellogenic greater amberjack (Seriola dumerili). The fish were exposed to short (8L:16D) or long (18L:6D) photoperiod.

Development of a homologous radioimmunoassay for red seabream follicle stimulating hormone and regulation of gonadotropins by GnRH in red seabream, Pagrus major.

Using a recombinant chimeric single-chain follicle stimulating hormone (FSH), we established a radioimmunoassay (RIA) for red seabream (Pagrus major) FSH (pmFSH) which became a powerful tool for studying reproductive physiology. We studied the profiles in plasma and pituitary concentrations of FSH and luteinizing hormone (LH) during sexual maturation. A pre-established RIA for red seabream LH was used for the LH measurements.

Greater amberjack Fsh, Lh, and their receptors: Plasma and mRNA profiles during ovarian development.

To understand the endocrine regulation of ovarian development in a multiple spawning fish, the relationship between gonadotropins (Gths; follicle-stimulating hormone [Fsh] and luteinizing hormone [Lh]) and their receptors (Gthrs; Fshr and Lhr) were investigated in greater amberjack (Seriola dumerili). cDNAs encoding the Gth subunits (Fshβ, Lhβ, and glycoprotein α [Gpα]) and Gthrs were cloned. The in vitro reporter gene assay using recombinant hormones revealed that greater amberjack Fshr and Lhr responded strongly to their own ligands.

High water temperature impairs ovarian activity and gene expression in the brain-pituitary-gonadal axis in female red seabream during the spawning season.

Temperature plays a pivotal role in the control of seasonal reproduction in temperate fish species. It is well known that temperatures that exceed a certain threshold impair gonadal development, maturation, and spawning. However, the endocrine mechanisms that underlie these effects are poorly understood.

Central distribution of kiss2 neurons and peri-pubertal changes in their expression in the brain of male and female red seabream Pagrus major.

kisspeptins that are encoded by kiss1 gene are now considered the key regulator of reproduction from a number of studies in mammals. In most vertebrates, a paralogue of kiss1, called kiss2, is also present, and the functional significance of kisspeptins is not known precisely. In the present study, we have cloned kiss2 from a perciform teleost, the red seabream Pagrus major.

Novel expression of importin alpha homologue in marine teleost, Pagrus major.

Importin alpha proteins are critical modulators of the classical nuclear protein import pathway. Although the physiological roles of importin alpha have been extensively studied in invertebrates and mammals, very little is known about their counterparts in lower vertebrates. In this study, to elucidate the roles of importin alpha in a teleost species, we isolated and characterized red seabream (Pagrus major) importin alpha cDNA derived from ovary and found changes in the mRNA levels of importin alpha in male and female red seabream during sexual maturation.

Effects of gonadotropin-releasing hormone on pituitary-ovarian axis of one-year old pre-pubertal red seabream.

To identify the pubertal development of the brain-pituitary-gonad (BPG) axis in female red seabream (Pagrus major), we investigated the effects of gonadotropin-releasing hormone agonist (GnRHa) on seabream (sb) GnRH mRNA levels in the brain, gonadotropin subunit mRNA levels in the pituitary, and serum concentrations of luteinizing hormone (LH), testosterone (T) and estradiol-17beta (E2) in pre-pubertal fish. Sexually immature 12-month-old fish were implanted with a cholesterol pellet containing GnRHa and maintained for 10-20 days. In the brain, GnRHa had no effect on sbGnRH mRNA levels.

Disturbance of plasma melatonin profile by high dose melatonin administration inhibits testicular maturation of precocious male masu salmon.

We have previously shown that the testicular development of underyearling male masu salmon Oncorhynchus masou reared under a long photoperiod was accelerated by oral melatonin treatment (0.5 mg melatonin/kg body weight/day), suggesting that melatonin mediates photoperiodic signaling. In this study, we further examined the effects of a disturbance in the plasma melatonin profile on gonadal development in underyearling male masu salmon by administering a higher dose of melatonin.

Effects of gonadotropin-releasing hormone agonist and dopamine antagonist on hypothalamus-pituitary-gonadal axis of pre-pubertal female red seabream (Pagrus major).

The effects of GnRH agonist (GnRHa) on the hypothalamus-pituitary-gonadal axis were studied in female pre-pubertal red seabream. Sexually immature 16-month-old fish were implanted intramuscularly with cholesterol pellets containing GnRHa or GnRHa in combination with domperidone, putative dopamine antagonist, and reared for 10-20 days. In both GnRHa and GnRHa+domperidone implanted groups, vitellogenesis was observed on Day 10 and ovulation was observed on Day 20, while ovarian development was not observed in the control fish throughout the experimental period.

Effects of luteinizing hormone and follicle-stimulating hormone and insulin-like growth factor-I on aromatase activity and P450 aromatase gene expression in the ovarian follicles of red seabream, Pagrus major.

To clarify the mechanism of estradiol-17beta production in the ovarian follicle of red seabream, in vitro effects of luteinizing hormone (LH), follicle-stimulating hormone (FSH), and insulin-like growth factor (IGF-I) on aromatase activity (conversion of testosterone to estradiol-17beta) and cytochrome P450 aromatase (P450arom) mRNA expression in ovarian fragments of red seabream were investigated. Of the growth factors used in the present study, only IGF-I stimulated both aromatase activity and P450arom gene expression in the ovarian fragments of red seabream. LH from red seabream pituitary, but not FSH, stimulated both aromatase activity and P450arom gene expression.

Seasonal variation of the three native gonadotropin-releasing hormone messenger ribonucleic acids levels in the brain of female red seabream.

We studied the seasonal variation of the expression of genes encoding the three native gonadotropin-releasing hormones (GnRHs), namely salmon(s) GnRH, chicken(c) GnRH-II, and seabream(sb) GnRH in red seabream, Pagrus (Chrysophrys) major, in order to better understand the regulatory mechanisms of GnRH gene expression by environmental and endocrine factors. Female red seabream, reared under natural conditions, were collected monthly or bimonthly from October to June, and the levels of the three distinct GnRH messenger ribonucleic acids (mRNAs) in the brains of those fish (n = 4-6) were determined by ribonuclease (RNase) protection analysis. The levels of sbGnRH mRNA correlated well with the observed ovarian histology; the levels of sbGnRH mRNA of immature fish in October and December were low, and increased in February and March in conjunction with active vitellogenesis.

Ontogenic origin of salmon GnRH neurons in the ventral telencephalon and the preoptic area in masu salmon.

During the ontogeny of masu salmon Oncorhynchus masou, neurons producing the salmon type of gonadotropin-releasing hormone (sGnRH) were first detected in the olfactory epithelium of the eyed egg and, subsequently, in the brain, suggesting a migration of these cells. Among sGnRH neurons distributed from the olfactory nerve (ON) through the preoptic area (POA), those in the ventral telencephalon (VT) and the POA are indicated to regulate gonadotropin secretion. Thus, it is of interest to know whether all the sGnRH neurons originate from the olfactory epithelium.

Biosynthesis of steroids in ovarian follicles of red seabream, Pagrus major (Sparidae, Teleostei) during final oocyte maturation and the relative effectiveness of steroid metabolites for germinal vesicle breakdown in vitro.

The steroid synthesis pathway in the ovarian follicles of the red seabream during final oocyte maturation (FOM) was investigated by incubating intact follicles with different radioactively labeled steroid precursors. During FOM, the steroidogenic shift from estradiol-17beta to 20 beta-hydroxylated progestin production occurred mainly due to a combination of inactivation of C 1720-lyase and activation of 20 beta-hydroxysteroid dehydrogenase. Of the steroids produced, 1720 beta-dihydroxy-4-pregnen-3-one (1720 beta-P) and 1720 beta,21-trihydroxy-4-pregnen-3-one (20 beta-S) exhibited the greatest effect on germinal vesicle breakdown (GVBD) in vitro.

Three GnRH systems in the brain and pituitary of a pleuronectiform fish, the barfin flounder Verasper moseri.

To clarify the possible function of gonadotropin-releasing hormone (GnRH) in the brain of a pleuronectiform fish, the barfin flounder Verasper moseri, the distribution of three forms of GnRH in various areas of the brain was examined by radioimmunoassay, and the localization of GnRH-immunoreactive (ir) cell bodies and fibers in the brain and pituitary was determined by immunocytochemistry. The dominant form in the pituitary was seabream GnRH (sbGnRH), levels of which were much higher than those of salmon GnRH (sGnRH) and chicken GnRH-II (cGnRH-II). In contrast, sbGnRH levels were extremely low in all other brain areas examined.

Maturational changes in brain contents of salmon GnRH in rainbow trout as measured by a newly developed time-resolved fluoroimmunoassay.

A newly developed time-resolved fluoroimmunoassay (TR-FIA) for salmon gonadotropin-releasing hormone (sGnRH) was applied to investigate changes in sGnRH content in discrete brain areas at three different gonadal stages in the rainbow trout, Oncorhynchus mykiss. The sensitivity (6.8 pg/well), specificity, intraassay coefficients of variation (<7.