Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 19 de 19
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Gen Comp Endocrinol ; 271: 30-38, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30408482

RESUMO

Chemiluminescent immunoassays (CLIAs) were developed for each of three subtypes of vitellogenin (VtgAa, VtgAb and VtgC) in grey mullet, primarily for use in monitoring estrogenic pollution of the environment. The working range of VtgAa-CLIA and VtgAb-CLIA was from 0.975 to 1,000 ng/ml, while that of VtgC-CLIA was from 0.487 to 1,000 ng/ml. Each CLIA appeared to be specific to the targeted Vtg subtype. Intra- and inter-assay coefficients of variation in the developed CLIAs were lower than 10%. In male serum, VtgAa, VtgAb and VtgC were detected in ranges from 0.01 to 0.38, 0.02 to 1.01, and 0.01 to 3.12 µg/ml, respectively, during various sampling periods. In vitellogenic females (October), serum VtgAb levels (1,192.05 ±â€¯237.81 µg/ml) were significantly higher than levels of the other two Vtg subtypes (120.82 ±â€¯30.42 and 119.23 ±â€¯16.95 µg/ml for VtgAa and VtgC, respectively). When immature mullet were fed diets containing 17α-ethinylestradiol (EE2) at three different doses (0.4, 40 and 4,000 ng/g body weight), all Vtg subtypes were induced by 40 ng/g and 4,000 ng/g EE2. The VtgC (610.30 ±â€¯150.18 µg/ml) was most highly expressed among the three Vtgs in fish fed 40 ng/g EE2, while VtgAb (33.25 ±â€¯13.58 mg/ml) was highest in expression in fish fed 4,000 ng/g EE2. The present study provided practical subtype-specific Vtg assays for the first time in grey mullet, providing the necessary means to evaluate estrogenic activities in aquatic environments.


Assuntos
Imunoensaio/métodos , Medições Luminescentes/métodos , Smegmamorpha/metabolismo , Vitelogeninas/metabolismo , Animais , Reações Cruzadas , Etinilestradiol/farmacologia , Feminino , Soros Imunes/metabolismo , Masculino , Padrões de Referência , Smegmamorpha/sangue , Vitelogeninas/sangue
2.
Artigo em Inglês | MEDLINE | ID: mdl-30017911

RESUMO

Glycogen, as an intracellular deposit of polysaccharide, takes important roles in energy balance of many animals. In fish, however, the role of glycogen during development is poorly understood. In the present study, we assessed changes in glycogen concentration and gene expression patterns of glycogen-metabolizing enzymes in developing masu salmon (Oncorhynchus masou masou), a salmonid species inhabiting west side of North Pacific Ocean. As we measured glycogen levels in the bodies and yolk sacs containing the liver separately, the glycogen concentration increased in both parts as the fish developed, whereas it transiently decreased in the yolk sac after hatching, implying glycogen synthesis and breakdown in these tissues. Immunofluorescence staining using anti-glycogen monoclonal antibody revealed localization of glycogen in the liver, muscle and yolk syncytial layer of the pre-hatching embryos and hatched larvae. In order to estimate glycogen metabolism in the fish, the genes encoding homologs of glycogen synthase (gys1 and gys2) and glycogen phosphorylase (pygma, pygmb and pygl) were cloned, and their expression patterns were assessed by quantitative PCR and in situ hybridization. In the fish, gys1 and gys2 were robustly expressed in the muscle and liver, respectively. Also, expression of pyg isoforms was found in muscle, liver and yolk syncytial layer during hatching. With changes in glycogen concentration and expression patterns of relevant genes, our results suggest, for the first time, possible involvement of glycogen in energy balance of fish embryos, especially during hatching.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Glicogênio/metabolismo , Fígado/enzimologia , Músculos/enzimologia , Salmão/metabolismo , Animais , Clonagem Molecular , Feminino , Imunofluorescência , Glicogênio Fosforilase/metabolismo , Fígado/crescimento & desenvolvimento , Masculino , Desenvolvimento Muscular , Filogenia , RNA Mensageiro/genética , Salmão/genética , Salmão/crescimento & desenvolvimento
3.
Gen Comp Endocrinol ; 251: 74-84, 2017 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-27720752

RESUMO

Gonadal maturation is a critical event wherein gonads, under the influence of several hormones and factors, undergo cyclic morphological and physiological changes to produce functional gametes during the spawning phase. However, artificial induction can be effectively used to advance the maturation of gonad vis-à-vis spawning like behavior in seasonal breeders during the off-breeding season. In the present study, osmotic pumps loaded with 5000IU of human chorionic gonadotropin (hCG) or saline as control were implanted intraperitoneally for 21days during the pre-spawning phase (May-June) in catfish Clarias batrachus and C. gariepinus. Significant increase in gonado-somatic index and sperm motility, and in the levels of certain sex steroids were observed in the hCG treated catfish when compared to control while estradiol-17ß (E2) was low. Histological analysis in hCG treated testis revealed densely packed sperm and/or spermatids inside the lumen wherein the control testis displayed normal characteristics of the pre-spawning phase. In females, histological analysis showed a significant increase in post-vitellogenic full-grown immature follicles as seen in the spawning phase. In accordance with this, the steroid hormone profile correlated well with steroidogenic shift from E2 to 17α,20ß-DP indicating oocyte maturation. However, in the control ovaries of C. batrachus, perinucleolar and pre-vitellogenic oocytes were seen to be predominant. In addition, when compared with the control, the hCG treated group displayed a significant increase in the transcripts of several genes associated with gonadal growth. Taken together, artificial induction by slow release of hCG is an effective strategy to advance sexual maturation in catfish in a programmed manner.


Assuntos
Peixes-Gato/fisiologia , Gonadotropina Coriônica/farmacologia , Fertilização/efeitos dos fármacos , Osmose , Maturidade Sexual/efeitos dos fármacos , Animais , Peixes-Gato/sangue , Estradiol/sangue , Feminino , Hormônios Esteroides Gonadais/sangue , Hormônios Esteroides Gonadais/genética , Hormônios Esteroides Gonadais/metabolismo , Humanos , Masculino , Oócitos/efeitos dos fármacos , Oócitos/metabolismo , Oogênese/efeitos dos fármacos , Ovário/efeitos dos fármacos , Ovário/metabolismo , Cloreto de Sódio/farmacologia , Motilidade Espermática/efeitos dos fármacos , Testículo/efeitos dos fármacos , Testículo/metabolismo , Testosterona/sangue , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
4.
Reprod Biol Endocrinol ; 9: 71, 2011 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-21615964

RESUMO

To elucidate the molecular mechanisms underling hydration during oocyte maturation, we characterized the structure of Japanese eel (Anguilla japonica) novel-water selective aquaporin 1 (AQP1b) that thought to be involved in oocyte hydration. The aqp1b cDNA encodes a 263 amino acid protein that includes the six potential transmembrane domains and two Asn-Pro-Ala motifs. Reverse transcription-polymerase chain reaction showed transcription of Japanese eel aqp1b in ovary and testis but not in the other tissues. In situ hybridization studies with the eel aqp1b cRNA probe revealed intense eel aqp1b signal in the oocytes at the perinucleolus stage and the signals became faint during the process of oocyte development. Light microscopic immunocytochemical analysis of ovary revealed that the Japanese eel AQP1b was expressed in the cytoplasm around the yolk globules which were located in the peripheral region of oocytes during the primary yolk globule stage; thereafter, the immunoreactivity was observed throughout the cytoplasm of oocyte as vitellogenesis progressed. The immunoreactivity became localized around the large membrane-limited yolk masses which were formed by the fusion of yolk globules during the oocyte maturation phase. These results together indicate that AQP1b, which is synthesized in the oocyte during the process of oocyte growth, is essential for mediating water uptake into eel oocytes.


Assuntos
Aquaporina 1/genética , Aquaporina 1/metabolismo , Enguias , Oócitos/metabolismo , Sequência de Aminoácidos , Animais , Aquicultura , Aquaporina 1/isolamento & purificação , Clonagem Molecular , Enguias/genética , Enguias/metabolismo , Enguias/fisiologia , Feminino , Expressão Gênica , Dados de Sequência Molecular , Oócitos/fisiologia , Oogênese/genética , Oogênese/fisiologia , Filogenia , Homologia de Sequência de Aminoácidos , Distribuição Tecidual
5.
Gen Comp Endocrinol ; 167(3): 379-86, 2010 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-20064515

RESUMO

Recombinant follicle-stimulating hormone (reFSH) and luteinizing hormone (reLH) of the Japanese eel Anguilla japonica were produced by baculovirus in silkworm Bombyx mori larvae. cDNAs encoding Japanese eel gonadotropin subunits (i.e., FSH beta, LH beta, and common alpha) were introduced into the baculovirus, which was infected into silkworm larvae after propagation of the recombinant virus in B. mori culture cells. A 100ml solution of pooled hemolymph from silkworm larvae containing reFSH or reLH were obtained from approximately 250 infected larvae. Ten milliliters of hemolymph were applied to Ni-affinity choromatography, and 5.6 and 3.5mg of partially purified reFSH and reLH were obtained, respectively. Using Western blot analysis concentrations of reFSH and reLH in the original hemolymph was estimated to be 2.2 and 1.1mg/ml, respectively. Biological activities of reFSH and reLH were assessed in vitro and in vivo. Purified reFSH and reLH induced eel oocyte maturation in vitro, and administration of hemolymph containing reFSH or reLH induced spermatogenesis in vivo in sexually immature Japanese eel. The present study indicates that a baculovirus-silkworm system could produce large amounts of biologically active recombinant fish gonadotropins for use in investigations in reproductive endocrinology and/or aquaculture of fish.


Assuntos
Baculoviridae , Bombyx/metabolismo , Enguias/genética , Gonadotropinas , Proteínas Recombinantes , Animais , Baculoviridae/genética , Baculoviridae/metabolismo , Bombyx/crescimento & desenvolvimento , Células Cultivadas , Clonagem Molecular , Avaliação Pré-Clínica de Medicamentos , Feminino , Vetores Genéticos/administração & dosagem , Gonadotropinas/genética , Gonadotropinas/isolamento & purificação , Gonadotropinas/metabolismo , Gonadotropinas/farmacologia , Larva/metabolismo , Masculino , Modelos Biológicos , Oócitos/efeitos dos fármacos , Oócitos/fisiologia , Oogênese/efeitos dos fármacos , Oogênese/fisiologia , Engenharia de Proteínas/métodos , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacologia , Espermatogênese/efeitos dos fármacos , Espermatogênese/fisiologia , Transdução Genética/métodos
6.
J Exp Zool A Ecol Genet Physiol ; 311(10): 752-62, 2009 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-19623568

RESUMO

In this study, to elucidate the mechanisms of oocyte hydration in the Japanese eel (Anguilla japonica), we examined the in vivo and in vitro morphological changes and hydration process occurring during oocyte maturation and ovulation. We also investigated the effects of the presence of ovarian follicles, aquaporin water permeability (HgCl(2)), and yolk proteolysis (bafilomycin A1) inhibitors, gap junction uncouplers (carbenoxolone and 1-octanol), a 3beta-hydroxysteroid dehydrogenase inhibitor (trilostane), and a P450 antagonist (aminoglutethimide) on oocyte hydration during in vitro oocyte maturation and ovulation. The oocytes underwent more than threefold increase in volume during maturation and ovulation, which was artificially induced by injecting salmon pituitary extracts and 17,20beta-dihydroxy-4-pregnen-3-one (DHP). Wet and dry weight measurements indicated that water accumulation during oocyte maturation is the major factor contributing to the follicular diameter increase, suggesting that follicular diameter measurements can be used as a hydration index. In the in vitro experiments, human chorionic gonadotropin (HCG) and DHP caused an increase in the diameter of follicle-enclosed oocytes but not defolliculated oocytes. Addition of HgCl(2) and bafilomycin A1 to the incubation media inhibited the HCG- and DHP-induced increase in the follicular diameter in a dose-dependent manner. Neither carbenoxolone nor 1-octanol influenced the HCG-induced increase in the follicular diameter. Trilostane and aminoglutethimide slightly but significantly inhibited HCG-induced oocyte hydration. Consequently, we concluded that ovarian follicles are essential for HCG- and DHP-induced oocyte hydration. Furthermore, aquaporin facilitates water uptake by acting as a water channel, and yolk proteolysis is essential for water influx into oocytes via osmotic mechanisms.


Assuntos
Anguilla/fisiologia , Meiose/fisiologia , Oócitos/fisiologia , Ovulação/fisiologia , Água/metabolismo , Análise de Variância , Animais , Aquaporinas/fisiologia , Feminino , Junções Comunicantes/fisiologia , Oogênese/fisiologia , Folículo Ovariano/metabolismo , Equilíbrio Hidroeletrolítico/fisiologia
7.
Artigo em Inglês | MEDLINE | ID: mdl-18789395

RESUMO

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. The 1846-bp cDNA encodes a 520 amino acid protein that includes the importin beta-binding domain, a short acidic domain, and an armadillo (arm) repeat domain. Northern blot analysis and reverse transcription-polymerase chain reaction (RT-PCR) showed transcription of red seabream importin alpha in testis and ovary but not in the other tissues. The importin alpha mRNA levels in males increase in association with testicular development, whereas those in females remain high throughout sexual maturation. These findings suggest that red seabream ovary-derived importin alpha may be controlled in a tissue-specific manner and may perform unique functions in the gonad in addition to its involvement in nuclear transport.


Assuntos
alfa Carioferinas/química , alfa Carioferinas/genética , Animais , DNA Complementar/isolamento & purificação , Feminino , Gônadas/crescimento & desenvolvimento , Masculino , Ovário/química , Estrutura Terciária de Proteína , RNA Mensageiro/análise , Dourada , Testículo/química , alfa Carioferinas/fisiologia
8.
Mol Reprod Dev ; 75(8): 1307-17, 2008 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-18324616

RESUMO

Disparate proteolytic patterns of yolk proteins, derived from three types of vitellogenin (VgA, VgB, and VgC), were observed in gray mullet. Immuno-biochemical analyses of extracts obtained from vitellogenic ovaries (VO) and ovulated eggs (OE) confirmed that a large proportion of VgA-derived lipovitellin (LvA) was degraded into free amino acids (FAAs) during ovarian follicle maturation. The maturation-associated alteration of VgB-derived Lv (LvB) involved only limited proteolysis; the heavy and light LvB chains were dissociated into at least three and one polypeptide fragments, respectively. The native mass of VgC-derived Lv (LvC) exhibited little difference between VO and OE, although it was apparent that the LvC was 'nicked' during maturation, resulting in the appearance of several bands in OE. Similar analyses confirmed that VgA-derived beta'-component (beta'-cA) and VgB-derived beta'-c (beta'-cB) decreased during maturation in both quantity and native mass, while phosvitin derived from either VgA (PvA) or VgB (PvB) appeared to be degraded into FAAs. The pattern of maturation-associated proteolysis of mullet yolk proteins is similar to that reported for other marine teleosts spawning pelagic eggs. However, the depository ratio of the three distinct types of Lv in the mullet VO appeared to be different from that estimated for another marine pelagophil, the barfin flounder. These results support a recent paradigm regarding the significance of Vg multiplicity upon successive physiological events in this group of fishes including the hydration of maturing oocytes, the acquisition of proper egg buoyancy, and the generation of requisite nutrient stocks for each stage of embryogenesis and larval development.


Assuntos
Proteínas do Ovo/metabolismo , Folículo Ovariano/crescimento & desenvolvimento , Peptídeo Hidrolases/metabolismo , Maturidade Sexual/fisiologia , Smegmamorpha/metabolismo , Vitelogeninas/metabolismo , Sequência de Aminoácidos , Animais , Cromatografia em Gel , Proteínas do Ovo/genética , Eletroforese em Gel de Poliacrilamida , Feminino , Imunoeletroforese , Dados de Sequência Molecular , Folículo Ovariano/metabolismo , Alinhamento de Sequência , Análise de Sequência de Proteína , Smegmamorpha/fisiologia
9.
J Exp Zool A Ecol Genet Physiol ; 307(6): 324-41, 2007 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-17480036

RESUMO

Seven yolk proteins (YPs), four large lipoproteins (YPs1-4) and three minor yolk components (YPs5-7) including one phosphoprotein (YP7), were purified from extracts of vitellogenic ovaries of grey mullet (Mugil cephalus) by combinations of hydroxylapatite, ion exchange, immunoadsorbent, and gel filtration chromatography. The molecular masses of native YP1, YP2, YP3, and YP4 were estimated to be 330, 325, 335, and 570 kDa, respectively. The tertiary structures of YP1, YP2, and YP3 revealed by sodium dodecyl sulfate polyacrylamide gel electrophoresis were typical of teleost lipovitellins (Lvs), consisting of a heavy chain ( approximately 110, approximately 99, and approximately 97 kDa, respectively) and a light chain ( approximately 30, approximately 29, and approximately 21.5 kDa, respectively), while YP4 exhibited a heavy chain ( approximately 110 kDa) and two more polypeptide bands ( approximately 70 and approximately 54 kDa). Mapping of N-terminal peptide sequences of the purified YPs to the primary structure of multiple mullet vitellogenins (Vgs) deduced from their respective complete cDNAs, which were cloned and sequenced, conclusively identified YP1, YP2, and YP3 as Lvs derived from mullet VgA, VgB, and VgC, respectively. The fourth YP (YP4) appeared to be a proteolytic variant consisting of Lv and phosvitin components of VgA. Two other YPs (YP5 and YP6) were identified as beta'-components derived from VgA and VgB based on their structures and common, but not identical, antigenicity to salmonid beta'-component, while purified YP7, a phosphoprotein with a high content of serine residues, was identified as a phosvitin derived from VgB. This is the first report, of which we are aware, on purification and molecular classification of three distinct forms of Lv from any oviparous vertebrate.


Assuntos
Proteínas do Ovo/isolamento & purificação , Smegmamorpha/genética , Vitelogeninas/genética , Sequência de Aminoácidos , Animais , Cromatografia , Clonagem Molecular , Proteínas do Ovo/química , Proteínas do Ovo/genética , Feminino , Dados de Sequência Molecular , Análise de Sequência de DNA , Smegmamorpha/metabolismo
10.
Artigo em Inglês | MEDLINE | ID: mdl-16828323

RESUMO

In a previous study, we identified cDNAs encoding the growth hormone receptor (eGHR1) and eGHR1 homologue (eGHR2) in Japanese eel (Anguilla japonica). In the present study, changes in the developmental expression of growth hormone (GH), eGHR1 and eGHR2 were investigated in the Japanese eel eggs and preleptocephali by RT-PCR and immunohistochemical methods in an attempt to examine the involvement of these proteins in larval growth. The GH transcripts and the production of GH protein were not detected in the newly hatched larvae and preleptocephali at day 3 post-hatch, however, these were detected at day 6 post-hatch, and also detected at higher levels at day 10 post-hatch. In contrast, prolactin and somatolactin transcripts could not be detected in all preleptocephalus specimens (newly hatched larvae and preleptocephali at day 3, 6 and 10 post-hatch). eGHR1 and eGHR2 transcripts were detected in all preleptocephalus specimens. Therefore, it is plausible that the actions of GH during the preleptocephalus stage are mediated through the eGHRs. The present data suggest that GHR-mediated actions of GH begin at the same time as the initiation of GH production, and that GH plays important roles in larval growth and survival to the leptocephalus stage. eGHR1 mRNA, which is thought to be of maternal origin, was also detected in ovulated eggs. However, the role of eGHR1 mRNA in eggs is not clear.


Assuntos
Anguilla/crescimento & desenvolvimento , Anguilla/metabolismo , Hormônio do Crescimento/metabolismo , Receptores da Somatotropina/metabolismo , Animais , Feminino , Regulação da Expressão Gênica , Imuno-Histoquímica , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
11.
Mol Reprod Dev ; 73(6): 719-36, 2006 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-16541459

RESUMO

Full-length cDNAs encoding three forms of vitellogenin (Vg) were obtained from a liver cDNA library of estrogen-treated red seabream, Pagrus major. Two of the three Vg sequences had high homology with type-A and -B Vgs (VgA and VgB) of other teleosts. The third red seabream Vg was classified as a type-C or phosvitinless (Pvl) Vg due to its lack of a phosvitin (Pv) domain. Two Vg preparations (610 and 340 kDa) from blood serum of estradiol-treated fish were biochemically characterized. Analyses of precursor-product relationships by examination of N-terminal amino acid sequences verified cleavage of the 610 kDa Vg into a 540 kDa lipovitellin (Lv) and a 32 kDa beta'-component. Each of these yolk preparations comprising both VgA- and VgB-derived polypeptides. The 340 kDa Vg, which was immunologically verified to be a PvlVg, was accumulated by vitellogenic oocytes with no alterations to its native molecular mass. During oocyte maturation, the VgA- and VgB-derived yolk proteins were differentially processed, presumably to generate a pool of free amino acids for oocyte hydration or for allocation of specific types of nutrients, amino acids, and proteins, to the developing embryo. Conversely, the 340 kDa Vg-derived yolk protein is unlikely to contribute to oocyte hydration or diffusible nutrients since the molecule underwent only minor proteolytic nicking during oogenesis. The present study elucidates for the first time specific functions of three different forms of Vg and their product yolk proteins in a higher taxonomic group of marine teleosts that spawn pelagic eggs.


Assuntos
Proteínas do Ovo/metabolismo , Desenvolvimento Embrionário , Oócitos/fisiologia , Perciformes , Fosvitina , Vitelogeninas , Sequência de Aminoácidos , Animais , DNA Complementar/metabolismo , Feminino , Dados de Sequência Molecular , Peso Molecular , Oócitos/citologia , Fosvitina/química , Fosvitina/genética , Fosvitina/metabolismo , Água do Mar , Alinhamento de Sequência , Vitelogeninas/química , Vitelogeninas/genética , Vitelogeninas/metabolismo
12.
Fish Physiol Biochem ; 31(2-3): 193-9, 2005 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-20035458

RESUMO

The eel has long been esteemed as an important food fish in the world, especially in Japan, and has been used as an experimental fish for many fields of fish physiology. However, the decreases in eel resources have been a serious concern in recent years. The catches of glass eels as seedlings for aquaculture have shown a long-term decrease in both Europe and East Asia. To increase eel resources, the development of techniques for artificial induction of maturation and spawning and rearing their larvae have been eagerly desired. Recent progress of reproductive physiology of fish, especially mechanisms of oocyte maturation and ovulation in female and of spermatozoa maturation in male, facilitate to establish techniques for hormonal induction of maturation and spawning in sexually immature eels. With persistent effort to development of rearing techniques of larvae, we have first succeeded to produce glass eel. These applied techniques are may contribute to understand the basic reproductive physiology of the eel.

13.
Gen Comp Endocrinol ; 138(2): 105-12, 2004 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-15302259

RESUMO

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. In the pituitary, although no marked changes were observed in follicle-stimulating hormone (FSH) beta subunit mRNA levels, the expression of glycoprotein (GP) alpha, and LHbeta subunit genes in the pituitary was drastically up-regulated (approximately 4- and 5-fold, respectively) and serum LH levels were also increased (approximately 3-fold) by GnRHa implantation. However, ovaries of GnRHa treated fish contained only oocytes at the peri-nucleolus stage, and oocyte development such as vitellogenesis and oocyte maturation did not occur throughout the experimental period. In these fish, even though LH was released, only slight increases in serum concentrations of T and E2 were observed. These results indicate that the pituitary gonadotropin cells of pre-pubertal 12-month-old fish were already receptive to GnRH stimulus, and acquired the ability to synthesize and release of LH as in the case of adult fish. Deficient factors for the onset of puberty by GnRHa treatment will be discussed.


Assuntos
Hormônio Liberador de Gonadotropina/fisiologia , Ovário/fisiologia , Perciformes/fisiologia , Hipófise/fisiologia , Maturidade Sexual/fisiologia , Fatores Etários , Animais , Estradiol/sangue , Feminino , Regulação da Expressão Gênica , Hormônio Liberador de Gonadotropina/agonistas , Hormônio Liberador de Gonadotropina/genética , Hormônio Luteinizante/sangue , Ovário/efeitos dos fármacos , Hipófise/efeitos dos fármacos , RNA Mensageiro/análise , Reprodução/fisiologia , Maturidade Sexual/efeitos dos fármacos , Testosterona/sangue
14.
Gen Comp Endocrinol ; 132(1): 35-45, 2003 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-12765642

RESUMO

Two distinct types of gonadotrophs, FSH (GtH I) cells and LH (GtH II) cells, were immunocytochemically identified from mummichog (Fundulus heteroclitus; Cyprinodontiformes, Acanthopterygii) pituitary using antisera raised against synthetic fragment peptides of FSHbeta and LHbeta. Both cell types were abundant during the spawning period (spring and early summer) and decreased in number during the post-spawning immature period. The number of FSH cells increased again during the early phases of gonadal development (cortical alveoli accumulation in the oocytes and basal spermatogenesis) in early winter, whereas the number of LH cells did not. Only FSH cells were abundant during the latter phases of gonadal development (vitellogenesis and active spermatogenesis) in early spring. These observations suggest that both GtHs have important yet different roles for reproduction in this species. Antisera against the conservative region of the FSHbeta and the LHbeta subunits immunostained FSH cells and LH cells, respectively, also in red seabream (Pagrus major; Perciformes, Acanthopterygii) and small mouth bass (Micropterus dolomieu; Perciformes, Acanthopterygii), suggesting the possibility of their use for other acanthopterygian fishes.


Assuntos
Hormônio Foliculoestimulante/metabolismo , Fundulidae/metabolismo , Hormônio Luteinizante/metabolismo , Hipófise/citologia , Hipófise/metabolismo , Estações do Ano , Animais , Bass/metabolismo , Feminino , Soros Imunes/imunologia , Imuno-Histoquímica , Masculino , Fragmentos de Peptídeos/imunologia , Dourada/metabolismo , Comportamento Sexual Animal/fisiologia
15.
Gen Comp Endocrinol ; 131(3): 264-73, 2003 May.
Artigo em Inglês | MEDLINE | ID: mdl-12714008

RESUMO

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. The levels of GnRH receptor mRNA were significantly higher in both GnRHa implanted groups than in the control. The expressions of all three gonadotropin subunit genes were up-regulated and serum luteinizing hormone levels were increased by the GnRHa implantation. Serum testosterone and estradiol-17beta levels were also increased on Day 10 and maintained high levels on Day 20. On the other hand, seabream (sb) GnRH mRNA levels in the brain were relatively low and unchanged in all experiment groups. The present study first shows that GnRH alone can induce precocious puberty in red seabream. These results indicate that the system of pituitary-gonadal axis has already been developed in 16-month-old fish and the commencement of sbGnRH secretion may be an important physiological event for the onset of puberty in the red seabream.


Assuntos
Antagonistas de Dopamina/farmacologia , Hormônio Liberador de Gonadotropina/agonistas , Sistema Hipotálamo-Hipofisário/efeitos dos fármacos , Ovário/efeitos dos fármacos , Dourada/fisiologia , Animais , Química Encefálica/efeitos dos fármacos , Química Encefálica/genética , Colesterol , Domperidona/administração & dosagem , Domperidona/farmacologia , Antagonistas de Dopamina/administração & dosagem , Implantes de Medicamento , Estradiol/sangue , Feminino , Expressão Gênica/efeitos dos fármacos , Hormônio Liberador de Gonadotropina/biossíntese , Gonadotropinas/biossíntese , Gonadotropinas/metabolismo , Sistema Hipotálamo-Hipofisário/metabolismo , Hormônio Luteinizante/sangue , Ensaios de Proteção de Nucleases , Ovário/crescimento & desenvolvimento , Ovário/metabolismo , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Radioimunoensaio , Maturidade Sexual , Testosterona/sangue
16.
Gen Comp Endocrinol ; 130(3): 324-32, 2003 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-12606275

RESUMO

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. The sbGnRH mRNA levels reached a maximum level in April (spawning season), after which they rapidly decreased together with the observed ovarian regression in June. In contrast, the levels of sGnRH mRNA showed no variation, while those of cGnRH-II mRNA were elevated only slightly in March and April. The increase in sbGnRH mRNA levels correlates with the increase in day length, water temperature and serum steroids levels, suggesting that these factors are candidates for regulators of sbGnRH synthesis.


Assuntos
Química Encefálica , Hormônio Liberador de Gonadotropina/genética , Perciformes/genética , RNA Mensageiro/análise , Estações do Ano , Animais , Estradiol/sangue , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/fisiologia , Hormônio Liberador de Gonadotropina/classificação , Ensaios de Proteção de Nucleases , Ovário/anatomia & histologia , Ovário/fisiologia , Testosterona/sangue
17.
Biol Reprod ; 68(5): 1562-8, 2003 May.
Artigo em Inglês | MEDLINE | ID: mdl-12606429

RESUMO

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. IGF-I slightly enhanced the LH-induced aromatase activity and P450arom gene expression. These data and our previous results indicate that LH, but not FSH, stimulates estradiol-17beta production in the ovarian follicle of red seabream through stimulation of aromatase activity and P450arom gene expression and IGF-I enhances the LH-stimulated P450arom gene expression.


Assuntos
Aromatase/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Hormônio Foliculoestimulante/farmacologia , Regulação Enzimológica da Expressão Gênica/genética , Fator de Crescimento Insulin-Like I/farmacologia , Hormônio Luteinizante/farmacologia , Folículo Ovariano/enzimologia , Dourada/fisiologia , Animais , Aromatase/genética , Northern Blotting , Sistema Enzimático do Citocromo P-450/genética , Relação Dose-Resposta a Droga , Estradiol/biossíntese , Feminino , Substâncias de Crescimento/farmacologia , Folículo Ovariano/efeitos dos fármacos , RNA Mensageiro/biossíntese
18.
Artigo em Inglês | MEDLINE | ID: mdl-12223211

RESUMO

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. 1720 beta-P was further converted to its 5 beta-reduced form, 1720 beta-dihydroxy-5 beta-pregnan-3-one (1720 beta-P-5 beta), which had lower GVBD activity, suggesting that 5 beta-reduction plays a role in the inactivation of the maturation-inducing ability of 1720 beta-P. In contrast, no 5 beta-reduced metabolite of 20 beta-S was found. Serum levels of 1720 beta-P and 20 beta-S, measured by ELISA, showed that circulating levels of both progestins increased during FOM, and 20 beta-S levels were approximately twice as high as 1720 beta-P levels. This study clarified the complete steroidogenesis pathway during FOM in red seabream ovarian follicles and showed that two 20 beta-hydroxylated progestins, 1720 beta-P and 20 beta-S, act as maturation-inducing hormones in this species. The catabolites of these two progestins and their physiological roles in reproduction are also discussed.


Assuntos
Folículo Ovariano/metabolismo , Dourada/metabolismo , Esteroides/biossíntese , Animais , Cortisona Redutase , Feminino , Oócitos/efeitos dos fármacos , Folículo Ovariano/efeitos dos fármacos , Pregnanolona/biossíntese , Pregnanolona/farmacologia , Progestinas/biossíntese , Progestinas/farmacologia , Traçadores Radioativos , Dourada/fisiologia , Trítio
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA