Perinucleolus Stage Research Articles

Ovarian Development and Sex Steriod Hormones in the Keeled Fish <i>Liza carinata</i> (Valenciennes, 1836) From Suez Bay, Egypt

This work aims to investigate the microscopic structures of ovaries of Liza carinata collected from Suez Bay during annual cycle combined with the relation between GSI (gonadosomatic index) and sex steroid hormones concentration for female during the study period. The oocyte maturation showed the following phases; first growth phase which includes chromatin-nucleolus stage, early and late perinucleolus stages. Second growth phase includes vacuolization stage (yolk vesicle stage), yolk stages (primary, secondary and tertiary), and ripe stage. The maturity stages of ovary recorded and classified into seven stages (Immature, Immature & Recovering, Developing, Maturing, Mature, Spawning and Spent). The cyclic changes in GSI indices are considered as further a proof that the spawning season is extending from November (8.88%) to March with value (0.62%). Sex steroids hormones play a role in controlling the maturation cycle in teleost especially during spawning times, which altered by environmental or hormonal manipulation. 17β - Estradiol levels in female L. carinata ranged from maximum value 0.199 ± 0.087 ng/ml recorded in maturing stage to 0.102 ± 0.016 ng/ml ml in December 2012. The lowest value of testosterone was observed in February with value (17.4 ± 1.054) in spawning and spent stages of maturity.

Molecular characterization of the gonadal kisspeptin system: Cloning, tissue distribution, gene expression analysis and localization in sablefish (Anoplopoma fimbria)

The kisspeptin system plays pivotal roles in the regulation of vertebrate reproduction. Classically, kisspeptin produced in the brain stimulates brain gonadotropin-releasing hormone signaling, which in turn activates the pituitary–gonad axis. Expression of the kisspeptin system has also been documented in peripheral tissues, including gonads of mammals and fishes. However, the fish gonadal kisspeptin system remained uncharacterized. Herein we report identification and characterization of four kisspeptin system mRNAs (kisspeptin 1 (kiss1), kiss2, and G protein-coupled receptor 54-1 (gpr54-1) and gpr54-2) in sablefish, Anoplopoma fimbria. Sablefish predicted protein sequences were highly similar to those of other marine teleosts, but less so to freshwater teleosts. Tissue distribution analyses revealed that all four kisspeptin-system transcripts were expressed in both brain and gonad. However, kiss2 was the predominant transcript in the gonads and the only transcript detected in ovulated eggs. Ontogenetic analysis of kiss2 expression in juvenile sablefish gonads demonstrated that levels were low during sex differentiation but increased with fish size and gonadal development. Dramatic increases in kiss2 mRNA occurred during primary oocyte growth, while levels remained relatively low in testes. In situ hybridization revealed that kiss2 mRNA was localized to cytoplasm of perinucleolus stage oocytes, suggesting it could play a local role in oogenesis or could be synthesized and stored within oocytes as maternal mRNA. This represents the first study to focus on the gonadal kisspeptin system in fishes and provides important tools for further investigation of both the gonadal and brain kisspeptin systems in sablefish.

Expression profiles of Fsh-regulated ovarian genes during oogenesis in coho salmon.

The function of follicle-stimulating hormone (Fsh) during oogenesis in fishes is poorly understood. Using coho salmon as a fish model, we recently identified a suite of genes regulated by Fsh in vitro and involved in ovarian processes mostly unexplored in fishes, like cell proliferation, differentiation, survival or extracellular matrix (ECM) remodeling. To better understand the role of these Fsh-regulated genes during oocyte growth in fishes, we characterized their mRNA levels at discrete stages of the ovarian development in coho salmon. While most of the transcripts were expressed at low levels during primary growth (perinucleolus stage), high expression of genes associated with cell proliferation (pim1, pcna, and mcm4) and survival (ddit4l) was found in follicles at this stage. The transition to secondary oocyte growth (cortical alveolus and lipid droplet stage ovarian follicles) was characterized by a marked increase in the expression of genes related to cell survival (clu1, clu2 and ivns1abpa). Expression of genes associated with cell differentiation and growth (wt2l and adh8l), growth factor signaling (inha), steroidogenesis (cyp19a1a) and the ECM (col1a1, col1a2 and dcn) peaked in vitellogenic follicles, showing a strong and positive correlation with transcripts for fshr. Other genes regulated by Fsh and associated with ECM function (ctgf, wapl and fn1) and growth factor signaling (bmp16 and smad5l) peaked in maturing follicles, along with increases in steroidogenesis-related gene transcripts. In conclusion, ovarian genes regulated by Fsh showed marked differences in their expression patterns during oogenesis in coho salmon. Our results suggest that Fsh regulates different ovarian processes at specific stages of development, likely through interaction with other intra- or extra-ovarian factors.

Cloning and expression of follistatin gene in half-smooth tongue sole Cynoglossus semilaevis during the reproduction cycle

Follistatin (FST) is a monomeric glycoprotein highly enriched in cysteines and belongs to TGF-β superfamily. FST can suppress the secretion of follicle-stimulating hormone and plays a vital role in the reproduction of vertebrates. We used rapid amplification of cDNA ends technology to clone the FST gene of half-smooth tongue sole, Cynoglossus semilaevis. We characterized its phylogenetic context and expression patterns to elucidate its function in the breeding season. The full-length sequence of FST is 1 455 bp and encodes a protein of 321 amino acids. We investigated the expression pattern of FST in C. semilaevis at different stages of reproduction using reverse transcription-polymerase chain reaction (RTPCR). FST mRNA was expressed in all 13 tissues analyzed, and was expressed at high levels in gonad and at slightly lower levels in gill and brain. During the reproductive cycle of C. semilaevis, the transcript level of FST was the highest in the perinucleolus stage, decreased in the primary yolk stage, slightly increased in the tertiary yolk stage, and then reduced to a minimal level in the atretic follicles stage of the ovary. We concluded that FST suppressed follicle-stimulating hormone, which stimulated oocyte development. However, no significant variation was observed across all stages of testis development, although the expression level in the spermatogenesis stage was relatively low, which may result from the regulation of FST by aromatase.

Induction of sex inversion in common snook (Centropomus undecimalis) males, using 17-β oestradiol implants

The common snook (Centropomus undecimalis) is a protandric hermaphrodite fish that has potential for aquaculture due to its high value and market acceptance. One of the difficulties to its reproduction in captivity is that females are older and bigger than males. The objective of this study was to use 17-β oestradiol (E2) hormone implants to induce sex inversion in adult males. Fish with an initial body weight of 383 ± 83 g (mean ± SD), individually tagged were used in the experiment. Four E2 dosages (n = 7) were tested (0.5, 1.0, 4.0 and 8.0 mg kg−1) in Ethylene-vinyl-acetate (EVAc) implants, and a control group (n = 7) implanted without hormone. The parameters evaluated were: survival, weight gain, hepatosomatic and gonadosomatic indices (GSI and HSI) and plasma levels of testosterone (T) and 17-β oestradiol (E2). Also, liver and gonad morphology was observed through histological sections. GSI was higher in E2 treated than in control fish. All E2 treated fish had completely developed ovaries with oocytes at the perinucleolar stage, while all fish in the control group remained males with evidence of active spermatogenesis. After 15 days, plasma levels of E2 were correlated with the hormone dosage. T levels in the control group were higher than in E2 treated fish, at all sampling times. In conclusion, 0.5 mg kg−1 of E2 in EVAc implants is effective to induce sexual inversion in common snook males, which could be useful to obtain broodstock females smaller than in the wild in a reduced time.

Efecto del pH sobre la proporción oocitaria de Paracheirodon axelrodi, en cautiverio

Paracheirodon axelrodi es la especie de pez ornamental más comercializado y exportado en Colombia. Los individuos comercializados son extraídos del ambiente afectando las poblaciones, lo que ha llevado a realizar estudios relacionados con su biología reproductiva para el desarrollo de protocolos de reproducción en cautiverio. En este estudio se evaluó el efecto de dos rangos de pH en el agua durante seis meses (T1: 5.5 - 5.8 y T2: 7.0 - 7.5), sobre las variables de crecimiento y la proporción de oocitos a nivel histológico en hembras adultas de P. axelrodi, para establecer si este parámetro afectaba el desarrollo gonadal y crecimiento a través del tiempo. De acuerdo a los resultados obtenidos, los rangos de pH no afectaron los porcentajes de los tipos de oocitos perinucleolar, alveolo cortical, vitelogénicos en etapa inicial y etapa tardía, ni en los oocitos en desarrollo de atresia; sin embargo, las condiciones mantenidas favorecieron las variables de crecimiento. Las proporciones oocitarias se mantuvieron similares durante el tiempo de experimentación. Los oocitos en vitelogénesis tardía son los que tuvieron la segunda mayor proporción celular desde el inicio hasta el final del experimento. Con este resultado se concluye que las condiciones a las que se sometieron los individuos de P. axelrodi, permitieron mantener hembras con un estado de madurez avanzado durante seis meses. Si se logra establecer el, o los parámetros desencadenantes de la maduración final y desove de las hembras, se tendrían individuos maduros por un periodo prolongado.

Stimulation of Ovarian Development in a Tropical Damselfish by Prolonged Photoperiod using Pellets Containing Long-afterglow Phosphorescent Pigment

The present study examined whether light emitted by long-afterglow phosphorescent pigments (LumiNova) would stimulate gonadal development in fish during the nonbreeding season. Pellets containing LumiNova powder (treatment group) were prepared and placed on the calvaria of specimens of the sapphire devil Chrysiptera cyanea, a reef-associated damselfish that requires long days for gonadal recrudescence. A pellet without LumiNova powder was placed on the calvaria of the control fish (control group). Fish were reared at <TEX>$26^{\circ}C$</TEX> under a light-dark cycle (12 h photophase, 12 h scotophase; LD 12:12) for 4 weeks. No difference in the gonadosomatic index (GSI) or ovarian histology was observed among the control, sham-operation, and treatment groups 1 week after the start of the experiment. After 4 weeks, the GSI of the control and sham-operation groups remained at low levels, and ovaries contained immature oocytes at the perinucleolus stage. In contrast, the treatment group exhibited significantly higher values of GSI as well as developed ovaries with fully vitellogenic oocytes. These results demonstrated that long-day conditions were produced by light emitted from the LumiNova pellets, thus stimulating ovarian development in the damselfish. Therefore, long-afterglow phosphorescent pigments can be used as an alternative to standard light sources for purposes of artificial stimulation of gonadal development in fish.

Oogenesis in Shabout Barbus grypus Heckel , 1843

The female Shabout were collected as samples from Dijla river in Baghdad. The oocyte developmental stages were divided in to nine stages, based on nucleus structure , cellular inclusions and ovarian follicle structure ,they are:-( Oogonia stage , Early Perinucleolar Stage , Advanced Perinucleolar Stage , Cortical Alveoli Stage , Primary Yolk Stage , Secondary Yolk Stage , Tertiary Yolk Stage , Migratory Nucleus Stage , Hydration Stage(

Oogenesis in Shabout Barbus grypus Heckel , 1843

The female Shabout were collected as samples from Dijla river in Baghdad. The oocyte developmental stages were divided in to nine stages, based on nucleus structure , cellular inclusions and ovarian follicle structure ,they are:-( Oogonia stage , Early Perinucleolar Stage , Advanced Perinucleolar Stage , Cortical Alveoli Stage , Primary Yolk Stage , Secondary Yolk Stage , Tertiary Yolk Stage , Migratory Nucleus Stage , Hydration Stage(

DIETARY SUPPLEMENTATION OF VITAMIN C, L-ASCORBATE 2-TRIPHOSPHATE CALCIUM (LATP-Ca) ON OOCYTE DEVELOPMENT OF MAJOR CARP ROHU (LABEO ROHITA) REARED IN A NATURAL CONCRETE POND

The aim of this study was to evaluate the effect of vitamin C L-ascorbate 2-triphosphate (LATP-Ca)on Indian major carp rohu (Labeo rohita) (Hamilton) during intensive aquaculture. Rohu were fedwith vitamin C (LATP-Ca) supplementation at doses of D1 (control) and D2 (500 mg/kg) diet for 10weeks. The effects were assessed by comparing the treated groups of fish to that of control group.Vitamin C (LATP-Ca) supplementation exerted significant (P&lt;0.05) effects on oocyte developmentof rohu (Labeo rohita). A number of distinct developmental stages of oocyte can be delineated andoocyte growth in major carp rohu (Labeo rohita) found in two distinct phases; primary growthphase (PGP) and the secondary growth phase (SGP). In PGP, oogonia, chromatin nucleolus, earlyand peri nucleolus stage oocyte with Balbiani’s vitelline body were observed while two types ofinclusions, lipid and protein, respectively were formed during vitellogenesis in the SGP. SGPdevelopment was found for a short period of time as compared to PGP. Similarly, development ofoocyte was very poor in fish fed with controlled diet.DOI: http://dx.doi.org/10.3126/eco.v19i0.9849EcoprintAn International Journal of EcologyVol. 19, 2012Page: 7-15Uploaded date: 2/14/2014

Efficacy of 17 α-methyl testosterone and letrozole on sex reversal of protogynous grouper,Epinephelus tauvina(Forskal, 1775) and spawning performance of sex-reversed males

This study aimed to test the efficacy of 17 α-methyl testosterone (17 α-MT) alone and in combination with letrozole, an aromatase inhibitor, for the induction of sex reversal in protogynous greasy grouper, Epinephelus tauvina. Further, the long-lasting effects of these treatments and spawning performance of sex-reversed males were also investigated. Greasy grouper with oocytes in the perinucleolus stage were implanted with 5 mg 17 α-MT kg−1 body weight (T1), 5 mg 17 α-MT and 0.2 mg letrozole kg−1 body weight (T2) and 5 mg 17 α-MT with 0.4 mg letrozole kg−1 body weight (T3) and no androgens/enzyme inhibitor implanted (C). The 17 α-MT alone and in combination of letrozole-induced sex reversal in greasy grouper, whereas untreated control fish (C) showed normal ovarian development. However, T2 and T3 group showed 100% sex reversal and completion of spermatogenesis up to functional male phase in 2 and 3 months, respectively, whereas T1 group resulted in only 66.67% functional male with motile spermatozoa after 4 months. Sex-reversed males successfully fertilized the eggs during induced spawning. There were significant differences on fertilization and hatching rates between T2 group (79.00 ± 4.36%; 77.67 ± 2.87%, respectively) and T1 group (57.67 ± 3.17%; 63.87 ± 2.91%, respectively). The result suggested that 17 α-MT (5.0 mg kg−1 BW) in combination with letrozole (0.2 mg kg−1 BW) has the potential to produce 100% sex-reversed male in short period in greasy grouper, which might greatly help in seed production of greasy grouper.

Seasonal changes in the histology of the ovaries of Nile tilapia (Oreochromis niloticus)

Basic Histology of Ovaries of Nile tilapia (Oreochromis niloticus) was studied. Sampling was initiated of a total 40 female sexually mature Nile tilapia fish were collected over the period from September 2009 to August 2010. Ovaries were processed by standard histological technique. Histological characteristics of ovarian tissues and oocyte stages were studied by light microscopy. It revealed different histological structure of each oocyte developmental stage: Oogonia stage; Chromatin nucleolar stage; Perinucleolar stage; Cortical alveoli formation stage; Vitellogenic (yolk) stage; Postvitellogenic (mature) stage. Yolk nucleus (Balbiani bodies) was noticed in the ooplasm of the perinucleolar stage . During breeding season the ovary was surrounded by thin tunica albuginea. The ovigerous lamellae contained oocyte in active vitellogenesis, In addition; atretic and post ovulatory follicles were increased. During winter, the tunica albuginea reached a maximuem thickness and contained oocytes in previtellogenic stages.

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. We evaluated the effect of high water temperature on the brain–pituitary–gonadal (B–P–G) axis of a perciform fish, red seabream, Pagrus (Chrysophrys) major during its spawning season (April–May). Fish were reared at either high (24°C: H-group) or optimal/control (17°C: C-group) temperatures for 5 or 10d. After 5d, the transcript abundance of gonadotropin-releasing hormone-1 (GnRH1) in the brain and GnRH receptor (GnRH-R) and FSH-β in the pituitary were significantly lower in H-group than in C-group. Conversely, there was no difference in pituitary LH-β mRNA levels, serum concentrations of estradiol-17β (E2), or the gonadosomatic index (GSIs) between H- and C-groups on Day 5. After 10d, the ovaries of all H-group fish had completely regressed and were filled with only perinucleolar stage primary oocytes and atretic oocytes. The brain GnRH1 expression, pituitary GnRH-R and pituitary LH-β expression, serum E2 concentrations, and the GSI were significantly lower in the H-group on Day 10. Our results suggest that high water temperature is the proximate driver of the termination of the spawning season of female red seabream. The effect appears to be mediated by suppression of gene expression in the B–P–G axis.

Molecular characterization and quantification of sablefish (Anoplopoma fimbria) gonadotropins and their receptors: Reproductive dysfunction in female captive broodstock

Efforts to establish an aquaculture industry for sablefish (Anoplopoma fimbria) are constrained by reproductive dysfunction in wild-caught fish and by lack of reproduction of F1 females. Toward a better understanding of the reproductive dysfunction of captive broodstock, full-length cDNAs encoding the sablefish gonadotropin subunits (fshb, lhb and cga) and their receptors (fshr and lhcgr) were cloned, sequenced and quantitative real-time PCR assays developed. Sablefish gonadotropin subunits display some unique features, such as two additional Cys residues in the N-terminal region of Fshb and a lack of potential N-glycosylation sites in Fshb and Lhb, whereas Fshr and Lhcgr possess conserved structural characteristics described in other vertebrates. Wild females captured in fall completed gametogenesis in captivity the next spawning season, whereas females captured three months earlier, during summer, failed to mature. Interestingly, these wild non-maturing females exhibited similar reproductive features as prepubertal F1 females, including low levels of pituitary gonadotropin and ovarian receptor mRNAs and plasma sex steroids, and ovarian follicles arrested at the perinucleolus stage. In conclusion, this study described the cloning, molecular characterization and development of qPCRs for sablefish gonadotropins and their receptors. Rearing conditions may impair vitellogenic growth of ovarian follicles in sablefish, compromising the reproductive success of broodstock.

Ovarian expression and localization of a vitellogenin receptor with eight ligand binding repeats in the cutthroat trout (Oncorhynchus clarki)

A cDNA encoding a vitellogenin receptor with 8 ligand binding repeats (vtgr) was cloned from ovaries of the cutthroat trout, Oncorhynchus clarki. In situ hybridization and quantitative PCR analyses revealed that the main site of vtgr mRNA expression was the oocytes. Expression was strongly detected in perinucleous stage oocytes, gradually decreased as oocytes grew, and became hardly detectable in vitellogenic oocytes. A rabbit antibody (a-Vtgr) was raised against a recombinant Vtgr protein in order to immunologically detect and localize Vtgr within the ovarian follicles. Western blotting using a-Vtgr detected a bold band with an apparent mass of ~95-105kDa in an ovarian preparation that also bound Sakhalin taimen, Hucho perryi, vitellogenin in ligand blots. Immunohistochemistry using a-Vtgr revealed that the Vtgr was uniformly distributed throughout the ooplasm of perinucleolus stage oocytes, subsequently translocated to the periphery of lipid droplet stage oocytes, and became localized to the oolemma during vitellogenesis. We provide the first characterization of Vtgr at both the transcriptional and the translational levels in the cutthroat trout, and our results suggest that this receptor is involved in uptake of Vtg by oocytes of this species.

Reproductive Biology of Feather Back, Chital (Notopterus chitala, Ham.) Cultured in a Pond of Bangladesh

Studies on Gonadosomatic index (GSI), ova diameter and histology of the gonad were performed to understand reproductive biology of Feather back, Chital (Notopterus chitala) for a period of 6 months from January to June 2010. Thirty live chital fish were used in this study. The mean GSI values for female chital were found to range between 0.20±0.013 and 4.63±0.50. The highest GSI value was found in June. The smallest diameter of ovum was recorded 0.04 mm (January) and the largest was 4.00 mm (June). During the experimental period, the fecundity was ranged from 8,238 to 18,569 (mean 13,052±4607) in fish samples with body weight range from 1,296 to 2,360 (mean 1,742.50±474.44 g) while the relative fecundity was 5.65 to 14.33. Histological study revealed that the percentage of late perinucleolus (LPN) stage was highest in April and Cortical alveoli (CA) stage appeared from April and reached to maximum in May. Vitellogenic stage (VG) appeared in the month of May and chronologically increased through June. Vitellogenic stage (VG) of oocyte as well as highest ova diameter reached to the peak in June. The variations in the gonad weight and GSI of the female fish reached to the peak during June indicating maturity of ovary and definite spawning season. DOI: http://dx.doi.org/10.3329/ijarit.v2i1.13991 Int. J. Agril. Res. Innov. &amp; Tech. 2 (1): 26-31, June, 2012

Molecular cloning and mRNA expression pattern of Sox10 in Paramisgurnus dabryanus

A number of genetic studies have established that Sox10 involved in a wide range of developmental processes including sex differentiation and neurogenesis in vertebrates. A Sox10 homologue was cloned from brain of Paramisgurnus dabryanus by using homologous cloning and RACE method, designated as PdSox10. The full-length cDNA of PdSox10 contains a 312 bp 5' UTR, a 1,476 bp open reading frame (ORF) encoding 492 amino acids and a 262 bp 3' UTR (Accession no.: JQ217143). The overall topology of the phylogenetic tree shows that the PdSox10 fits within the Sox10 clade. During embryogenesis, PdSox10 gene seemed to be de novo synthesized in the embryos from gastrulae stage. From the somitogenesis stage and thereafter, distinct expression of PdSox10 was observed in the medial neural tube, extending from the hindbrain through the posterior trunk. In adult, PdSox10 mRNA was detected primarily in the gonads, as well as in brain and heart by RT-PCR. In situ hybridization on gonadal sections further demonstrated that PdSox10 is expressed especially in premature germ cells, in early perinucleolus stage oocytes and cortical-alveolar stage oocytes in ovaries and in spermatogonia and spermatocytes in testes. These preliminary findings suggested that PdSox10 is highly conserved during vertebrate evolution and involved in a wide range of developmental processes including neurogenesis and sex differentiation in vertebrates.

Maturation-associated changes in the internal distribution of tetrodotoxin in the female goby Yongeichthys criniger

Maturation-associated changes in the internal distribution of tetrodotoxin (TTX) in the goby Yongeichthys criniger were investigated in 29 and 40 female specimens collected from Okinawa, Japan, from August 2008 to June 2009 (Group I), and from November 2009 to August 2010 (Group II), respectively. In Group I, based on changes in the gonadosomatic index (GSI) and histologic observation of the ovary, the period from October 2008 through January 2009 was estimated to be the ‘previtelline-forming period’, February through March 2009 the ‘vitelline-forming period’, April through June 2009 the ‘spawning period’, and August 2008 the ‘end of spawning period’ of the preceding year. The TTX content (mouse unit [MU] per gram tissue) of each Y. criniger tissue (skin, muscle, liver, and ovary) quantified by liquid chromatography/mass spectrometry (LC/MS) was generally high during the spawning period and continued to rise until the end of spawning period, especially in the ovary. Total TTX per individual increased considerably during the spawning period, most of which located in the ovary, indicating that Y. criniger obtains a high amount of TTX during the spawning period, and accumulates most of it in the ovary. In contrast, the TTX content of the skin was highest at the end of spawning period, and most of the total TTX located in the skin during this period as well as during the previtelline-forming period. In Group II, the maturation stage of the ovaries of all specimens was determined, and the specimens were grouped accordingly. In the perinucleolus stage, yolk vesicle stage, and yolk globule stage I, most of the TTX was localized in the skin, but the TTX in the ovary greatly increased as the maturation stage advanced from yolk globule stage I to yolk globule stage III.

Histología del ciclo reproductor de hembras del pejelagarto &lt;i&gt;Atractosteus tropicus&lt;/i&gt; (Lepisosteiformes: Lepisosteidae) enTabasco, México

In Southeast Mexico, A. tropicus is a species with great ecological, economic, cultural and aquaculture value, however wild populations have diminished due to diverse anthropological causes. The objective of this study was to characterize the reproductive cycle of A. tropicus, with a description of complete gonad structure and the changes in germinal and somatic elements during oogenesis, for better management and use of this species. Six sexually mature females (N=72) were captured monthly with drag nets during one year cycle (October 2003-October 2004) in wild populations of Pomposu lagoon, municipality of Jalpa de Mendez, Tabasco, Mexico. Females were sacrificed by decapitation and the ovaries were processed by standard histological methods; the diameter of 10-20 oocytes taken at random was determined in different stages of development. Gonadosmatic index (GSI), gonadal volume (gV) and follicule diameter (fD) were determined monthly. Among results we could determine eight oogenesis developmental stages: I: chromatin nucleolus stage, II: early perinucleolus stage, III: mid perinucleolus stage, IV: advanced perinucleolus stage, V: early vitellogenesis stage, VI: mid vitellogenesis stage, VII) advanced vitellogenesis stage and VIII: final maturation stage. The reproductive cycle could be divided in four phases: 1: proliferative phase, II: initial maturation phase, III: late maturation phase, IV: regressing phase. Analysis of the data obtained from the IGS, gV and fD values, oogenesis stages and reproductive phases, indicate that in December previtellogenesis phase restarts, from March through July yolk deposition (vitellogenesis) occurs, and from August to November the maximum reproductive activity takes place. Contrasting this pattern with local environmental factors such as temperature and precipitation, we suggested that these may serve as an environmental activator in the development of oogenesis and spawning of this species.

Stage-Specific Effects of Androgens and Estradiol-17beta on the Development of Late Primary and Early Secondary Ovarian Follicles of Coho Salmon (Oncorhynchus kisutch) In Vitro1

An in vitro system was used to analyze the effects of sex steroids on the development of primary (late perinucleolar stage) and early secondary, previtellogenic (early cortical alveolus stage) ovarian follicles of coho salmon cultured for up to 21 days. Late perinucleolar-stage follicles increased significantly in size after 7 days of treatment with low concentrations of 11-ketotestosterone (11-KT), a nonaromatizable androgen. An androgen receptor antagonist (flutamide) inhibited this growth-promoting effect, and the highest concentration resulted in atresia of follicles, implicating androgens as survival factors at this stage. Testosterone (T) was less effective than 11-KT in promoting growth, but blocking aromatization with exemestane resulted in a growth response similar to that of 11-KT. Estradiol-17beta (E2) had no effect on growth at this stage. After 21 days of culture, E2 was the most potent steroid in increasing the number of follicles containing cortical alveoli and the number of cortical alveoli within those follicles. At the early cortical alveolus stage, low doses of E2 promoted growth and strongly stimulated synthesis of cortical alveoli, actions that were inhibited by an estrogen receptor antagonist (tamoxifen). 11-KT displayed moderate growth-promoting effects, and 11-KT and T stimulated moderate to substantial increases in abundance of cortical alveoli. This study shows that the predominant role of androgens is the promotion of growth of late perinucleolar-stage follicles, while E2 stimulates both the growth and accumulation of cortical alveoli in early cortical alveolus-stage follicles.