Salmon Gonadotropin-releasing Hormone Research Articles

Effects of Exogenous Hormones on Spawning Performances, Serum Gonadotropin and Sex Steroid Hormone in Manchurian Trout (Brachymystax lenok) during Sexual Maturation

The objective of this study was to investigate the effects of salmon gonadotropin releasing hormone analog (S-GnRH-A) and human chorionic gonadotropin (HCG) on the serum levels of gonadotropins (GtHs) and sex steroid hormones in Manchurian trout during sexual maturity. Females in the treatment group were injected with 20 µg/kg S-GnRH-A and 400 IU/kg HCG, whilst males were injected with half the amount. Blood samples were collected at 0, 5, 10, and 20 days post injection (dpi). The results showed an increase in female follicle-stimulating hormone (FSH) levels in the treatment group at 5 dpi, and from 5 dpi onwards, a difference was observed between the groups, with higher FSH levels in the treatment group compared to the control group. In males, FSH levels showed group differences at only 5 dpi (p < 0.05) and were higher in the treatment group. In females, luteinizing hormone (LH) peaked at 10 dpi in the treatment group, and there were differences between the two groups at 10 dpi and 20 dpi. In males, LH was significantly higher in the treatment group than in the control group at 5 dpi and 10 dpi (p < 0.05). In females, estradiol (E2) was significantly higher in the treatment group than in the control group at 5 dpi and 10 dpi (p < 0.05), whereas in males, the difference between the two groups was only observed at 5 dpi (p < 0.05). The female testosterone (T) level in the treatment group was significantly higher than that of the control group at 5 dpi and 10 dpi (p < 0.05), while T levels were significantly higher in the male treatment group than in the control group at 5 dpi (p < 0.05). The level of 17α–hydroxyprogesterone (17α-OHP) in the females of the treatment group was significantly higher than that of the control group at 5 dpi and 10 dpi (p < 0.05), and 17α-OHP levels were significantly higher in the male treatment group than in the control group at 10 and 20 dpi (p < 0.05). The fish were successfully spawned after S-GnRH-A and HCG injections, and brood amount, fertilization rate, and hatching rate were significantly increased (p < 0.05). This study provides a framework for a better understanding of the mechanisms of exogenous hormone-mediated control of reproduction in Manchurian trout.

Chitosan nanoconjugation of GnRH analogue augments reproductive performance in Clarias magur (Hamilton, 1822)

A breeding trial was conducted to investigate the impact of a chitosan nanoconjugated formulation of Salmon gonadotropin-releasing hormone analogue (ChN-sGnRH-a) on the induced breeding performance of female Clarias magur (magur). A total of 72 magur breeders were divided into six treatment groups: Cn, the Negative Control (injected with bare chitosan nanoparticles); Cp, the Positive Control (injected with Gonopro-FH®, a commercially available inducing hormone, at a dose of 1 mL/kg body weight of fish); T1, injected with ChN-sGnRH-a at a dose of 1 mL/kg; T2, T1 + 10 mg/kg domperidone; T3, injected with ChN-sGnRH-a at a dose of 0.5 mL/kg; T4, T3 + 10 mg/kg domperidone. Blood was collected at 0 (before injection), 6, 12 and 18 h post-injection to determine the serum steroid profile. In the Cp treatment group, the serum steroid levels of testosterone, estradiol, and 17α, 20β-dihydroxy progesterone in the fish, displayed a sharp increase, peaking at the 6-h post-injection, followed by a significant decrease. However, the fish in the T2 and T4 treatment groups exhibited serum steroid peaks at the 12-h post-injection and maintained those levels up to the 18-h post-injection. The reproductive performance, as measured by spawning fecundity, fertilization rate, and hatching rate, was significantly higher in the T2 treatment group (p < 0.05). No significant differences (p > 0.05) were observed between the Cp and T4 treatment groups. Compared to the Cn treatment group, the mRNA expression levels of pituitary follicle stimulating hormone subunit beta and luteinizing hormone subunit beta, as well as ovarian 17β-hydroxysteroid dehydrogenase type 3, were significantly higher in the T2 treatment group, followed by Cp and T4 treatment groups. Fish in the T2 treatment group displayed fewer mature oocytes and numerous post-ovulatory follicles in their ovaries. Additionally, the ovarian histology of fish in the T4 treatment group closely resembled that of the Cp treatment group. The results indicate that the chitosan nanoconjugated formulation of Salmon GnRH analogue might contribute to maintaining elevated levels of gonadotropic and steroid concentrations, resulting in improved reproductive output in female magur breeders. It is also revealed that the induced breeding of magur cannot be achieved without using dopamine inhibitors in GnRH analogue injection. Using chitosan nanoparticles as a nano-delivery system could reduce the Salmon GnRH analogue dose without compromising the reproductive performance of female magur breeders.

Improved breeding and seed production of climbing perch (Anabas testudineus) in controlled tanks and cage systems

The climbing perch (Anabas testudineus) is a valuable fish species with significant potential for aquaculture. However, a low survival rate was observed at its early development stages, requiring an optimized protocol for sustainable aquaculture production. This paper presents an improved breeding and seed production technology for climbing perch. Mature climbing perch were induced to spawn using salmon gonadotropin-releasing hormone analog (sGnRHa) at a dose of 30 µg/kg-1 body weight of the breeder. The female broodfish spawn 9–10 hours after hormone injection. Developmental stages were defined based on the morphological features of the embryos from the first cleavage to the hatching stage. The first cleavage began at approximately 18–20 minutes after fertilization. Newly hatched larvae were observed within 18–20 h at 30°C, having an average larval size of 0.389±0.042 mm (total length). Successful breeding entails a selection of suitable algal species, an optimal aquaculture environment, regular feeding rates, consistent monitoring of larval development, and effective management of water quality. Survival rates from the larval stage to fry were achieved at 84.69% and 77.60%, respectively, while the survival rate to the fingerlings stage was 72.51%. Therefore, by implementing an optimized protocol, aquaculture practitioners can maximize the production potential of climbing perch while ensuring sustainable cultivation practices. The findings from this research contribute to the advancement of climbing perch aquaculture by providing valuable insights for the successful cultivation and increased economic profitability of A. testudineus species in hatchery productions.

Efficacy of salmon GnRHa, Ovaprim® and hCG for hormonal stimulation of spermiation in the Fowler's toad (Anaxyrus fowleri).

Ex situ amphibian populations can experience reproductive dysfunction due to the absence of environmental cues that trigger reproductive events. Assisted reproductive technologies (ART) for amphibians, specifically exogenous hormone regimens, can circumvent these external signals to induce gametogenesis and gamete release. Currently, the use of the mammalian reproductive hormones gonadotropin-releasing hormone (GnRH) and human chorionic gonadotropin (hCG) are used in a species-specific manner to stimulate amphibian breeding. Hormones or hormone mixtures that are effective in all breeding scenarios would provide the best option for conservation practitioners and some commercial products are already in use for breeding other ectotherms. Ovaprim®, which contains salmon GnRH analogue (sGnRHa) and the dopamine antagonist domperidone (DOM), is effective in fish aquaculture and may be effective for amphibians. To test this hypothesis, we treated Fowler's toads (Anaxyrus fowleri) with either sGnRHa alone, a high or low dose of Ovaprim® or hCG. We then compared spermiation response, sperm quantity and quality parameters, and changes in animal mass over time within each treatment. We found administration of Ovaprim® resulted in more males producing sperm with better motility compared to administration of sGnRHa alone. In addition, the Ovaprim® and sGnRHa treatments resulted in lower response rates, lower sperm motilities, more abnormal sperm, and higher aggregations of sperm compared to the hCG treatment. Furthermore, Ovaprim®-treated males gained significant mass, suggesting an anti-diuretic effect of DOM. Together, these results show that neither Ovaprim® nor sGnRHa, at the concentrations tested, are likely suitable replacements for hCG in ex situ bufonid breeding programmes and that hormone mixtures developed for fish may have limited transferability to new world toad species.

Chitosan- hypothalamic hormonal analogue nanoconjugates enhanced the reproductive performance in Indian major carp, Labeo rohita

The current study reports the potential of chitosan nanoparticles for the efficient delivery of hypothalamic hormonal analogue in Labeo rohita, aiming to improve reproductive performance. Here salmon gonadotropin-releasing hormone analogue (sGnRH-a) was conjugated with chitosan nanoparticles (ChN-sGnRH-a) to evaluate its efficiency in enhancing the reproductive gene expression, hormones as well as the overall reproductive output in L. rohita. A total of 54 pairs of brooders were selected and divided into six treatments viz; C0: Negative Control (fish injected with bare chitosan nanoparticles), C: Positive Control (fish injected with Gonopro-FH®, a commercially available inducing hormone, at a dose of 0.2 ml/kg body weight of fish), T1: fish injected with ChN-sGnRH-a at a dose of 0.2 ml/kg, T2: T1 + 10 mg/ml domperidone, T3: fish injected with ChN-sGnRH-a at a dose of 0.1 ml/kg, T4: T3 + 10 mg/ml domperidone. In T2 and T4 treatments, serum hormones, Testosterone (T); Estradiol (E2); Vitellogenin (Vtg); and 17α, 20β-dihydroxyprogesterone (17α, 20β-DHP) showed the sustained elevation. The means of reproductive traits like fecundity, fertilization rate, pseudo-gonadosomatic index and spawning rate were significantly (P&amp;lt;0.05) higher in T2 treatment, while no significant difference was found between C and T4 treatment. The mRNA expression level of follicle-stimulating hormone (fshβ), luteinizing hormone (lhβ) and their cognate receptors was significantly better in T2 treatment, while no significant difference was found between T4 and C treatments. Further, the histological analysis of ovaries showed increased post-ovulatory follicles in C, T2 and T4 treatments. The results indicate that ChN-sGnRH-a could help in reducing the recommended dose of sGnRH-a without affecting the reproductive performance in L. rohita females. The sustained releasing mechanism of this formulation may be used as an induced breeding strategy in female L. rohita broodstock.

Functional evaluation of a novel kisspeptin analogue on the reproduction of female goldfish

Kisspeptin (kp) is a key regulator of reproduction, which stimulates sexual maturation and gametogenesis in mammals, amphibians, and teleosts. In the present study, to enhance the biological activity of kp10, a novel analog (referred to as M-kp10) was designed based on the endogenous goldfish variant, in which phenylalanine 6 was substituted by tryptophan and the N-terminus was acetylated. Compared with the native kp-10 and salmon gonadotropin-releasing hormone (GnRH3), the effect of M-kp10 on sexual hormones and reproductive indices as well as the expression of kiss1, cyp19a1, and kiss1ra genes in goldfish (Carassius auratus) was investigated. In practice, peptides were synthesized based on the standard Fmoc-solid-phase peptide synthesis and purified by employing RP-HPLC, followed by approving their structure using ESI-MS. The results showed that M-kp10 increased significantly 17,20β-DHP, LH, FSH and E2 as well as fecundity, hatching and fertilization percentages than the other peptides. Histological studies revealed that M-kp10 led to the faster growth of ovarian follicles compared to the kp-10 and GnRH3. The genes of cyp19a1, kiss1ra, and kiss1 were remarkably more expressed after treatment with M-kp10. In conclusion, the results indicated the superiority of M-kp10 over kp-10 in inducing sexual maturation and accelerating the percentage of fecundity, suggesting that M-kp10 could be a promising candidate for application in the artificial breeding of fish.

Effects of exogenous hormones treatment on spermiation and plasma levels of gonadal steroids in Roughscale sole, Clidoderma asperrimum

Roughscale sole (Clidoderma asperrimum) is only wild caught because basic reproductive research on this species is lacking and gamete production in an artificial setting has not been successful. Exogenous hormone treatment has been used to induce gonadal maturation and final spermiation in wild-caught individuals. In this study, the effects of an exogenous hormone on spermiation in roughscale sole was investigated by implanting different concentrations of a salmon gonadotrophin-releasing hormone analog (sGnRH; 0, 25, 50, and 100 μg/kg body weight) into male fishes. The control group did not produce sperm after 21 days post-implantation, and the duration of spermiation was shorter compared to the other groups. The spermiation period and milt amount differed among the hormone-treated groups according to the hormone concentration used. Milt volumes in the groups treated with 25 and 100 μg/kg sGnRH increased compared to the control group, whereas exogenous hormone treatment had no effect on the movable sperm ratio. The spermatocrit was high at the beginning of spermiation in all groups and then tended to decrease gradually over time except in the experimental group treated with 100 μg/kg sGnRH. Plasma levels of testosterone, 11-ketotestosterone, 17α, 20β-dihydroxy-4-pregnen-3-one were not significantly affected by the sGnRH treatments. Our results demonstrate that it is possible to prolong the spermiation period and increase milt volume by treating male roughscale soles with an exogenous hormone. In addition, the artificial hormone treatment did not affect sperm motility.

Identification and functional characterization of gonadotropin -releasing hormone in pompano (Trachinotus ovatus)

Gonadotropin-releasing hormone (GnRH) is an important neuropeptide in the reproductive system. Although GnRH analogues have been used to artificially spawn pompano (Trachinotus sp.), the native forms of GnRH have not been described in this species. In this study three GnRH subtypes [sea bream GnRH (sbGnRH), chicken GnRH-Ⅱ (cGnRH-Ⅱ) and salmon GnRH (sGnRH)] were identified in pompano (Trachinotus ovatus). cgnrh-Ⅱ and sgnrh were mainly expressed in the brain of male and female fish, showing a tissue-specific expression pattern, while sbgnrh was expressed at different transcriptional levels in all tested tissues. In vivo injection experiment showed that sbGnRH significantly increased fsh and lh genes expression in a dose-dependent manner, but a high concentration of sbGnRH could desensitize the expression of lh. High concentrations of cGnRH-Ⅱ and sGnRH could induce the expression of fsh and lh. In addition, the results of in vitro incubation experiments showed that the high concentration of sbGnRH peptide could induce the expression of fsh and lh, while cGnRH-Ⅱ and sGnRH peptides could only induce the expression of fsh. 17β-estradiol (E2) and 17α-methyltestosterone (MT) significantly inhibited sbgnrh mRNA expression in a dose-dependent manner, but did not affect the expression of cgnrh-Ⅱ and sgnrh mRNA. sbGnRH is the main GnRH subtype in pompano. E2 and MT can play a negative role in the regulation of sbgnrh. This study provides a theoretical basis for the reproductive endocrinology of pompano.

Mid-autumn spermiation in outdoor-cultured pikeperch (Sander lucioperca) using different gonadoliberin application strategies

This study aimed to find the optimal strategy for the application of salmon gonadotropin-releasing hormone analogue (sGnRHa) in outdoor-cultured pikeperch males in an exceptionally early term in early November. According to the histological examination, on the day of hormonal treatment, pikeperch testes were in the late stadium of spermatogenesis, with 78.7% of germ cells attributed to sperm cells. Five experimental groups were established varying on hormonal preparations and water temperature: (1) sGnRHa (25 μg kg−1) was applied to fish at pre-warming at water temperature of 11 °C (GnRH-LOW); (2) sGnRHa (25 μg kg−1) was applied to fish post-warming at water temperature of 15 °C (GnRH-HIGH); (3) priming group (5 μg kg−1 at 11 °C +25 μg kg−1 at 15 °C); (4) reference group – fish were injected with human chorionic gonadotropin; (5) control group - fish were injected with saline solution. Sperm was obtained from all fish in hormonally treated groups, whereas the fish treated with saline did not spermiate. The greatest values of the straight-line velocity (VSL), the curvilinear velocity (VCL), and the angular path velocity (VAP) of sperm cells, as well as the amounts of Mg2+, K+, and Na+ in the sperm seminal plasma, were obtained in GnRH-HIGH, whereas the GnRH-LOW group yielded the greatest sperm volume. Among the ions in seminal plasma, K+ and Na+ dominated and showed a relationship with sperm kinematic parameters. According to the obtained data, sGnRHa can be considered as an appropriate hormonal preparation to induce spermiation at an early preseason. Either before or after warming, sGnRHa application can yield sperm of proper quantity and quality as early as 5 months before the natural spawning season. A comprehensive study is recommended to evaluate the full period of sperm availability in outdoor-reared males, followed by direct evaluation of the fertilizing capacity.

Sex Dependent Action of Aroclor 1254 on Basal and sGnRHa-Stimulated Secretion of LH from the Pituitary Cells of Common Carp, Cyprinus carpio L.

Abstract Polychlorinated biphenyls (PCBs) affect the hypothalamic-pituitary-gonadal axis in many vertebrates, changing the hormonal regulation of reproduction. To identify one of the possible sites of action of PCBs on gonadotropin release in common carp, the direct effects of Aroclor 1254 on luteinizing hormone (LH) secretion from dispersed pituitary cells were investigated. Pituitary cells were obtained from sexually mature male and female common carp (Cyprinus carpio L.) at the time of natural spawning. The cells were incubated with different concentrations of Aroclor 1254 (5, 10, 50 and 100 ng mL–1 medium) and/or salmon gonadotropin-releasing hormone analogue (sGnRHa) at a concentration of 10−8 M. LH levels were measured in the cultured medium by the ELISA method after 10 hours of cell incubation. Incubation of male pituitary cells in the presence of tested concentrations of Aroclor did not change the basal LH secretion to the media. In the female pituitary cell incubations Aroclor (5, 10 and 100 ng mL–1 medium) caused a significant increase in LH concentrations in comparison to control incubations. In the case of sGnRHastimulated LH secretion in incubations of cells of both sexes, all the concentrations of Aroclor significantly stimulated LH release and potentiated stimulatory effects of sGnRH analogue. These results indicate that endocrine disrupters, such as Aroclor 1254, may affect reproduction in fish, acting also directly on gonadotrophs at the level of the pituitary gland, changing LH secretion.

The use of S-GnRHa (salmon gonadotropin releasing hormone analogue) in induced breeding and early embryonic development of Gulsha, Mystus cavasius

The current study was carried out to optimize the dose of the synthetic hormone for induction, and to observe the embryonic and larval developmental in Gulsha, Mystus cavasius. Induced breeding was conducted by using Ovupin (S-GnRHa) hormone (each 1.5 ml vial contain 0.2 mg of an analogue of S-GnRHa) at four different doses i.e., 0.25, 0.5, 1.0- and 1.50-ml kg-1 body weight (BW) for females, and the half of these doses were applied to males. Among the applied doses, 0.5 ml kg-1 BW for female and 0.25 ml kg-1 BW for male provided the maximum fertilization (83.66%) and hatching (80.0%) rates. The eggs of M. cavasius were strongly adhesive, with covering on egg surface. The average diameter of fertilized eggs just after spawning was 85.58±5.87 µm. After fertilization, the first, second, and third cleavage stages occurred within 20-25, 35-40 and 60-65 min, respectively. The identity of blastomeres was gradually lost and appeared at 64-cell stage to 128-cell stage onwards. The 64-cell stage appeared at 150-160 min and the morula stage 3:00-3:20 h (blastomeres completely lost), respectively. The gastrula stage appeared at 5:0-5:30 h in which the blastoderm spread in both the sides covering about 60-70% area, together with a thread-like germinal ring. Afterward, twisting locomotion was recorded at 23:30 h. The larvae started hatching at 24:00 to 25:00 h. The barbells were partially visible when the larvae were 10-12 h of age. Finally, the yolk sac was fully absorbed in the end of Day 3.

The stimulation effects of green light on the growth, testicular development and stress of olive flounder Paralichthys olivaceus

The light spectrum is one of the photic environmental factors. To investigate the effects of the light spectrum on the growth, testicular development and stress in fish, the olive flounder (Paralichthys olivaceus) from total length 1.5 cm were exposed to green light or white light for 205 days. The total length and body weight of the flounder reared under green light were significantly higher than those under white light. No significant difference in survival rate was found between the two groups. Levels of growth hormone (GH) and insulin-like growth factor (IGF) in serum were higher in flounders exposed to green light than those to white light. The gene expression levels were analyzed by quantitative-real time PCR. In hypothalamus, the mRNA levels of salmon gonadotropin-releasing hormone 3 (gnrh3) were significantly higher in the flounders exposed to green light than white light (p < 0.01); while melatonin receptor 1 and melatonin receptor 2 mRNA level was significantly lower in the green group (p < 0.05). In pituitary, luteinizing hormone β-subunit mRNA level was significantly higher under green light (p < 0.05). Gonadal histology analysis showed that there was significant difference in the testicular development between the two groups. Cortisol level was higher in flounders exposed to green light than white light both in the whole body and in serum. In conclusion, green light could promote growth by elevating GH and IGF levels in serum and expression levels of gnrh3 in hypothalamus, stimulate testicular development via hypothalamus-pituitary-gonad axis, and cause higher levels of cortisol in flounder. These results indicate green light could be used in aquaculture to increase production and be helpful for breeding.

Effects of hormonal treatment before water warming on synchronisation of spawning time, oocyte size, and egg quality in pikeperch (Sander lucioperca)

Inducing reproduction during periods of the year when spawning typically does not occur is an important goal for the feasibility of commercial fish farming. Pre-seasonal propagation of pikeperch generally occurs about 3 months before the natural spawning season. The objective of this study was to assess effects of imposing a thermal schedule for control of water temperature and differing salmon gonadotropin releasing hormone analogue (sGnRHa) dosages on final stages of oocyte growth, and egg quality by optimizing protocol duration and synchronizing spawning time. In Experiment 1, there was analysis of thermal schedule effects for water temperature control when hormonal administrations occurred before or after water warming (WARMING and STABLE, respectively). In Experiment 2, there was assessment of the sGnRHa dosage effects during the warming schedule. In both experiments there was analysis of oocyte diameter from time of sGnRHa administration until the late stages of maturation. There was greater synchrony in time of spawning in specimens of the WARMING group with lesser variability in time from sGnRHa administration to spawning. In Experiment 2, values for reproductive variables were variable among the different groups, without any differences between treatments. Oocyte diameter at the time of sGnRHa administration was correlated with embryo survival. For effective pre-seasonal pikeperch propagation, the selection of breeders based on oocyte diameter, and administration of 5 μg/kg sGnRHa immediately upon transport to hatchery, followed by a 1 °C/d temperature increase to 10 °C, are effective methods for induction of spawning during periods when spawning does not naturally occur.

Use of Synthetic Salmon GnRH and Domperidone (Ovaprim®) in Sharks: Preparation for ex situ Conservation

Shark populations are constantly decreasing owing to environmental destruction and overfishing; thus, sharks are now at a risk of extinction, with 27.9% of shark species classified as endangered on the International Union for Conservation of Nature’s Red List. Sharks are apex predators and a keystone species in balancing the marine food chain; their extinction will create an imbalance of the entire marine ecosystem. Assisted reproductive technology is the last resort for protecting animals facing severe extinction. Here, as a proactive effort toward building a hormone-induced artificial insemination protocol for endangered wild sharks, we identified the possibility of germ cell maturation by administration of Ovaprim®, a commercially produced synthetic salmon gonadotropin-releasing hormone, and calculated its optimum dosage and injection timing. The experiment was conducted on two shark species—Triakis scyllium and Triaenodon obesus. We found that intramuscular injections of 0.2 mL/kg of Ovaprim® for male T. scyllium and T. obesus, 0.2 mL/kg + 0.5 mL/kg at a 24 h interval for female T. scyllium, and 0.2 mL/kg + 0.2 mL/kg or 0.2 mL/kg + 0.3 mL/kg at a 24 h interval for female T. obesus were optimal dose protocols. These doses effectively induced the maturation and ovulation of oocytes and the release of semen. Our results confirm that Ovaprim® is a suitable tool for shark hormone-induced artificial insemination and indicate that this method may enable the conservation of the endangered shark species.

Induced spawning and embryonic development of striped murrel Channa striata (Bloch, 1793) under indoor conditions using salmon gonadotropin releasing hormone analogue (sGnRHa)

Ten successful trials on induced breeding of the striped murrel Channa striata (Bloch, 1793) under indoor conditions were carried out during July-August 2019. Brooders reared in cement tanks,(males weighing in the range of 290-700 g and females 220-600 g), were used in a set of 2:2 male to female ratio for induced breeding with salmon gonadotropin releasing hormone anologue (sGnRHa) and dopamine antagonist commercial formulation. Each brooder set was kept in an FRP tank (13’x3’x14’’) along with submerged macrophyte Hydrilla verticillata, which was sparsely spread all around the tank. Maturity of brooders was screened on the basis of secondary sexual traits. The brooders were given hormonal injection after 6 h of acclimatisation under indoor conditions @ 1 ml kg-1 body weight (containing sGnRHa 20 µg ml-1 and domperidone 10 µg ml-1) to both the sexes. Natural spawning occurred in all ten sets within 21.5+3.2 h (17-26 h) of injection. In all the tanks, two separate egg masses were observed with 30-95% fertilisation. Approximately 0.14 million eggs were scooped out from the tanks and reared in round plastic tanks (70 l) for hatching, where hatching occurred within 17-26 h post-fertilisation (hpf) with hatching rates of 50.76 to 93.18% at 28+2oC. The embryonic development was recorded under a stereo-zoom microscope in live condition under controlled temperature conditions and described with digital images. The study revealed that induced breeding of C. striata can be undertaken in FRP tanks under indoor conditions for mass-scale seed production, which will provide a platform for diversification of this high valued fish species for large scale farming practices in ponds as well a new species for culture in recirculatory aquaculture system (RAS) and biofloc technology (BFT).

Reproductive performance of lambari ( Astyanax altiparanae ) in a seminatural system using different protocols

We evaluated lambari (Astyanax altiparanae) reproductive performance by administration of gradually increasing doses of analogue of salmon gonadotropin-releasing hormone (sGnRHa). We performed two consecutive experiments applying six sGnRHa single doses in three replicates (ranging from 1 to 160 µg/kg) (Exp. 1) and three sGnRHa-fractioned doses in four replicates (10% + 90% – 12 hr interval) (10, 100 and 1,000 µg/kg) (Exp. 2) in 6 months old breeders. In both experiments, 6 mg of Carp pituitary extract (CPE)/kg and saline solution (without hormonal therapy) were used as positive and negative controls, respectively. Reproductive performance was recorded for both experiments; and quantification of 17α-20β-dihydroxy-4-pregnen-3-one (17,20β-P) and prostaglandin F2α plasma levels and ovarian histological evaluations were introduced in Exp. 2. Using a single dose, the spawning rate was low and similar between all groups (from 0% to 20%). Using fractioned doses, the spawning rate of CPE (60%) was higher than other groups (from 10% to 25%), but similar to 1,000 µg of sGnRHa/kg (40%). 17,20β-P increased at the final sampling regardless of treatment (including controls) or successful spawning, suggesting that the simple grouping of fish caused this increase. 17,20β-P and PGF2α plasma levels did not explain ovulation failures in this species. CPE-fractioned doses improved lambari ovulation and provided final maturation in all evaluated females; however, hormonal induction was not conditional for obtaining ovulation since spawning was registered in negative control. Concluding, CPE-fractioned doses protocol can be applied to increase reproductive performance of this species.

Nesfatin-1 suppresses fish reproductive axis and gonadal steroidogenesis.

Nesfatin-1 is a naturally occurring orphan ligand in fish and mammals. Research in our lab resulted in the identification of an inhibitory role for nesfatin-1 on pituitary hormones (goldfish) and oocyte maturation (zebrafish). The present study is an extension of these original findings and aimed to determine whether nesfatin-1 has any additional effects on HPG genes in male and female goldfish. We found that a single i.p. injection of synthetic nesfatin-1 (50 ng/g body weight) downregulated the expression of salmon gonadotropin-releasing hormone (sgnrh), chicken gnrh-II (cgnrh-II), kisspeptin receptor (gpr54a) and brain aromatase (cyp19a1b) mRNAs in the hypothalamus of both male and female goldfish at 15 min post-administration. In the pituitary of both males and females, nesfatin-1 reduced luteinizing hormone beta (lhβ) and follicle stimulating hormone beta (fshβ) mRNA expression at 60 min and gpr54a mRNA at 15 min. Similarly, the gonadotropin receptors lhr and fshr were downregulated in the gonads. Meanwhile, gonadotropin inhibiting hormone (gnih), gnih receptor, kisspeptin 1 (kiss1) and gpr54a mRNA expression in the gonads were increased post-nesfatin-1 treatment. Nesfatin-1 negatively influences the star, cytochrome P450 family 11 subfamily A member 1, anti-mullerian hormone and aromatase mRNAs. In agreement with these results, nesfatin-1 reduced plasma estradiol and testosterone in female and male goldfish circulation at 60 min post-injection. The information generated through this research further solidified nesfatin-1 as an inhibitor of reproductive hormones in fish. Targeting nesfatin-1 and related peptides could yield beneficial effects in fish reproduction and aquaculture.

Comparison of artificial spawning effectiveness of hCG, CPH and GnRHa in combination with dopamine inhibitors in a wild strain of ide Leuciscus idus (L.) in hatchery conditions

Optimization of artificial reproduction, by increasing the survival rate of embryos and hatching rate, is of major importance for reducing genetic diversity, especially in fish captured from their natural habitat that subsequently spawn in hatcheries. The artificial reproduction of ide, Leuciscus idus (L.) was conducted in controlled conditions. The spawning agents included: different doses of human chorionic gonadotropin (hCG) that was compared with common carp pituitary homogenate (CPH), Ovopel – a commercial agent for induction of spawning that contains mammalian gonadotropin releasing hormone analogue (mGnRHa) with dopamine antagonists (DA): metoclopramide (MET) and Ovaprim – a commercial agent containing salmon gonadotropin releasing hormone analogue (sGnRHa) and a dopamine antagonists (DA): domperidone (DOM). There were no ovulations in females from control groups. There were no differences between the ovulation rates (90 %) or embryo survival (> 92 %) and hatching rates (> 91 %) when there was administration of hCG doses between 500 and 1000 IU/kg. When there was comparison of different spawning agents, the ovulation rate was 100 % for all treated groups. There were the shortest and longest latency times to the time of ovulation after administration of CPH (26 h) and hCG (79 h), respectively. The greatest embryo survival (> 93 %) and hatching (> 91 %) rates occurred as a result of hCG administration with these values being slightly greater than when there was treatment with Ovaprim. The association between latency time and hatching rate indicated that when there was a slower final oocyte maturation (FOM) there were greater hatching rates.

Phoenixin-20 Stimulates mRNAs Encoding Hypothalamo-Pituitary-Gonadal Hormones, is Pro-Vitellogenic, and Promotes Oocyte Maturation in Zebrafish

Phoenixin-20 (PNX-20) is a bioactive peptide with hormone-like actions in vertebrates. In mammals, PNX stimulates hypothalamo-pituitary-gonadal hormones and regulate reproductive processes. Our immunohisto/cytochemical studies show PNX-like and the putative PNX receptor, SREB3-like immunoreactivity in the gonads of zebrafish, and in zebrafish liver (ZFL) cells. Intraperitoneal injection of zebrafish PNX-20 upregulates mRNAs encoding both salmon gonadotropin-releasing hormone (GnRH), and chicken GnRH-II and kisspeptin and its receptor in zebrafish hypothalamus. Similarly, luteinizing hormone receptor mRNA expression in the testis, follicle-stimulating hormone receptor in the ovary, and the kisspeptin system were upregulated in the gonads of PNX-20 injected fish. We also observed the upregulation of genes involved in the sex steroidogenic pathway (cyp11a1, cyp17a1, 17βhsd, cyp19a1a) in the gonads of PNX-20 administered fish. PNX-20 upregulates the expression of vitellogenin isoforms and estrogen receptor (esr2a and 2b) mRNAs in ZFL cells in vitro. Meanwhile, siRNA-mediated knockdown of PNX-20 resulted in the downregulation of all vitellogenin transcripts, further suggesting its possible role in vitellogenesis. PNX-20 treatment resulted in a significant increase in germinal vesicle breakdown in zebrafish follicles in vitro. Collectively, these results provide strong evidence for PNX-20 effects on the HPG axis and liver to promote reproduction in zebrafish.

Evaluation of induced spawning on oocyte characteristics and serum biochemistry of African catfish (Clarias gariepinus)

The effects of induced spawning using salmon gonadotropin-releasing hormone analogue (GnRHa) and domperidone proprietary drug (ovaprim) injected (0.4 ml/kg) intramuscularly on oocyte characteristics, sex hormones, thyroid hormones, and serum biochemical response were evaluated at 0, 12, 24, and 36 h post-induction. Twenty-eight 8 months old female Clarias gariepinus were used. At the designated hour, 7 of these fish were randomly selected for sampling. The response to the induction of spawning using ovaprim was time-dependent. The onset of response was observed from 12 h for increase in GSI, oocyte flotation, and oocyte diameter. The mean diameter of the different categories of the oocytes was significantly higher than those of the control at 24 h post-induction. Spawning was associated with a significant increase in serum estrogen and calcium level from 12 h post-induction. Furthermore, induction of spawning resulted in significantly higher serum levels of cholesterol, testosterone, and triiodothyronine as well as significantly higher activities of ALT and AST at different time-points. However, the BUN and serum levels of chloride and sodium were significantly lower compared with the control. From this study, the time for stripping of roe for optimal spawning outcome may be between 12 and 24 h.