The relationship between plasma and ovarian levels of gonadal steroids in the repeat spawning marine fishesPagrus auratus (Sparidae) andChromis dispilus (Pomacentridae)

The relationship between plasma and ovarian levels of gonadal steroids was examined in two New Zealand fish species with multiple spawning cycles of differing length. Snapper (Pagrus auratus) have a daily cycle of oocyte development, ovulation and spawning, whereas demoiselles (Chromis dispilus) spawn over 2–3 days during a repeat spawning cycle of 7–9 days. Ovarian and plasma levels of the gonadal steroids 17β-estradiol (E₂), testosterone (T), 17-hydroxyprogesterone (17P) and 17,20β-dihydroxy-4-pregnen-3-one (17,20βP) were measured in reproductively active fish captured from the wild. Ovarian levels of E₂, T and 17P changed in relation to spawning cycle and gonad stage in both snapper and demoiselles. E₂ and T levels were detectable at all times, but highest during vitellogenesis in both species. Cyclic changes of 17P occurred in both species, and levels appeared to depend on the rate of conversion of 17P to other hormones. No changes in ovarian levels of 17,20βP were detected in relation to stage of the spawning cycle in snapper; however, ovarian levels of 17,20βP were highest in demoiselles before spawning when fish undergoing final oocyte maturation predominated. Plasma levels of E₂ and T were strongly correlated with ovarian concentrations (r=0.850 and r=0.819 for E₂ and T respectively) in demoiselles but there was poor correlation between ovarian and plasma levels of 17P and 17,20βP (r=0.004 and 0.273 respectively), or between ovarian and plasma levels of E₂, T, 17P or 17,20βP of snapper (r=0.135, 0.277, 0.131 and 0.279). The poor correlation between plasma and ovarian levels of some steroid hormones suggests that plasma concentrations of steroids may not adequately reflect the reproductive status of the fish during short-term cyclic ovarian changes. It is suggested that this disparity is likely to be most marked in species with ovulatory periodicity of short duration.     

Hormonal and pheromonal control of spawning behavior in the goldfish

Species that employ sexual reproduction must synchronize gamete maturity with behavior within and between genders. Teleost fishes solve this challenge by using reproductive hormones both as endogenous signals to synchronize sexual behavior with gamete maturation, and as exogenous signals (pheromones) to synchronize spawning interactions between fish. This dual role of hormonal products is best understood in the goldfish, an external fertilizer with a promiscuous mating system. Female gonadal growth and vitellogenesis is stimulated by 17β-estradiol (E2) which also evokes release of a recrudescent pheromone. At the completion of vitellogenesis, ovarian E2 production drops and plasma testosterone increases, sensitizing the female gonadotropin II (luteinizing hormone; LH) system to environmental cues (temperature, spawning substrate, pheromones). These cues eventually trigger a LH surge that alters steroidogenesic pathways to favor the production of progestins including 17,20β-dihydroxy-4-pregnen-3-one (17,20β-P). Plasma 17,20β-P stimulates oocyte maturation but is also released to the water along with sulfated 17,20β-P and androstenedione to serve as a preovulatory pheromone. This pheromone stimulates male behavior, LH release, and sperm production. At the time of ovulation, females become sexually active in response to prostaglandin F2α (PGF2α) synthesized in the oviduct. PGF2α and its metabolites are released as a postovulatory pheromone that induces male spawning behavior which further increases male LH and sperm production. Androgenic hormones are required for male behavior and LH release. Although goldfish are gonochorists, hormone treatments can induce heterotypical functions in adults. Similar findings in other fish demonstrate that a sexually bipotential brain is not restricted to hermaphroditic fishes. This revised version was published online in July 2006 with corrections to the Cover Date.     

The use of luteinizing hormone releasing hormone analogue for ovulation induction in black sea bass (Centropristis striata)

Interest in commercial production of black sea bass has increased in recent years, but reliable spawning methods remain problematic. The objective of this study was to evaluate the effects of oocyte size and luteinizing hormone releasing hormone analogue (LHRHa) dosage and delivery systems on ovulatory success for in vitro fertilization. Vitellogenic females with maximum oocyte diameters of 400–625 μm were implanted with a 95% cholesterol–5% cellulose pellet containing 50 μg of LHRHa. Fish with maximum oocyte diameters < 450 μm failed to ovulate. In contrast, 90% of fish with 500 μm oocytes spawned within 36 h and 40% of this group ovulated a second time. All of the females containing oocytes > 550 μm ovulated. In a second experiment, females with uniformly vitellogenic oocytes (> 500 μm) and implanted with 50 μg LHRHa ovulated substantial numbers of eggs (45,000–192,000 eggs/kg body weight (BW), but fertility was consistently low (0–15%). In a third experiment, 19 of 39 females receiving implants containing 6.3–23.6 μg LHRHa/kg BW during the spawning season ovulated, but fecundity (17,000–339,000 eggs/kg) and fertilization (0–98%) were highly variable. When fish were grouped by developmental index, calculated as the number of oocytes with diameters > 400 μm/total number of oocytes measured, there were no statistical differences among groups with respect to the number of spawns, fecundity or fertilization success. In a fourth experiment, 11 of 13 females with a clutch of fully vitellogenic oocytes that were injected with 20 or 100 μg/kg BW LHRHa ovulated between 1 and 2 times on consecutive days. Five of seven given an implant containing 12.5 μg LHRHa ovulated one or more times. Fish implanted with shams or injected with vehicle alone did not ovulate in any of the experiments. No differences were found in the number of spawns, fecundity or fertilization success from fish receiving different doses of injected or implanted LHRHa. Incubation of pooled eggs produced 155,000 larvae (60% hatch) and 95,000 one-gram juveniles. These results demonstrate that injected or implanted LHRHa is effective for inducing ovulation in black sea bass with maximum oocyte diameters > 500 μm.     

Effect of exogenous hormones on ovulation and gonadal steroid plasma levels in starry flounder, <Emphasis Type="Italic">Platichthys stellatus</Emphasis>

The present study was designed to examine the potential for inducing ovulation in starry flounder (Platichthys stellatus) using gonadotropin-releasing hormone analog (GnRHa) and human chorionic gonadotropin (hCG) to assess whether starry flounder are differentially responsive to GnRHa and hCG. Female starry flounder were injected or implanted with different doses of hCG or GnRHa pellets to examine their ovulation-inducing potential and effects on plasma levels of testosterone (T), 17β-estradiol (E2), and 17,20β-dihydroxy-4-pregnen-3-one (17,20βP). Blood samples were collected for up to 10 or 25 days post-injection or post-implantation in two separate experiments designed to mimic the early and middle stages of spawning, respectively. Fish treated with the GnRHa pellets (100 µg) showed a significant increase in the total number of stripped eggs relative to the controls. GnRHa administration had no effect on the floating rate or fertilization rate of ovulated eggs in the both experiments, whereas hCG injection affected both of these rates. Plasma T levels were not significantly different between the exogenous hormone-treated and control fish. In contrast, the plasma E2 level was elevated in those fish treated with GnRHa, regardless of injection or implantation, and was accompanied by increased numbers of stripped eggs in both experiments. Treatment with GnRHa resulted in higher 17,20βP levels compared to the controls, and there was a positive relationship between elevated plasma 17,20βP and an increase in ovulated eggs in response to GnRHa treatment. The implantation of starry flounder with GnRHa-containing pellets was effective at inducing sustained ovulation compared to hCG treatment.     

Reproductive performance of common dentex, Dentex dentex, broodstock held under different photoperiod and constant temperature conditions

The effect of photoperiod and temperature on the timing and the quality of spawning, and on associated endocrine changes in circulating 17β-oestradiol, 11-ketotestoserone (11-KT) and vitellogenin (Vtg) were investigated in common dentex, Dentex dentex, undergoing their second reproductive cycle. The possibility was also explored of using the measurement of steroids in the culture water of broodstock tanks, rather than in individual blood plasma samples, as a potentially useful tool for assessing the physiological state of a fish without disturbing them. One group of fish was exposed to a simulated natural seasonal cycle and ambient temperature (CONTROL). The other two groups were exposed to simulated seasonal photoperiod cycles (12 month-long) but which were phase-shifted either three months before (ADVANCED) or after (DELAYED) the natural cycle. Temperature was kept at 19.4±0.9 °C all-year-round. In the CONTROL, spawning started in mid-April and lasted until mid-June, while in the ADVANCED group, spawning started 4 months earlier and in the DELAYED group 2 months later than the CONTROL. The total egg production, egg quality, hatching rate, relative fecundity, and spawning index of the experimental groups were similar to the controls. The differences in spawning time induced by photothermal manipulation were associated with a difference in the timing of peak concentrations of plasma E₂, 11-KT and Vtg. In all three groups, the amounts of conjugated 17,20β-dihydroxy-4-pregnen-3-one (17,20β-P) and free and sulfated 11-KT which could be extracted from the water during the spawning period were significantly higher than those found in the preovulatory period. However, the differences were mostly less than 2-fold suggesting that, at least under the conditions employed in this study, the method was of limited use for non-intrusive detection of gonadogenesis and spawning (as had been hoped). The observed differences in spawning time and in the seasonal changes of sex steroids and Vtg confirm and extend the findings on marine fish. This revised version was published online in August 2006 with corrections to the Cover Date.     

Effect of Oxygen Saturation in Water on Reproductive Performances of Pacu Piaractus brachypomus

Broodstock pacu Piaractus brachypomus as well as their eggs during their embryonic development were exposed to either normoxia (5.5–7.5 mg O₂/L) or hypoxia (2.0–4.5 mg O²/L) conditions. The plasma concentrations of 11-ketotestosterone in males and estradiol-17β in females, as well as that of their precursor testosterone (T) were significantly ( P < 0.01) higher in fish maintained under normoxic conditions than in fish exposed to hypoxia. After ovulation and spermiation induced by hormonal treatments, the plasma concentrations of T and 17,20β-dihydroxy-4-pregnen-3-one (17,20βP) significantly ( P 0.05) increased in both sexes under both normoxic and hypoxic conditions. The plasma levels of T and 17,20βP achieved under normoxic conditions were higher than the ones recorded under hypoxia, except for those of 17,20βP in males. Males responded positively to the hormonal treatments, and the concentration of spermatozoa was 10.5 ± 0.8 10⁹/mL under both oxygen conditions. Hypoxia resulted in significantly lower survival of embryos (17.3 ± 28%) in comparison to normoxic conditions (68.5 ± 25%). Moreover, larval deformities were found when exposed to hypoxia (91.6 ± 6%). During embryonic development of this species 4 mg/L of oxygen is tolerated at 26–27 C without negative impact. We conclude that despite the highly adaptable nature of adult pacu to environmental hypoxia, oxygen concentrations below 4 mg/L severely impacted survival of embryos.     

Determination of hormones inducing oocyte maturation in <Emphasis Type="Italic">Chalcalburnus tarichi</Emphasis> (Pallas, 1811)

Chalcalburnus tarichi is an endemic cyprinid species living in the Lake Van basin, in eastern Anatolia, Turkey. The present study was undertaken to determine which hormones induce oocyte maturation in C. tarichi. The levels of 17α,20β,21-trihydroxyprogesterone (20β-S), progesterone (P), 17α-hydroxyprogesterone (17α-HOP), 11-deoxycortisol (11-DOC), and 17α-hydroxy-20β-dihydroprogesterone (17,20β-P) were measured in fish caught from Lake Van and the Karasu River, and injected with human chorionic hormone (hCG) (1,000 and 1,500 IU/kg). Oocytes of fish caught from the lake were also incubated in vitro with different doses (50, 200, and 1,000 ng/ml) of 20β-S, 17α-HOP, 11-DOC, and 17,20β-P. 11-DOC was found to be the most effective hormone among those measured for inducing oocyte maturation in vivo and in vitro. 17,20β-P could not be determined in the plasma of any fish in vivo (P < 0.05). 1,000 IU/kg dose of hCG given by injection caused a statistically significant increase in all plasma hormone levels (P < 0.05). It was found that there was a significant decrease in the P level only at 1,500 IU/kg dose of hCG injected (P < 0.05), while the level of other hormones increased at this dose (P < 0.05). It was also determined that all the hormones were effective in germinal vesicle breakdown (GVBD) in in vitro oocyte culture (P < 0.05). However, 11-DOC was found to be the most effective hormone in GVBD at a dose of 200 ng/ml (70% GVBD). In conclusion, 11-DOC synthesized during final oocyte maturation in C. tarichi was found to be a potent inducer of GVBD, which shows that 11-DOC may be described as an oocyte maturation steroid in this species.     

LHRHa‐induced ovulation of the endangered‐Caspian brown trout (Salmo trutta caspius) and its effect on egg quality and two sex steroids: testosterone and 17α‐hydroxyprogestrone

To induce synchronized ovulation, migrating wild Caspian brown trout (Salmo trutta caspius) females were treated with two interperitoneal injections of Des‐Gly¹⁰, d ‐Ala⁶ LHRH (LHRHa), given 3 days apart. Two injections of 100 μg kg^?1 body weight of this hormone effectively induced ovulation. Within 27 days from the second injection, all fish injected with 100 μg kg^?1 LHRHa had ovulated compared with 54.5% of the controls. The mean time to ovulation was reduced significantly (P<0.05) from 31.67±4.84 days in control fish and 28.83±7.31 days in sham‐treated fish to 16.36±1.61 days in fish injected with 100 μg kg^?1 LHRHa. The fertilization rate in 50 and 100 μg kg^?1 LHRHa‐injected fish was significantly lower than that in the control fish (P<0.05). In fish injected with 50 and 100 μg kg^?1 LHRHa, significant (P<0.05) changes in testosterone (T) and 17α‐hydroxyprogestrone (OHP) levels were observed. After the second LHRHa injection, the fish injected with 100 μg kg^?1 showed the highest serum levels of testosterone and OHP. These results demonstrate that the use of LHRHa can effectively reduce the mean time to ovulation and induce synchronized ovulation in Caspian brown trout.     

Induction of ovulation in ornamental common carp (Koi, Cyprinus carpio L.) using LHRHa ([d-Ser(tBu)6, Pro9-NEt]-LHRH) combined with haloperidol and carp pituitary extract

The effects of a single injection of mammalian superactive analogue [d ‐Ser(tBu)⁶,Pro⁹‐NEt]‐LHRHa (20 μg/ kg^?1) combined with the dopamine antagonist, haloperidol (HAL, 0.5 mg kg^?1), for induction of ovulation in the koi carp broodstocks were determined under routine hatchery conditions. The results were compared with classic carp pituitary extract (CPE, double injection) application (water temperature 22 °C). Physiological saline (0.7% NaCl)‐injected fish were used as a control group and no ovulation occurred in this group. The spawning ratio was high in the LHRHa+HAL treatment group and in the CPE treatment group (6/7 and 5/7 respectively). The latency period was 14–16 h in the LHRHa+HAL treatment group and 12–14 h in the CPE treatment group (after the second injection). There was no difference between the two ovulating groups with respect to the spawning index (the weight of eggs as a percentage of female body weight) and fertilization rate of eggs (P>0.05). As a result, ovulation can be induced successfully in koi carp broodstocks with 20 μg kg^?1 [d ‐Ser(tBu)⁶,Pro⁹‐NEt]‐LHRHa+0.5 mg kg^?1 HAL treatment in a single injection without decreasing the egg quality. Application of this combination can be useful for hatchery and broodstock management in koi carp culture.     

The ovarian regulation of ovulation in teleost fish

Ovulation is the release of a mature oocyte from its follicle wall enclosure in the ovary. This process requires the separation of the oocyte from the granulosa layer, the rupture of the follicle wall and the active expulsion of the oocyte through the rupture site. Results of experiments on various vertebrates, including fish, have shown that the control of these processes may involve the cooperative action of a number of ovarian regulators including proteases, protease inhibitors, progestational steroids, eicosanoids, catecholamines and vasoactive peptides. We have used two teleost models, the brook trout (Salvelinus fontinalis) and the yellow perch (Perca flavescens) to study the mechanism and control of ovulation in fish. Using subtractive cDNA cloning, a family of ovarian and ovulation specific mRNAs (TOPs = trout ovulatory proteins) was isolated from the brook trout ovary. These mRNAs have not previously been observed in the ovary of any vertebrate species; however, the proteins they code for have significant sequence homology to a group of mammalian protease inhibitors called antileukoproteinases. These inhibitors have been isolated from several mammalian mucosal tissues and their function may be to protect the mucosal lining from nonspecific degradation by proteases released from infiltrating leukocytes. The ovarian proteins encoded by the TOP mRNAs have now been characterized by Western blotting using antibodies derived against recombinant TOPs. Given the similarity of TOPs to antileukoproteinases, one function of TOPs may be to regulate proteolysis at the time of ovulation. In yellow perch, the maturational steroid, 17,20-dihydroxy-4-pregnen-3-one (17,20-PG), stimulates both germinal vesicle breakdown and ovulation in vitro. The stimulation of ovulation can be blocked by indomethacin, a prostaglandin endoperoxide synthase inhibitor. Thus, it appears that 17,20-PG acts through the production of an eicosanoid that is most likely a primary prostaglandin. This hypothesis is further supported by the observations that (1) a direct correlation exists between indomethacin levels that block ovulation and those that block primary prostaglandin synthesis in the ovary; (2) ovulation can be restored in indomethacin-blocked incubates with primary prostaglandins; (3) PGF levels increase at the time of ovulation in incubations of yellow perch follicles stimulated with 17,20-PG; and (4) the stimulation of PGF by steroids in the ovary is specific for 17,20-PG. Finally, 17,20-PG-stimulated ovulation and follicular prostaglandin synthesis requires the close interaction of extrafollicular tissue and other follicle wall layers.     

The combined effects of temperature and GnRHa treatment on the final stages of sexual maturation in Atlantic salmon (Salmo salar L.) females

Atlantic salmon (Salmo salar L.) females (2 SW), maturing for the first time, were reared under one of three temperature regimes (high: 14.3 ± 0.5°C; natural: 10.6 ± 1.0°C; and cold: 6.9 ± 1.0°C) in combination with one of two experimental treatments; an injection of GnRH analogue (GnRHa) contained in biodegradable microspheres, or a sham injection (microspheres only). The six experimental groups were then reared under simulated natural photoperiod for 4 weeks. Blood samples were drawn for analysis of plasma steroid levels and the fish were inspected for ovulation weekly. Batches of stripped eggs were incubated in triplicate incubators in raceways until the eyed stage. Treatment with GnRHa resulted in a substantial advancement and synchronization of ovulation at all temperatures, while exposure to cold water also appeared to advance ovulation slightly. While 75% (warm and cold) to 90% (natural) of GnRHa fish ovulated during the 4-week trial, only 30% of sham-treated females exposed to cold water, and none of the sham-treated fish held at higher temperatures, ovulated during this period. Survival rates of embryos to the eyed-stage were significantly higher for broodstock exposed to cold water. Plasma levels of testosterone (T), 17β-oestradiol (E₂), and 17α,20β-dihydroxy-4-pregnen-3-one (17,20βP) were all significantly affected by treatment with GnRHa and, to a lesser extent, temperature. The efficiency of GnRHa in counteracting the negative effects of high temperature on ovulation and the associated changes in circulating sex steroids suggest that temperature inhibition operates at least in part at the brain or pituitary.     

Endocrine and gonadal changes during the annual reproductive cycle of the freshwater teleost,Stizostedion vitreum

The annual reproductive cycle of walleye (Stizostedion vitreum) was characterized by documenting changes in gonadal development and serum levels of estradiol-17β (E₂), testosterone (T), 17α,20β-dihydroxy-4-pregnen-3-one (17,20-P), and 11-ketotestosterone (11-KT) in wild fish captured from upper midwestern lakes and rivers throughout the year. Fish from the populations used in this study spawn annually in early- to mid-April. Walleye showed group synchronous ovarian development with exogenous vitellogenesis beginning in autumn. Oocyte diameters increased rapidly from ∼ 200 μm in October to ∼ 1,000 μm in November, and reached a maximum of 1,500 μm just prior to spawning. Changes in gonadosomatic indices (GSIs) paralleled changes in oocyte diameters. Serum E₂ levels in females increased rapidly from low values in October (< 0.1 ng ml⁻¹) to peak levels of 3.7 ng ml⁻¹ in November, coinciding with the period of the most rapid ovarian growth. Subsequently, E₂ levels decreased from December through spawning. Serum T levels exhibited a bimodal pattern, increasing to 1.6 ng ml⁻¹ in November, and peaking again at 3.3 ng ml⁻¹ just prior to spawning. We detected 11-KT in the serum of some females at concentrations up to 5.6 ng ml⁻¹, but no seasonal pattern was apparent. In this study (unlike our results in a related study) 17,20-P was not detected. In males, differentiation of spermatogonia began in late August, and by January the testes were filled (> 95% of germ cells) with spermatozoa. Mature spermatozoa could be expressed from males from January through April. GSIs ranged from 0.2% (post-spawn) to 3.2% (pre-spawn). Serum T levels rose from undetectable levels in post-spawn males to 1.6 ng ml⁻¹ by November, remained elevated throughout the winter, and peaked at 2.8 ng ml⁻¹ I prior to spawning. Levels of 11-KT in males remained low (< 10 ng ml⁻¹, from post-spawning through January, then increased significantly by March and peaked just prior to spawning at 39.7 ng ml⁻¹. Our results indicate that vitellogenesis and spermatogenesis are complete or nearly so, in walleye by early winter, and suggest that it may be possible to induce spawning in this species several months prior to the normal spawning season by subjecting fish to relatively simple environmental and hormonal treatments.     

The role of prostaglandins in the control of ovulation in yellow perch,Perca flavescens

Previous studies have shown that 17α,20β-dihydroxy-4-pregnen-3-one (17α,20β-P) can induce both germinal vesicle breakdown and ovulationin vitro of yellow perch (Perca flavescens) oocytes. The stimulation of ovulation can be blocked by indomethacin and restored by the subsequent addition of several primary prostaglandins (Goetz and Theofan 1979). In the present investigation, medium levels of prostaglandin F (PGF) and E (PGE) were measured by radioimmunoassay duringin vitro 17α,20β-P-induced ovulation of perch oocytes. PGF levels increased significantly (compared to controls) from 30 to 36h of incubation. Hourly samples taken through the time of ovulation revealed that the increase in PGF was very closely correlated to the time of ovulation though it did not preceed it. Cortisol, testosterone, estradiol-17β, 17α,20α-dihydroxy-4-pregnen-3-one and 17α-hydorxyprogesterone did not increase PGF levels by 48h of incubation, however, several other progestational steroids including 20β-dihydroprogesterone (20β-P) and progesterone did. 17α,20β-P, 20β-P and progesterone also stimulated an increase in PGF in spontaneously ovulating oocytes (in which all oocytes ovulated including controls), indicating that the increase in PGF was not merely a result of the physical process of ovulation but was related to the presence of the steroid. Based on work supported by the National Science Foundation under grant DCB-8517718 and DCB-8718178.     

Hormonally Induced Spawning, Embryonic Development, and Larval Rearing of the Southern Temperate Banded Morwong, Cheilodactylus spectabilis

Banded morwong (Cheilodactylus spectabilis) are of interest for marine finfish aquaculture in temperate southern Australia. To improve their ovulatory response, adult females were implanted during the autumn spawning season with slow‐release pellets containing 0–400 μg luteinizing‐hormone‐releasing hormone analogue (LHRHa)/kg body weight within 24 h of capture from the wild. Compared to the sham control group, animals treated with LHRHa produced significantly more eggs on each day after implantation for the following 7 d (91 ± 39 and 290 ± 38 mL) and a higher proportion ovulated (8/12 and 27/27). Of fish treated with LHRHa, 93% ovulated 2 d after implantation and 79% ovulated three times at 2‐d intervals, whereas control animals showed no cyclicity of ovulation and few ovulated more than once. Egg production was highest at the first ovulation after LHRHa treatment and declined at subsequent ovulations. In a second experiment investigating the range 100–400 μg LHRHa, there was no effect of dose rate on ovulation parameters, which additionally examined implantation either immediately after capture or after a 5‐d delay. Compared to immediate implantation, a delay resulted in a lower proportion of animals that could be stripped after implantation (100 and 50%, respectively) and the volume of eggs was lower (135 ± 15 and 107 ± 10 mL). The egg quality was poor following delayed implantation, resulting in no fertilization after artificial insemination compared with immediate implantation in which fertilization and hatch rates were higher for eggs collected on Day 2 after implantation (79 ± 8% and 58 ± 9%) than on Day 4 (23 ± 7% and 15 ± 6%). Thus, it is important to implant animals as soon as possible after capture to ensure optimum egg quality. Good‐quality eggs were buoyant and spherical and had a diameter of 1050 ± 25 μm with a single pigmented oil droplet of 190 ± 9 μm. When a separate large batch of eggs collected 2 d after implantation with 100 μg LHRHa was inseminated and cultured at 18 C, larvae hatched after 63 ± 2 h at a standard length of 2.6 ± 0.4 mm. Newly hatched larvae were buoyant and transparent with only a few melanophores, eyes were nonpigmented and jaws were nonfunctional. By the fourth day, jaws were functional and eyes were fully pigmented. Utilization of the endogenous yolk and oil was completed by Day 6, and swimming commenced with exogenous feeding. Larvae, initially fed lipid‐enriched rotifers followed by Artemia, reached 8.9 ± 0.7 mm length on Day 55, after which they metamorphosed to the postlarval paperfish stage of development, 22 ± 0.9 mm on Day 100, and 43 ± 1.0 mm at 6 mo of age. The results show that treatment of wild‐caught females with slow‐release pellets containing LHRHa is effective for the production of eggs for hatchery rearing.     

Plasma cortisol and hypothalamic monoamine responses in yellow perch <Emphasis Type="Italic">Perca flavescens</Emphasis> after intraperitoneal injection of lipopolysaccharide

The concentrations of monoamines in the hypothalamus were determined in yellow perch Perca flavescens before and after injection with lipopolysaccharide (LPS; 3 mg kg⁻¹ fish weight) or saline to test for the presence of neurochemical changes potentially associated with changes in plasma cortisol characteristic of intraperitoneal (ip) challenge with LPS. In the first experiment, yellow perch were injected with saline or LPS and the hypothalamus removed and plasma sampled before and at 0.5, 1.5, 3.0, and 6 h after injection. Plasma cortisol was elevated in both saline- and LPS-injected fish through 1.5 h after injection and returned to levels resembling pre-injection by 3 h after injection. Significantly higher amounts of cortisol in plasma from LPS-injected relative to saline-injected fish were observed 6 h following injection. A significant decrease relative to levels observed 0.5–3 h after handling was observed in serotonin concentrations at 6 h following LPS and saline injection with a concomitant increase in the ratio of 5-hydroxyindoleacetic acid:serotonin. In the second experiment, hypothalamic monoamines were sampled before and at 3, 6, 9, 12, and 24 h after injection with LPS or saline. Significant increases from pre-injection levels were observed in the ratio 5-hydroxyindoleacetic acid:serotonin at 9, 12, and 24 h after injection, but no differences were detected between LPS- and saline-injected fish. These results support a model linking serotonergic system activation following handling stress, but no correlations with the sustained elevations of plasma cortisol associated with inflammatory challenge were observed.     

Effects of Temperature and Photoperiod on Reproduction of Female Yellow Perch Perca flavescens: Plasma Concentrations of Steroid Hormones, Spontaneous and Induced Ovulation, and Quality of Eggs

The involvement of photoperiod and temperature in the regulation of reproductive processes was investigated in female yellow perch. Initially, all fish kept indoors were exposed to the same water temperature (22°C) and photoperiod (15L:9D). By the end of August, following the first sampling, fish were submitted to different photothermal regimes. Group A₃ was maintained under photothermal conditions characteristic for southern Ohio. Group B₃ was submitted to a condensed light/temperature regime designed to accelerate photothermal changes. The mean gonadosomatic index in group A₃ gradually increased throughout the experiment. Gonadosomatic index in group A₃ was higher than that in group B₃ from February through April. The highest plasma concentrations of estradiol in group A₃ occurred in November and December and exceeded those in group B₃. Plasma concentrations of testosterone in group A₃ were highest in December and tended to be higher in October and April than in other months. There was no difference in plasma testosterone between groups A₃ and B₃ except in March when concentrations of this androgen were extremely high in group B₃. Ovulation occurred earlier in B regime fish in comparison to that of A regime fish. However, egg quality of fish from regime B was lower than that of fish from regime A. We conclude that the condensed photothermal cycle is not an entirely effective method of inducing out-of-season spawning in female yellow perch. These data suggest that although the compression of the photothermal cycle slightly accelerated spawning, it also caused disturbances in patterns and levels of plasma steroids, diminished ovarian and hepatic growth, and reduced the quality of eggs. Thus, temperature and/or photoperiod may have only a modifying effect on the onset of perch spawning, but these environmental cues seem to markedly influence earlier stages of gonadal growth and development.     

A comparison of human chorionic gonadotropin and luteinizing hormone releasing hormone analogue for ovulation induction in black sea bass Centropristis striata (Linnaeus, 1758)

Mature black sea bass, Centropristis striata L. (200–800 g), were captured in coastal South Carolina during the spawning season and administered hormones for ovulation induction and strip spawning. During both study years, control groups of females were incorporated into the study design and administered sham injections containing physiological saline solution. In 2004, females received a single intramuscular injection of human chorionic gonadotropin (hCG) (330 IU kg⁻¹) (n=8) or two injections of hCG at 24‐h intervals (n=8). In 2005, females received a single injection of hCG (n=10) or an analogue of luteinizing hormone releasing hormone (LHRHa) (n=10). In 2004, all fish administered a single dose of hCG ovulated at least once. Six fish ovulated on two consecutive days and one fish ovulated on 3 days consecutively. In contrast, six of eight fish receiving two doses of hCG ovulated once, five ovulated on 2 days successively and three fish ovulated 3 days in succession. Of the fish that spawned, no differences were found in any reproductive parameters. In 2005, all fish administered hCG or LHRHa ovulated at least once. Three fish administered hCG ovulated twice, four fish ovulated on three consecutive days and one fish 4 days successively. All fish administered LHRHa spawned at least twice, six fish ovulated thrice and three fish ovulated 4 days, successively. A significant difference in fertility was found between hCG (75.6±11.4%) and LHRHa (55.6±27.4%). The results of this study indicate that both hCG and LHRHa are effective for ovulation induction in prespawning black sea bass.     

Additive effects of advanced temperature and photoperiod regimes and LHRHa injection on ovulation in Atlantic salmon (Salmo salar)

In order to evaluate manipulation of spawning time as a potential means to extend 0+ Atlantic salmon (Salmo salar) smolt production in Tasmania, Australia, female salmon were exposed to a natural/simulated natural (42°S) photoperiod or an advanced (L:D 9:15) photoperiod from the austral summer solstice (20 December) under natural or advanced (~ 6 °C below natural temperature) temperature conditions. In late summer (26 February) injections of a commercial LHRHa preparation or vehicle (propylene glycol) commenced. Regular ovulation checks were conducted and ova were fertilised using milt from LHRHa-injected males held under matching photo-thermal conditions. Plasma levels of 17β-estradiol (E₂) and testosterone (T) were monitored and reproductive success (cumulative % ovulation, % fertilisation and % survival to the eyed-egg stage) was recorded. Ovulations commenced first (09 March) in LHRHa-treated fish that experienced advanced photoperiod and thermal regimes whereas sham-treated fish exposed to natural photoperiod and temperature conditions where the last to ovulate (22 May-08 June). Treatment-related sequential changes in the timing of ovulations were reflected by sequential advances in the timing of peaks in plasma levels of E₂ and T. The fertilisation of ova from LHRHa-treated fish that experienced advanced photoperiod and thermal regimes was significantly reduced (~ 52%) relative to all other treatments (> 80%) but there were no significant treatment-related differences in the survivals of eggs to the eyed stage (~ 50-90%). Consequently, a maximum advance in the timing of median ovulation of 71 days and commercially acceptable eyed-egg yields were generated, demonstrating that combinations of photoperiod, thermal and hormone treatments may be employed to significantly extend spawning and thereafter increase the availability of 0+ smolts for grow-out.     

Dietary administration of an LHRH analogue induces spawning of spotted seatrout (Cynoscion nebulosus)

The effects of a single oral administration of des-Gly¹⁰-[D-Ala⁶]-luteinizing hormone-releasing hormone (1–9)-ethylamide (LHRHa) in the diet (0.2–2.5 mg/kg body weight) on ovulation and spawning of spotted seatrout (Cynoscion nebulosus) were examined. Oral administration of 1.0–2.5 mg LHRHa/kg to females in four separate experiments resulted in successful spawning 32–38 h later with mean fertilization and hatching success rates of 93.3% and 74.6%, respectively. A lower dose of LHRHa (0.2 mg/kg) was ineffective in two subsequent experiments. The data suggest that oral administration of 1 mg LHRHa/kg is a reliable method of inducing ovulation in spotted seatrout, resulting in predictable spawning within 38 h of feeding. This noninvasive treatment may be particularly useful for induced spawning of teleosts which are susceptible to handling stress.     

Spawning induction and hormonal levels during final oocyte maturation in the silver perch (Bidyanus bidyanus)

The silver perch (Bidyanus bidyanus Mitchell) (Teraponidae), is a native Australian freshwater fish that, due to its high potential for aquaculture, was introduced to Israeli fish farming. The objective of this study was to find an optimal method for inducing spawning of silver perch. The agents tested for this purpose were: human chorionic gonadotropin (hCG; 150 or 200 IU/kg BW); salmon gonadotropin releasing hormone analogue (sGnRHa at 10, 20, 30, or 40 μg/kg BW); mammalian GnRH analogue (mGnRHa at 30 μg/kg) and the combination of sGnRHa at 20 μg/kg and domperidone at 5 mg/kg BW. Based on spawning success and relative fecundity, sGnRHa at the dose of 30 μg/kg was found to be more efficient than hCG, mGnRHa or sGnRHa with domperidone. Since domperidone does not improve the GnRHa effect on final oocyte maturation (FOM) and spawning, it is suggested that the dopaminergic inhibition during the stages of FOM in the silver perch is weak. Therefore, the use of GnRHa alone is sufficient to induce spawning in this fish. Immunoreactive gonadotropin (IR-GtH) and estradiol levels increased after a single injection of sGnRHa, and peaked after 24 h. Plasma levels of 17α,20β-dihydroxy-4-pregnen-3-one (17,20-P) also increased significantly 24 h after the injection of mGnRHa, 12 h before spawning, suggesting that 17,20-P is the maturation-inducing steroid in silver perch. In order to reveal whether the heterologous gonadotropin may elicit an immunological reaction, silver perch was subjected to prolonged treatment with hCG. This treatment resulted in no detectable titer of antibodies against the mammalian gonadotropin. In conclusion, although hCG has no deleterious effects in this fish, and is the more commonly used for spawning induction, sGnRHa at 30 μg/kg is the recommended treatment for spawning induction of female silver perch under the conditions prevailing in Israeli aquaculture.