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.     

Reproductive performance in induced and spontaneous spawning of the mangrove red snapper, Lutjanus argentimaculatus: a potential candidate species for sustainable aquaculture

A reliable breeding technique was developed for the mangrove red snapper, Lutjanus argentimaculatus (Forsskal 1775), to help sustain the aquaculture of this immensely popular species in Southeast Asia. Using standardized indices of female maturity (based on mean oocyte diameter of ≥0.40 mm), time of injection (1000–1130) and sex ratio (one female to two males), a single injection of 100 μg kg^?1 luteinizing hormone‐releasing hormone analogue (LHRHa) (n=16 fish), but not 50 μg kg^?1 (n=five fish), successfully induced egg (62.5% success rate) and larval (43.8%) production. Human chorionic gonadotropin (hCG) at 500 IU kg^?1 (n=five fish) also failed to induce spawning, but doses of 1000 (n=22 fish) and 1500 IU kg^?1 (n=15 fish) gave spawning (77.3% and 80.0% respectively) and hatching success rates (72.7% and 60.0% respectively) that were not significantly different from those of 100 μg kg^?1 LHRHa. No spawning was observed in saline‐injected controls (n=seven fish). While mean spawning latency, egg diameter, egg production per spawn, percent egg viability, hatching rate, percent of normal larvae and cumulative survival of eggs to normal larvae did not differ significantly among the effective hormones and doses, 1000 IU kg^?1 hCG had a higher percentage (76.5%) of total spawns with egg production per spawn in excess of one million than those of 1500 IU kg^?1 hCG (50.0%) and 100 μg kg^?1 LHRHa (40.0%). Mangrove red snapper spontaneously spawned from March–April to November–December with a peak of egg collection and spawning in May–June. Egg collection per spawn ranged from 0.05 to 6.35 million. Spontaneous spawning of mangrove red snapper exhibited lunar periodicity with spawns mostly occurring 3 days before or after the last quarter and new moon phases and occurred consistently between 02:00 and 04:00 hours. High fecundity and good egg quality, coupled with the ability to respond to induce spawning or natural spawning in captivity, provide a sound basis for improving the sustainability of red snapper aquaculture in Southeast Asia.     

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.     

Reproduction of striped bass, Morone saxatilis (Walbaum), broodstock: monitoring maturation and hormonal induction of spawning

Abstract Levels of gonadal steroid hormones were quantified in an adult striped bass, Morone saxatilis (Walbaum), broodstock during their gametogenic cycle. Blood plasma concentrations of Estradiol (E₂) and testosterone in females, or 11-ketotestosterone(11-KT) and testosterone (T) in males, were used as indicators of maturation. In both sexes, hormone levels were low in summer but increased significantly by late October to intermediate levels which were then maintained until late January. They then increased again rapidly to maximum pre-spawning values attained in late February or March, and subsequently decreased during the spawning period (April and May) with an increased incidence of spent fish with low hormone levels. The changes in blood hormone concentrations coincided with annual changes in photoperiod and water temperature that may be useful landmarks for maturation in captive broodstock. Mature females were implanted with pellets containing a dose of approximately 20 μg/kg body weight of [D-Ala⁶-Pro⁹-Net]-LHRH (GnRHa) in a matrix of cholesterol (CH) and cellulose. In April, they had not yet begun final oocyte maturation (FOM) and were too immature for conventional induction of spawning by injection with human chorionic gonadotropin (hCG). In early April, females given two 95% CH (slow hormone-release) GnRHa pellets (95/95) or females given one 80% CH (fast hormone-release) GnRHa pellet and one 95% CH GnRHa pellet (80/95) spawned within 13 days treatment (n= 4) with good egg fertility (76 ± 7% of total) and hatch rates (62 ± 15% of fertile). Females given dual fast-release GnRHa pellets (80/80) or control (Sham) pellets did not spawn or show evidence of increased oocyte diameter or development. In late April, four of six females given the 80/95 GnRHa pellet combination spawned within 9 days. Three fish produced fertile eggs (54 ± 18%). one spawned overripe eggs, and the remaining two increased oocyte diameter and maturation. Three corresponding controls did not spawn, and two of these showed clear signs of atresia within 11 days. In early May, some females were undergoing early FOM and were mature enough to be spawned by hCG injection. Three were given a single 80% CH GnRHa pellet and spawned within 6 days of treatment to produce fertile eggs (44 ± 6%). Of two other females given dual 80% CH GnRHa pellets, one spawned infertile eggs and the other failed to spawn within 9 days. GnRHa implants show promise as a technique for inducing spawning of captive striped bass broodstock although the optimum hormone delivery systems, dosages and release rates should be verified for fish at specific maturational stages.     

Effects of hormonal manipulation on stress responses in male and female broodstocks of pikeperch Sander lucioperca

This study was designed to determine the effects of hormonal manipulation on stress responses in female and male pikeperch. Two-year-old cultured female and male broodstocks with an average weight of 337.4 ± 20.1 (mean ± SE; n = 16) and 318.7 ± 15.1 g (n = 16), respectively, were randomly allocated into four hormonal treatments each containing 4 fish. Two sexual groups of 16 fish for each gender were considered. Sexually mature male and female pikeperch were injected with either physiological saline solution (as control group), common carp pituitary extract (CPE), human chorionic gonadotropin (hCG) and or luteinizing hormone-releasing hormone analog (LHRHa₂). The blood samples were taken before hormonal injection and after ovulation and spermiation. Then the plasma levels of stress indices (cortisol, glucose, and lactate) were determined. The results showed that all CPE-, HCG-, and LHRHa_2- injected males produced sperm. In females treated with CPE and hCG, three of four ovulated, but none of LHRHa_2- and saline-injected fish spawned. Significant changes in cortisol, glucose, and lactate levels were observed among the females injected with different hormones. Plasma cortisol and glucose levels increased significantly in males injected with CPE and females injected with hCG, but no significant change was observed in lactate levels before and after hormonal induction. Comparison of two sexes revealed significant differences in glucose levels for females in some groups before injection, while CPE-injected sexes showed significant changes in cortisol and lactate concentrations. The results indicated that the induction of ovulation or spermiation stimulated stress responses especially in female pikeperch, and therefore, all the procedures should be made to minimize the disturbance during the artificial spawning.     

Induced spawning in bream, Abramis brama (L.), using carp and bream pituitary extract and hCG

Induced spawning in bream, Abramis brama (L), was studied using acetone-dried common carp pituitary (CP) and bream pituitary (BP) with or without the addition of human chorionic gonadotrophin (hCG). The total dose administered to fish was of 5.0 mg kg^?1 BP or 4.0 mg kg^?1 CP with or without 2000-2200 IU hCG kg^?1 for females and 2.5 mg kg^?1 BP or 2.0 mg kg^?1 CP with or without 1000–1100 IU HCG kg^?1 for males. In all male treated groups 100% of spermiation was observed: in females the most effective method was a triple injection with hCG and carp pituitary, resulting in 79% of females ovulated (over 68% of eyed eggs). Biological quality of eggs, expressed as a percentage of eyed eggs, was negatively correlated with time elapsing between resolving (final) injection and ovulation. Spawning success, expressed as a value of S_e (spawning effectiveness coefficient), was higher in fish treated with triple injection.     

Preliminary Studies of Artificial Spawning of Channel Catfish as Male-Female Pairs or All-Female Groups in Recirculating Systems

Abstract— Channel catfish Icralurus punctatus are commonly spawned for research purposes by pairing of a hormonally treated female with a male in flow-through aquaria. A technique that allows hormonal induction of ovulation in females without pairing would accelerate genetic improvement and production of hybrid catfish. Over a 3-yr period (1994, 1995, and 1996) we conducted a series of trials to demonstrate the potential for artificial spawning in recirculating systems, and in 1996 we included trials with grouped females in addition to male-female pairs. Females were induced to spawn with injection of synthetic leuteinizing hormone-releasing hormone, and those that ovulated were stripped and the eggs were artificially fertilized. During 1994 and 1995, all fish were spawned by pairing, and in 1996, half of the females were spawned by pairing and half were grouped in tanks without males. Spawning success (percent of females that produced eggs), latency (time between injection and ovulation), and percent fertilization were observed for the paired and grouped trials. Spawning success was 36% in 1994 (N= 36). 22% in 1995 (N= 54). 41% in 1996 (N= 27). and 58% for grouped females (N= 26). The latency period was 113 ± 69 h in 1994, 109 ± 57 h in 1995, 44 ± 8 h in 1996, and 50 ± 9 h for grouped females. Percent fertilization was 16 ± 26% for eggs stripped in 1994, 72 ± 25% in 1995, 43 ± 20% in 1996, and 16 ± 37% for grouped females. In 1995, water quality problems were associated with high mortality of females (24 of 44 females; 4 of 44 males). The metabolic demands of final oocyte maturation in combination with methemoglobinemia caused by high nitrite levels could account for the increased vulnerability of females. These trials indicate that with adequate biofiltration, artificial spawning is possible in recirculating systems and with females grouped rather than paired. Further research on hormone dosage and timing of egg stripping will increase the utility of grouped spawning of channel catfish.     

Spawning induction of pejerrey Odontesthes bonariensis in captivity using sustained‐release gonadotropin releasing hormone agonist implants

The aim of this study was to induce and synchronize spawning of pejerrey Odontesthes bonariensis (Valenciennes, 1835), using gonadotropin releasing hormone agonist (GnRHa) implants. In the first experiment, the ovarian condition was assessed by ovarian biopsies and the measurement of the genital pore width (GPW). Females having the leading clutch of oocytes with a diameter of around 800–900 μm and a GPW between 4.5 and 5.5 mm were treated with GnRHa implants. Eighty per cent of females spawned between 2 and 9 days after treatment, 12 days earlier than 20% of the fish in the control group that presented signs of spawning activity. In order to avoid any possible ovarian injury and/or stress by the catheterization procedure, in a second experiment, females were selected only by visual inspection of the abdomen and GPW measurement. As in experiment 1, 80% of females spawned between 2 and 8 days after treatment, 8 days earlier than 30% of the fish that spawned in the control group. In both experiments, fertilization and hatching success were similar between control and GnRHa‐treated groups. These results clearly demonstrated that GnRHa implantation can advance and synchronize ovulation and spawning in pejerrey without affecting egg quality.     

Changes of plasma steroid concentrations associated with spontaneous or induced ovulation in yellow perch Perca flavescens

This study examined the changes in plasma steroids during natural (Experiment 1) and induced (Experiment 2) final maturation in yellow perch Perca flavescens. In experiment 1, ovulating yellow perch were stripped of eggs and blood samples collected to determine the concentrations of testosterone (T), estradiol-17β (E2), and 17,20β-dihydroxy-4-pregnen-3-one (17,20βP). Eggs from individual females were weighed and fertilized. Fertilization rate was determined at the embryo eyed stage. In experiment 2, females were randomly assigned to one of the following treatment groups: (1) saline (0.7% NaCl), (2) des-Gly¹⁰[D-Ala⁶] LHRH-ethylamide (100 μg LHRHa/kg), and (3) LHRHa plus 17,20βP (100 μg LHRHa/kg + 2 mg 17,20βP/kg). Fish were injected intraperitoneally with two doses at a two-day interval. Blood was collected prior to injections and at the time of ovulation/spawning and concentrations of T, E2, and 17,20βP (free and conjugated) were determined. In experiment 1, low concentrations of 17,20βP were recorded at spawning. In experiment 2, all surviving fish injected with LHRHa (5 of 5) released their eggs spontaneously during the week following injections. None of the surviving control fish (0 of 5) ovulated during this period, whereas only 1 of 3 surviving fish injected with LHRHa + 17,20βP released eggs. In the control group, concentrations of E2 and 17,20βP did not show significant differences over the experimental period, whereas plasma T concentrations increased significantly. In fish injected with LHRHa, the concentrations of T and 17,20βP increased significantly after the first injection but then declined at ovulation/spawning. It also appears that 17,20βP was conjugated to its sulfated form. Mortality reached 62.5% in the group injected with LHRHa + 17,20βP indicating that this treatment was severe. Thus, LHRHa alone appears highly effective in inducing ovulation in yellow perch. This revised version was published online in July 2006 with corrections to the Cover Date.     

Hormone-induced ovulation and spawning of captive and wild broodfish of the catadromous Australian bass, Macquaria novemaculeata (Steindachner), (Percichthyidae)

Australian bass, Macquaria novemacideata (Steindachner),mature but do not spawn in fresh or brackish water ponds. Ovulation and spawning of captive (n = 158)and wild Australian bass (n = 123) was induced in the normal breeding season by single injections of human chorionic gonadotropin (hCG). Doses of 100-4000IU kg-“ hCG induced ovulation and the optimum dosage was 500 IU kg” hCG. The breeding season was from mid-May to late August for wild fish,and extended into September for captive fish. There was a tendency for mean fertilization and hatching success to decline over the breeding season. Greater fertilization and hatching success was obtained from fish which spawned naturally than from stripped fish. Fish spawned after 34,2 ± 0.4 h (mean ± SE, n = 74). Ovulating fish that failed to spawn were stripped after 40.2 ± 0.3 h (n = 76). The timing of stripping and fertilization was an important factor determining hatching success. There was no apparent difference in latency periods or the number of eggs spawned between captive and wild fish. However, the mean number of eggs obtained from naturally spawned fish was higher than for stripped fish. The techniques described in this paper will assist the largescale production of Australian bass by increasing the quality and quantity of larvae from hCG-induced spawnings.     

The Effect of Luteinizing Hormone Releasing Hormone Analog Regime and Stage of Oocyte Maturity for Induced Ovulation of Channel Catfish,Ictalurus punctatus

This study addresses rapid methods to identify mature channel catfish, Ictalurus punctatus, females for induced spawning with luteinizing hormone releasing hormone analog (LHRHa) and common carp pituitary extract (CPE) and the effects of stage of maturity, hormone dose, season, and cannulation before hormone injection. Hormonal intervention is the most effective method for inducing maturation and spawning in channel catfish × blue catfish, Ictalurus furcatus, hybrids. A total of 80 mature channel catfish were staged for maturity based on secondary sexual characteristics and 20–30 oocytes cannulated to ascertain their maturation stage based on the position of gonadal vesicle. Twenty females were randomly assigned to one of the four hormone regimes (priming + resolving dose): CPE 2 + 8 mg/kg (CPEC); LHRHa 20 + 40 µg/kg (LHRHaL); LHRHa 20 + 60 µg/kg (LHRHaM); and LHRHa 20 + 80 µg/kg (LHRHaH) and a fifth treatment consisted of 20 females selected based on apparent maturity as evidenced by external appearance were injected with CPE, 2 + 8 mg/kg without cannulation (CPEO). Two trials were conducted in early part of the spawning season and two trials in peak season. External methods to identify maturity correlated (r = 0.63) with that of “germinal vesicle position” method to identify the stage of maturity in females. Mean percent of females that ovulated, hatched, and fry produced per kg body weight did not differ (P > 0.05) among the five hormone treatments. The mean percent females that ovulated was higher (P < 0.05) for CPE‐induced females that were identified as “excellent” females based on external examination. The performance of cannulated females did not differ (P > 0.05) with that of non‐cannulated catfish. The mean egg quality of hormone‐induced females and percent of females ovulated in response to hormone treatment established a weak but significant linear relationship (Y = 3.85 + 0.008 × ovulation, r² = 0.39, P < 0.05).     

Induced maturation and spawning of domestic and wild striped bass, Morone saxatilis (Walbaum), broodstock with implanted GnRH analogue and injected hCG

Abstract. A domestic striped bass. Morone saxatilis (Walbaum), broodstock was established by rearing fish to sexual maturity in ponds. A method was developed to reproduce the domestic females, and also wild females too immature to be successfully induced to spawn with injected human chorionic gonadotropin (hCG). The fish were implanted with pellets containing 100–150μg of a synthetic analogue of mammalian gonadotropin reieasing-hormone, [D-Ala⁶- Pro⁹-NEt]-LHRH (mGnRHa), in a matrix of cholesterol (CH) and cellulose. They were implanted with one fast hormone-release (80% CH) pellet and one slow hormone-release (95% CH) pellet and allowed to mature for 1–3 days, until they entered the process of final oocyte maturation and were induced to ovulate or spawn with an hCG injection. The secondary hCG injection was found to be necessary to speed the maturation of the wild fish; they otherwise would succumb to the stresses of capture, handling and confinement before they could be spawned. The total mGnRHa dosages used ranged from 33 to 111μg mGnRHa/kg body weight, and the hCG doses were either 165 or 330 IU hCG/kg body weight. Using the combined mGnRHa implant-hCG injection technique, fry production rates were comparable to those obtained using fully mature wild females taken directly from their spawning grounds.     

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.     

Improvement of ovulation induction by additive injection of 17,20β‐Dihydroxy‐4‐pregnen‐3‐one after human chorionic gonadotropin administration in a pelagic egg spawning marine teleost,nibe croaker Nibea mitsukurii (Jordan & Snyder)

This study aimed to develop the consistent ovulation induction method in a pelagic egg spawning marine teleost, nibe croaker Nibea mitsukurii. Attempts to induce oocyte maturation and ovulation in nibe croaker using human chorionic gonadotropin (hCG; 0.5 IU g^?1) resulted in the normal progression of oocyte maturation and hydration, but a failure to induce ovulation in many individuals. This ovulation disorder was similarly observed even when the dose of hCG was increased 10 times (5 IU g^?1) or decreased to one tenth (0.05 IU g^?1), indicating that it cannot be completely overcome solely by hCG administration. However, this ovulation disorder could be completely overcome by subsequent administration of 17,20β‐dihydroxy‐4‐pregnen‐3‐one (DHP) at the appropriate dose (0.5 μg g^?1) and time (20 h after hCG administration). An increase in the number of individuals that ovulated due to DHP administration led to an increase in individuals producing larvae, resulting in an approximately threefold increase in the estimated number of larvae produced compared with the group of fish administered hCG alone. Thus, this ovulation induction method using DHP administration after hCG was demonstrated to overcome the ovulation disorder in nibe croaker and could be applicable to commercially important species with similar ovulation problems.     

Induced spawning in perch, Perca fluviatilis L. using carp pituitary extract and HCG

Induced spawning in Eurasian perch, Perca fluviatilis L., was studied using human chorionic gonadotrophs (hCG) or acetone-dried common carp pituitary with or without hCG. The dose administered to fish was 5700 IU hCG kg^-1 or 4.0 mg kg^-1 carp pituitary with or without 500-700 IU hCG kg^-1 for females and 2850 IU hCG kg^-1 or 2.0 mg kg^-1 carp pituitary with or without 250-350 IU hCG kg^-1 for males. There were no statistically significant differences in quantity of milt in treated and control groups, although the best result was observed when males were treated with a triple injection of hCG and carp pituitary extract. Male spermiating success, expressed as a quantity of milt, was negatively correlated with fish weight. All females treated in this experiment had oocytes at the same division, so time of ovulation was highly synchronous. Spawning success, expressed as a spawning effectivity coefficient (S_c), was highest in fish treated with the triple injection. Spawning methods described in this paper were successful, even though hormones from another order of fish were used.     

Induced ovulation using LHRHa in anadromous obscure puffer Takifugu obscurus cultured entirely in freshwater

Puffer fishes of the order Tetradontidae exist in both anadromous and non-anadromous seawater resident forms. The obscure puffer akifugu obscurus is an anadromous species whose intensive culture in freshwater has developed rapidly in China. To mass produce larvae for intensive culture, induction of ovulation of 3-year-old obscure puffer cultured entirely in freshwater was attempted by giving the fish multiple injections of LHRHa. Twenty 3-yearold cultured obscure puffers were treated with multiple injections of LHRHa at a constant dosage of 30 g of LHRHa kg-1 of body weight with 36 h interval between injections. Beginning two days after hormonal treatment, abdominal palpation was performed every day to check expansion and hardening of the abdomen of the fish due to hydration of oocytes that indicates the completion of final oocyte maturation. The first four fish ovulated after the second LHRHa injection on day 3, but most females ovulated after the fourth hormone injection. A total of 18 fish (90%) ovulated over a period of 7 days, while no fish ovulated in the control group. The mean fertilization rate and the mean hatch rate was 67.1% and 87.6%, respectively. The viability of newly hatched larvae was very good, with a high survival rate of 98.6% 24 h post-hatch. These results indicate that cultured obscure puffer, which are not allowed to undergo diadromous migration and are entirely cultured in freshwater, could be induced to complete maturation and ovulate by simple multiple injections of LHRHa and that the eggs could be successfully hatched into viable larvae.     

Early out-of-season induced spawning of channel catfish Ictalurus punctatus (Rafinesque) conditioned in heated earthen ponds

This study documents early out-of-season induced spawning of channel catfish Ictalurus punctatus. During the early spring (February–April) of 1999, 2000 and 2001, ponds containing (1) male and female channel catfish (mixed-sex ponds) or (2) male channel and blue catfish I. furcatus only, or female channel catfish only (single-sex ponds) were heated to 24–30°C to encourage gonadal maturation and spawning. Unheated ponds were stocked with males and females and were monitored during the duration of heating. When natural spawning occurred in the heated ponds, the fish were captured by seining and unspawned females were injected with 100 μg kg⁻¹ of synthetic leutenizing hormone-releasing hormone. Injected females were either paired with males or held in communal all-female groups, and monitored for ovulation. Eggs were collected and fertilized with sperm of channel catfish or blue catfish. Females paired with males were induced to spawn 44 days (mixed-sex ponds) and 50 days (single-sex ponds) before natural spawning occurred in unheated ponds. Spawning latency (the time between injection and ovulation) and the percentage of neurulated embryos from eggs fertilized using channel catfish sperm was not different between spawning before the natural season (P=0.68) and during the natural season in fish from mixed-sex ponds (P=0.57). Females held in all-female groups produced eggs 34 days before the onset of spawning in unheated ponds. Spawning latency was not different between spawns before and during the natural season (P=0.16), and the percentages of neurulated embryos from eggs fertilized with channel catfish sperm (P=0.76) or blue catfish sperm (P=0.77) before or during the natural season were not different. This study demonstrates the feasibility of conditioning of channel catfish females for early out-of-season induced spawning in the laboratory.     

Spawning frequency of the Tsushima Current subpopulation of chub mackerel <Emphasis Type="Italic">Scomber japonicus</Emphasis> off Kyushu,Japan

Female Japanese chub mackerel Scomber japonicus of the Tsushima Current subpopulation were collected during the spawning season from March to May 2001. A total of 137 adult females were caught between midnight and daybreak. A considerable number of fish displayed new postovulatory follicles (POF), whereas there was no evidence of germinal vesicle breakdown or hydrated oocytes in any of the fish collected. This suggests a daily spawning synchronicity toward midnight. To estimate the spawning frequency (S), the female reproductive state was classified into four criteria based on the degenerative stage of the POFs and the developmental stage of the oocytes. To stage the POFs according to age and determine the stage duration, ovaries from S. japonicus were induced to spawn in the laboratory and were sampled 0–72 h after ovulation at appropriate intervals. The average S, which is evaluated from four different indices, was 16.9%, corresponding to the average female chub mackerel spawned every 5.9 days (8.5 times) during the 50 days.     

Induction of Out-of-Season Spawning of Pikeperch, Sander Lucioperca (L.)

Pikeperch were induced to spawn 3 months prior to the natural spawning period through photothermal and hormonal stimulation. Females (five specimens in each group) were stimulated with injection of human chorionic gonadotropin (hCG) once (200 IU kg^–1), twice (200 IU kg^–1, second dose after 48 h–400 IU kg^–1) or three times (200 IU kg^–1, after 24 h–200 IU kg^–1 and after another 24 h–200 IU kg^–1). The control group was injected once with 0.9% NaCl. The males were stimulated with a single hormone dose of 200 IU kg^–1. Eggs were obtained from all the hormonally treated fish. None of the control group females, which were only stimulated photothermally, ovulated any eggs. The time of ovulation was 66–71 h following the first injection, and the eggs viability until the eyed stage (from 71.5 to 77.5%) did not depend on the number of hormone doses (P > 0.05). The out-of-season spawning method described in this paper could be used to provide pikeperch larvae for intensive culture systems (recirculating water systems) before natural spawning season and to produce larger-sized pikeperch fingerlings for stocking.     

Induced final maturation and ovulation in a small anabantoid teleost, the dwarf gourami, Colisa lalia. I. The effects of gonadotrophic preparations, LHRHa, steroids and thyroid hormones

Colisa lalia (Ham.) has been used as a model for the development of techniques for induced spawning that are applicable to small teleosts where ovulation requires prolonged exposure to suitable breeding conditions.Human chorionic gonadotrophin (hCG; 4 IU per fish) induced ovulation within 24 hours, whereas homologous pituitary extracts were relatively ineffective. When administered in saline ( 20 g per fish per injection), des-Gly10,[D-Ala6]- LHRH N-ethylamide (LHRHa) was ineffective, but it stimulated ovulation in a proportion of fish when administered ( 1.6 g per fish) as an emulsion in Freund's incomplete adjuvant (FIA). Together, these results suggest that ovulation requires the synthesis as well as the secretion of gonadotrophin in C. lalia.Long-term treatment with thyroid hormones appeared to enhance the ovulatory response to LHRHa in FIA, possibly by effects on the ovary; whereas the various steroids tested were ineffective at the dosages used