Fisheries proceedings iii

Comparison of the effects of buserelin in combination with various forms of domperidone and pituitary gland on the induction of spawning and gonadal development in the Thai carp, Barbonymus gonionotus Naruepon Sukumasavin*, Pongsai Chansri and Mali Lamanthieng Inland Fisheries Research and Development Bureau, Thailand and Aquaculture of Indigenous Mekong Fish Species Component, MRC Fisheries Programme This study set compares the effects of buserelin (BUS) in combination with various forms of domperidone (DOM) to the effects of extract of pituitary gland (PG) on the induction of spawning and gonadal development in the Thai carp, Barbonymus gonionotus. The results indicated that injection of 10 mg/kg BUS in combination with 10 mg/kg DOM (in the form of the human preparation, Motilium) dissolved in distilled water, dimethylsulfoxide (DMSO) or N,N-dimethylformamide (DMFM) had similar effects on the number of fish spawned, fertilization, hatching and survival rates of seven-day-old fry (p>0.05) to those seen after the injection of one dose of pituitary gland. Repeated treatments with BUS in combination with all forms of DOM used in this study had no negative effect on gonad development in females, which was completed within three weeks of each spawning. Fish were induced to spawn at least three times during the season and the same was true of fish that were induced to spawn by repeated use of pituitary gland. KEY WORDS: Barbonymus gonionotus, Puntius gonionotus, induced spawning, buserelin,
domperidone
In most teleost fish, including the Thai carp (Barbonymus gonionotus), a surge of gonadotropin (GtH II) secreted by the pituitary initiates the final maturation of gonads and ovulation, and triggers the onset of spawning. The hypothalamus regulates this surge through the interaction of two other hormones, gonadotropin-releasing hormone (GnRH) and dopamine (an agent that inhibits the release of GtH). Fish reared artificially, will spawn if dosed with GnRH, or its one of its analogues (GnRHa), in combination with a dopamine antagonist (DA) (Peter et al. 1988). In Thailand, this proved a very effective, and reliable, way of inducing several freshwater fish species to spawn (Sukumasavin and Leelapatra 1988). Sukumasavin and Leelapatra (1993) also found that buserelin (BUS), a mammalian luteinising hormone, is the most effective of the several forms of GnRHa. To date, domperidone (DOM) is the only DA used in spawning experiments. DOM is insoluble in water but dissolves in organic solvents such as DMSO (Gissis et al. 1991) and DMFM (Gissis et al. 1991). It is also partly soluble in propylene glycol (PROP). However, the most common practice is to use Motilium suspension (MOT), a DOM preparation designed for use in humans. However, the effect of repeated use of a BUS-DOM combination on the development the Thai carp’s gonads is unknown. In this study, therefore, we set out to investigate the consequences of using BUS-DOM (in various * Inland Fisheries Research and Development Bureau, DoF, Chatuchak, Bangkok, Thailand, 10900. Email: naruepos@fisheries.go.th Comparison of the effects of buserelin in combination with various forms of domperidone and pituitary gland preparations), and extract of pituitary gland (PG), over several spawning cycles. The study was carried out at Surin Inland Fisheries Station from February to September 2001. Thai carp were reared in earthen ponds at a stocking density of 1 kg/4 m2 and fed once daily with pellets containing 30% protein at 1.5% body weight. The fish used in this experiment were ripe to spawn, i.e. females with 80% of their oocytes at the germinal vesicle migration stage and males that produced milt when gentle pressure is applied to the abdomen. BUS: Buserelin (Hoechst) was diluted in distilled water at a concentration of 100µg/ml. DOM: Two types of DOM were used i.e. the pure chemical (Sigma) and the human drug, MOT (Jenssen Pharmaceutica). Pure DOM was dissolved in: DMSO (Sigma) at a concentration of 25 mg/ml (DMSO-DOM) DMFM (Sigma) at a concentration of 25 mg/ml (DMFM-DOM) PROP (Sigma) at a concentration of 25 mg/ml (PROP-DOM) MOT: MOT was ground and dissolved in distilled water at a concentration of 25 mg/ml (MOT-DOM). PG: PG, collected from mature common carp (average body weight 500 g) about one hour prior to injection was homogenised and mixed with distilled water. The injection volume was adjusted to 0.5 ml/ kg of fish. The dosage was determined using the following formula: 1 dose = weight of donor/weight of recipient Experiment 1: Effects of various forms of DOM in combination with BUS on inducing Thai carp to Sexually mature female Thai carp weighing an average of 300 g were divided into five groups of 80 fish each. Fish in each group were identified using spaghetti tags. Each group received an intra-peritoneal Proceedings of the 6th Technical Symposium on Mekong Fisheries, Pakse, Lao PDR 26-28 November 2003 Comparison of the effects of buserelin in combination with various forms of domperidone and pituitary gland Group 2: 10µg/kg BUS in combination with 10 mg/kg MOT-DOM Group 3: 10µg/kg BUS in combination with 10 mg/kg DMSO-DOM Group 4: 10µg/kg BUS in combination with 10 mg/kg DMFM-DOM Group 5: 10µg/kg BUS in combination with 10 mg/kg PROP-DOM Following this treatment, the groups of female fish were transferred to 10 m3 cement tanks containing the same number of untreated males. The fish then spawned naturally. The resultant eggs incubated in hatching jars and the rates of fertilisation and hatching were counted. Newly hatched larvae were moved to cement tanks. The following data were collected: number of fish that spawning during the 8 hr induction period survival rate of larvae at seven days old Experiment 2: Effects of various forms of DOM in combination with BUS on gonadal development in The fish resulting from the spawning during Experiment 1 were reared together in a 400 m2 earthen pond. Every day each fish was fed with 30% protein pellets equivalent to 1.5% its body weight. On the first day, five fish were taken from each group; these were weighed, killed and their gonads removed. This procedure was repeated every seven days. The gonadosomatic index (GSI = gonad weight*100/ body weight) was calculated for each fish. After four weeks, the surviving fish were used to repeat Experiment 1. After the fish had spawned, Experiment 2 was repeated for another cycle. Experiment 1: Effects of various forms of DOM in combination with BUS on the induction of spawning of the Thai carp (Appendix I, Tables 1, 2 and 3). The form of DOM is not a critical factor in determining the number of fish that spawn (Table 1). The difference in the number of fish that spawned following injections of PG, of DOM in the form of MOT, or various preparations of DOM, is not statistically significant. This is true for all three induction-cycles. In each cycle, latency periods were all between 4.15 - 6.15 hours and there is no significant difference Proceedings of the 6th Technical Symposium on Mekong Fisheries, Pakse, Lao PDR 26-28 November 2003 Comparison of the effects of buserelin in combination with various forms of domperidone and pituitary gland (p>0.05) in the rates of fertilisation and hatching, and the proportion of larvae that survived to seven Experiment 2: Effects of various forms of DOM in combination with BUS on gonad development in the Within spawning cycles, the GCI varied little between groups of fish that had been injected with PG, different preparations of DOM and DOM in the form of MOT. At the time of injection, the index varied within the range 17.02±0.42% (MOT-DOM) and 21.13±3.99% (PG). In all instances the index fell sharply after spawning (to within the range 5.56±0.68% (PG) to 7.8±1.8% (DMSO-DOM)); thereafter it increased until, by week three, the index returned to the levels recorded at the time of the first injection. This pattern repeated during the second and third spawning cycles. Overall, there was no statistically significant variation in the levels of the GSI within, or between, induction-cycles. The results of this study show that BUS in combination with DOM is as efficient in inducing spawning in the Thai carp as PG (c.f. Sukumasavin and Leelapatra, 1988). Furthermore, BUS in combination with DOM dissolved in DMSO, DMFM or PROP did not affect the number of fish that spawned, fertilisation rate, hatching rate or survival rate of seven-day-old larvae when compared with PG or DOM in the form of the human preparation, MOT (Sukumasavin 1994). Sirikul (1987) demonstrated that Thai carp induced to spawn with PG would, if induced again, spawn for a second time one month later and three or more times in a spawning season. In this study, Thai carp induced to spawn with BUS in combination with DOM, if induced again, also spawned at least three times in the spawning season at intervals of one month. This study uses GSI as an indicator of gonadal development because in the Thai carp GSI is highly correlated with mature oocytes (r=0.9, p<0.01) (Sukumasavin and Leelapatra 1994). GSI has been used as an indicator for gonadal development in several cyprinids, including goldfish (Clemens and Reed 1967; Munkittrick and Leatherland 1984). The peak GSI at the beginning of the study was between 17% and 21%, similar to that of fish reared in earthen ponds reported by Sukumasavin (1992). Furthermore, three weeks after spawning the GSI of fish treated with all forms of DOM in combination with BUS were not significantly different. This indicates that the gonadal development in the Thai carp can complete within three weeks under culture conditions. In addition, there were no significant differences in GSI between the two reproductive cycles (p>0.05). The two sets experiments demonstrate repeated use of DOM in combination with BUS has no negative effect on the induction of spawning or gonadal Table 5 (Appendix I) gives the costs of using PG and BUS in all forms of DOM to induce spawning of 1 kg Thai carp. PG is the most expensive at 10 baht/kg. BUS and MOT-DOM cost 4.65 baht/kg while the costs for DMSO-DOM, DMFM-DOM or PROP-DOM are lower. PROP-DOM is the cheapest overall at Proceedings of the 6th Technical Symposium on Mekong Fisheries, Pakse, Lao PDR 26-28 November 2003 Comparison of the effects of buserelin in combination with various forms of domperidone and pituitary gland 2.27 baht/kg. These results show that PROP-DOM is the most cost effective method of inducing the Clemens, H.P. and Reed, C.A. (1967) Long term gonadal growth and maturation of goldfish (Carassius auratus) with pituitary injections. Copeia. pp. 465-466. Gissis, A., Levavi-Sivan, B., Rubin-Kedem, H., Ofir, M. and Yaron, Z. (1991) The effect of gonadotropin releasing hormone superactive analog and dopamine antagonists on gonadotropin level and ovulation in tilapia hybrids. Israeli J. Aquacult. Bamidgeh. 43(4), 123-136.
Munkittrick, K.R. and Letherland, J.F. (1984) Seasonal changes in the pituitary-gonad axis of feral goldfish, Carassius auratus L., from Ontario, Canada. J. Fish Biol. 24, 75-90.
Peter, R.E., Chang, J.P., Narhorniak C.S., Omeljaniuk, R.S., Solowska, M., Shis, S.H. and Billard, R. (1986) Interactions of catecholamines and GnRH in regulation of gonadotropin secretion in teleost fish. Recent Prog. Horm. Res. 42, 513-548.
Peter, R.E., Lin, H.R. and Van Der Kraak, G. (1988) Induced ovulation and spawning of cultured freshwater fish in China: advances in application of GnRH analogues and dompamine antagonist. Aquaculture 74, 1-10.
Sukumasavin, N. (1994) Effect of using high dosages of Buserelin on the induction of spawning in the Thai carp, Puntius gonionotus Bleeker. Thai Fish. Gaz. 47(5), 415-419.
Sukumasavin, N. (1992) Aspects of the reproductive endocrinology of the Thai carp, Puntius gonionotus Bleeker. Department of Zoology, University of British Columbia, Vancouver, B.C., Sukumasavin, N. and Leelapatra, W. (1988) The application of gonadotropin releasing hormone analogue and domperidone for fish seed production in Northeast Thailand. Northeast Fisheries Project Sukumasavin, N. and Leelapatra, W. (1993) Comparison on the biological activities of gonadotropin releasing hormone and its analogs in combination with domperidone on the induction of gonadotropin secretion and spawning in the Thai carp, Puntius gonionotus (Bleeker). Thai Fish. Gaz. 46(6), 511-518.
Sukumasavin, N. and Leelapatra, W., (1994) Annual reproductive cycle of the female Thai carp, Puntius gonionotus (Bleeker). Thai Fish. Gaz. 47(1), 21-30.
Sirikul, C., Lanamttieng, M., Munsiri P., Sonkogsoong, P. and Tonetoh, K., (1987) Ability to spawn of the Thai carp in one year. Proceedings of the Technical Symposium 1987. Department of Fisheries. pp Proceedings of the 6th Technical Symposium on Mekong Fisheries, Pakse, Lao PDR 26-28 November 2003 Comparison of the effects of buserelin in combination with various forms of domperidone and pituitary gland rvival rat
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Proceedings of the 6th Technical Symposium on Mekong Fisheries, Pakse, Lao PDR 26-28 November 2003 Comparison of the effects of buserelin in combination with various forms of domperidone and pituitary gland Proceedings of the 6th Technical Symposium on Mekong Fisheries, Pakse, Lao PDR 26-28 November 2003

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