Dariusz K u char c z y k1, Roman K uj a w a l, Andrzej M a m car z 1, Elzbieta W y s z o m i r s k a 2

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1 POLSKIE ARCHIWUM HYDROB!OLOGII (Pol. Arch. Hydrobiol.) Dariusz K u char c z y k1, Roman K uj a w a l, Andrzej M a m car z 1, Elzbieta W y s z o m i r s k a 2 Induced spawning in rudd (Scardinius erythrophthalmus L.) 1 Department of Fisheries, Olsztyn University of Agriculture and Technology, Oczapowskiego 5, Olsztyn-Kortowo, Poland 2 Department of Chemical Analysis, Drug Institute, 30/34 Chelmska St., Warsaw, Poland Abstract The effect of administration different kind of hormones to induced ovulation and spermiation in rudd (Scardinius e!ytlzroplztlwlmus) was studied. We tested acetone dried common carp pituitary with or without HCG at total dose 4 mg kg- t, HCG at total dose 5000 IU kg- 1 and FSH+LH at total both dose 75 IU kg - 1 for with 10 mg kg- 1 of metoclopramide or pimozide for females and acetone dried common carp pituitary with or without HCG at total dose 2 mg kg-\ HCG at total dose 2500 IU kg- 1 and FSH+LH at total both dose 25 IU kg- 1 for with 10 mg kg- 1 of mctoclopramide or pimozidc for males. There is no statistically significant differences of quantity of milt in treated and control groups, although the best result was observed when males were treated with HCG and carp pituitary extract. All females treated in this experiment had oocytes in the same division, so time of ovulation was very synchronous. Spawning success, expressed as a value of Se (spawning effectiveness coefficient) was higher in fish treated with HCG + carp pituitary extract and FSH+LH with pimozide. Key words: rudd, Scardinius erythrophthalmus, artificial spawning 1. Introduction One of the main problems in aquaculture is obtaining mature gametes. Most of fish species from cultured and wild stocks arc incapable of spontaneous maturation under artificial conditions (A hma d Taju ddi n et al. 1986; Glu bokov et al. 1991) or there arc problems of synchrony in time of spawning (D ab rows k i et al. 1994; K u char c z y k et al. 1996), so it is a need of hormonal stimulation for gametes maturation. In this paper we present a method of artificial spawning of wild sexually mature S. erytlzroplztlwlmus using *Proceedings of the international conference "Fish Reproduction '96", 9-12 September 1996, Ccske Budejovicc, Czech Republic. - -

2 210 D. Kucharczyk et al. HCG with or without carp pttujtary extract, carp pttuttary extract and FSH+LH with two dopamine antagonists: pimozide and metoclopramide. 2. Material and methods S. e1ytltrophthalmus broodstock was caught in Kortowskie Lake (Olsztyn, Poland) in June Fish were selected according to the following criteria: the belly of females had to be fully distended and bulging, soft and resilient to touch; males had to be spermiating. The selected males and females were kept in separate 1000 dm 3 tanks in a hatchery with controlled temperature and photoperiod. After one day of acclimation, the water temperature was raised from 17 to 20 C and kept oscillating between 20 and 21 o C throughout the experiment; the light was similar to natural photoperiod (18 h light, 6 h dark). Five days later the fish were individually marked and oocytes were taken from females using the method described by K u jaw a, K u char c z y k (1996). Oocytes were sampled in vivo and placed in Serra's solution for clarification of the cytoplasm. After 5 minutes the position of oocyte nucleus was determined using a 4-division scale (B r z us k a, B i en i a r z 1977). For further experiment we used only those females which oocyte maturation was between 2-3 and 3 division. Before manipulations fish were anaesthetised with 2-phenoxyethanol (0.2 mg dm- 3 ). Then the fish were treated with hormonal injections (Table I). Hormones and drugs were dissolved in 0.9% NaCl (G 1 u b o k o v et al. 1991; Dab rows k i et al. 1994). Eggs were stripped from the females and were fertilized using "dry" method with sperm pooled from three males from the respectively treated group. Two replicates of eggs per trial were incubated in Petri dishes, at a temperature oscillating between 19 and 21 o C. Eggs were incubated up to the eyed egg stage (embryos eyes were pigmented). For showing effectiveness of spawning success in each group we used spawning effectiveness coefficient (S,) (K u char c z y k et al. 1996), which is describing by the formula: Se= 0, I, where: Os -ovulating success (No. of ovulating females/no. of total females), Is- incubation success (No. of live embryos/no. of total embryos). Statistical differences between groups (spermiation success and incubation success) were analysed with Duncan's multiple range test (P < 0.0 5) (PI at t 1977). 3. Results and discussion All S. erythrophthalmus males were spermiating when transported to the hatchery, and hormonal treatment did not cause a statistically significant change in quantity of milt obtained from males in treated and control groups (Table I). Such situation was differ than observed for many fish species, such as Cyprinus earpio (T ak a s h i m a et al. 1984), Aristiehthys nobilis, Hypophthalmiehthys molitrix, Tinea tinea (K o z I o w ski 1994) or Abramis brama (K u char c z y k et al. 1996). From cyprinids, similar situation as in case of S. erythrophthalmus was observed in Gobio gobio, where injection in males from LHRH with addition of pimozide did not provide to obtained more milt than from "control" males (Kestemont 1989).

3 Table I. Effectiveness of hormonal treatment applied to induce artificial spawning in rudd (Scardinius erythrophthalmus L.) (j: S.E.). In each group No. of males 4; No. of females 6. Data marked with the same capital letters did not differ significantly (Duncan's multiple range test, P > 0.05) 5' "' "0 "' :;;: 2. ::J Ot> ::J c: 8: N Survival to Quantity of mil Ovulation Latency eyed egg stage Group Resolving Sex Hormone Priming dose dose (hours after (% of (cm 3 kg- 1 ) resolving (%) females) injection) 1 Male 0.9%NaCI j: 0.20 A (control) Female 0.9%NaCI Male HCG IUkg j: 0.16 A Female HCG - SOOOIUkg j: 1.05 A 3 Male CPE - 2.0mgkg j: 0.11 A Female HCG 250IUkg j:Q j: 0.88 c CPE 0.4mgkg mgkg- 1 4 Male FSH+LH IUkf j: 0.05 A metoclopramide - 10mg kg- Female FSH+LH IUk{ j: j: 0.81 AB metoclopramide - 10mgkg- 5 Male FSH+LH IUk{ 3.8 j: A pimozide - 10 mg kg j: j: c Female FSH+LH IUkg pimozide - 10 mgkg- 1 6 Male CPE 0.4 mgkg mg kg ± 0.09 A Female CPE mgkg j: j: 0.47 B '

4 212 D. Kucharczyk et al. The biological quality of eggs, expressed as a survival of the embryos up to the eyed egg stage were highest in groups: 3 (triple injection from HCG and CPE) and 5 (injection from FSH+LH with addition of pimozide). The lowest survival and lowest percentage of ovulated females was noted in group 2 (injection from HCG) (Table I). 0,3 0,25 0,2 Jl 0,15 0,1 0,05,-- 2,,. 3 4 Group r-,. 5 6 Fig. 1. Spawning success (Se) of rudd (Scardinius erythrophtltalmus L.) in control (1) and treated (2{)) groups (hormonal treatment described in Table I) In all treated groups ovulation was synchronous. The shortest latency was observed in groups, in which fish were injected from CPE (with or without HCG), what was similar to situation observed for C. carpio (D r o r i et al. 1994),A. brama (Kucharczyk et a!. 1996) and Perca fluviatilis (K u char c z y k et al. 1997). Y a r on (1995) suggested that latency was always shorter in fish treated with pituitary extract than in other hormones or drugs. The spawning effectivenes coefficient (Se), combining results of ovulation and incubation success, showed that the best hormonal treatments from this experiment were triple injection from HCG and CPE (group 3) and injection from FSH+LH with pimozide (group 5) (Fig. 1). Acknowledgments This study was supported by the Project No financed by Olsztyn University of Agriculture & Technology, Olsztyn, Poland. 4. References A h m a d T a j u d d i n, A.Z., C h u a h, H.P., Ha r on, A., Z a ha r i, A Induced breeding techniques for some of the cyprinid foodfishes in Malacca. In: Proc. Jnt. Conf DeFelopment Management of Tropical Lil'ing Aquatic Resources. Serdang, Malaysia. 2-5 Aug., 1983, B r z us k a, E., B i c n i a r z, K Metoda przyzyciowego okreslania dojrzalosci plciowej samic karpia w zwiqzku z iniekcjami homogenatu przysadki m6zgowej kapia [A method of in Fivo designating oocytes maturation of carp females with connection of injections with common carp pituitary extract]. Broszura lnstytutu Rybactwa Sr6dlqdowego, 105, 1-27.

5 Induced spawning in rudd 213 D 1\ brows k i, K., Ci eres zko, A., Ram se ye r, L., Cu 1 v er, D., Ke stem on t, P Effects of hormonal treatment on induced spermation and ovulation in the yellow perch (Perca flavescens). Aquaculture, 120, Drori, S., Ofir, M., Levavi-Sivan, B., Ya ron, Z Spawning induction in common carp (Cyprinus carpio) using pituitary extract or GnRH superactive analogue combined with metoclopramidae: analysis of hormone profile, progress of oocyte maturation and dependence on temperature. Aquaculture, 119, Gl ubokov, A.I., Motl och, N.N., Sedova, M.A Effect of a synthetic LHRH analogue and dopamine antagonists on the maturation of bream, Abramis brama L. Aquaculture, 95, Ke stem on t, P Hormonal induction of the ovulation and spermiation of the gudgeon Gobio gobio L. In: De P a u w, N., Jaspers, E., A c k er fors, H., W i 1 kin s, N. [Eds] Aquaculture - a biotechnology in progress, Belgium, Bredene, K o z lows k i, B Praktyka stymulacji hormonalnej w sztucznym rozrodzie ryb karpiowatych [Practise of hormonal stimulation of artificial spawning in cyprinid fishes]. Broszura Instytutu Rybactwa Sr6dlqdowego, 162, Kucharczyk, D., Ku jawa, R., Luczynski, M., Gl ogowski, J., Babiak, I., W y s z o m i r s k a, E Induced spawning in bream (Abramis brama L.) using bream and carp pituitary extract and HCG. Aquacult. Res., 28, Ku charczyk, D., Kujawa, R., Mamcarz, A., Skrzypczak, A., Wyszomirska, E Induced spawning in perch (Perca fluviatilis L.) using carp pituitary extract and HCG. Aquacult. Res. 27, 847--S52. K u ja w a, R., K u char c z y k, D Przyzyciowe pobieranie oocyt6w ryb karpiowatych i okoniowatych za pomocq katetera [Collection of oocytes from alive cyprinids and percids using catheter]. Komunikaty Rybackie, 4, P 1 at t, C Problemy raclwnku prawdopodobiefzstwa [Problems of calculus of probability and statistical mathematics]. Warszawa. Panstwowe Wydawnictwo Naukowe. T aka s h i m a, F., We i l, C., B i 11 a r d, R., Cri m, L.W., F os tier, A Stimulation of spermiation by LHRH analogue in carp. Bull. Jpn. Soc. Sci. Fish., 50, Ya r on, Z Endocrine control of gametogenesis and spawning induction in the carp. Aquaculture, 129,