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Ing in a Number of Bivalve Species by Injection of Serotonin Solution

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Ing in a Number of Bivalve Species by Injection of Serotonin Solution ;. C.M. 1985/F:61 Mariculture Committee/ A RAPID METHOD TO INDUCE SPA\~ING IN A NUMBER OF BIVALVE SPECIES BY INJECTION OF SEROTONIN SOLUTION l l 2 M. Castagna • M. C. Gibbons and J. Goodsell lVirginia Institute of Marine Science, School of Marine Science, College cf William and Mary Eastern Shore Laboratory. Wachapreague, VA 23480. 2Rutgers Shellfish Research Laboratory. P.O. Box 587, Port Norris, NJ 08349. ABSTRACT Injection of serotonin (5-hydroxytryptamine, creatinine sulfate complex) • solution was used to induce spawning in a number of bivalve species. Studies at the Virginia Institute of Marine Science demonstrated that intergonadal injection induced spawning in Argopecten irradians, Crassostrea virginica, Mercenaria mercenaria and Spisula solidissima. Intermuscular injection of serotonin solution induced spawning of Arctica islandica, Chione cancellata, Cyrtopleura costata, Geukensia demissa, Hiatella arctica, Mercenaria mercenaria, Mesodesma arctatum, Mya arenaria, Mya truncata, Mytilus edulis and Rangia cuneata. \"'., ... -2- Patinopecten yessoensis, Patinopecten caurinus, Hinnites multirugosus, Argopecten circularis, Pinctada mazatlantica, Protothaca asperrima, Ensis sp., Tridacna gigas, Tridacna derasa, Tridacna crocea, Hippopus hippopus have also been induced to spawn by injection of serotonin (Hatsutani and Nomura, 1982; personal communication - Neil Boume, Pacific Biological Station, Nanaimo, B.C., Canada; John W. Ewart, University of Delaware, School of Marine Science, Lewes, Delaware, USA; and Richard D. Braley, 4It School of Zoology, University of South Wales, P.O. Box 1, Kensington, New South Wales 2033, Australia). Concentration of serotonin was 0.2 to 2.0 mM solution with 2.0 ~1 most eommonly used. Dosages were adjusted to size of the specimen with 0.4 ml used for average sized 4 to 8 cm in length oysters or elams. Symptomatic behavior was observed almost immediately following injection of the solution. Spawning usually began within 15 minutes. I NTRODUCTI ON Many methods of indueing bivalve mollusks to spawn have been described. • Temperature cyeling and the addition of sperm or ova are the most eommonly used methods (Loosanoff and Davis, 1963). The chemical serotonin, 5-hydroxytryptamine (5-BT), has been shown to be an effeetive inducer of spawning for many bivalves (Matsutani and Nomura, 1982; Gibbons et al., in press; Gibbons and Castagna, 1984). Injection of serotonin into the gonad or adductor museie of ripe bivalves will often induee spawning without additional stimuli. ~1ale bivalves respond and spawn in greater number than females treated with serotonin, and unripe individuals who cannot spawn will display the usual behavioral response to thc serotonin. ..L.... .....,.,.'.:.•,,·,4 .~ -3- Serotonin is a neurotransmitter naturally occurring in the nervous system of mollusks (I'lelsh and Moorhead, 1960). It stimulates heart beating and ciliary beating of gills in bivalves (Leake and Walker, 1980). However, the role of serotonin in spawning of bivalves is unknown. Studies were carried out to test the effectiveness of serotonin on different species. MATERIALS AND METHODS • Serotonin (S-hydroxytryptamine, ereatinine sulfate eomplex) was dissolved in 1 pm fi1tered seawater to the desired eoneentration before use. Onee a solution is mixed it is kept refrigerated since it deteriorates at room temperature or in light. Matsutani and Nomura (1982) found the optimal eoncentration for Patinopecten yessoensis was 2 Illi-l so this was the· concentration common1y used. When using this chemica1, the biva1ves to be spawned are either placed in individual dishes or in a common spawning trough filled with filtered seawater. The serotonin was injected either into the gonad or into the • adductor museie or on occasion into the foot. A new needle was used for each bivalve to prevent transference of gonadal products, blood, or other tissue which may act as a stimuli. A small nick was made into the edge of the shell to allow insertion of the needle in species which close tightly such as oysters. This was accomp1ished with a sma11 stee1 file. Most of the bivalves were injected with 0.4 ml of 2 mr-t solution. I'lhen smaller sized .species were used such as Mulinia lateralis, a smaller amount was injected. All controls were injected with the same amount of 1 pm filtered seawater as the test animals. / -4- Testsofindependence using the g-statistic and Williams correction for a 2 x 2 contingency table were conducted to determine whether or not spawning and injection with serotonin were independent (SokaI and Rohlf, 1981). The level of statistical reference was chosen to be 0(. = 0.01. RE SU LTS In these studies, serotonin was effective in inducing spawning in 83% 4It of bay scallops Argopecten irradians, 70% of oysters Crassostrea virginica, 60% of surf clams Spisula solidissima, 45% of ribbed musseIs Geukensia demissa, 45% of hard clams Mercenaria mercenaria and 27% of ocean quahogs Arctica islandica. In these bivalves males were much more sensitive to serotonin since they predominated the spawners. For the hermaphroditic bay scallops, 97% released sperm while for dioecious bivalves between 67 and 100% of the spawners in each test lot were males. This data was not extracted from tests with other species due to insufficent numbers of spawners. Reaction of ripe male bivalves to serotonin is almost immediate. • Oysters gaped and spawned as quickly as 2-5 minutes after injection and most of them had spawned within 30 minutes. Ribbed musseIs spawned within 15 minutes. The behavior of biva1ves injected with serotonin was to extend the foot and probe the bottom, gape and usually pump more vigorous1y. All broodstock survived the stress of injection with serotonin solution or the filtered seawater used for the controls. Often the notches in the shell showed new growth within one week. .. -5- The authors successfully demonstrated that intergonadal or inter­ muscular injection of serotonin solution induced spawning in 14 species (Table I).' It was not successful in two species of blood arks, Noetia ponderosa and Anadara ovalis. 80th species released quantities of blood cells but no gonadal products were observed. Other investigators have reported that injection of serotonin solution successfully induced spawning in 11 additional species (Table 11). Spawning induced by serotonin has the advantages of ease of application, does not require temperature cycling, and speed and synchronization of induction. Serotonin may also be used to successfully induce spawning in other bivalves which cannot be spawned using traditional spawning stimuli . • . -6- LITERATURE CITED Boume, Nei1. Personal communication. Bra1ey, Richard D. Personal communication. Ewart, John W. Personal communication. Gibbons, M.C., J.G. Goodse11, M. Castagna, and R.A. Lutz. Chemica1 induction of spawning by serotonin in the ocean quahog Arctica is1andica. J. of She11fish Research. In press. Gibbons, M.C. and M.. Castagna. 1984. Serotonin as an inducer of spawning in six biva1ve species. Aquacu1ture 40: 189-191. Leake, L.D. and R.J. Walker. 1980. Invertebrate Neuropharmaco1ogy. John Wi1ey, New York, pp. 102-143. Loosanoff, V.L. and H.C. Davis. 1963. Rearing of biva1ve mo11usks. Adv. Mar. Bio1. 1: 1-136. Matsutani, T. and T. Nomura. 1982. Induction of spawning by serotonin in the sca110p Patinopecten yessoensis (Jay). Mar. Bio1. Lett. 3: 353-358. Soka1, R.R. and F.J. Roh1f. 1981. Biometry. W.H. Freeman, San Francisco, CA, 2nd edition, 859 pp . • We1sh, J.H. and M. Moorhead. 1960. The quantitative distribution of 5­ hydroxytryptamine in the invertebrates, especia11y in their nervous systems. J. Neurochem. 6: 146-169. · . Table I. Species of bivalves induced to spawn by injection with serotonin. Common Name Scientific Name ocean quahog Arctica islandica bay scallop Argopecten irradians cross-barred venus Chione cancellata American oyster Crassostrea virginica angel wing Cyrtopleura costata ribbed musseI Geukensia demissa arctic rock borer Hiatella arctica hard clam Mercenaria mercenaria arctic wedge clam Mesodesma arctatum soft-shell clam Mya arenaria truncate soft-shell clam Mya truncata blue musseI MytHus edulis common rangia Rangia cuneata surf clam Spisula solidissima · . Tab1e 11. Additional species of biva1ves induced to spawn by injection with serotonin. Species Reported by Patinopecten yessoensis Matsutani and Nomura. 1982 Patinopecten caurinus Nei1 Bourne. personal communication Hinnites mu1tirugosus Nei1 Boume, personal communication Argopecten circu1aris John W. Ewart, personal communication Pinctada mazat1antica John W. Ewart. personal communication Protothaca asperrima John W. Ewart, personal communication Ensis sp. John W. Ewart, personal communication Tridacna gigas Richard D. Bra1ey, personal communication Tridacna derasa Richard D. Bra1ey, personal communication Tridacna crocea Richard D. Bra1ey. personal communication Hippopus hippopus Richard D. Braley, personal communication.
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