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Serotonin as an inducer of spawning in six bivalve species
Aquaculture, 40 (1984) 189-191 Elsevier Science Publishers B.V., Amsterdam
189 - Printed
in The Netherlands
Short Communication SEROTONIN SPECIES*
AS AN INDUCER OF SPAWNING IN SIX BIVALVE
M.C. GIBBONS
and M. CASTAGNA
Virginia Institute of Marine Science and School of Marine Science, William and Mary, Wachapreague, VA 23480 (U.S.A.) *Contribution (Accepted
No. 1183 from Virginia 4 February
Institute
College of
of Marine Science
1984)
ABSTRACT Gibbons, M.C. and Castagna, M., 1984. Serotonin species. Aquaculture, 40: 189-191.
as an inducer
of spawning
in six bivalve
Serotonin induced spawning in six bivalve species using individual spawning techniques without any additional stimuli. Intragonadal injection of serotonin induced spawning in the bay scallop Argopecten irradians, the American oyster Crassostrea uirginica, and the surf clam Spisula solidissima. Injection of serotonin into the anterior adductor muscle of the ocean quahog Arctica islandica, the ribbed mussel Geukensia demissa, and the hard clam Mercenaria mercenaria induced spawning. The dosage of 0.4 ml of 2 mM serotonin solution used in this study stimulated bivalves to spawn within 15 min.
INTRODUCTION
Many methods of artificially inducing bivalve molluscs to spawn have been described. Temperature cycling and/or the addition of sperm or ova are methods most commonly used to induce spawning in bivalves’(Loosanoff and Davis, 1963). In this preliminary study, spawning was induced by injection of serotonin in the following bivalve species: the ocean quahog Arctica ishndica, the bay scallop Argopecten irradians, the American oyster Crassostrea virginica, the ribbed mussel Geukensia demissa, the hard clam Mercenaria mercenaria, and the surf clam Spisula solidissima. MATERIALS
AND METHODS
The technique of individual spawning was used to spawn the six bivalve species without any additional stimuli (Castagna and Kraeuter, 1981). Individual bivalves were placed in glass dishes containing 1 1 of 1 pm filtered seawater (32%,) and held at desired temperatures in water baths. Crystalline serotonin (5hydroxytryptamine, creatinine sulfate com-
0044~8486/84/$03.00
0 1984 Elsevier
Science
Publishers
B.V.
190
plex, Sigma Chemical Company) was dissolved in 1 pm filtered seawater to prepare a 2 rniV solution. This concentration is similar to the optimal concentration used by Matsutani and Nomura (1982) to induce spawning in the scallop Pa tinopec ten yessoensis. Small notches were filed into the valve margins adjacent to anterior adductor muscles of ocean quahogs, hard clams, and ribbed mussels. Serotonin solution was injected into the muscles. In scallops, oysters, and surf clams the serotonin solution was injected directly into the gonad. A new needle was used for each bivalve to prevent transference of gonadal products. Bay scallops were held open by hand and injected into the middle of the gonad. A small notch was filed in the valve margin of each oyster to allow insertion of the needle. Surf clams were injected at the umbone posterior to the resilium. All bivalves received 0.4 ml of 2 miV serotonin solution, while controls were injected with 0.4 ml of 1 pm filtered seawater. Statistical analyses using the g-statistic and Williams’ correction for a 2 X 2 contingency table were conducted to determine whether spawning was independent of serotonin injection (Sokal and Rohlf, 1981). RESULTS
AND DISCUSSION
Serotonin induced the ocean quahog, bay scallop, oyster, ribbed mussel, hard clam, and surf clam to spawn (Table I). Ocean quahogs have not been spawned by other investigators. Reaction of ripe bivalves to serotonin was almost immediate. Ocean quahogs, hard clams, and surf clams injected TABLE
I
Numbers
of bivalves induced
to spawn
by serotonin
Species
Temperature
Treatment
A rctica islandica
15-16°C
Serotonin Control
70 70
19a 0
15 0
4 0
Argopec ten irradians
ZO-21°C
Serotonin Control
35 35
29a 3
29 2
1 1
Crassostrea virginica
25°C
Serotonin Control
30 30
21a 0
21 0
0 0
Geukensia demissa
28°C
Serotonin Control
20 20
9a 1
8 1
1 0
Mercenaria mercenaria
28-29°C
Serotonin Control
329 70
137a 0
116 0
21 0
Spisula solidissima
19°C
Serotonin Control
45 45
27a 1
18 0
9 1
Vignificant
at P < 0.005.
Number tested
injection Number spawned
Number males
of
Number females
of
191
with serotonin gaped, extended siphons, probed with feet, and began spawning within 15 min. Bay scallops reacted to serotonin by gaping, foot probing, and initiated spawning 10 min after injection; the majority spawning within 15 min. Oysters gaped and began spawning 5 min after injection with the majority spawning within 30 min. Ribbed mussels injected with serotonin gaped and spawned after 15 min. None of the controls injected with filtered seawater exhibited any behavior such as foot probing, gaping, or increased pumping. For the ocean quahog, bay scallop, and hard clam, the gametes released by broodstock injected with serotonin were competent, fertilization occurred, and larvae grew to metamorphosis. All broodstock survived the stress of injection with filtered seawater or serotonin solution. The notches filed in the shells of hard clams and oysters exhibited new growth after 1 week. Serotonin is a neurotransmitter present in the nervous systems of mollusts (Welsh and Moorhead, 1960). It stimulates heart beating and ciliary beating of gills in bivalves (Leake and Walker, 1980). However, the role of serotonin in the spawning of marine bivalves is unknown. Spawning induced by serotonin has the advantages of ease of application, lack of need to cycle temperature, and speed and synchronization of induction. As broodstock may be spawned individually, this method may be applicable to techniques to manipulate stocks genetically. Serotonin may also be used successfully to induce spawning in other bivalve species which are resistant to traditional spawning stimuli.
REFERENCES Castagna, M. and Kraeuter, J.N., 1981. Manual for growing the hard clam Mercenariu. VIMS Special Report in Applied Marine Science and Ocean Engineering No. 249, 110 pp. Leake, L.D. and Walker, R.J., 1980. Invertebrate Neuropharmacology. John Wiley, New York, pp. 102-143. Loosanoff, V.L. and Davis, H.C., 1963. Rearing of bivalve molluscs. Adv. Mar. Biol., 1: l-136. Matsutani, T. and Nomura, T., 1982. Induction of spawning by serotonin in the scallop Patinopecten yessoensis (Jay). Mar. Biol. Lett., 3: 353-358. Sokal, R.R. and Rohlf, F.J., 1981. Biometry. W.H. Freeman, San Francisco, CA, 2nd edn., 859 pp. Welsh, J.H. and Moorhead, M., 1960. The quantitative distribution of 5-hydroxytryptamine in the invertebrates, especially in their nervous systems. J. Neurochem., 6: 146-169.
189 - Printed
in The Netherlands
Short Communication SEROTONIN SPECIES*
AS AN INDUCER OF SPAWNING IN SIX BIVALVE
M.C. GIBBONS
and M. CASTAGNA
Virginia Institute of Marine Science and School of Marine Science, William and Mary, Wachapreague, VA 23480 (U.S.A.) *Contribution (Accepted
No. 1183 from Virginia 4 February
Institute
College of
of Marine Science
1984)
ABSTRACT Gibbons, M.C. and Castagna, M., 1984. Serotonin species. Aquaculture, 40: 189-191.
as an inducer
of spawning
in six bivalve
Serotonin induced spawning in six bivalve species using individual spawning techniques without any additional stimuli. Intragonadal injection of serotonin induced spawning in the bay scallop Argopecten irradians, the American oyster Crassostrea uirginica, and the surf clam Spisula solidissima. Injection of serotonin into the anterior adductor muscle of the ocean quahog Arctica islandica, the ribbed mussel Geukensia demissa, and the hard clam Mercenaria mercenaria induced spawning. The dosage of 0.4 ml of 2 mM serotonin solution used in this study stimulated bivalves to spawn within 15 min.
INTRODUCTION
Many methods of artificially inducing bivalve molluscs to spawn have been described. Temperature cycling and/or the addition of sperm or ova are methods most commonly used to induce spawning in bivalves’(Loosanoff and Davis, 1963). In this preliminary study, spawning was induced by injection of serotonin in the following bivalve species: the ocean quahog Arctica ishndica, the bay scallop Argopecten irradians, the American oyster Crassostrea virginica, the ribbed mussel Geukensia demissa, the hard clam Mercenaria mercenaria, and the surf clam Spisula solidissima. MATERIALS
AND METHODS
The technique of individual spawning was used to spawn the six bivalve species without any additional stimuli (Castagna and Kraeuter, 1981). Individual bivalves were placed in glass dishes containing 1 1 of 1 pm filtered seawater (32%,) and held at desired temperatures in water baths. Crystalline serotonin (5hydroxytryptamine, creatinine sulfate com-
0044~8486/84/$03.00
0 1984 Elsevier
Science
Publishers
B.V.
190
plex, Sigma Chemical Company) was dissolved in 1 pm filtered seawater to prepare a 2 rniV solution. This concentration is similar to the optimal concentration used by Matsutani and Nomura (1982) to induce spawning in the scallop Pa tinopec ten yessoensis. Small notches were filed into the valve margins adjacent to anterior adductor muscles of ocean quahogs, hard clams, and ribbed mussels. Serotonin solution was injected into the muscles. In scallops, oysters, and surf clams the serotonin solution was injected directly into the gonad. A new needle was used for each bivalve to prevent transference of gonadal products. Bay scallops were held open by hand and injected into the middle of the gonad. A small notch was filed in the valve margin of each oyster to allow insertion of the needle. Surf clams were injected at the umbone posterior to the resilium. All bivalves received 0.4 ml of 2 miV serotonin solution, while controls were injected with 0.4 ml of 1 pm filtered seawater. Statistical analyses using the g-statistic and Williams’ correction for a 2 X 2 contingency table were conducted to determine whether spawning was independent of serotonin injection (Sokal and Rohlf, 1981). RESULTS
AND DISCUSSION
Serotonin induced the ocean quahog, bay scallop, oyster, ribbed mussel, hard clam, and surf clam to spawn (Table I). Ocean quahogs have not been spawned by other investigators. Reaction of ripe bivalves to serotonin was almost immediate. Ocean quahogs, hard clams, and surf clams injected TABLE
I
Numbers
of bivalves induced
to spawn
by serotonin
Species
Temperature
Treatment
A rctica islandica
15-16°C
Serotonin Control
70 70
19a 0
15 0
4 0
Argopec ten irradians
ZO-21°C
Serotonin Control
35 35
29a 3
29 2
1 1
Crassostrea virginica
25°C
Serotonin Control
30 30
21a 0
21 0
0 0
Geukensia demissa
28°C
Serotonin Control
20 20
9a 1
8 1
1 0
Mercenaria mercenaria
28-29°C
Serotonin Control
329 70
137a 0
116 0
21 0
Spisula solidissima
19°C
Serotonin Control
45 45
27a 1
18 0
9 1
Vignificant
at P < 0.005.
Number tested
injection Number spawned
Number males
of
Number females
of
191
with serotonin gaped, extended siphons, probed with feet, and began spawning within 15 min. Bay scallops reacted to serotonin by gaping, foot probing, and initiated spawning 10 min after injection; the majority spawning within 15 min. Oysters gaped and began spawning 5 min after injection with the majority spawning within 30 min. Ribbed mussels injected with serotonin gaped and spawned after 15 min. None of the controls injected with filtered seawater exhibited any behavior such as foot probing, gaping, or increased pumping. For the ocean quahog, bay scallop, and hard clam, the gametes released by broodstock injected with serotonin were competent, fertilization occurred, and larvae grew to metamorphosis. All broodstock survived the stress of injection with filtered seawater or serotonin solution. The notches filed in the shells of hard clams and oysters exhibited new growth after 1 week. Serotonin is a neurotransmitter present in the nervous systems of mollusts (Welsh and Moorhead, 1960). It stimulates heart beating and ciliary beating of gills in bivalves (Leake and Walker, 1980). However, the role of serotonin in the spawning of marine bivalves is unknown. Spawning induced by serotonin has the advantages of ease of application, lack of need to cycle temperature, and speed and synchronization of induction. As broodstock may be spawned individually, this method may be applicable to techniques to manipulate stocks genetically. Serotonin may also be used successfully to induce spawning in other bivalve species which are resistant to traditional spawning stimuli.
REFERENCES Castagna, M. and Kraeuter, J.N., 1981. Manual for growing the hard clam Mercenariu. VIMS Special Report in Applied Marine Science and Ocean Engineering No. 249, 110 pp. Leake, L.D. and Walker, R.J., 1980. Invertebrate Neuropharmacology. John Wiley, New York, pp. 102-143. Loosanoff, V.L. and Davis, H.C., 1963. Rearing of bivalve molluscs. Adv. Mar. Biol., 1: l-136. Matsutani, T. and Nomura, T., 1982. Induction of spawning by serotonin in the scallop Patinopecten yessoensis (Jay). Mar. Biol. Lett., 3: 353-358. Sokal, R.R. and Rohlf, F.J., 1981. Biometry. W.H. Freeman, San Francisco, CA, 2nd edn., 859 pp. Welsh, J.H. and Moorhead, M., 1960. The quantitative distribution of 5-hydroxytryptamine in the invertebrates, especially in their nervous systems. J. Neurochem., 6: 146-169.