Induction of apoptosis in testis of the marine teleost mummichog Fundulus heteroclitus after in vivo exposure to the antifouling biocide 4,5-dichloro-2-n-octyl-3(2H)-isothiazolone (Sea-Nine 211)

摘要

4,5-dichloro-2-n-octyl-3(2H)-isothiazolone (Sea-Nine 211) has been widely used as an effective antifouling biocide. However, little is known about its reproductive toxicity in fish. Here we investigated testic-ular toxicity in a marine teleost, the mummichog Fundulus heteroclitus, after exposure to Sea-Nine 211 for 28 d. Although Sea-Nine 211 exposure did not affect germ cell proliferation in testis, terminal deoxynu-cleotidyl transferase-mediated deoxy-UTP nick-end labeling revealed that the number of apoptotic sper-matocytes was increased in the 1.0- and 3.0-μg L~(-1) groups, and significant differences emerged between the 1.0-μg L~(-1) group and control groups. Immunohistochemistry showed that the numbers of cysts expressing caspases 2, 3, 6, and 8 (apoptosis-associated proteins) were significantly increased in the 1.0-μg L~(-1) group, whereas the signal intensity of an anti-apoptotic protein Bcl-xL was reduced in a dose-dependent manner. Moreover, the number of cysts positive for neuronal nitric oxide synthase was twofold higher in the 1.0-μg L~(-1) group than in the control groups. These results suggest that long-term exposure to Sea-Nine 211 induces apoptosis in the testicular germ cells of mummichogs via a cas-pase-dependent pathway and that oxidative stress via nitric oxide synthesized by neuronal nitric oxide synthase is involved in this induction.