Effects Of Octylphenol On Spermatogenesis And Reproductive Behavior Of Rana Chensinensis

Octylphenol(OP),one of the degradation products of alkylphenol ethoxylates,belongs to non-ionic surfactants widely used in industrial and agricultural production and daily life.OP as a kind of environmental pollutants with weak estrogenic activity,can affect the growth,development and reproduction of wild animals through accumulation in the body.The aim of the present study is to evaluate the effects of OP on testis function,reproductive behavior and embryonic development of amphibians.The Chinese brown frogs(Rana chensinensis)were considered as experimental animal in this study.The male adults were exposed to 10-8,10-7,10-6,10-5 mol/L OP for 10,20 and 30 days,respectively.The testes were got from R.chensinensis at certain days.To study effects of OP on the histopathological and cytological changes in testis,the microscopic and ultrastructural changes of the testis were observed by light and transmission electron microscopy,and apoptosis cells of seminiferous tubule and the Leydig cells are tested by Tunel technique,and the apoptosis index(AI)was counted.In order to evaluate the changes of reproductive behavior and reproductive function of R.chensinensis after OP exposure,the male R.chensinensis exposed to OP 30 days,with untreated female frogs pairing in accordance with the proportion of 1:1,then its pairing behavior was observed and the number of pairing,laying and hatchability was computed.In addition,to investigate the effects of OP on embryonic hatching,post-embryonic growth and development through interfering the embryonic development,the embryos at Gs 9-10 were continuously exposed to 10-8,10-7 and 10-6 mol/L OP until the completion of metamorphosis.The hatchability was computed and the number of experimental days needed from Gs 10 to Gs 38,Gs 42 and Gs 46 were recorded respectively.In addition,the body length,weight and sex ratio of tadpoles were measured on 30?50?70 days and Gs 46 after exposure to OP.The experimental results and conclusions are listed as follows:1.After exposure to OP,the microstructures of testis in the male R.chensinensis were observed through tissue section.The non-metamorphic spermatogenic cells were observed in the seminiferous tubule of testis,and the number of spermatozoa was decreased.In addition,the hyperplasia of some Leydig cells were observed in the part of testis,the abnormal expansions were also observed in some seminiferous tubules,and a few of spermatozoa accompanied by Sertoli cells were scattered in the lumen after exposure 10-5 mol/L OP for 20 and 30 days,respectively.Electron microscopic investigation showed that the terochromatin were increased in the nucleus of some Leydig cells,and the lipid droplets were embedded in the nucleus,and the nuclear membrane gap was expanded in the testis after OP exposure.Besides,the number of lipid droplets and mitochondria with tubular christae were increased dramatically in the cytoplasm of some Leydig cells.This indicate that low dose of OP exposure can partly disorder the spermatogenesis process in testis of R.chensinensis.In addition,the compensatory number of some Leyding cells was increased while OP causes the change of Leydig cells' structure and function.2.Tunel detection showed that apoptotic cells in the testis were mainly Leyding cells.Because in the present study the testis were in sperm storage overwintering stage,so a little of sperm cells apoptosis were observed.As compared to the control,the AI of Leydig cells was significantly elevated after exposure to 10-6 mol/L OP for 30 days.In 10-5 mol/L OP group,AI was significantly higher than the control at each of exposure period.The AI of Leydig cell suggested OP have significant dose-dependent and time-cumulative effects,and also suggested that cell apoptosis is induced by OP.3.Pairing of OP-exposed males with untreated female frogs showed that OP treatment resulted in decreased behavior of male and female frogs and a significant OP dose-dependent effect.The fecundity and hatching rate of the embryos showed that,compared with control,the hatching rate of 10-5 mol/L OP group was significantly reduced.Thsed indicated that OP exposure can affect the male reproductive behaviour and fertilization.4.The hatchability was counted after all embryos hatching.The result of hatchability showed that exposure to 10-8 mol/L OP did not caused any affection in hatchability compared with control.In contrast,the hatchability was reduced significantly in 10-7 and 10-6 mol/L OP group compared to control.These indicated that OP exposure could affect the embryonic development of R.chensinensis.5.As compared with the control group,continuous exposure to OP caused different degree of reduction in the body length and weight of tadpoles,and the development was also delayed at different degree.The difference indicated that OP has a dose-dependent and time cumulative effect.These suggested that OP continuous exposure can affect the growth and development in post-embryonic tadpoles.6.The statistics of sex ratio in larvae showed that compared with control group the proportion of female was elevated after continuous exposure to 10-7 mol/L and 10-6 mol/L OP.The results indicated that OP exposure had an estrogenic effect on the sex ratio of larvae.The above results indicated that OP is an environmental endocrine disrupter with weak estrogenic effects.The OP exposure has a toxic and estrogenic effect on the testis in the male adult of R.chensinensis in a longer time.In addition,OP exposure can cause cell damage and apoptosis in the part of testis,change the reproductive behavior of the pairs of adult R.chensinensis.In addition,The OP exposure affects the incubation of embryo and the growth and development,disrupt gonadal differentiation of tadpoles.