Studies On Reproduction Physiological Function Of The P450arom In Rockfish (Sebastes Schlegeli)

In this thesis, takes the rockfish(Sebastes schlegeli) as study objects. Development of the gonad, sexual steroid hormone, two types of aromatase gene and its gene organization expression, are taken and studied by the methods of histological staining methods, immunohistochemical techniques, radioimmunoassay and biochemical methods in details. The main results and conclusions are as follows:1. Histological study on the development and annual change in the testis of Sebastes schlegeli.Sebastes schlegeli has a pair of testical, two leaf asymmetry, belong to lobular type. Spermatogenesis through 6 typical developmental steps:the primary spermatogonium, secondary spermatogonium, primary spermatocytes, secondary spermatocytes, spoermatid and sperm.The anniversary change from the testicle is it can be seen, testicle growth and seasonal characteristic breed the synchronism.The testis is the proliferation of primary spermatogonia from Murch to May. Spermatogenesis period is from July to November and the sperm degeneration and absorption stage is from January to early May.2 Study on the changes of plasma sex steroid hormone levels in Sebastes schlegeli in season and the reproductive cycle.Plasma testosterone,progesterone and 17β-Estradiol in female and male was detected with radio immunoassay(HIA).The results show that plasma progesterone and 17β-Estradiol in female are significantly increased during the reproductive period.Then E2 and Psignificantly descend and fall to the annual minimum in the May, but testosterone levels increased significantly during the embryonic period (P<0.05) to peak.All these results show the increase of female Sebastes schlegeli's plasma 17β-Estradiol contents, have satisfied the vitellus' formation and accumulation, even promoted the ovary's maturation. T and P levels associated with ovary development, suggesting that they are a certain stage of ovarian development necessary.In male Sebastes schlegeli, the variation of plasma progesterone and 17β-Estradiol are similarity with female. The annual chang of T lagged than E2 and P levels.These results indicate that male reproductive maintained at a high level of testosterone to promote the occurrence and formation of sperm.3 Immunohistochemical study on aromatase in gonadal and sexual cellsAromatase immunopositive reaction is found in each period of ovary's development. In each phase of the follicular oocytes, the cytoplasm of oocyte and the Follicular membrane showed immunopositive reaction to aromatase. The nucleus of oocyte shows negative reactinon.Embryos showed strong immunoreactivity. Spermatogonia, spermatocytes, sperm cells and stromal cells in the developing testis showed aromatase immunoreactivity in different levels, but the nucleolar of spermatogonia and sperm showed negative immune response.During the testicle's development, strong immunopositive reaction of aromatase happens in mesangial cell and basement membrane. Negative expression is found in sertoli cell immunonegative reaction to ER.The above results show that aromatase plays an important physiological role in the gonad development. Aromatase expression providing evidence that sex steroid hormones has important status in gonadal development and maturation.4 Molecular cloning, characterization and expression analysis of ovarian P450arom(P450aromA) and brain P450arom(P450aromB) in Sebastes schlegeli.The 1781 bp full-length ovarian P450 aromatase(P450aromA)cDNA encoding a deduced protein of 516 residues was isolated from the ovary of Sebastes schlegeli by using homologous cloning and RACE.The RT-PCR analysis reveals that P450aromA mRNA was expressed highly in the ovary and weakly in the testis,but not other tissues.These results indicate that P450aromA is involved in gonad development and sex determination.The CYP19B gene cloned 1109bp long cDNA fragment, encoding 369 amino acids. The results from RT-PCR analysis reveals that P450aromB mRNA was expressed highly in the brain and gonad, and low expressed in other tissues.These results suggested that the P450aromB is involved in course of the gonad development and sex determination in Sebastes schlegeli. And may be involved in endocrine regulation of nervous system.