|
Günther (Cynoglossus semilaevis Günther) underlying the large luxury economy of our coastal fish. In this paper, the artificial breeding male half slip tongue sole object of study, the use of histology, immunohistochemistry chemistry, hormone radiation immunity determination method and molecular biology methods, semi slippery tongue sole of the testis, plasma sex class of steroid hormones and their receptors carry out the research and analysis, obtained results and conclusions are as follows: 1. breeding half-smooth tongue sole testis development anniversary of the changes in the histological Günther testis one pair, like most bony fish of the testis, half-smooth tongue sole testis as lobular-shaped structure, compact capsule structure is not obvious. The male half-smooth tongue sole has the structure of the seminal vesicle, seminal vesicle testes back forward extending out the length of the testis quite fine lobular structures in the seminal vesicle, surrounded by smooth muscle and connective tissue cavities of different sizes, and the cavity filled with mature sperm. Testis output tube together in one side of the testis, and the mature sperm lobular cavity remitted to the output tube, then the output tube into the seminal vesicle. Experience Günther sperm occurred spermatogonia, primary spermatocytes, secondary spermatocytes, spermatids and sperm of five typical developmental stage, but the development of sperm cells in the testis is not synchronized. Breeding half-smooth tongue sole testicular development and wild male lag compared with the breeding season lasted from October to next February, after which the testis into the repeat developmental stages, started in August spermatogenic activity. 2 and a half smooth tongue sole testis estrogen receptor alpha (ERα), estrogen receptor β (ERβ), androgen receptor (AR) immunohistochemical study 2.1 estrogen receptor α (ERα) in testis and sperm cells in the immune Organization chemical positioning sexually mature half slip tongue sole testis of various developmental stages, of ERα in spermatogonia, spermatocytes, sperm cells and sperm lobular quality at showed immunopositive reaction, whereas in the development of each interim the essence of the original nucleus Ren the spermatocyte nucleus and sperm showed immunonegative reaction. 2.1 estrogen receptor β (ERβ) in the testis and sperm cells Immunohistochemical localization of spermatogonia, spermatocytes, sperm cells and cells of mesenchymal different levels of ERβ immunopositive reaction in the testis of the developmental period, but the spermatogonial nuclei Ren and sperm showed immunonegative reaction. 2.2 male hormone receptor (AR) in the testis and sperm cells immunohistochemistry chemical positioning of the half-smooth tongue sole testes of androgen receptor (AR) Immunohistochemical localization of discovery, AR in half-smooth tongue sole testis of all developmental period of spermatogonial cells primary spermatocytes and fine lobular stromal expression. These results suggest that the estrogen receptor, plays an important physiological role in the proliferation of androgen receptors in testicular development and sperm cells, and provide evidence for proof of sex steroid hormones in gonadal development and maturation process. 3. Semi-smooth tongue sole in the androgen receptor (AR) gene cloning and expression of this experiment to clone the androgen receptor AR gene cDNA sequence (434bp). Tissue expression analysis showed that the expression of the AR gene in 11 kinds of organizations Günther gonads, liver, stomach, spleen, kidney, head kidney, intestine, gills, heart, brain and muscle. The expression of different expression levels in the kidney is the most abundant, at least gills. Determined by hormones, the male half smooth tongue sole plasma T levels showed the cyclical changes in the gonads, the highest peak in the February to April, and then begin to decline, 11 to January, the lowest for the whole year. AR mRNA expression in the male Günther testis, liver, spleen, kidney and brain tissues by RT-PCR method for the first time the annual change analysis. The results show that the expression of the AR mRNA in the five kinds of tissues showed a periodic variation. Plasma T content, immunohistochemistry results confirm each other, revealing the AR in male Günther in. 4 and a half smooth tongue sole estrogen receptor α (ERα) gene cloning and expression of the experimental cloning estrogen receptor ERα gene cDNA partial sequence (454bp). ERα gene expression in 11 tissues of male Günther testis, liver, stomach, spleen, kidney, head kidney, intestine, gills, heart, brain and muscle were detected by RT-PCR method. The results show that of ERα gene expression has nine kinds of gills and muscle tissue in just the expression of different expression is most abundant in the liver, head kidney least. Determined by hormones, the male gonads showed the cyclical changes in plasma E2 levels Günther. The content of the highest peak appears in the August to October, and then began a significant decline, while 2 to 4 drops to the lowest level of the year, 5 to July plasma E2 levels increased significantly. The expression of ERαmRNA in the male Günther testis, liver, spleen, kidney and brain tissues by RT-PCR method for the first time the annual variation analysis. The results showed that of ERα gene mRNA expression in five tissues showed a certain regularity changes, negative correlation between expression in the testis, liver and spleen trend curves of E2 and GSI and HSI consistent. 5. Günther estrogen receptor β (ERβ) gene cloning and expression by RT-PCR, amplified from the testis Günther the ERβ fragment (525bp). The results of tissue expression of ERβ gene expression, which have 11 kinds of tissue of the testis, liver, stomach, spleen, kidney, head kidney, intestine, gills, heart, brain and muscle richest expression in the liver, head kidney, at least. Analysis of the annual variation expression of ERβmRNA in the male Günther testis, liver, spleen, kidney and brain tissues, results showed that ERβ and ERα expression pattern is very similar.
|