STUDY ON TEH MECHANISM OF GONADOTROPIN SYNTHESIS IN JAPANESE EEL

建立了一种灵敏的反转录-聚合酶链式反应定量体系，用于多次注射鲑鱼脑垂体匀浆(SPH)诱导雌性日本鳗促性腺激素释放激素(GTH)、Ibeta和IIbeta基因的表达。注射SPH后，GTH Ibeta mRNA水平降低，而GTH IIbeta mRNA水平升高。使用GTH Ibeta和IIbeta寡肽抗体的免疫组织化学分析也支持这一结果。此外，还建立了鳗鱼脑垂体器官培养系统，以分析GTH Ibeta和IIbeta表达的调节机制。分离的未成熟雄性脑垂体腺在雌二醇-17β(E_2)存在或不存在的情况下孵育1周。经E2孵育的脑垂体中可见大量的GTH II细胞，而对照组仅有少量GTH II细胞。为了测定培养液或血液中GTH I和II的浓度，以大肠杆菌为载体制备和纯化重组GTH Iβ和IIβ。然而，建立灵敏和特异的完整GTH I和II定量体系仍需进一步研究。

A sensitive quantitation system using reverse transcription-polymerase chain reaction was developed to measure pituitary GTH Ibeta and IIbeta mRNA levels in female Japanese eel where artificial maturation was induced by multiple injections of salmon pituitary homogenate ( SPH). The GTH Ibeta mRNA levels decreased after SPH injections, while the GTH IIbeta mRNA levels increased. Immunohistochemical analysis using oligopeptide antibodies for GTH Ibeta and IIbeta also supported this result. In addition, an organ culture system for eel pituitary was developed to analyze the mechanisms for the regulation of GTH Ibeta and IIbeta expressions. Isolated pituitaries from immature males were incubated for 1 week in the presence or absence of estradiol-17beta (E2 ). A large number of GTH II cells were observed in the pituitary incubated with E2.In contrast, few GTH II cells were found in controls. To measure concentrations of GTH I and II in incubation media or blood, recombinant GTH Ibeta and IIbeta using E.coli were produced and purified as standards. However, establishment of a sensitive and specific quantitation system for intact GTH I and IIstill requires further research.