Study on the Gonadotropin Receptors Expression in the Reproductive Endocrine System

本研究旨在阐明脊椎动物生殖内分泌系统中激素靶器官上的肽激素受体的行为，得到以下结果：1)促性腺激素释放激素(GnRH)对性腺发育的影响：在培养的前24 h，用10-9M GnRH或不加GnRH的胎鼠脑垂体原基进行培养，分析激素的合成和释放。结果提示，妊娠13.5天的大鼠垂体原基能够对GnRH产生反应，GnRH能有效地刺激促性腺激素细胞对GnRH的反应。2)大鼠GnRH受体mDNA的克隆与表达：从一个大鼠GnRH的cDNA文库中克隆了一个全长1317bp的GnRH基因。该克隆含有完整的氨基酸编码区，编码区5‘端和3’端分别有172和161个非编码区。然后在…中分析受体基因的表达在发情周期和卵巢切除后，大鼠的脑垂体率较高。该基因只在促性腺激素中表达。动情期的mRNA水平显著低于周期的其他阶段。3)GnRH受体在卵巢中的表达：在卵巢中，GnRH受体在正常卵巢和排卵前期卵泡上高表达，在其壁上颗粒细胞中有强信号。黄体呈弱信号，而黄体卵泡可能在闭锁后呈中等信号。这些数据表明GnRH受体参与了卵巢的各种功能，包括卵泡发育、闭锁、排卵以及排卵和卵泡闭锁后的黄体化。4)各种动物的促性腺激素受体激素结合结构域的特征：在本研究中，我们设计了几对可以从不同动物物种中扩增该结构域的聚合酶链式反应引物。逆转录聚合酶链式反应后进行核苷酸和氨基酸序列分析。哺乳动物和禽类的黄体生成素受体和卵泡刺激素受体的同源性始终高于其配体激素的同源性，提示激素和受体的进化是强相互作用的，受体比配体更为保守。5)隐睾症对促性腺激素受体基因表达的影响：采用Northern印迹法检测单侧隐睾症诱导的成年大鼠睾丸中促性腺激素和促黄体生成素受体mRNAs的表达模式。术后4周，与阴囊睾丸相比，腹侧睾丸中两种受体mRNAs的信号均显著增强。黄体生成素受体转录本的大小模式也发生了明显的变化。这一发现表明，黄体生成素受体mRNA的表达在睾丸内受到局部调控。较少

This research has been started for clarifing the behavior of the peptide hormone receptors on the hormone target organs in the reproduction endocrine system of the vertebrate, and got following results :1) THE EFFECT OF THE GnRH ON THE DEVELOPMENT OF GONADOTROPES : Pituitary primordia from rat fetuses were cultured with or without 10-9 M GnRH during the first 24 h of the culture, and the hormone synthesis and releasing were analyzed. The results suggest that pituitary primordia at 13.5 days of gestation are capable to respond to GnRH,and that GnRH is effective in stimulating the responsiveness of gonadotropes to GnRH during early pituitary cytodifferentiation.2) CLONING AND EXPRESSION OF THE RAT GnRH RECEPTOR mDNA : A rat GnRH cDNA was cloned with 1317 bp from a pituitary cDNA library. This clone had a full amino acid coding region, with 172 and 161 bp noncoding sequences in 5'-and 3'-flanking regions of the coding region, respectively. Then the receptor mRNA expression was analyze in … More rat pituitary during estrous cycle and after ovariectomy. The mRNA was expressed only in the gonadotropes. The mRNA level was significantly smaller in estrus than in other phases of the cycle. In ovariectomized rats, the mRNA expression was significantly increased 4 days after the operation, and maintained until 21 days after ovariectomy.3) GnRH RECEPTOR EXPRESSION IN THE OVARY : In the ovary, GnRH receptor was highly expressed on the healthy Graafian and preovulatory follicles with intense signals in their mural granulosa cells. Corpora lutea showed weak signals, but luteinizing follicles probably after atresia exhibited signals of moderate intensity. These data suggest that the GnRH receptor is involved in the various ovarian functions including follicular deveolopment, atresia, ovulation, and luteinization after ovulation and follicular atresia.4) CHARACTERIZATION OF THE HORMONE BINDING DOMAIN OF THE GONADOTROPIN RECEPTOR IN VARIOUS ANIMALS : For this study, we disigned several pairs of PCR primers which could amplify the domain from various animal species. After RT-PCR,the nucleotide and amino acid sequences were analysized. The values of homology of LH or FSH receptors between mammal and avian were always higher than those of their ligand hormone, suggesting that the hormone and receptor were evolved with strong interaction, and receptor was more conservative than ligand.5) EFFECTS OF CRYPTORCHIDISM ON THE EXPRESSION OF GONADOTROPIN RECEPTOR mRNA : Expression patterns of mRNAs of FSH and LH receptors were examined by Northern blot in adult rat testes after induction of unilateral cryptorchidism. At 4 weeks after the operation, signals were markedly increased in both receptor mRNAs in the abdominal testes as compared to those in the scrotal testes. There was also a remakable change in size pattern of LH receptor transcripts. This finding suggest that the expression of LH receptor mRNA is regulated locally within testis. Less