Molecular mechanisms of oviposition control

我们提出了一个关于产卵机制的假设：卵巢前列腺素被释放到外周血浆中，刺激子宫收缩，进而触发神经垂体释放AVT。本课题通过提供以下分子基础上的证据来证明上述假设。在排卵前和排卵后最大卵泡的卵膜层中产生前列腺素，组织PG水平最高，同时在排卵前血浆PGF浓度升高。高效液相色谱法鉴定卵巢-产卵诱导因子为PG。这些结果表明卵巢PG在排卵调节中起着关键作用。血浆AVT浓度也在排卵时升高。AVT释放响应子宫收缩，但这种增加被抑制的吲哚美辛治疗延迟排卵。免疫细胞化学研究显示，在视前区和第三脑室浅表区检测到较多含avt的神经细胞。克隆AVT cDNA并确定其碱基序列。以562个碱基的cDNA为探针，Northern杂交分析在下丘脑中检测到约700个碱基的AVT mRNA。缺水4 d后，mRNA水平升高4倍。2 .原位杂交分析显示AVT mRNA定位于下丘脑脑室周围核。最后，AVT受体cDNA的克隆工程仍在进行中，但我们纯化了鸡子宫AVT受体的溶解部分，并对其进行了电泳和分离。目前，氨基酸测序正在研究中，将用于PCR克隆引物分离该基因。根据人催产素受体cDNA同源区和大鼠精氨酸加压素受体同源区氨基酸序列，合成了鸡子宫RNA的RT-PCR引物。以452个碱基的RT-PCR cDNA产物为探针，对鸡子宫进行北方杂交。分别鉴定出5.6、4.6和2.1 kd的杂交mRNA为AVT受体mRNA。少

We have proposed a hypothesis for oviposition mechanism as follows ; ovarian prostaglandin is released into peripheral plasma to stimulating uterine contractions, which in turn, triggers AVT release from the neurohypophysis. The present project was conducted to prove the above hypothesis by providing following evidence on the molecular basis.1. Prostaglandins were produced in the theca layers of the largest pre- and postovulatory follicles and tissue PG level was highest with concurrent increase in plasma PGF concentration immediately before oviposition. Ovarian-oviposition inducing factor was identified as PG by HPLC analysis. These results indicate that ovarian PG plays a pivotal role in the regulation of oviposition.2. Plasma AVT concentration also increased at the time of oviposition. AVT was released in response to uterine contractions but this increase was suppressed by indomethacin treatment which delayed oviposition. Immunocytochemical studies revealed that AVT-containing neuro … More ns were detected at the preoptic region and superficial area of the third ventricle. AVT cDNA was cloned and its base sequence was determined. Using the 562-base cDNA as a probe Northern hybridization analysis detected about 700 bases of AVT mRNA in the hypothalamus. The mRNA levels increased 4-fold after 4-day of water deprivation. Analysis of in situ hybridization revealed AVT mRNA localized in the periventricular nuclei of the hypothalamus.3. Finally, the project of cloning of AVT receptor cDNA is stil in progress but solubilized fraction of AVT receptor of the chicken uterus was purified, electophoresed and this fraction was isolated. At present, amino acid sequencing is under an investigation and will be used for primers in PCR cloning to isolate the gene. On the basis of amino acid sequences of homologous region of human oxytocin receptor cDNA and rat arginine vasoporessin receptor, primers were synthesized for RT-PCR of chicken uterus RNA. The 452-bases RT-PCR cDNA product was used as a probe for Northern hybridization of the chicken uterus. Hybridizable mRNA species of 5.6, 4.6 and 2.1 kd were identified as AVT receptor mRNA. Less