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Localization and dynamics of oxytocin mRNA as revealed by in situ hybridization histochemistry.

原位杂交组织化学揭示催产素 mRNA 的定位和动态。

基本信息

批准号：
63570032
负责人：
KAWATA Mitsuhiro
金额：
$ 1.34万
依托单位：
Kyoto Prefectural University of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1988
资助国家：
日本
起止时间：
1988 至 1989
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-63570032/
关键词：
In situ hybridization Oxytocin mRNA Gene expression Semiquantitative analysis Paraventricular nucleus Supraoptic nucleus Sex steroids Naloxone オピオイドラットコンピュ-タ-ホモロジ-検索 in situ ハイブリダイゼーションオキシトシン mRNA

项目摘要

Mechanisms of the gene expression of oxytocin neuron system were investigated by using in situ hybridization histochemistry. The first attempt was done to make specific probes for oxytocin mRNA because of the similar sequence of nucleotides to vasopressin mRNA, and this reflected the evaluation of specific hybridization signals to analyze. After designing the specific nucleotide sequence for oxytocin mRNA, 25-30mer oligonucleotides were synthesized. A particular attention was paid to make probes having least homology to mRNAs of vasopressin and other bioactive substances. The probes were labeled by radioactive mononucleotides such as ^3H・dCTP, ^<35>S・dATP. Second attempt was done to investigate the effects of estrogen and/or progesterone to oxytocin gene expression. Estrogen treatment did not induce changes of oxytociin mRNA both in neurons of the paraventricular and supraoptic nuclei. However, estrogen and progesterone treatment caused a significant increase of oxytocin mRNA in neurons, particularly in the ventromedial subnucleus of the paraventricular nucleus. Neurons in the supraoptic nucleus did not show any changes of oxytocin mRNA by estrogen and progesterone treatment. Third experiment was carried out to investigate whether a substained increase in oxytocin secretion, with or without enhanced electrical activity of the cell bodies of oxytocin neurons, leads to a rapid increase in oxytocin mRNA content in these neurons. To stimulate oxytocin release, naloxone was given to female rats after intracerebroventricular morphine infusion. Naloxone increased oxytocin content in plasma 40 fold for at least 40 min in morphine-infused rats. Naloxone had no significant effect on the oxytocin mRNA content in labeled cells in the supraoptic nucleus, and no effect on the proportion of labeled cells.We concluded that there exist differences in mechanisms of gene expression of oxytocin neurons at the nuclear, cell body and terminal levels.

应用原位杂交组织化学方法研究催产素神经元系统基因表达的机制。第一次尝试是制作催产素mRNA的特异性探针，因为催产素mRNA的核苷酸序列与加压素mRNA相似，这反映了对待分析的特异性杂交信号的评价。在设计催产素mRNA的特异性核苷酸序列后，合成了25- 30聚体的寡核苷酸。特别注意制备与加压素和其他生物活性物质的mRNA具有最小同源性的探针。探针用放射性单核苷酸标记，如^3 H·dCTP、^<35>S·dATP。第二个尝试是研究雌激素和/或孕激素对催产素基因表达的影响。雌激素处理不引起室旁核和视上核神经元催产素mRNA的变化。然而，雌激素和孕激素治疗引起的神经元，特别是在室旁核腹内侧亚核的催产素mRNA的显着增加。雌激素和孕激素对视上核神经元催产素mRNA表达无影响。进行第三个实验以研究催产素分泌的持续增加，伴随或不伴随催产素神经元的细胞体的电活性增强，是否导致这些神经元中催产素mRNA含量的快速增加。为了刺激催产素的释放，雌性大鼠侧脑室注射吗啡后给予纳洛酮。纳洛酮增加血浆中催产素含量40倍，至少40分钟吗啡输注大鼠。纳洛酮对视上核标记细胞中催产素mRNA的含量无明显影响，对标记细胞的比例也无明显影响，说明催产素神经元在核、胞体和终末水平的基因表达机制存在差异。