Specific expression of an oxytocin-enhanced cyan fluorescent protein fusion transgene in the rat hypothalamus and posterior pituitary
Specific expression of an oxytocin-enhanced cyan fluorescent protein fusion transgene in the rat hypothalamus and posterior pituitary
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DOI：
10.1677/joe-09-0289
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发表时间：
2010-03-01
影响因子：
4
通讯作者：
Ueta, Yoichi
中科院分区：
文献类型：
--
作者：
Katoh, Akiko;Fujihara, Hiroaki;Ueta, Yoichi
我们已经产生了携带催产素（OXT）增强的青色荧光蛋白（eCFP）融合转基因的大鼠，该转基因是由先前显示在转基因小鼠中忠实表达的小鼠结构设计的。原位杂交组织化学结果显示，Oxt-eCfp融合基因在视上核（SON）和室旁核（PVN）中表达。eCFP仅在SON、PVN、正中隆起内层和垂体后叶（PP）可见荧光。在体外制备中，从SON和轴突末端新解离的细胞显示出清晰的eCFP荧光。OXT和精氨酸抗利尿素（AVP）的免疫组化结果显示，在SON和PVN中，eCFP荧光与OXT免疫荧光共定位，而与AVP免疫荧光不共定位。虽然转基因大鼠SON和PVN中Oxt-eCfp融合基因的表达水平存在较大的差异，但长期盐负荷后，SON和PVN中eCFP荧光明显增强，而PP中eCFP荧光减弱。在非转基因和转基因大鼠的SON和PVN中，慢性盐负荷后Oxt基因的表达均显著增加。与野生型动物相比，脱水和盐负荷的雄性和雌性转基因大鼠血浆渗透压、钠浓度、OXT和AVP水平均无显著差异，表明融合基因的表达没有干扰任何生理过程。这些结果表明，我们的新转基因大鼠是鉴定产生oxt的神经元及其终端的有价值的新工具。内分泌学杂志（2010）204,275-285
We have generated rats bearing an oxytocin (OXT)-enhanced cyan fluorescent protein (eCFP) fusion transgene designed from a murine construct previously shown to be faithfully expressed in transgenic mice. In situ hybridisation histochemistry revealed that the Oxt-eCfp fusion gene was expressed in the supraoptic nucleus (SON) and the paraventricular nucleus (PVN) in these rats. The fluorescence emanating from eCFP was observed only in the SON, the PVN, the internal layer of the median eminence and the posterior pituitary (PP). In in vitro preparations, freshly dissociated cells from the SON and axon terminals showed clear eCFP fluorescence. Immunohistochemistry for OXT and arginine vasopressin (AVP) revealed that the eCFP fluorescence co-localises with OXT immunofluorescence, but not with AVP immunofluorescence in the SON and the PVN. Although the expression levels of the Oxt-eCfp fusion gene in the SON and the PVN showed a wide range of variations in transgenic rats, eCFP fluorescence was markedly increased in the SON and the PVN, but decreased in the PP after chronic salt loading. The expression of the Oxt gene was significantly increased in the SON and the PVN after chronic salt loading in both non-transgenic and transgenic rats. Compared with wild-type animals, euhydrated and salt-loaded male and female transgenic rats showed no significant differences in plasma osmolality, sodium concentration and OXT and AVP levels, suggesting that the fusion gene expression did not disturb any physiological processes. These results suggest that our new transgenic rats are a valuable new tool to identify OXT-producing neurones and their terminals. Journal of Endocrinology (2010) 204, 275-285