Dopamine D2-like receptors (DRD2 and DRD4) in chickens: Tissue distribution, functional analysis, and their involvement in dopamine inhibition of pituitary prolactin expression
Dopamine D2-like receptors (DRD2 and DRD4) in chickens: Tissue distribution, functional analysis, and their involvement in dopamine inhibition of pituitary prolactin expression
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鸡中多巴胺 D2 样受体（DRD2 和 DRD4）：组织分布、功能分析及其参与多巴胺抑制垂体催乳素表达
DOI：
10.1016/j.gene.2018.01.087
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发表时间：
2018-04-20
期刊：
影响因子：
3.5
通讯作者：
Wang, Yajun
中科院分区：
文献类型：
--
作者：
Lv, Can;Mo, Chunheng;Wang, Yajun
多巴胺（DA）D_2样（和D_1样）受体介导鸟类垂体前叶和/或中枢神经系统的多巴胺作用。然而，关于鸟类D2样受体的结构、功能和表达的信息尚未完全表征。本研究利用RACE PCR技术从鸡脑中克隆了两个D2样受体（CDRD 2、CDRD 4）。克隆的CDRD 4是一个378个氨基酸的受体，其显示57%的氨基酸（a.a.）与小鼠DRD 4相同。如在哺乳动物中，两种cDRD 2同种型，cDRD 2L（长同种型，437 a.a.）和cDRD 2S（短亚型，408 a.a.），它们的第三胞内环不同，在鸡中被鉴定。利用基于细胞的荧光素酶报告基因分析或Western blot，我们证明了多巴胺和喹吡罗（一种D2样受体激动剂）可以剂量依赖性地激活cDRD 4、cDRD 2L和cDRD 2S，并且它们的激活抑制cAMP信号通路并刺激MAPK/ERK信号级联，表明它们是能够介导多巴胺作用的功能性受体。实时荧光定量PCR结果显示，CDRD 2和CDRD 4在鸡组织中广泛表达，在垂体前叶中表达丰富，其表达可能受其外显子1附近的启动子控制。与垂体中cDRD 2/cDRD 4的表达一致，DA或喹吡罗可部分抑制血管活性肠肽诱导的鸡垂体细胞催乳素的表达。总之，我们的数据证明了DRD 2和DRD 4在鸟类中的功能，并有助于揭示脊椎动物中DA/D2样受体系统的保守作用，例如其对垂体的作用。
Dopamine (DA) D2-like (and D1-like) receptors are suggested to mediate the dopamine actions in the anterior pituitary and/or CNS of birds. However, the information regarding the structure, functionality, and expression of avian D2-like receptors have not been fully characterized. In this study, we cloned two D2-like receptors (cDRD2, cDRD4) from chicken brain using RACE PCR. The cloned cDRD4 is a 378-amino acid receptor, which shows 57% amino acid (a.a.) identity with mouse DRD4. As in mammals, two cDRD2 isoforms, cDRD2L (long isoform, 437 a.a.) and cDRD2S (short isoform, 408 a.a.), which differ in their third intracellular loop, were identified in chickens. Using cell-based luciferase reporter assays or Western blot, we demonstrated that cDRD4, cDRD2L and cDRD2S could be activated by dopamine and quinpirole (a D2-like receptor agonist) dose-dependently, and their activation inhibits cAMP signaling pathway and stimulates MAPK/ERK signaling cascade, indicating that they are functional receptors capable of mediating dopamine actions. Quantitative real-time PCR revealed that cDRD2 and cDRD4 are widely expressed in chicken tissues with abundant expression noted in anterior pituitary, and their expressions are likely controlled by their promoters near exon 1, as demonstrated by dual-luciferase reporter assays in DF-1 cells. In accordance with cDRD2/cDRD4 expression in the pituitary, DA or quinpirole could partially inhibit vasoactive intestinal peptide-induced prolactin expression in cultured chick pituitary cells. Together, our data proves the functionality of DRD2 and DRD4 in birds and aids to uncover the conserved roles of DA/D2-like receptor system in vertebrates, such as its action on the pituitary.