Generation of transgenic mice to investigate the expression patterns, protein interactions and physiological functions of P2X4, P2X6 and P2X7 receptors

P2X受体(P2XRs)是ATP激活的钙离子通透性阳离子通道，已被证明影响多种细胞功能。通常，七个已知的P2X亚基(P2X1-P2X7)中的一个以上在给定的组织或细胞类型中表达。在天然组织中，由于激动剂的快速分解代谢，不同的P2XR亚型和代谢性的P2Y受体同时激活，以及缺乏亚型特异性配体，P2XRs的功能特征一直受到阻碍。因此，P2XRs在体内的功能在很大程度上是未知的。小鼠单个P2X基因的缺失揭示了P2XRs参与了成骨、白细胞产生、排尿、疼痛传递和生殖等多种生理过程。尽管P2XRs在大脑中广泛表达，但到目前为止，在可用的基因敲除动物中还没有发现对运动功能或行为的影响。然而，电生理学研究表明，P2XRs在突触传递的调节和神经病理性疼痛的发生中起着重要作用。在这个项目中，我们建议建立在其内源性启动子下过度表达表位标记的显性阴性的P2X亚基的转基因小鼠，以研究P2X4、P2X6和P2X7亚基在哺乳动物中枢神经系统神经胶质细胞和神经元中的表达模式、功能和蛋白质相互作用。

P2X receptors (P2XRs) are ATP-activated Ca2+-permeable cation channels that have been shown to influence a variety of cell functions. Usually more than one of the seven known P2X subunits (P2X1-P2X7) are expressed in a given tissue or cell type. The functional characterization of P2XRs in native tissues has been hampered by the rapid catabolism of the agonist, the simultaneous activation of different P2XR subtypes and metabotropic P2Y receptors as well as the lack of subtype-specific ligands. Therefore, the in vivo functions of P2XRs are largely unknown. Deletion of single P2X genes in mice revealed the involvement of P2XRs in such diverse physiological processes as osteogenesis, interleukine production, micturition, pain transmission and reproduction. Despite the wide-spread expression of P2XRs in brain, no effects on motor function or behavior have been identified so far in the available knockout animals. However, electrophysiological studies indicate an important role of P2XRs in the modulation of synaptic transmission and the development of neuropathic pain. In this project, we propose to generate transgenic mice that over-express epitope-tagged and dominant-negative P2X subunits under their endogenous promoters to investigate the expression patterns, functions and protein interactions of P2X4, P2X6 and P2X7 subunits in glia cells and neurons of the mammalian central nervous system.