Gao N, Hu H-Z, Zhu M X, Fang X, Liu S, Gao C, Wood J D
Department of Physiology and Cell Biology, The Ohio State University, Columbus, 43210, USA.
Neurogastroenterol Motil. 2006 Apr;18(4):316-23. doi: 10.1111/j.1365-2982.2005.00754.x.
Electrophysiological recording methods provided evidence for presynaptic release of ATP from enteric neurones and postganglionic sympathetic fibres in the enteric nervous system (ENS) of guinea-pig intestine (J Physiol Lond 2003; 550: 493-504). The released ATP acted at postsynaptic P2Y(1) receptors to evoke slow synaptic excitation in neurones in the submucosal division of the ENS. Here, we report the cloning and characterization of the P2Y(1) receptor, which was found in the guinea-pig submucosal layer. A 1178 bp cDNA clone was isolated from guinea-pig submucosal RNA by reverse transcription polymerase chain reaction (RT-PCR). The cDNA contained an open-reading frame of 1119 bp, encoding a 373 amino acid polypeptide of the same length and with 95% identity to the human P2Y(1) receptor. Stable expression of the guinea-pig cDNA in human embryonic kidney (HEK)293 cells was accompanied by a marked increase in sensitivity for elevation of free intracellular calcium evoked by ATP or related nucleotides. The potency order for ATP and its analogues was: 2-methio-adenosine diphosphate > 2-methio-adenosine triphosphate > ADP > ATP-gamma-S > ATP. The selective P2Y(1) receptor antagonist, MRS2179, was a competitive antagonist for the receptor with a pA(2) value of 6.5. The results add to existing evidence for expression of a functional P2Y(1) purinergic receptor in neurones of the submucosal division of the ENS.
电生理记录方法为豚鼠肠道肠神经系统（ENS）中肠神经元和节后交感神经纤维突触前释放ATP提供了证据（《伦敦生理学杂志》2003年；550:493 - 504）。释放的ATP作用于突触后P2Y(1)受体，在ENS黏膜下部分的神经元中引发缓慢的突触兴奋。在此，我们报告了在豚鼠黏膜下层发现的P2Y(1)受体的克隆和特性。通过逆转录聚合酶链反应（RT-PCR）从豚鼠黏膜下RNA中分离出一个1178 bp的cDNA克隆。该cDNA包含一个1119 bp的开放阅读框，编码一个长度相同的373个氨基酸的多肽，与人P2Y(1)受体具有95%的同一性。豚鼠cDNA在人胚肾（HEK）293细胞中的稳定表达伴随着对ATP或相关核苷酸引起的细胞内游离钙升高的敏感性显著增加。ATP及其类似物的效价顺序为：2-甲硫基二磷酸腺苷＞2-甲硫基三磷酸腺苷＞二磷酸腺苷＞ATP-γ-S＞ATP。选择性P2Y(1)受体拮抗剂MRS2179是该受体的竞争性拮抗剂，pA(2)值为6.5。这些结果为ENS黏膜下部分神经元中功能性P2Y(1)嘌呤能受体的表达增添了现有证据。
