Molecular characterization and functional expression of a substance P receptor from the sympathetic ganglion of Rana catesbeiana.
Molecular characterization and functional expression of a substance P receptor from the sympathetic ganglion of Rana catesbeiana.
复制标题
林蛙交感神经节 P 物质受体的分子特征和功能表达。
DOI：
10.1016/s0306-4522(97)00027-4
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发表时间：
1997
期刊：
影响因子：
3.3
通讯作者：
Krause,JE
中科院分区：
文献类型：
--
作者：
Simmons,MA;Brodbeck,RM;Karpitskiy,VV;Schneider,CR;Neff,DP;Krause,JE
P物质是中枢神经系统、自主神经系统和肠神经系统中重要的神经肽类神经递质。在交感神经节中，P物质被认为在调节突触传递中起作用。通过神经元刺激释放P物质或将P物质直接应用于神经节神经元增加神经元兴奋性。从牛蛙（Rana catesbeiana）交感神经节互补DNA文库中克隆了两栖类P物质受体互补DNA。推导的一级结构包含的功能指示七个跨膜结构域G蛋白偶联受体。推导的蛋白质序列与之前克隆的哺乳动物P物质受体具有69%的同一性。使用地高辛标记的互补RNA探针对牛蛙交感神经节进行的原位杂交分析表明，约75%的主要神经元显示高于背景水平的反应产物。放射性配体结合研究进行稳定转染细胞与[125 I]Tyr-1-P物质作为配体。P物质的IC_（50）为16 nM，激动剂效力曲线为P物质>神经激肽A>>神经激肽B。当应用于稳定转染的克隆细胞系时，三种速激肽增加细胞内钙的效力顺序为P物质>神经激肽A>>神经激肽B。该顺序的激动剂效力也保持了对完整牛蛙交感神经元中的M型钾电流的抑制。在阻断哺乳动物P物质受体的浓度下，非肽P物质拮抗剂CP-96345和RP-67580对牛蛙受体对P物质的反应几乎没有影响。总之，这些结果证明克隆的序列具有与P物质受体一致的特征和P物质受体的特征。参考牛蛙P物质受体的药理学和已知的哺乳动物速激肽受体的结构-活性关系的结果进行了讨论。
Substance P is an important neuropeptide neurotransmitter in the central, autonomic and enteric nervous systems. In sympathetic ganglia, substance P is thought to play a role in modulating synaptic transmission. Release of substance P by neuronal stimulation or direct application of substance P to ganglionic neurons increases neuronal excitability. An amphibian substance P receptor complementary DNA has been cloned and characterized from bullfrog, Rana catesbeiana, sympathetic ganglion complementary DNA libraries. The deduced primary structure contains features indicative of a seven transmembrane domain G-protein-coupled receptor. The deduced protein sequence shows 69% identity to previously cloned mammalian substance P receptors. In situ hybridization analysis performed on bullfrog sympathetic ganglia using digoxigenin-labelled complementary RNA probe demonstrated that approximately 75% of the principal neurons displayed reaction product above background levels. Radioligand binding studies were performed on stably transfected cells with [125I]Tyr-1-substance P as the ligand. Substance P had an IC50of 16nM and the agonist potency profile was substance P>neurokinin A>>neurokinin B. The order of potency for three tachykinins to increase intracellular calcium when applied to a stably transfected clonal cell line was substance P>neurokinin A>>neurokinin B. This order of agonist potency also held for inhibition of the M-type potassium current in intact bullfrog sympathetic neurons. The non-peptide substance P antagonists CP-96345 and RP-67580 at concentrations that block mammalian substance P receptors had little or no effect on the responses to substance P at the bullfrog receptor. Overall, these results demonstrate that the cloned sequence has the features consistent with and characteristic of a substance P receptor. The results are discussed with reference to the established pharmacology of the bullfrog substance P receptor and known structure–activity relationships of mammalian tachykinin receptors.