FUNCTIONAL EXPRESSION OF HUMAN 5-HT1A-RECEPTORS AND DIFFERENTIAL COUPLING TO 2ND MESSENGERS IN CHO CELLS
FUNCTIONAL EXPRESSION OF HUMAN 5-HT1A-RECEPTORS AND DIFFERENTIAL COUPLING TO 2ND MESSENGERS IN CHO CELLS
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DOI：
10.1007/bf00165293
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发表时间：
1992-08-01
影响因子：
3.6
通讯作者：
MIDDLETON, JP
中科院分区：
文献类型：
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作者：
RAYMOND, JR;ALBERS, FJ;MIDDLETON, JP
研究了在CHO细胞中稳定表达的克隆的人5-HT 1A受体的信号转导联系。表达900 +/- 36 fmol 5-HT 1A受体/mg蛋白的转染克隆细胞系（命名为CHO-5-HT 1A/WT-27）通过偶联几种第二信使途径对5-HT和/或8-OH-DPAT产生应答。5-HT 1A受体以剂量依赖性方式抑制但不刺激膜腺苷酸环化酶活性和全细胞cAMP蓄积（对于5-HT，IC 50分别为146 +/- 27和55 +/- 12 nM）。受体的激活与其他信号转导联系有关：（i）肌醇磷酸的水解增加40 - 50%（对于5-HT，EC 50 = 1.33 +/- 0.15-mu-M），（ii）胞质Ca 2+水平的瞬时升高（在1 - 100 μ M 5-HT下明显），其不受EGTA对细胞外Ca 2+的螯合作用的影响，和（iii）用钙离子载体A23187或通过激活内源性凝血酶或P2嘌呤能受体来增加[H-3]-花生四烯酸的释放（对于5-HT，EC 50 = 1.22 +/- 0.17-mu-M）。该途径可能是在具有高浓度的几种神经递质、激素或autacoids的解剖区域中的信号传导的放大机制，例如在神经元连接处或血小板聚集区域附近。所有连接对百日咳毒素预处理敏感（所有途径的IC 50几乎等于0.5 - 0.6 ng/ml x 4.5 h），表明G（i）蛋白参与这些信号转导途径。在单细胞系统中与不同的信号转导途径偶联可能是经典抑制腺苷酸环化酶的受体如5-HT 1A受体的共同特征。
The signal transduction linkages of the cloned human 5-HT1A receptor as expressed stably in CHO cells were studied. A transfected clonal cell line which expresses 900 +/- 36 fmol 5-HT1A receptor/mg protein (designated CHO-5-HT1A/WT-27) responded to 5-HT and/or 8-OH-DPAT by coupling to several second messenger pathways. The 5-HT1A receptor inhibited, but did not stimulate, membrane adenylyl cyclase activity and whole cell cAMP accumulation in a dose-dependent manner (for 5-HT, IC50 = 146 +/- 27 and 55 +/- 12 nM, respectively). Activation of the receptor was associated with other signal transduction linkages: (i) a 40 - 50% increase in hydrolysis of inositol phosphates (for 5-HT, EC50 = 1.33 +/- 0.15-mu-M for 5-HT), (ii) a transient elevation of cytosolic Ca2+ levels (apparent at 1 - 100-mu-M 5-HT) which was not affected by chelation of extracellular Ca2+ by EGTA, and (iii) an augmentation of [H-3]-arachidonic acid release pharmacologically with the calcium ionophore A23187 or by activation of endogenous thrombin or P2 purinergic receptors (for 5-HT, EC50 = 1.22 +/- 0.17-mu-M). This pathway may be an amplification mechanism for signaling in anatomic regions with high concentrations of several neuro-transmitters, hormones or autacoids, such as at neuronal junctions or near areas of platelet aggregation. All linkages were sensitive to pertussis toxin pre-treatment (IC50 almost-equal-to 0.5 - 0.6 ng/ml x 4.5 h for all pathways), suggesting the involvement of G(i) protein(s) in these signal transduction pathways. Coupling to varied signal transduction pathways in a single cell system may be a common feature of receptors which classically inhibit adenylyl cyclase such as the 5-HT1A receptor.