Increased basal phosphorylation of the constitutively active serotonin 2C receptor accompanies agonist-mediated desensitization.
Increased basal phosphorylation of the constitutively active serotonin 2C receptor accompanies agonist-mediated desensitization.
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发表时间：
1995-08
影响因子：
3.6
通讯作者：
R. Westphal;J. Backstrom;E. sanders-Bush
中科院分区：
文献类型：
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作者：
R. Westphal;J. Backstrom;E. sanders-Bush
5-羟色胺（5-HT）2C受体是一种G蛋白偶联受体，表现出组成性受体活化，定义为信号转导途径的激动剂非依赖性受体活化。进行本研究以确定表达5-HT 2C受体的NIH/3 T3成纤维细胞是否表现出激动剂介导的磷酸肌醇水解的脱敏。此外，5-HT 2C受体特异性抗体用于确定在不存在激动剂的情况下5-HT 2C受体是否被磷酸化，以及用激动剂或反向激动剂处理是否改变了受体磷酸化。时间过程的基础和阿托伐他汀刺激的磷酸肌醇水解的研究表明，基础值以线性方式增加，而对血清素的反应在60分钟内达到稳定。此外，与血清素预处理导致随后确定的血清素剂量-反应曲线的快速移动。为了确定5-HT 2C受体的磷酸化状态，使用特异性抗体从32 P标记的成纤维细胞制备的裂解物中免疫沉淀受体。磷酸化的5-HT 2C受体是明显的基础条件下，和5-羟色胺治疗增加受体磷酸化。当单独加入时，反向激动剂米安色林对5-HT 2C受体磷酸化没有可检测的影响，但阻断了阿托伐他汀刺激的5-HT 2C受体磷酸化增加。本研究首次证明5-HT 2C受体在基础条件下磷酸化，而激动剂处理条件下磷酸化增加，导致受体信号转导脱敏。因此，这些研究表明，表现出高水平的组成型5-HT 2C受体活性的细胞系具有经历激动剂介导的脱敏的能力，这与G蛋白偶联受体调节的当前模型一致。
The 5-hydroxytryptamine (5-HT)2C receptor is a G protein-coupled receptor that exhibits constitutive receptor activation, defined as agonist-independent receptor activation of the signal transduction pathway. The present studies were performed to determine whether NIH/3T3 fibroblasts expressing the 5-HT2C receptor exhibited desensitization of agonist-mediated phosphoinositide hydrolysis. Furthermore, 5-HT2C receptor-specific antibodies were used to determine whether the 5-HT2C receptor was phosphorylated in the absence of agonist and whether treatment with an agonist or an inverse agonist altered receptor phosphorylation. Time course studies of basal and serotonin-stimulated phosphoinositide hydrolysis demonstrated that basal values increased in a linear manner, whereas the response to serotonin plateaued within 60 min. In addition, pretreatment with serotonin resulted in a rightward shift of the subsequently determined serotonin dose-response curve. To determine the phosphorylation state of the 5-HT2C receptor, specific antibodies were used to immunoprecipitate the receptor from lysates prepared from 32P-labeled fibroblasts. Phosphorylation of the 5-HT2C receptor was evident under basal conditions, and serotonin treatment increased receptor phosphorylation. The inverse agonist mianserin had no detectable effect on 5-HT2C receptor phosphorylation when added alone but blocked the serotonin-stimulated increase in 5-HT2C receptor phosphorylation. The present study is the first to demonstrate that the 5-HT2C receptor is phosphorylated under basal conditions and phosphorylation is increased by agonist treatment conditions that result in desensitization of receptor signaling. Thus, these studies demonstrate that a cell line exhibiting a high level of constitutive 5-HT2C receptor activity has the ability to undergo agonist-mediated desensitization, consistent with current models of G protein-coupled receptor regulation.