MUSCARINIC RECEPTOR-OPERATED CA2+ INFLUX IN TRANSFECTED FIBROBLAST CELLS IS INDEPENDENT OF INOSITOL PHOSPHATES AND RELEASE OF INTRACELLULAR CA2+
MUSCARINIC RECEPTOR-OPERATED CA2+ INFLUX IN TRANSFECTED FIBROBLAST CELLS IS INDEPENDENT OF INOSITOL PHOSPHATES AND RELEASE OF INTRACELLULAR CA2+
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DOI：
10.1073/pnas.89.2.509
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发表时间：
1992-01-15
影响因子：
11.1
通讯作者：
WESS, J
中科院分区：
文献类型：
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作者：
FELDER, CC;POULTER, MO;WESS, J
受体介导的细胞质钙浓度变化通过细胞内钙库的释放或质膜通道的打开而发生，从而允许钙从细胞外液流入。卡巴胆碱，毒蕈碱受体激动剂，刺激钙内流和肌醇1，4，5-三磷酸（InsP 3）介导的细胞内钙释放A9成纤维细胞表达m3毒蕈碱受体克隆。即使用佛波醇12-肉豆蔻酸酯13-乙酸酯预处理细胞后，钙内流仍持续存在，这完全阻止了肌醇磷酸和细胞内钙水平的升高。钙内流被二价阳离子阻断，但不受电压依赖性钙通道或高钾去极化抑制剂的影响，表明这些细胞中存在受体操纵和电压不敏感的钙通道。钙内流没有刺激的cAMP类似物或花生四烯酸的加入。为了研究G蛋白可能参与M3受体激活的钙离子内流，两个嵌合M2和M3毒蕈碱受体在A9细胞中表达，其中第三个细胞质环（毒蕈碱受体的G蛋白偶联选择性的主要结构决定因素）已在M2受体（对钙离子内流没有影响）和M3受体之间交换。钙离子内流被发现与第三细胞质环以外的m3毒蕈碱受体的结构成分。
Receptor-mediated changes in cytoplasmic calcium concentrations occur either through release from intracellular calcium stores or hy the opening of channels in the plasma membrane, allowing influx of calcium from the extracellular fluid. Carbachol, a muscarinic receptor agonist, stimulated both calcium influx and inositol 1,4,5-trisphosphate (InsP3)-mediated intracellular calcium release in A9 fibroblast cells expressing a m3 muscarinic receptor clone. The calcium influx persisted even after pretreatment of cells with phorbol 12-myristate 13-acetate, which completely prevented the rise in inositol phosphates and intracellular calcium levels. The calcium influx was blocked by divalent cations but was not affected by inhibitors of voltage-dependent calcium channels or high potassium depolarization, indicating the presence of a receptor-operated and voltage-insensitive calcium channel in these cells. Calcium influx was not stimulated by the addition of cAMP analogs or arachidonic acid. To examine the possible involvement of G proteins in m3 receptor-activated calcium influx, two chimeric m2 and m3 muscarinic receptors were expressed in A9 cells in which the third cytoplasmic loop (the primary structural determinant in G protein coupling selectivity of muscarinic receptors) had been exchanged between the m2 receptor, which has no effect on calcium influx, and the m3 receptor. Calcium influx was found to be associated with a structural component of the m3 muscarinic receptor other than the third cytoplasmic loop.