Biodesign Center for Bioelectronics and Biosensors, Arizona State University , Tempe, Arizona 85287, United States.
Department of Chemistry, Beijing Key Laboratory for Microanalytical Methods and Instrumentation, Tsinghua University , Beijing 100084, China.
Anal Chem. 2016 Dec 6;88(23):11498-11503. doi: 10.1021/acs.analchem.6b02677. Epub 2016 Nov 15.
G protein-coupled receptors (GPCRs) are the largest protein family for cell signal transduction, and most of them are crucial drug targets. Conventional label-free assays lack the spatial information to address the heterogeneous response from single cells after GPCRs activation. Here, we reported a GPCRs study in live cells using plasmonic-based electrochemical impedance microscopy. This label-free optical imaging platform is able to resolve responses from individual cells with subcellular resolution. Using this platform, we studied the histamine mediated GPCRs activation and revealed spatiotemporal heterogeneity of cellular downstream responses. Triphasic responses were observed from individual HeLa cells upon histamine stimulation. A quick peak P1 in less than 10 s was attributed to the GPCRs triggered calcium release. An inverted P2 phase within 1 min was attributed to the alternations of cell-matrix adhesion after the activation of Protein Kinase C (PKC). The main peak (P3) around 3-6 min after the histamine treatment was due to dynamic mass redistribution and showed a dose-dependent response with a half-maximal effective concentration (EC) of 3.9 ± 1.2 μM. Heterogeneous P3 responses among individual cells were observed, particularly at high histamine concentration, indicating diverse histamine H1 receptor expression level in the cell population.
G 蛋白偶联受体（GPCRs）是细胞信号转导中最大的蛋白质家族，其中大多数是至关重要的药物靶点。传统的无标记检测方法缺乏空间信息，无法解决 GPCR 激活后单个细胞的异质反应。在这里，我们使用基于等离子体的电化学阻抗显微镜报告了活细胞中的 GPCRs 研究。这个无标记的光学成像平台能够以亚细胞分辨率解析来自单个细胞的反应。使用该平台，我们研究了组胺介导的 GPCRs 激活，并揭示了细胞下游反应的时空异质性。组胺刺激单个 HeLa 细胞时观察到三相反应。不到 10 秒的快速峰 P1 归因于 GPCR 触发的钙释放。1 分钟内的倒置 P2 相归因于蛋白激酶 C（PKC）激活后细胞-基质粘附的改变。组胺处理后 3-6 分钟左右的主要峰（P3）是由于动态质量重新分布，并且表现出与半最大有效浓度（EC）为 3.9 ± 1.2 μM 的剂量依赖性反应。在高组胺浓度下观察到单个细胞之间的异质 P3 反应，表明细胞群体中存在不同的组胺 H1 受体表达水平。
