Exploring G Protein-Coupled Receptor Signaling in Primary Pancreatic Islets.

IF 3.7 3区生物学 Q1 BIOCHEMICAL RESEARCH METHODS

Biological Procedures Online Pub Date : 2020-02-15 eCollection Date: 2020-01-01 DOI:10.1186/s12575-019-0116-y

Juliane Röthe, Robert Kraft, Torsten Schöneberg, Doreen Thor

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引用次数: 7

Abstract

Background: Targeting G protein-coupled receptors (GPCRs) in pancreatic cells is feasible to modulate glucose-induced insulin secretion. Because pancreatic islets consist of several cell types and GPCRs can couple to more than one G-protein family, results obtained in pancreatic cell lines do not always match the response in primary cells or intact islets. Therefore, we set out to establish a protocol to analyze second messenger activation in mouse pancreatic islets.

Results: Activation of Gq/11-coupled receptor expressed in primary β cells increased the second messenger IP1 in an accumulation assay. Applying a Gq/11 protein inhibitor completely abolished this signal. Activation of the V1 vasopressin and ghrelin receptors, predominantly expressed in the less abundant alpha and delta cells, was not sufficient to induce a significant IP1 increase in this assay. However, fura-2-based fluorescence imaging showed calcium signals upon application of arginine vasopressin or ghrelin within intact pancreatic islets. Using the here established protocol we were also able to determine changes in intracellular cAMP levels induced by receptors coupling to Gs and Gi/o proteins.

Conclusions: Detection of the second messengers IP1, cAMP, and calcium, can be used to reliably analyze GPCR activation in intact islets.

Abstract Image

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原发性胰岛中G蛋白偶联受体信号传导的研究。

背景:靶向胰腺细胞中的G蛋白偶联受体(gpcr)调节葡萄糖诱导的胰岛素分泌是可行的。由于胰岛由几种细胞类型组成，并且gpcr可以与不止一种g蛋白家族偶联，因此在胰腺细胞系中获得的结果并不总是与原代细胞或完整胰岛的反应相匹配。因此，我们着手建立一种方案来分析小鼠胰岛中的第二信使激活。结果:在积累实验中，激活原代β细胞中表达的Gq/11偶联受体增加了第二信使IP1。使用Gq/11蛋白抑制剂完全消除了该信号。V1抗利尿激素和胃饥饿素受体(主要表达于较少的α和δ细胞)的激活不足以诱导IP1显著增加。然而，在完整的胰岛内应用精氨酸加压素或胃饥饿素后，fura-2荧光成像显示钙信号。使用这里建立的方案，我们还能够确定由受体偶联到Gs和Gi/o蛋白引起的细胞内cAMP水平的变化。结论:检测第二信使IP1、cAMP和钙可以可靠地分析完整胰岛中GPCR的激活情况。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Biological Procedures Online 生物-生化研究方法

CiteScore

10.50

自引率

0.00%

发文量

审稿时长

>12 weeks

期刊介绍： iological Procedures Online publishes articles that improve access to techniques and methods in the medical and biological sciences. We are also interested in short but important research discoveries, such as new animal disease models. Topics of interest include, but are not limited to: Reports of new research techniques and applications of existing techniques Technical analyses of research techniques and published reports Validity analyses of research methods and approaches to judging the validity of research reports Application of common research methods Reviews of existing techniques Novel/important product information Biological Procedures Online places emphasis on multidisciplinary approaches that integrate methodologies from medicine, biology, chemistry, imaging, engineering, bioinformatics, computer science, and systems analysis.