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Subcellular glucose exposure biases the spatial distribution of insulin granules in single pancreatic beta cells.

ABSTRACT: In living tissues, a cell is exposed to chemical substances delivered partially to its surface. Such a heterogeneous chemical environment potentially induces cell polarity. To evaluate this effect, we developed a microfluidic device that realizes spatially confined delivery of chemical substances at subcellular resolution. Our microfluidic device allows simple setup and stable operation for over 4 h to deliver chemicals partially to a single cell. Using the device, we showed that subcellular glucose exposure triggers an intracellular [Ca(2+)] change in the ?-cells. In addition, the imaging of a cell expressing GFP-tagged insulin showed that continuous subcellular exposure to glucose biased the spatial distribution of insulin granules toward the site where the glucose was delivered. Our approach illustrates an experimental technique that will be applicable to many biological experiments for imaging the response to subcellular chemical exposure and will also provide new insights about the development of polarity of ?-cells.

SUBMITTER: Terao K

PROVIDER: S-EPMC3927211 | biostudies-other | 2014

REPOSITORIES: biostudies-other

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Subcellular glucose exposure biases the spatial distribution of insulin granules in single pancreatic beta cells.

Terao Kyohei K Gel Murat M Okonogi Atsuhito A Fuke Ariko A Okitsu Teru T Tada Takashi T Suzuki Takaaki T Nagamatsu Shinya S Washizu Masao M Kotera Hidetoshi H

Scientific reports 20140218

In living tissues, a cell is exposed to chemical substances delivered partially to its surface. Such a heterogeneous chemical environment potentially induces cell polarity. To evaluate this effect, we developed a microfluidic device that realizes spatially confined delivery of chemical substances at subcellular resolution. Our microfluidic device allows simple setup and stable operation for over 4 h to deliver chemicals partially to a single cell. Using the device, we showed that subcellular glu ...[more]

PMID: 24535122

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Project description:Aims/introductionSrc, a non-receptor tyrosine kinase, regulates a wide range of cellular functions, and hyperactivity of Src is involved in impaired glucose metabolism in pancreatic ?-cells. However, the physiological role of Src in glucose metabolism in normal, unstressed ?-cells remains unclear. In the present study, we investigated the role of Src in insulin secretion and glucose metabolism.Materials and methodsSrc was downregulated using small interfering ribonucleic acid in INS-1 cells, and glucose-induced insulin secretion, adenosine triphosphate content, intracellular calcium concentration, glucose utilization and glucokinase activity were measured. Expression levels of messenger ribonucleic acid and protein of glucokinase were examined by semiquantitative real-time polymerase chain reaction and immunoblotting, respectively. Cells were fractionated by digitonin treatment, and subcellular localization of glucokinase was examined by immunoblotting. Interaction between glucokinase and neuronal nitric oxide synthase was estimated by immunoprecipitation.ResultsIn Src downregulated INS-1 cells, glucose-induced insulin secretion was impaired, whereas insulin secretion induced by high K(+) was not affected. Intracellular adenosine triphosphate content and elevation of intracellular calcium concentration by glucose stimulation were suppressed by Src downregulation. Src downregulation reduced glucose utilization in the presence of high glucose, which was accompanied by a reduction in glucokinase activity without affecting its expression. However, Src downregulation reduced glucokinase in soluble, cytoplasmic fraction, and increased it in pellet containing intaracellular organelles. In addition, interaction between glucokinase and neuronal nitric oxide synthase was facilitated by Src downregulation.ConclusionsSrc plays an important role in glucose-induced insulin secretion in pancreatic ?-cells through maintaining subcellular localization and activity of glucokinase.

Dataset Information

Subcellular glucose exposure biases the spatial distribution of insulin granules in single pancreatic beta cells.

Publications

Subcellular glucose exposure biases the spatial distribution of insulin granules in single pancreatic beta cells.

Similar Datasets

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