BIOPHYSICS AND ELECTROPHYSIOLOGY OF BETA CELLS--DEFECTS IN CELL SIGNALLING

该项目的总体目标是确定和描述 相关β细胞的电生理功能异常 患有糖尿病或因接触高浓度葡萄糖而引起的。我们 将检验以下假设，即这些条件与特定的 调节去极化的离子通道中的缺陷和 β细胞膜或钙稳态过程的复极化 在β细胞内。Fura-2荧光技术和数字化技术 图像分析将结合整个细胞和 穿孔贴片记录和单通道测量以研究 单个胰岛素分泌细胞内的钙信号模式和 细胞群。实验将在分散的胰岛β细胞中进行。 从正常动物或有β细胞功能障碍的动物分离 由输注葡萄糖引起的或由以下原因引起的自发性糖尿病 β细胞质量的减少(GK大鼠)或自身免疫的β细胞 破坏(易患糖尿病的BB/WOR大鼠)。的各个组件 正常的β细胞和β细胞衍生的株系与 将研究钙信号的调节，包括钙通道，K 细胞内钙稳态的各种途径和机制 如钙储存库、泵和交换机制。我们将尝试 确定这些因素如何促进脑内钙信号的产生 β细胞对包括葡萄糖、氨基酸在内的各种调节剂的反应 酸、磺脲类和神经递质。一种分泌β-的胰岛素- 细胞系，保留了许多分泌物的BetaTC3细胞系 正常β细胞的特性已被证明可以增加其 低糖孵育后对分泌刺激的分泌反应 与高糖相比。与此相关的电生理学 贝塔细胞功能的增加将被研究。最后是 表达的新离子通道基因的电生理特性 将使用异源表达系统对β细胞进行鉴定。 使用反义寡核苷酸和抗体对这些基因的影响 对细胞电生理和钙信号的影响将被确定。它 预计这些研究将为深入了解 离子通道在正常β细胞胰岛素分泌调节中的作用 在早期IDDM的生理和分泌功能障碍中。

The overall goal of this project is to identify and characterize abnormalities in the electrophysiological function of beta-cells associated with diabetes or induced by exposure to high concentrations of glucose. We will test the hypothesis that these conditions are associated with specific defects in the ion channels which regulate depolarization and repolarization of the beta-cell membrane or of Ca2+ homeostatic processes within the beta-cell. The techniques of fura-2 fluorimetry and digital image analysis will be applied in combination with whole cell and perforated patch recordings and single channel measurements to study the patterns of Ca2+ signalling within individual insulin secreting cells and cell groups. Experiments will be conducted in dispersed islet beta-cells isolated from normal animals or from animals with beta-cell dysfunction induced by infusion of glucose or with spontaneous diabetes resulting from a reduction in beta-cell mass (the GK rat) or autoimmune beta-cell destruction (the diabetes prone BB/Wor rat). The various components of the normal beta-cell and beta-cell derived lines which are concerned in the regulation of Ca2+ signalling will be studied including Ca2+ channels, K+ channels and various types of intracellular Ca2+ homeostatic mechanisms such as Ca2+ stores, pumps and exchange mechanisms. We shall attempt to determine how these factors contribute to the production of Ca2+ signals in beta-cells in response to various regulators including glucose, amino acids, sulfonuylureas and neurotransmitters. An insulin secreting beta- cell line, the betaTC3 cell line which retains many of the secretory properties of normal beta-cells has been demonstrated to increase its secretory response to a secretory stimulus after incubation in low glucose compared with high glucose. The electrophysiologic correlates of this increase in beta-cell function will be studied. Finally the electrophysiologic properties of novel ion channel genes expressed in the beta-cell will be characterized using heterologous expression systems. Using antisense oligonucleotides and antibodies the effects of these genes on cellular electrophysiology and Ca2+ signalling will be determined. It is anticipated that these studies will provide insights into the role of the ion channels in the regulation of insulin secretion in normal beta-cell physiology and in the secretory dysfunction of early IDDM.