Metabolic Oscillations in Pancreatic Beta-Cells

描述（由申请方提供）：胰岛β细胞分泌的胰岛素为双相和脉冲式。第一时相分泌丧失和稳态血浆胰岛素波动是2型糖尿病的早期发展。尽管对葡萄糖刺激胰岛素分泌的信号级联进行了广泛的研究，但仍不清楚胰岛素分泌的振荡是如何产生的。我们的总体假设是胰岛代谢和胰岛素分泌的缓慢波动反映了糖酵解的缓慢波动。基于这一假设，已经开发了一个计算模型，其中由磷酸果糖激酶-1（PFK 1）介导的缓慢糖酵解振荡与由膜电活性和Ca 2+引起的快速振荡相互作用（“双振荡器模型”，或DOM）。 虽然DOM可以唯一地解释在体外和体内胰岛中观察到的振荡模式的多样性，但缺乏β细胞糖酵解振荡的直接证据，这留下了一个悬而未决的问题：代谢振荡是否是内在的，正如DOM预测的那样，或者是由Ca 2+振荡驱动的，正如竞争模型中提出的那样。要测试的DOM在代谢活动水平的预测，我们建议检查是否葡萄糖激酶和磷酸果糖-2-激酶/果糖-2，6-二磷酸酶（PFKFB），这是调节PFK 1，代表振荡参数，调节代谢分泌耦合在胰岛。使用新型代谢传感器的动态FRET成像将与膜片钳电生理学、数学建模和Ca 2+和NAD（P）H的光学测量相结合，以解决以下具体目标： 1.定义葡萄糖激酶激活的空间和时间特性，包括通过其结合伴侣PFKFB的调节，并评估它们各自对代谢振荡调节的贡献。 2.修改双振荡器模型，以纳入葡萄糖激酶/PFKFB相互作用和PFKFB产生的果糖-2，6-二磷酸诱导的糖酵解通量和胰岛振荡行为的变化。 3.确定Ca 2+是否驱动代谢的变化，代谢驱动Ca 2+，或两者都发生。 这些研究旨在为胰岛素分泌振荡的起源提供新的见解。2型糖尿病患者及其近亲的胰岛素波动受到抑制。了解这些缺陷可能会导致新的方法来诊断和治疗2型糖尿病和相关的代谢疾病。

DESCRIPTION (provided by applicant): Secretion of insulin from beta-cells of pancreatic islets is biphasic and pulsatile. Loss of first phase secretion and steady-state plasma insulin oscillations are an early development in Type 2 diabetes mellitus. Despite extensive study of the signaling cascades underlying glucose-stimulated insulin secretion, it remains unclear how oscillations in insulin secretion arise. Our overall hypothesis is that slow oscillations in islet metabolism and insulin secretion reflect slow oscillations in glycolyis. Based on this hypothesis, a computational model has been developed in which slow glycolytic oscillations mediated by phosphofructokinase-1 (PFK1) interact with fast oscillations arising from membrane electrical activity and Ca2+ (the 'Dual Oscillator Model', or DOM). Although the DOM can uniquely account for the diversity of oscillatory patterns observed in islets in vitro and in vivo, direct evidence for beta-cell glycolytic oscillations is lacking, leaving the open question of whether metabolic oscillations are intrinsic, as the DOM predicts, or driven by Ca2+ oscillations, as proposed in competing models. To test the predictions of the DOM at the level of metabolic activity, we propose to examine whether glucokinase and phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFKFB), which are regulators of PFK1, represent oscillatory parameters that modulate metabolism-secretion coupling in islets. Dynamic FRET imaging using novel metabolic sensors will be combined with patch-clamp electrophysiology, mathematical modeling, and optical measurements of Ca2+ and NAD(P)H, to address the following Specific Aims: 1. Define the spatial and temporal properties of glucokinase activation, including regulation via its binding partner PFKFB, and assess their respective contributions to the modulation of metabolic oscillations. 2. Modify the Dual Oscillator Model to incorporate changes in glycolytic flux and islet oscillatory behavior induced by glucokinase/PFKFB interaction and fructose-2,6-bisphosphate production by PFKFB. 3. Determine whether Ca2+ drives changes in metabolism, metabolism drives Ca2+, or both occur. These studies are designed to provide novel insights into the origin of oscillations in insulin secretion. Insulin oscillations are suppressed in patients with Type 2 diabetes mellitus and their near relatives. Understanding these defects may lead to new approaches for the diagnosis and treatment of Type 2 diabetes mellitus and related metabolic diseases.