Metabolic Oscillations in Pancreatic Beta-Cells

描述(申请人提供)：从胰岛的β细胞分泌的胰岛素是双相的和搏动性的。第一时相分泌丧失和稳态血浆胰岛素振荡是2型糖尿病的早期发展。尽管对葡萄糖刺激的胰岛素分泌背后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，胰岛新陈代谢和胰岛素分泌的缓慢振荡反映了糖酵素的缓慢振荡。基于这一假设，建立了一个计算模型，在该模型中，由磷酸果糖激酶-1(PFK1)介导的缓慢糖酵解振荡与由膜电活动和钙离子引起的快速振荡相互作用(‘双振荡器模型’，或DOM)。 尽管DOM可以唯一地解释在体外和体内观察到的胰岛振荡模式的多样性，但缺乏关于β细胞糖酵解振荡的直接证据，留下了一个悬而未决的问题，即代谢振荡是像DOM预测的那样是内在的，还是像竞争模型中提出的那样由钙振荡驱动的。为了在代谢活动水平上测试DOM的预测，我们建议检查作为PFK1的调节者的葡萄糖激酶和phosphofructo-2-kinase/fructose-2，6-bisphosphatase是否代表调节胰岛新陈代谢-分泌耦合的振荡参数。使用新型代谢传感器的动态FRET成像将与膜片钳电生理学、数学建模以及对钙和NAD(P)H的光学测量相结合，以解决以下具体目标： 1.确定葡糖激酶激活的空间和时间特性，包括通过其结合伙伴PFKFB的调节，并评估它们在代谢振荡调节中的各自贡献。 2.修改双振子模型，考虑葡萄糖激酶/PFKFB相互作用引起的糖酵解通量和胰岛振荡行为以及PFKFB产生2，6-二磷酸果糖的变化。 3.确定钙是驱动新陈代谢的变化，还是代谢驱动钙的变化，还是两者兼而有之。 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振荡受到抑制。了解这些缺陷可能会导致诊断和治疗2型糖尿病和相关代谢性疾病的新方法。

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.