Stimulus secretion coupling in pancreatic beta-cells

β细胞： 我们在过去几年的主要活动之一是开发了一个全面的膜电位和钙振荡模型，其时间尺度从几秒到几分钟不等。这些都会导致相应的胰岛素分泌振荡。该模型的基本假设是，较快的振荡(几十秒)源于钙对离子通道的反馈，可能是钙激活的钾(K(Ca)通道)和ATP依赖的钾(K(ATP)通道)，而较慢的振荡(5分钟)来自代谢的振荡。代谢振荡通过K(ATP)通道转化为电振荡。因此，该模型由电子振荡器(EO)和代谢振荡器(MO)组成，称为双振荡器模型(DOM)。在我们的模型中，MO是糖酵解振荡器，但如果代谢振荡发生在其他地方，如线粒体，系统的许多特征仍然存在。 K(ATP)通道具有临床意义，因为它们是胰岛素刺激药物的一线靶点，如用于治疗2型糖尿病的磺脲类药物甲苯磺脲和格列本脲。严重的K(ATP)功能获得突变是新生儿糖尿病的主要原因，而在全基因组关联研究(GWAS)中，适度的功能获得突变与较温和但更常见的疾病-成人2型糖尿病有关。相反，K(ATP)功能丧失突变是家族性高胰岛素血症的主要原因，这是一种在儿童中发现的遗传性疾病，在正常或低血糖面前，β细胞持续电活动并分泌胰岛素，导致危及生命的低血糖。 有关胰岛素分泌振荡对健康和疾病的重要性的综述，请参阅2015年报告中的参考文献#3。 多年来，我们积累了大量支持该模型的间接证据，但我们认为重要的是设计一个中心特征的直接测试，即糖酵解振荡。我们的实验合作者通过修饰丙酮酸激酶(PK)开发了一种基于FRET的传感器。PK结合果糖-1，6-二磷酸(FBP)，这是一种关键的糖酵解代谢产物，DOM预测它会振荡。以前的一篇论文证实，传感器(PKAR，丙酮酸激酶活性报告)确实振荡。在当前时期，通过与膜电位同步记录来更严格地探测振荡的特征，以确定它们之间的相位关系。实验(参考文献)#1)再次证实，PKAR是振荡的，而且即使在钙离子不振荡的情况下也可以这样做，正如模型所预测的那样。然而，相位关系与模型预测不一致：PKAR在振荡的活跃期下降，而由于糖酵解活动的脉冲，预测在整个活跃期都很高。另外两种代谢物NAD(P)H和ATP也被成像，并显示出类似的模式。 预测和实验之间的差异是惊人的，但只需要对模型进行一次重大修改就能解决。添加了一种先前已知但被忽视的钙加速柠檬酸的作用。这增加了线粒体对丙酮酸的消耗，以足够快的速度消耗胞浆中的FBP，以克服糖酵解中FBP产生的激增。为了与ATP在活动阶段也下降的观察结果相匹配，有必要另外假设活动阶段钙泵对ATP的消耗足够大，足以克服由于柠檬酸循环的刺激而导致的ATP产生的增加。这种对ATP的强烈消耗有助于加速活动阶段的终止，因为它允许K(ATP)通道重新开放。还需要进一步的工作，以检验经修订的模型产生的新预测，并充分了解这一更复杂安排的好处。 Alpha单元格： 胰腺细胞中高血糖素分泌的调节还不是很清楚。已有研究表明，葡萄糖可直接通过细胞内的内在机制或通过外在机制间接抑制胰升糖素的分泌。我们之前描述了一个分离的胰腺α细胞的数学模型，并用它来研究调节胰升糖素分泌的可能的内在机制。我们证明葡萄糖可以通过依赖于ATP的钾通道(K(ATP))和存储操作电流(SOC)来抑制胰高血糖素的分泌。现在，我们开发了一个胰岛模型，它将先前发表的α细胞和β细胞的数学模型与一个新的三角洲细胞模型结合起来，并用它来探索胰岛素和生长抑素对胰升糖素分泌的影响。我们证明，该模型可以重现实验观察结果，即即使旁分泌调节剂不再作用于细胞，葡萄糖的抑制作用仍然存在。我们演示了旁分泌相互作用如何使细胞同步化，从而在胰升糖素和生长抑素的分泌中产生搏动性振荡，或者无法做到这一点。该模型还可以解释一个矛盾的观察，即胰高血糖素可能与胰岛素异相，而阿尔法细胞钙与胰岛素同相。我们的结论是，旁分泌相互作用和阿尔法细胞的内在机制都需要解释胰升糖素分泌对葡萄糖的反应。

Beta Cells: One of our main activities over the last few years has been the development of a comprehensive model for oscillations of membrane potential and calcium on time scales ranging from seconds to minutes. These lead to corresponding oscillations of insulin secretion. The basic hypothesis of the model is that the faster oscillations (tens of seconds) stem from feedback of calcium onto ion channels, likely calcium-activated potassium (K(Ca)) channels and ATP-dependent potassium (K(ATP)) channels, whereas the slower oscillations (five minutes) stem from oscillations in metabolism. The metabolic oscillations are transduced into electrical oscillations via the K(ATP) channels. The model thus consists of an electrical oscillator (EO) and a metabolic oscillator (MO) and is referred to as the Dual Oscillator Model (DOM). In our model, the MO is a glycolytic oscillator, but many of the features of the system would still hold if the metabolic oscillation arose elsewhere, such as the mitochondria. K(ATP) channels are of clinical significance as they are a first-line target of insulin-stimulating drugs, such as the sulfonylureas tolbutamide and glyburide, used in the treatment of Type 2 Diabetes. Severe gain-of-function mutations of K(ATP) are a major cause of neo-natal diabetes mellitus, whereas moderate gain-of-function mutations have been linked in genome-wide association studies (GWAS) to the milder but more common disease, adult-onset type 2 diabetes. Conversely, loss-of-function mutations of K(ATP) are a major cause of familial hyperinsulinism, a hereditary disease found in children in which beta cells are persistently electrically active and secrete insulin in the face of normal or low glucose, causing life-threatening hypoglycemia. For a review of the importance of oscillations of insulin secretion for health and disease see Reference # 3 in the 2015 report. Over a period of years we have accumulated a good deal of indirect evidence supporting the model, but we felt it important to devise a direct test of the central feature, namely that glycolysis oscillates. Our experimental collaborators developed a FRET-based sensor by modifiying pyruvate kinase (PK). PK binds fructose-1,6-bisphosphate (FBP), a key glycolytic metabolite, which was predicted to oscillate by the DOM. A previous paper confirmed that the sensor (PKAR, for pyruvate kinase activity reporter) does oscillate. In the current period the characteristics of the oscillations were probed more stringently by recording them simultaneously with membrane potential in order to ascertain the phase relationship between them. The experiments (Ref. # 1) again confirmed that PKAR oscillates and that it can do so even when calcium does not oscillate, as also predicted by the model. However, the phase relationship did not agree with the model prediction: PKAR declined during the active phase of the oscillation, whereas it was predicted to be high throughout the active phase because of a pulse of glycolytic activity. Two other metabolites, NAD(P)H and ATP, were also imaged and revealed a similar pattern. The discrepancy between prediction and experiment was striking but required only one major revision to the model to resolve. A previously known but neglected effect of calcium to accelerate the citric acid was added. This increases mitochondrial consumption of pyruvate, drawing down cytosolic FBP rapidly enough to overcome the surge in FBP production in glycolysis. In order to match the observation that ATP also declined during the active phase, it was necessary to assume in addition that the consumption of ATP by calcium pumps during the active phase was great enough to overcome the increased ATP production due to the stimulation of the citric acid cycle. This potent consumption of ATP serves to hasten the termination of the active phase because it allows K(ATP) channels to reopen. Further work will be needed to test the new predictions generated by the revised model and to fully understand the benefits of this more complicated arrangement. Alpha Cells: The regulation of glucagon secretion in the pancreatic -cell is not well understood. It has been proposed that glucose suppresses glucagon secretion either directly through an intrinsic mechanism, within the -cell, or indirectly through an extrinsic mechanism. We previously described a mathematical model for isolated pancreatic alpha-cells and used it to investigate possible intrinsic mechanisms of regulating glucagon secretion. We demonstrated that glucose can suppress glucagon secretion through both ATP-dependent potassium channels (K(ATP)) and a store-operated current (SOC). We now develop an islet model that combines previously published mathematical models of alpha- and beta-cells with a new model of delta-cells and use it to explore the effects of insulin and somatostatin on glucagon secretion. We show that the model can reproduce experimental observations that the inhibitory effect of glucose remains even when paracrine modulators are no longer acting on the -cell. We demonstrate how paracrine interactions can either synchronize - and -cells to produce pulsatile oscillations in glucagon and somatostatin secretion or fail to do so. The model can also account for the paradoxical observation that glucagon can be out of phase with insulin while alpha-cell calcium is in phase with insulin. We conclude that both paracrine interactions and the alpha-cell's intrinsic mechanisms are needed to explain the response of glucagon secretion to glucose.