The role of innervation for hormone secretion in human islets

描述（由申请人提供）：在了解朗格汉斯胰岛在体内的功能方面，无论是在胰腺的天然环境中还是移植后治疗1型糖尿病，都存在根本性的差距。大多数关于胰岛功能的研究都是在体外进行的，因此对神经支配对胰岛激素分泌的作用知之甚少。我们研究的长期目标是了解活生物体中朗格汉斯岛的细胞生物学。这一特殊应用的目的是确定神经支配在胰岛激素分泌中的作用，使用一种新的技术平台，允许移植后血管化和再神经化的胰岛的体内成像。在这项新技术中，胰岛被移植到眼睛的前房，在操纵眼睛的神经输入后，局部和系统地记录它们的功能。核心假设是自主神经系统通过调节胰岛内的血流来调节人体胰岛功能。与小鼠胰岛不同，人类内分泌细胞不受自主神经支配的直接影响。在提出的机制中，自主神经纤维强烈支配可收缩的周细胞，这些周细胞作为括约肌改变局部血流，从而改变葡萄糖感知和激素释放到循环中的效率。提出这项研究的基本原理是，研究结果将为基础知识提供一个缺失的基本要素，没有这些要素，人类胰岛的生物学就无法理解。因此，拟议的研究与NIH的使命有关，即追求有关生命系统的性质和行为的基本知识。在强有力的初步数据的指导下，这一假设将通过追求三个具体目标来验证：1)体外胰岛神经支配模式的物种差异；2)眼内胰岛移植物决定了其自身的神经支配模式；3)神经输入对体内葡萄糖稳态的影响。在第一个目标下，将使用免疫组织化学和细胞内[Ca2+]成像系统地检查小鼠和人类胰岛的神经支配模式和神经支配内分泌细胞类型的神经递质受体谱。在第二个目标下，将比较人与小鼠眼内胰岛移植在免疫组织化学染色模式和体内自主神经纤维示踪方面的差异。在第三个目标下，通过激活瞳孔反射的副交感神经和交感神经成分、药物阻断和选择性消除神经输入，小鼠和人类胰岛移植物的局部胰岛细胞反应和葡萄糖稳态调节将受到挑战。提出的工作是创新的，因为它利用了一种新的技术平台，允许首次在体内成像受神经支配的人类和小鼠胰岛的功能。这项研究具有重要意义，因为它有望推进和扩展当前胰岛调节葡萄糖稳态的模型。

DESCRIPTION (provided by applicant): There is a fundamental gap in understanding how the islets of Langerhans function in vivo, either in the native environment in the pancreas or after transplantation to treat type 1 diabetes. Most studies on islet function have been performed in vitro, and as a consequence little is known about the role of innervation on islet hormone secretion. The long-term goal of our research is to understand the cell biology of islets of Langerhans in the living organism. The objective of this particular application is to determine the role innervation plays in the secretion of islet hormones using a new technological platform allowing in vivo imaging of vascularized and reinnervated islets after transplantation. In this new technology, islets are transplanted into the anterior chamber of the eye, and their function is recorded locally and systemically after manipulation of the eye's neural input. The central hypothesis is that the autonomic nervous system modulates human islet function by regulating blood flow within the islet. In contrast to mouse islets, human endocrine cells are not affected directly by the autonomic innervation. In the proposed mechanism, autonomic nerve fibers strongly innervate contractile pericytes that work as sphincters to change blood flow locally, thus changing the efficiency of glucose sensing and of hormone release into the circulation. The rationale for the proposed research is that the results will contribute a missing, fundamental element to basic knowledge, without which the biology of human islets cannot be understood. The proposed research is therefore relevant to the mission of the NIH that pertains to the pursuit of fundamental knowledge about the nature and behavior of living systems. Guided by strong preliminary data, this hypothesis will be tested by pursuing three specific aims: 1) Species differences in the innervation patterns of pancreatic islets in vitro; 2) Intraocular islet grafts determine their own innervation pattern; and 3) The role of neural input on glucose homeostasis in vivo. Under the first aim, the innervation patterns of islets and the neurotransmitter receptor profiles of the innervated endocrine cell types will be systematically examined in mouse and human islets using immunohistochemistry and imaging of intracellular [Ca2+]. Under the second aim, intraocular human and mouse islet grafts will be compared in terms of immunohistochemical staining patterns and in vivo tracing of autonomic nervous fibers. Under the third aim, local islet cell responses and regulation of glucose homeostasis by mouse and human islet grafts will be challenged by activating the parasympathetic and sympathetic components of the pupillary reflex, by pharmacological blockade, and by selective elimination of the neural input. The proposed work is innovative because it capitalizes on a new technological platform that allows for the first time in vivo imaging the function of innervated human and mouse islet. The proposed research is significant because it is expected to advance and expand current models of the regulation of glucose homeostasis by pancreatic islets. PUBLIC HEALTH RELEVANCE: The proposed work will demonstrate the role of innervation for human islet cell function. It capitalizes on a new technological platform developed by our group allowing for the first time in vivo imaging of innervated human and mouse islets. As a major expected outcome we will obtain a comprehensive picture of islet innervation, which will have a strong impact on current models of islet regulation of glucose homeostasis.