TRPV1-Dependent Autonomic Control in Diabetes

描述（由申请人提供）：全身葡萄糖稳态基本上由中枢自主神经回路调节，如果存在自主神经功能障碍，则发生2型糖尿病的风险很高。因此，如果不充分了解自主前神经元的控制机制，就很难理解血糖状态的控制。这一建议的总体长期目标是阐明中枢自主控制和葡萄糖稳态之间的基本关系。下丘脑室旁核（PVN）是控制自主神经流出的关键指挥中心，从而影响葡萄糖和能量稳态。由于糖尿病患者的葡萄糖稳态受损涉及控制自主神经输出的中枢回路，因此本研究的直接目的是通过体内和体外方法确定瞬时受体电位香草样蛋白1 （TRPV1）依赖性机制，该机制参与调节自主神经前PVN神经元控制血糖状态。TRPV1与糖尿病的关系已经确立，我的实验室最近的研究表明，TRPV1通过增加兴奋性神经递质释放，是自主神经前PVN神经元的重要控制者。此外，这种TRPV1依赖性的肝脏相关PVN神经元的兴奋在糖尿病小鼠模型中不存在。我们的初步观察表明，激活PVN中的TRPV1可能通过增加肌肉的葡萄糖摄取和减少肝脏的糖异生来降低对照小鼠的全身血糖水平。这些观察结果导致了一个中心假设，即自主神经前PVN神经元接受trpv1表达的输入，这些输入被整合到协调的自主神经输出信号中，以产生适当的血糖反应。拟议的体内研究将确定PVN回路中TRPV1激活对自主终点的影响。PVN中TRPV1激活对葡萄糖稳态的影响将通过清醒小鼠的高胰岛素-正血糖钳夹研究来确定，并通过遥测技术来确定对血压和心率的影响。使用光遗传学的系统水平研究将确定TRPV1输入的选择性刺激和抑制对TRPV1cre小鼠葡萄糖稳态的影响。全细胞膜片钳研究将揭示TRPV1输入对PVN神经元的激活和抑制的细胞效应，我们将确定PVN中受TRPV1表达预测调节的自主神经群。此外，TRPV1激活对神经传递的影响将在TRPV1cre小鼠中通过光刺激来确定。这些研究将明确中枢TRPV1通过自主神经前PVN神经元对全身葡萄糖稳态的作用，并将区分TRPV1在该系统中的突触前和突触后位置的重要性。所提出的研究结果有望为通过自主控制改善糖尿病患者的血糖状态开辟创新的临床策略和药物靶点。

DESCRIPTION (provided by applicant): Systemic glucose homeostasis is substantially regulated by central autonomic circuits and there is a high risk of developing type 2 diabetes if autonomic dysfunction is present. Therefore, without a full understanding of the mechanisms governing the preautonomic neurons, there is a barrier to the understanding of the control of glycemic status. The overall long term goal of this proposal is to elucidate the fundamental relationship between central autonomic control and glucose homeostasis. The paraventricular nucleus (PVN) of the hypothalamus is a critical command center controlling autonomic outflow and, thereby, influencing glucose and energy homeostasis. Since impaired glucose homeostasis in diabetic patients involves central circuits controlling autonomic output, the immediate objective of this proposal is to identify transient receptor potential vanilloid type 1 (TRPV1)-dependent mechanisms involved in the regulation of preautonomic PVN neurons in the control of glycemic status using in vivo and in vitro approaches. The involvement of TRPV1 in diabetes has been established and recent studies from my laboratory have indicated that TRPV1 is a vital controller of preautonomic PVN neurons through increasing excitatory neurotransmitter release. Furthermore, this TRPV1- dependent excitation of liver-related PVN neurons was absent in a diabetic mouse model. Our preliminary observations demonstrate that activation of TRPV1 in the PVN lowers systemic blood glucose levels in control mice likely through increasing glucose uptake of the muscle and decreasing gluconeogenesis by the liver. These observations lead to the central hypothesis that preautonomic PVN neurons receive TRPV1-expressing inputs and these inputs are integrated into a coordinated autonomic output signal to generate an appropriate glycemic response. The proposed in vivo studies will determine the effect of TRPV1 activation in PVN circuits on autonomic endpoints. The effect of TRPV1 activation in PVN on glucose homeostasis will be determined using hyperinsulinemic-euglycemic clamp studies in conscious mice, and on blood pressure and heart rate using telemetry. Systems level studies using optogenetics will determine the effect of selective stimulation and inhibition of TRPV1 inputs on glucose homeostasis using TRPV1cre mice. Whole-cell, patch-clamp studies will reveal the cellular effects of activation and inhibition of TRPV1 inputs on PVN neurons, and we will identify preautonomic neuronal populations in the PVN regulated by TRPV1-expressing projections. Furthermore, the effect of TRPV1 activation on neurotransmission will be determined using photostimulation in TRPV1cre mice. These studies will define the role of central TRPV1 action on whole body glucose homeostasis through preautonomic PVN neurons, and will differentiate the importance of pre- and postsynaptic locations of TRPV1 on this system. The outcomes of the proposed studies hold the promise of opening innovative clinical strategies and drug targets for the improvement of glycemic status in diabetic patients via autonomic control.