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，没有充分了解的机制，支配的前自主神经元，有一个障碍，以了解血糖状态的控制。这项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的稳态。由于糖尿病患者的葡萄糖稳态受损涉及控制自主输出的中央回路，本提案的直接目标是确定瞬时受体电位香草酸1型（TRPV 1）依赖性机制参与调节前自主PVN神经元在血糖状态的控制使用体内和体外方法。TRPV 1在糖尿病中的参与已经确立，并且我的实验室最近的研究表明，TRPV 1是通过增加兴奋性神经递质释放的前自主PVN神经元的重要控制器。此外，在糖尿病小鼠模型中，肝脏相关PVN神经元的TRPV 1依赖性兴奋不存在。我们的初步观察表明，PVN中TRPV 1的激活可能通过增加肌肉的葡萄糖摄取和减少肝脏的葡萄糖生成来降低对照小鼠的全身血糖水平。这些观察结果导致中心假设，即前自主PVN神经元接收TRPV 1表达的输入，这些输入被整合到一个协调的自主输出信号，以产生适当的血糖反应。拟议的体内研究将确定PVN回路中TRPV 1激活对自主终点的影响。PVN中TRPV 1激活对葡萄糖稳态的影响将使用清醒小鼠中的高胰岛素-正葡萄糖钳夹研究来确定，并使用遥测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使用TRPV 1cre小鼠确定选择性刺激和抑制TRPV 1输入对葡萄糖稳态的影响。全细胞膜片钳研究将揭示TRPV 1输入对PVN神经元的激活和抑制的细胞效应，并且我们将确定由TRPV 1表达投射调控的PVN中的前自主神经元群体。此外，TRPV 1激活对神经传递的影响将使用TRPV 1cre小鼠中的光刺激来确定。这些研究将通过前自主PVN神经元定义中央TRPV 1对全身葡萄糖稳态的作用，并将区分TRPV 1在该系统中的突触前和突触后位置的重要性。这些研究的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.