Pathways of Insulin Transport across the Blood-Brain Barrier

 描述(申请人提供)：胰岛素抵抗与心血管疾病、血管功能障碍和神经退行性疾病高度相关(17，21，48)。虽然关于胰岛素抵抗的大部分研究都集中在它在肝脏中的表现上， 肌肉和脂肪，大脑正成为一个越来越重要的焦点。与肌肉和脂肪不同，大脑不需要胰岛素来促进葡萄糖的吸收。相反，大脑中的胰岛素似乎具有神经调节作用，如饱腹感信号(27)和体重调节(8)。此外，对阿尔茨海默病患者进行鼻腔注射胰岛素可改善认知功能(15)。虽然胰岛素在大脑中的作用很多，但对于胰岛素如何穿过血脑屏障(BBB)高度限制性的内皮细胞，人们知之甚少。从脑脊液(CSF)中抽取的胰岛素作为脑间质液(BISF)的替代物，对胰岛素进入大脑的研究有限，并检测到极低的胰岛素浓度(3，54)；然而，最近的新发现表明，脑脊液是一个不同于围绕神经元的BISF的池(32)。然而，肥胖者血浆和脑脊液胰岛素之间的梯度比瘦人更大(33)，这表明肥胖期间胰岛素的跨血管内皮细胞转运(TET)减少。在外周胰岛素Tet(56)中也观察到这种现象，那里的血管系统比血脑屏障开窗数多得多。尚未对穿过脑微血管的胰岛素Tet进行检测。我们实验室以前的工作表明，胰岛素在肌肉血管中的Tet依赖于胰岛素受体(IR)信号，由一氧化氮介导，并通过激活小窝的主要蛋白质成分小窝1(Cav1)来促进。该项目的目标是确定血脑屏障血管系统的胰岛素运输是否依赖于由Cav1介导的IR信号，以及在胰岛素抵抗期间这种信号是否被改变。我的第一个目标将使用原代大鼠脑微血管内皮细胞(RBMVECs)来确定IR、IR信号和Cav1之间的机制关系，以确定它们在胰岛素摄取、信号传递和跨内皮运输中的作用。我的第二个目标将使用高脂饮食(HFD)喂养的大鼠模型来确定胰岛素抵抗对胰岛素向大脑运输的影响，并继续关注新鲜分离的脑内皮细胞和固定脑片中IR和Cav1信号通路的功能和调节。综上所述，这些实验将阐明胰岛素通过血脑屏障运输并对大脑神经元发挥作用的途径(S)。最终，这将为胰岛素抵抗的治疗提供新的药理靶点。

 DESCRIPTION (provided by applicant): Insulin resistance is highly correlated with cardiovascular disease, vascular dysfunction, and neurodegenerative disorders (17,21,48). While the bulk of research regarding insulin resistance has focused on its manifestations in liver, muscle, and adipose, the brain is becoming an increasingly important focal point. Unlike muscle and adipose, the brain does not require insulin to facilitate glucose uptake. Instead, insulin in te brain appears to have a neuroregulatory role as a satiety signal (27) and regulator of body weight (8). Additionally, administration of intranasal insulin to individuals with Alzheimer's disease improves cognitive function (15). While there are many roles for insulin action in the brain, there is little understanding as to how insulin crosses the highly-restrictive endothelium o the blood-brain barrier (BBB). Limited research investigating insulin transport into the brain sampled from the cerebrospinal fluid (CSF) as a surrogate for brain interstitial fluid (BISF) and detected very low insulin concentrations (3,54); however, recent novel findings have demonstrated that the CSF is a different pool than the BISF that surrounds neurons (32). Nevertheless, the gradient between plasma and CSF insulin is greater in obese than in lean individuals (33), suggesting decreased transendothelial transport (TET) of insulin during obesity. This phenomenon is also observed in peripheral insulin TET (56), where vasculature is considerably more fenestrated than the BBB. Insulin TET across the brain microvasculature has not been examined. Previous work in our laboratory has demonstrated insulin's TET in muscle vasculature is dependent on insulin receptor (IR) signaling, mediated by nitric oxide, and facilitated by activation of caveolin-1 (Cav1), the main protein component of caveolae. The goal of this project is to determine whether insulin transport by the BBB vasculature is dependent on IR signaling, mediated by Cav1, and whether this is modified during insulin resistance. My first aim will employ primary rat brain microvascular endothelial cells (RBMVECs) to determine mechanistic relationships between IR, IR signaling, and Cav1 to determine their role in insulin uptake, signaling, and trans endothelial transport. My second aim will use the high-fat diet (HFD) fed rat model to determine the effect of insulin resistance on insulin transport into the brain, with continued focus on function and regulation of IR and Cav1 signaling pathways in freshly-isolated brain endothelial cells and in fixed brain slices. Taken together, these experiments will elucidate the pathway(s) through which insulin is transported across the BBB to exert its actions on neurons in the brain. Ultimately, this will provide novel pharmacologic target for the treatment of insulin resistance.