Regulatory Role of Growth Hormone Secretagogue Receptor

描述：(摘自申请者的摘要)长期目标是 了解生长激素促分泌素受体(GHS-R)在 内分泌学。这种G蛋白偶联受体已经被鉴定，克隆， 并被确定为脉冲性生长激素(GH)的重要调节因子 放手。在人类中，使用合成的GHS-R配体MK-0677进行日常治疗 逆转与年龄相关的脉动性生长激素释放和血清IGF-1的下降 级别。这种年轻化的功能优势包括改进 睡眠质量，身体成分得到改善， 力量。因此，GHS-R通路是内分泌学中的一个重要因素。 衰老的问题。本研究的重点是GHS-R在下丘脑调节中的作用 生长激素释放。GHS-R与生长激素释放激素的相互作用 生长激素释放激素神经元、生长激素、生长抑素和神经肽Y在调节中的作用 将评估生长激素搏动性的夹带情况。NPY和SST的作用 将使用NPY缺失小鼠和SSTR2缺失小鼠进行评估。神经肽Y的特异性 将使用NPY受体缺失的小鼠和拮抗剂来确定效果 对NPY受体亚型有选择性。MK-0677对生长激素释放激素表达的影响 弓状神经元将通过利用转基因小鼠进行视觉识别，其中 GHRH调控下的增强型绿色荧光蛋白的表达 推动者。这些GHRH神经元将被表征为离子通道表达 采用实时荧光定量RT-PCR和电生理学及药理学方法进行活性检测。 将测试MK-0677对特定通道活性的修改。至 直接决定底物参与的机制和磷酸化 GHS-R信令，如Kvl.2和PYK2，组件将被重组 在非洲爪哇卵母细胞和垂体来源的GC细胞中稳定表达GHS-R。 GHS-R与下丘脑其他神经元之间的信号通路研究 在体内，下丘脑GHS-R神经元的投射将被绘制出来。这 将通过以下方式将IREStau-Lacz整合到小鼠GHS-R基因座中来实现 同源重组。GHS-R下丘脑神经元的连接将是 以β-半乳糖苷酶免疫染色为特征。这些研究的结果 实验将提供有关中枢机制的重要信息 在衰老的内分泌学中。

DESCRIPTION: (Scanned from the applicant's abstract) The long-term objective is to understand the role of the Growth Hormone Secretagogue Receptor (GHS-R) in endocrinology. This G-protein coupled receptor has been characterized, cloned, and established as an important regulator of pulsatile Growth Hormone (GH) release. In humans, daily treatment with the synthetic GHS-R ligand MK-0677 reverses the age-related decline in both pulsatile GH release and serum IGF-1 levels. Functional benefits of this rejuvenation include improvements in quality of sleep, improved body composition and modest improvements in strength. Hence, the GHS-R pathway is an important factor in the endocrinology of aging. This proposal focuses on the role of GHS-R on hypothalamic regulation of GH release. The interplay between GHS-R, growth hormone releasing hormone (GHRH) neurons, GH, somatostatin (sst) and neuropeptide Y (NPY) in regulating the entrainment of GH pulsatility will be evaluated. The roles of NPY and sst will be evaluated using NPY null mice and sstr2 null mice. Specificity of NPY effects will be determined using NPY-receptor null mice and antagonists selective for NPY-receptor subtypes. Effects of MK-0677 on GHRH expressing arcuate neurons will be identified visually by exploiting transgenic mice where enhanced green fluorescent protein is expressed under the control of the GHRH promoter. These GHRH neurons will be characterized for ion channel expression using real time RT-PCR and activity by electro physiology and pharmacology. Modification of specific channel activity by MK-0677 will be tested. To directly determine the mechanism and phosphorylation of substrates involved in GHS-R signaling, such as a Kvl.2 and PYK2, the components will be reconstituted in Xenopus oocytes and in pituitary derived GC cells that stably express GHS-R. To investigate signaling pathways between GHS-R and other hypothalamic neurons in vivo, the projections of hypothalamic GHS-R neurons will be mapped. This will be accomplished by integrating IREStau-lacZ into the mouse GHS-R locus by homologous recombination. The wiring of GHS-R hypothalamic neurons will be characterized by immunostaining for beta-galactosidase. The results of these experiments will provide important information in central mechanisms involved in the endocrinology of aging.