Effect of Killing or Removing GABA from NPY/AgRP Neurons

杀死或去除 NPY/AgRP 神经元中 GABA 的效果

• 批准号: 7652490
• 负责人: Richard D. Palmiter
• 金额: $ 15.29万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-25 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7652490
• 关键词: ART protein; Ablation; Address; Adult; Affect; Alleles; Aminobutyric Acids; Anabolism; Animals; Behavioral; Body Weight; Brain; Brain region; Cells; Desire for food; Diphtheria Toxin; Enzymes; Feeding behaviors; Financial compensation; Gene Silencing; Genes; Genetic; Genetic Techniques; Glutamate Decarboxylase; Goals; Hormonal; Hypothalamic structure; Knockout Mice; Lead; Leptin; Lesion; Masks; Mediator of activation protein; Melanocortin 4 Receptor; Messenger RNA; Metabolism; Methods; Middle Hypothalamus; Molecular; Monitor; Mus; Neonatal; Neurons; Obesity; Peptides; Phenotype; Physiological; Population; Pro-Opiomelanocortin; Protein Isoforms; Regulation; Role; Signal Pathway; Signal Transduction; Signaling Molecule; Starvation; Structure of nucleus infundibularis hypothalami; Synapses; Tamoxifen; Techniques; Testing; Up-Regulation; base; diphtheria toxin receptor; energy balance; feeding; gamma-Aminobutyric Acid; insight; killings; leptin receptor; neonate; neural circuit; neuropeptide Y; paraventricular nucleus; pup; recombinase; research study; response

DESCRIPTION (provided by applicant): A small population of neurons (~ 5000 cells) in the arcuate region of hypothalamus that make neuropeptide Y (NPY) and agouti-related protein (AgRP) become essential for survival in adult mice. We targeted the diphtheria toxin receptor (DTR) to these NPY/AgRP neurons to allow their ablation by administration of diphtheria toxin (DT). Ablation of these neurons in adult mice results in starvation. However, their ablation in neonatal mice results in circuit-based compensation such that mice grow almost normally. Although neonatally DT-lesioned mice grow to normal size, we experimentally explore whether they manifest deficits in physiological responses that normally depend on NPY/AgRP neurons. We propose to determine the critical signaling molecules made by NPY/AgRP neurons and whether they function primarily by regulating the melanocortin-signaling pathway or other circuits. A prime candidate for the critical signaling molecule is ?-aminobutyric acid (GABA); thus, we propose to make conditional alleles of the two genes, GAD1 and GAD2 encoding the biosynthetic enzymes such that they can be inactivated in NPY/AgRP neurons and temporally controlled manner by the action of Cre recombinase. If GABA is the critical signaling molecule, then we predict that inactivation of GABA biosynthesis in NPY/AgRP neurons of neonatal mice will lead to compensation such that the cells are no longer necessary for survival. In contrast, inactivation of GABA biosynthesis in NPY/AgRP neurons of adult mice may promote starvation. NPY/AgRP neurons are known to inhibit neighboring neurons in the arcuate nucleus that make proopiomelanocortin (POMC) and their target neurons in the paraventricular nucleus that express melanocortin-4 receptor. Activation of POMC neurons inhibits feeding. Thus, we include experiments to directly test whether up-regulation of this melanocortin-signaling pathway after sudden ablation of NPY/AgRP neurons is critically involved in the starvation phenotype. However, because NPY/AgRP neurons also project to several other brain regions, we include experiments that address which of these projection regions are affected the most and whether they contribute to feeding behavior. These experiments should provide insight into the neural circuits and signaling molecules that are critical for feeding.

描述（由申请人提供）：下丘脑弓状区域的一小群神经元（约 5000 个细胞）使神经肽 Y (NPY) 和刺鼠相关蛋白 (AgRP) 成为成年小鼠生存所必需的。我们将白喉毒素受体 (DTR) 靶向这些 NPY/AgRP 神经元，以便通过施用白喉毒素 (DT) 来消除它们。成年小鼠中这些神经元的消融会导致饥饿。然而，它们在新生小​​鼠中的消融会导致基于电路的补偿，使小鼠几乎正常生长。尽管新生 DT 损伤小鼠生长至正常大小，但我们通过实验探索它们是否表现出通常依赖于 NPY/AgRP 神经元的生理反应缺陷。我们建议确定 NPY/AgRP 神经元产生的关键信号分子，以及它们是否主要通过调节黑皮质素信号通路或其他回路发挥作用。关键信号分子的主要候选者是 γ-氨基丁酸 (GABA)；因此，我们建议制作编码生物合成酶的两个基因 GAD1 和 GAD2 的条件等位基因，以便它们可以在 NPY/AgRP 神经元中失活，并通过 Cre 重组酶的作用进行暂时控制。如果 GABA 是关键的信号分子，那么我们预测新生小鼠 NPY/AgRP 神经元中 GABA 生物合成的失活将导致补偿，使细胞不再是生存所必需的。相反，成年小鼠 NPY/AgRP 神经元中 GABA 生物合成的失活可能会促进饥饿。已知 NPY/AgRP 神经元会抑制弓状核中产生阿片黑皮质素原 (POMC) 的邻近神经元，以及室旁核中表达黑皮质素 4 受体的目标神经元。 POMC 神经元的激活会抑制进食。因此，我们进行了直接测试 NPY/AgRP 神经元突然消融后黑皮质素信号通路上调是否与饥饿表型密切相关的实验。然而，由于 NPY/AgRP 神经元也投射到其他几个大脑区域，因此我们进行了一些实验来解决这些投射区域中哪些区域受到的影响最大以及它们是否有助于进食行为。这些实验应该可以深入了解对进食至关重要的神经回路和信号分子。