Phenotypic and Functional Determination of Central Extended Amygdala Cell Groups

描述（由申请人提供）：本提案侧重于作为中央扩展杏仁核一部分的大脑区域：杏仁核中央核（CEA）。CEA参与对恐惧、压力和焦虑的一系列反应，并包含不同的细胞群。然而，目前尚不清楚细胞群是否具有不同的功能，目前也不可能用现有的工具在体内确定这一点。本研究旨在测试以神经肽B和W型1受体（NPBW1）为靶点，在CEA中开发特异性神经元群选择性神经毒性病变的可行性。这种受体非常分散地表达，特别是在CEA外侧，而不是在周围区域。此外，该受体在含有促肾上腺皮质激素释放激素（CRH）和促啡肽的外侧CEA细胞上表达，但在含有脑啡肽的细胞或内侧CEA细胞上不表达，内侧CEA是杏仁核输出神经元的主要来源。该受体的选择性配体（神经肽W-30）将与皂苷定制偶联（通过先进的靶向系统）。目的1侧重于开发一种有效和选择性的CEA神经毒性病变。该配体将与NPBW1受体结合，内化后将皂苷释放到细胞内，导致细胞死亡。这种方法在其他受体系统中也有应用，包括使用mu阿片受体作为靶标，选择性靶向杏仁核嵌入核细胞。NPBW1受体定位的特异性表明，这种损伤方法将选择性地破坏含有CRH和dynorphin的细胞，但不会破坏外侧CEA中含有脑啡肽的细胞和内侧CEA中的杏仁核输出细胞，这些细胞被认为对调节几种恐惧和焦虑反应（例如恐惧增强的惊吓反应）至关重要。在Aim 2中，将在成年雄性大鼠中评估中央延伸杏仁核的CRH/dynorphin细胞的潜在功能。这些实验将初步评估CEA神经毒性病变对A)升高+迷宫和防御性退缩范式中基础焦虑样行为的影响，以及B)恐惧增强的声惊反射。人们希望开发一种工具来选择性地针对CEA的特定人群，这将有助于确定其功能，最初的重点是细胞在恐惧和焦虑样反应中可能发挥的作用。希望随着时间的推移，这项工作将最终导致中央延伸杏仁核中几种不同细胞类型的神经毒性病变的发展，包括终纹床核，总的来说，这将导致更好地理解与许多恐惧和焦虑相关疾病的发展和恶化相关的负面情绪的处理。这项提议旨在选择性地移除大脑中被认为与压力、恐惧和/或焦虑反应有关的特定神经元群。希望有选择地针对特定人群（而不是目前可用的更全面的方法）将有助于更好地了解大脑如何处理这些负面情绪。这反过来将有助于设计焦虑和恐惧相关障碍的治疗策略。

DESCRIPTION (provided by applicant): This proposal focuses on a brain area that is part of the central extended amygdala: the central nucleus of the amygdala (CEA). The CEA is involved in a number of responses to fear, stress and anxiety and contains distinct cell populations. However, it is not known whether the cell populations have distinct functions, and it is currently impossible to ascertain this in vivo with the tools available. This proposal aims to test the feasibility of developing a selective neurotoxic lesion of a specific neuronal population in the CEA using the neuropeptide B and W type 1 receptor (NPBW1) as a target. This receptor is very discretely expressed, specifically in the lateral CEA, but not in the surrounding areas. Furthermore, this receptor is expressed on cells of the lateral CEA that contain corticotropin releasing hormone (CRH) and dynorphin, but is not expressed on cells that contain enkephalin, or on cells of the medial CEA, a major source of amygdaloid output neurons. A selective ligand for this receptor (neuropeptide W-30) will be custom conjugated with saporin (by Advanced Targeting Systems). Aim 1 focuses on the development of an effective and selective neurotoxic lesion of the CEA. The ligand will bind to the NPBW1 receptor and when internalized will release saporin into the cell, causing the death of the cell. The utility of this approach has been shown for other receptor systems, including selective targeting of cells of the intercalated nuclei of the amygdala, using the mu opioid receptor as a target. The specificity of the localization of the NPBW1 receptor suggests that this lesioning method will selectively destroy cells containing CRH and dynorphin, but spare both enkephalin containing cells in the lateral CEA, and amygdaloid output cells within the medial CEA that are thought to be critical for modulation of several fear and anxiety responses (e.g. fear potentiated startle response). In Aim 2, the potential functions of CRH/dynorphin cells of the central extended amygdala will be assessed in adult male rats. These experiments will initially evaluate the effects of this neurotoxic lesion of the CEA on A) basal anxiety-like behaviors in the elevated plus maze and defensive withdrawal paradigm, and B) fear-potentiated acoustic startle reflexes. It is hoped that the development of a tool to selectively target a specific population of the CEA will help to define its functions, with an initial emphasis on the roles that the cells may play in fear and anxiety-like responses. It is hoped that with time, this work will ultimately lead to the development of neurotoxic lesions of several distinct cell types in the central extended amygdala, including the bed nucleus of the stria terminalis, and collectively, this will lead to a better understanding of the processing of negative emotions that are associated with the development and exacerbation of many fear and anxiety-related disorders. This proposal aims to selectively remove a specific neuronal population in the brain that is thought to be involved in stress, fear and/or anxiety responses. It is hoped that selectively targeting a specific population (rather than the more global approach currently available) will lead to a better understanding of how the brain processes these negative emotions. This in turn will be helpful in the design of treatment strategies for anxiety and fear-related disorders.