Mouse Models of Habenula Development and Function in Mental Illness and Addiction

描述(申请人提供)：这是一个探索性的R21方案，重点是内缰核的发育和功能。缰核是一对位于丘脑背侧(上皮层)的核。它由参与大体不同神经回路的外侧和内侧亚核组成。缰核的两个部分都通过突出的缰核延髓束(FR)将它们的输出纤维发送到腹侧中脑。外侧缰核直接支配中脑的多巴胺和5-羟色胺系统，而内侧缰核向脚间核发出显著的胆碱能投射，而脚间核又支配中脑腹侧。在最近的工作中，我们发现缰核具有不同于其他前脑或丘脑区域的基因表达模式，并且同源结构域转录因子Brn3a(Pou4f1)调节内侧缰核和外侧缰核神经元的许多特征分子特征。人们对内侧缰核的功能知之甚少，但它位于边缘/基底前脑和中脑单胺能系统之间的交界处，表明它在奖赏、注意力、学习和药物依赖中发挥作用。出于这个原因，有人假设缰核功能障碍可能在精神分裂症中发挥作用，治疗尼古丁滥用的药物开发应该针对缰核通路。目的1利用Nurr1(Nr4a2)基因座Cre重组酶的表达，通过靶向缺失Brn3a来阻断缰核-延髓通路。我们将测试特定的假设，即在Brn3a基因敲除小鼠中观察到的缰核投射的发育缺陷是由于对Brn3a的细胞自主需求。目的2将使用混合遗传/立体定向注射策略，基于可诱导白喉毒素受体(IDTR)的靶向表达，特异性地消融成年小鼠内侧缰核的胆碱能成分。我们将验证一种假设，即这种损伤将导致缰核对中枢IPN的胆碱能和谷氨酸能神经支配完全丧失，而对外侧IPN的肽能神经支配则不受影响。目的3将发展转基因小鼠，允许光诱导的，可逆的抑制缰核活性，通过靶向表达光激活的氯泵卤视紫红质，融合到YFP报告。我们将使用YFP荧光来识别切片制备中的缰核神经元，并测试Brn3a神经元表现出先前在缰核中描述的低频自发去极化的假设。我们将确定这种活动和诱导的动作电位是否可以被光抑制。除了提案的特定目标外，这些实验还将生成小鼠模型，用于未来的R01提案，检查与精神分裂症和尼古丁滥用有关的复杂行为的作用。 与公共卫生相关：缰核是所有哺乳动物的丘脑背侧核，它连接着与动机、奖励、注意力和运动有关的前脑和中脑中心。尽管它位于中心位置，但人们对这个核的功能知之甚少。目前的建议将创造转基因小鼠，在其中：1)缰核与其目标发育脱节，或2)遗传定义的内侧缰核神经元可通过注射毒素选择性地消融，或3)缰核功能可通过植入的光纤探头的光关闭。这些小鼠将被用来解决缰核如何发展其独特特性的具体问题，并在未来的实验中确定缰核在记忆、学习、注意力以及精神分裂症和尼古丁依赖模型中的功能。

DESCRIPTION (provided by applicant): This is an exploratory R21 proposal focused on the development and function of the medial habenula. The habenula is a paired nucleus residing in the dorsal thalamus (epithalamus). It consists of lateral and medial subnuclei which participate in largely distinct neural circuits. Both parts of the habenula send their output fibers to the ventral midbrain via the prominent habenulopeduncular tract or fasciculus retroflexus (FR). The lateral habenula directly innervates the midbrain dopamine and serotonin systems, while the medial habenula sends a prominent cholinergic projection to the interpeduncular nucleus, which in turn innervates the ventral midbrain. In recent work we have shown that the habenula has a pattern of gene expression quite distinct from other forebrain or thalamic areas, and that a homeodomain transcription factor, Brn3a (pou4f1), regulates many of the characteristic molecular features of the medial habenula and a subset of lateral habenula neurons. Little is known about the function of the medial habenula, but its position at the interface between the limbic/basal forebrain and midbrain monoaminergic systems suggests role in reward, attention, learning and drug dependence. For this reason it has been hypothesized that habenular dysfunction may play a role in schizophrenia, and that drug development for nicotine abuse should be targeted to the habenular pathway. Aim 1 will use genetic methods to disrupt the habenulopeduncular pathway by targeted deletion of Brn3a, using expression of Cre recombinase from the Nurr1 (nr4a2) gene locus. We will test the specific hypothesis that the developmental defects in the habenular projections observed in Brn3a knockout mice are due to a cell-autonomous requirement for Brn3a. Aim 2 will use a mixed-genetic/stereotactic injection strategy to specifically ablate the cholinergic component of the medial habenula in adult mice, based on the targeted expression of inducible diphtheria toxin receptor (iDTR). We will test the hypothesis that such lesions will result in a complete loss of cholinergic and glutamatergic innervation of the central IPN by the habenula, while peptidergic innervation of the lateral IPN will be spared. Aim 3 will develop transgenic mice permitting the light inducible, reversible inhibition of habenula activity using targeted expression of the light-activated chloride pump halorhodopsin, fused to a YFP reporter. We will use YFP fluorescence to identify habenula neurons in slice preparations, and test the hypothesis that Brn3a+ neurons exhibit the low-frequency spontaneous depolarizations previously described in the habenula. We will determine whether this activity and induced action potentials can be inhibited by light. Beyond the specific aims of the proposal, these experiments will generate mouse models to be used in future R01 proposals examining the role of complex behaviors related to schizophrenia and nicotine abuse. PUBLIC HEALTH RELEVANCE: The habenula is brain nucleus in the dorsal thalamus of all mammals that connects the forebrain to midbrain centers associated with motivation, reward, attention, and movement. Despite its central location, remarkably little is known about the function of this nucleus. The present proposal will create transgenic mice in which: 1) the habenula is developmentally disconnected from its targets, or 2) genetically defined medial habenula neurons can be selectively ablated by the injection of a toxin, or 3) habenula function can be turned off with light from an implanted fiber optic probe. These mice will be used to address specific questions about how the habenula develops its unique properties, and in future experiments to determine habenula function in memory, learning, attention, and models of schizophrenia and nicotine dependence.