Function-specific serotonergic neurons, discrete brain targets, and addiction

描述(申请人提供)：产生5-羟色胺(5-羟色胺)的神经元被认为是可卡因寻求行为的关键调节器。由于寻找可卡因的行为反映了与可卡因配对的刺激的激励动机效应，5-羟色胺被认为有助于形成与复吸有关的成瘾记忆。改变5-羟色胺的指令是治疗成瘾的一个新兴的治疗方向，努力主要集中在受体的操纵上。在这里，我们提出了一个新的，补充的方法，重点是突触前，在5-羟色胺神经元本身。最近的研究表明，可卡因寻找涉及5-羟色胺在5-HT2A和5-HT2C受体上的相反活动，这两个受体是多巴胺(DA)输出的关键调节器。在此指导下，我们将确定小鼠不同亚型的5-羟色胺神经元通过条件性位置偏爱程序(目标1)对与可卡因配对的刺激的兴奋兴奋效应进行多大程度的差异调制，并将探索它们的前脑投射靶点，特别是与突触后5-HT2A和5-HT2C受体表达(目标2)有关的目标。我们的出发点是Dymecki实验室正在组装的5-羟色胺能神经系统的新出现的结构-功能-连通性图，其中5-羟色胺神经元根据它们独特的基因组合的表达进行分类，从而可能具有独特的功能。根据它们的轴突靶区以及在体内选择性沉默后的行为和生理缺陷的评估，它们被进一步分型。在这里，我们建议探索这些5-羟色胺神经元亚型在成瘾相关行为中的作用，首先集中在5-羟色胺神经元亚型，这些亚型神经元亚类型支配与增强或抑制可卡因寻找行为有关的脑区。有三种5-羟色胺神经元亚型：R1-EN1 5-羟色胺神经元亚型，根据其起源于菱形球(R)1和转录因子Engrailed1的表达而命名；R2-Hoxa2亚型，由其在R2中的起源和转录因子Hoxa2的表达定义；以及Drd1a 5-羟色胺神经元亚型，由1a型DA受体的表达定义。由于后两者在成瘾相关的中脑边缘系统中表现出更多受限的神经分布，可能暗示了在行为调节中的特殊作用，这两个将在本R21应用中首先探讨。我们将分子基因技术与成熟的小鼠行为测试相结合的方法在技术和概念上都是创新的。这些不同学科的整合现在才有可能，通过开发跨部门的遗传工具，使小鼠中分子上不同的5-羟色胺神经元亚型明显，并可用于行为探测和全基因组分子图谱。识别关键的5-羟色胺神经元亚型及其对成瘾相关行为的影响的性质，以及拥有选择性分离和分子探测的分子遗传学工具，是发现新的、可能具有行为选择性的治疗线索的基础。

DESCRIPTION (provided by applicant): Serotonin (5-HT)-producing neurons are recognized as key modulators of cocaine-seeking behavior. As cocaine-seeking behavior reflects the incentive motivational effect of stimuli paired with cocaine, 5-HT is thought to contribute to the formation of addiction-related memory that underlies relapse. Alteration of 5-HT commands is an emerging therapeutic direction for addiction treatment, with efforts focusing largely on receptor manipulations. Here we propose a novel, complementary approach that focuses presynaptically, on the 5-HT neurons themselves. Guided by recent research showing that cocaine seeking involves opposing activity of 5-HT at 5-HT2A and 5-HT2C receptors, which are key modulators of dopamine (DA) output, we will determine to what degree different subtypes of 5-HT neurons in mice differentially modulate the incentive motivational effects of stimuli paired with cocaine via the conditioned place preference procedure (Aim 1) and will explore their forebrain projection targets, especially as relates to postsynaptic 5-HT2A and 5-HT2C receptor expression (Aim 2). Our starting point is an emerging structure-function-connectivity map of the serotonergic neural system being assembled by the Dymecki lab, in which 5-HT neurons are classified by their expression of unique gene combinations and thus likely unique functionalities. They are further typed by their axonal target regions and by assessment of behavioral and physiological deficits following their selective silencing in vivo. Here we propose probing these 5-HT neuron subtypes for their role in addiction-related behavior, focusing first on 5-HT neuron subtypes that innervate brain regions implicated in enhancing or suppressing cocaine-seeking behavior. Three 5-HT neuron subtypes stand out: the r1-En1 5-HT neuron subtype, named by its origin in rhombomere (r) 1 and expression of the transcription factor Engrailed1; r2-Hoxa2 subtype, defined by its origin in r2 and expression of the transcription factor Hoxa2; and the Drd1a 5-HT neuron subtype, by expression of the type 1a DA receptor. Because the latter two show more restricted innervation profiles within the addiction-relevant mesolimbic system, perhaps suggestive of specialized roles in behavior modulation, these two will be explored first in this R21 application. Our approach of partnering molecular genetic techniques with a well-established behavioral assay in mice is both technically and conceptually innovative. The integration of these distinct disciplines is made possible only now through development of intersectional genetic tools that make molecularly distinct subtypes of 5-HT neurons apparent and accessible in mice for behavioral probing and genome-wide molecular profiling. Identifying the key 5-HT neuron subtypes involved and the nature of their effects on addiction-related behavior, along with having molecular genetic tools for their selective isolation and molecular probing is foundational for discovering new, possibly behaviorally-selective, therapeutic leads.