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

描述（申请人提供）：产生5-羟色胺（5-HT）的神经元被认为是可卡因寻求行为的关键调节剂。由于可卡因寻求行为反映了与可卡因配对的刺激的激励效应，5-HT被认为有助于形成成瘾相关记忆，这是复发的基础。改变5-羟色胺指令是成瘾治疗的一个新兴治疗方向，主要集中在受体操纵上。在这里，我们提出了一种新颖的，互补的方法，集中在突触前，对5-HT神经元本身。最近的研究表明，可卡因寻求涉及5-HT在5-HT2A和5-HT2C受体上的相反活性，这是多巴胺（DA）输出的关键调节剂，我们将确定小鼠中不同亚型的5-HT神经元通过条件位置偏好程序（Aim 1）在多大程度上调节与可卡因配对的刺激的激励动机效应（Aim 1），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受体表达有关（Aim 2）。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兴结构-功能-连接图，其中5-羟色胺神经元根据其独特基因组合的表达进行分类，因此可能具有独特的功能。它们通过轴突靶区和在体内选择性沉默后的行为和生理缺陷评估进一步分型。在这里，我们建议探索这些5-HT神经元亚型在成瘾相关行为中的作用，首先关注支配与增强或抑制可卡因寻求行为有关的大脑区域的5-HT神经元亚型。5-HT神经元有三种亚型：r1-En1 5-HT神经元亚型，因其起源于菱形球(r) 1和转录因子Engrailed1的表达而命名；r2-Hoxa2亚型，由其起源于r2和转录因子Hoxa2的表达定义；和Drd1a 5-HT神经元亚型，通过1a型DA受体的表达。由于后两者在成瘾相关的中边缘系统中显示出更有限的神经支配谱，可能暗示了在行为调节中的特殊作用，因此这两者将首先在R21应用中进行探讨。我们将分子遗传技术与小鼠行为分析相结合的方法在技术上和概念上都是创新的。只有通过交叉遗传工具的发展，这些不同学科的整合才有可能实现，这些交叉遗传工具使5-HT神经元的分子不同亚型在小鼠中清晰可见，并可用于行为探测和全基因组分子谱分析。确定关键的5-HT神经元亚型及其对成瘾相关行为影响的性质，以及对其选择性分离和分子探测的分子遗传工具，是发现新的、可能是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.