Norepinephrine and dopamine dynamics during conditioned place preference and aversion in dorsal hippocampus

摘要：2017年，1970万美国成年人与药物使用障碍作斗争，2020年，近 7万人死于阿片类药物过量。在阿片依赖者中，环境应激 持续的渴望，复发与强烈的压力反应和对药物线索的反应性有关。 意外的奖赏和厌恶刺激影响腹侧被盖区(VTA)多巴胺(DA)神经元的活动，以及 神经药理学研究表明，背侧海马区(CA1)的VTA释放DA 规范情景学习。蓝斑去甲肾上腺素(LC-NE)神经元也可释放DA，它具有 潜在地影响CA1的背景记忆和可塑性。要解决令人反感和有益的环境 影响阿片类药物的寻找，必须了解去甲肾上腺素(NE)和阿片的神经药理学特性 阿片类药物相关背景记忆中的DA。形成记忆的最初过程是习得，而之后是习得 记忆被重新激活，在被重新巩固为长期记忆之前，它们被认为是多才多艺的。 虽然我们知道去甲肾上腺素和多巴胺能受体与阿片类药物的上下文加工有关， 然而，目前还没有明确的药理学方法来检测内源性单胺的释放。 在上下文行为期间。这一提议的中心假设是，CA1中的DA主要从 VTA和仅VTA-DA是记忆获取和再巩固所必需的，而LC-NE只是增强记忆获取和再巩固。 Aim 1A使用带有单胺传感器的双光子体外成像和光遗传学来确定 DCA1中VTA终末DA释放的时空特性，而Aim 1B将决定时空特性 CA1期LC释放NE和DA的特性利用体外药理学，我们将阐明 CA1的多巴胺和去甲肾上腺素能机制影响DA和NE的来源释放 地区。Aim 2将测试NE和DA来源对吗啡或纳洛酮位置偏爱的必要性 再整合以及去甲肾上腺素和去甲肾上腺素获得偏好的必要性。在Aim 2A中，我们将同时 用化学遗传学方法测定NE和DA在药物抑制其胞体时的动态变化 VTA和LC在条件性位置偏好或厌恶记忆再巩固中的必要性。在AIM 2B，我们将检验去甲肾上腺素和多巴胺对条件性偏爱或厌恶记忆获得的必要性 利用CRISPR基因编辑技术敲除LC和Th中的去甲肾上腺素产生酶 在VTA。这一系列的体外和体内神经药理学实验将阐明单胺的充分性。 以及语境加工的必要性，以加深我们对语境对阿片类药物寻找的影响的理解。 这项提议将作为神经药理学、双光子光学成像、光遗传学、 化学遗传学、基因编辑和行为药理学方法。最终，这篇文章中概述的培训 提案将加强我的职业发展计划，因为我追求一个独立的，学术的职位 研究员。

SUMMARY: In 2017, 19.7 million American adults struggled with substance use disorders, and in 2020, nearly 70,000 people died from opioid-involved overdoses. In opioid dependent individuals, environmental stress perpetuates cravings, and relapse is associated with strong stress responses and reactivity to drug cues. Unexpected reward and aversive stimuli affect ventral tegmental area (VTA) dopamine (DA) neuron activity, and studies neuropharmacological studies have revealed DA released from VTA in dorsal hippocampus (CA1) regulating contextual learning. Locus coeruleus noradrenergic (LC-NE) neurons may also release DA, which has the potential to affect contextual memory and plasticity in CA1. To address how aversive and rewarding contexts affect the opioid seeking, we must understand the neuropharmacological properties of norepinephrine (NE) and DA in opioid-associated contextual memory. The initial process of forming memories is acquisition, while after memories are reactivated, they are considered versatile before being reconsolidated for long term memory. While we know noradrenergic and dopaminergic receptors have been implicated in opioid contextual processing, yet there have not yet been clear pharmacological approaches examining endogenous monoamine release during contextual behavior. The central hypothesis of this proposal is that DA in CA1 is primarily released from the VTA, and only VTA-DA is necessary for, and LC-NE only enhances, memory acquisition and reconsolidation. Aim 1A uses two-photon ex vivo imaging and optogenetics with monoamine sensors to determine spatiotemporal properties of DA release from VTA terminals in dCA1, while Aim 1B will determine spatiotemporal properties of NE and DA release from LC in CA1. Using ex vivo pharmacology, we will elucidate which dopaminergic and noradrenergic mechanisms in CA1 influence the release of DA and NE from their source regions. Aim 2 will tests the necessity of NE and DA sources for morphine or naloxone place preference reconsolidation and the necessity of NE and DA for preference acquisition. In Aim 2A, we will simultaneously measure NE and DA dynamics as we pharmacologically inhibit their soma using chemogenetics to determine the necessity of VTA and LC in memory reconsolidation of a conditioned place preference or aversion. In Aim 2B, we will test the necessity of NE and DA for memory acquisition of conditioned preference or aversion by using CRISPR gene-editing to knockdown Dbh (NE-producing enzyme) in LC and Th (DA-producing enzyme) in VTA. This series of in vitro and in vivo neuropharmacology experiments will elucidate monoamine sufficiency and necessity in contextual processing to further our understanding of the influence of context in opioid seeking. This proposal will serve as training in neuropharmacology, two-photon optical imaging, optogenetics, chemogenetics, gene-editing, and behavioral pharmacology approaches. Ultimately, the training outlined in this proposal will enhance my career development plan as I pursue a position as an independent, academic researcher.