Mechanisms of nicotinic acetylcholine receptor modulation of cocaine reward

烟碱乙酰胆碱受体调节可卡因奖赏的机制

基本信息

批准号：
10672207
负责人：
Janna K Moen
金额：
$ 1.33万
依托单位：
ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-07-01 至 2023-08-15
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10672207
关键词：
Abstinence Acetylcholine Acute Affect Alleles Attenuated Award Back Behavior Behavioral Brain CRISPR/Cas technology Cells Chronic Cocaine Cocaine Dependence Cocaine use disorder Code Compensation Complex Corpus striatum structure Data Development Development Plans Dopamine Dose Electrophysiology (science)Feedback Fiber Fluorescence Foundations Future Genes Genetic Glutamates Goals Health Human Human Genome Hyperactivity Individual Infusion procedures Interneurons Intravenous Knock-out Knockout Mice Laboratories Measures Medial Mediating Mediator Methodology Methods Molecular Monitor Mus Neurons Nicotinic Receptors Nucleus Accumbens Opsin Optics Pathway interactions Patients Pattern Phenotype Photometry Physiological Physiology Play Positioning Attribute Prefrontal Cortex Procedures Property Psychological reinforcement Receptor Signaling Refractory Reporter Research Personnel Research Proposals Rewards Role Self Administration Self Stimulation Shapes Signal Transduction Site Slice Source Stimulus Testing Training Transcript Transgenic Organisms Variant Viral Wild Type Mouse abuse liability addiction career development cell type cholinergic cocaine reward cocaine self-administration experimental study gamma-Aminobutyric Acid genetic approach genome wide association study in vivo innovation interest neurotransmission optogenetics permissiveness sensor transmission process

项目摘要

PROJECT SUMMARY Large-scale human genome-wide association studies have identified alleles in CHRNA5, the gene that encodes for the α5 nicotinic acetylcholine receptor (nAChR) subunit, that are protective against developing cocaine use disorder. Preliminary data from our group show that α5 knockout mice are less sensitive than wild-type mice to the rewarding effects of cocaine as measured using intravenous self-administration and intracranial self- stimulation procedures. Cholinergic interneurons (CINs) in the nucleus accumbens (NAc) shape reward-related behaviors, and inhibitory effects of cocaine on CIN activity are considered critical to its rewarding properties. Preliminary electrophysiology data collected in support of this application show that inhibitory GABAergic transmission onto CINs in the NAc is deficient in α5 knockout mice, which results in increased spontaneous firing activity of CINs. Further, CINs in α5 knockout mice are refractory to the inhibitory actions of cocaine. While Chrna5 transcript is not expressed in CINs or any other cell type in the NAc, Chrna5 transcripts are detected in the medial prefrontal cortex (mPFC) and a small number of other brain sites that provide long-range input to the NAc. Based on these exciting findings, I hypothesize that CIN-derived cholinergic transmission in the NAc stimulates α5-containing (α5*) nAChRs located on the terminals of long-range inputs from the mPFC, which enhances GABAergic transmission back onto CINs to inhibit their activity. Further, I propose that α5* nAChR- mediated feedback inhibition plays a permissive role in cocaine reward. I propose to rigorously test this hypothesis using two specific aims. In AIM 1, I will determine how α5* nAChRs contribute to cholinergic mechanisms of cocaine reward. In vivo fiber photometry with a fluorescence-based reporter of acetylcholine signaling will be used to monitor cholinergic transmission in the NAc following acute cocaine to confirm that feedback inhibition of cholinergic signaling evoked by cocaine is deficient in α5 knockout mice. I will then use a chemogenetic approach to determine how inhibiting CINs influences cocaine self-administration in wild-type and α5 knockout mice. AIM 2 will directly test the role of α5 nAChR- expressing inputs to the NAc on cocaine reward and physiology. First, slice electrophysiology combined with targeted optogenetic manipulation will be used to assess how α5 nAChR-expressing inputs from mPFC regulate the activity of CINs as well as the actions of cocaine. Finally, I will use a CRISPR/Cas9 approach to cleave CHRNA5 in α5* nAChR-expressing mPFC→NAc inputs followed by an intravenous self-administration paradigm to assess the consequences of CHRNA5 knockout on cocaine reward in mice. This innovative research proposal will generate important new data relevant to cholinergic mechanisms of cocaine reward with direct relevance to genetic mechanisms of vulnerability to CUD in humans. Further, the tailored career development plan will provide me with training in cutting-edge methodologies and position me to submit a highly competitive application for a K99/R00 Pathway to Independence Award.

项目概要大规模人类全基因组关联研究已经确定了 CHRNA5 中的等位基因，该基因编码 α5 烟碱乙酰胆碱受体 (nAChR) 亚基，可防止可卡因使用我们小组的初步数据表明，α5 基因敲除小鼠对疾病的敏感性低于野生型小鼠。使用静脉内自我给药和颅内自我给药测量可卡因的奖励效果伏隔核 (NAc) 中的胆碱能中间神经元 (CIN) 形状与奖励相关。可卡因对 CIN 活性的行为和抑制作用被认为对其有益特性至关重要。为支持该应用而收集的初步电生理学数据表明，抑制性 GABAergic 在 α5 敲除小鼠中，NAc 中 CIN 的传递存在缺陷，这会导致自发放电增加此外，α5 基因敲除小鼠中的 CIN 对可卡因的抑制作用具有抵抗力。 Chrna5 转录本不在 CIN 或 NAc 中的任何其他细胞类型中表达，Chrna5 转录本在内侧前额皮质（mPFC）和少数其他大脑部位为大脑提供远程输入基于这些令人兴奋的发现，我勇敢地面对 NAc 中 CIN 衍生的胆碱能传递。刺激位于 mPFC 远程输入末端的含 α5 (α5*) nAChR，增强 GABA 能传输回 CIN 以抑制其活性。此外，我建议 α5* nAChR-。介导的反馈抑制在可卡因奖励中发挥着许可作用，我建议对此进行严格的测试。在 AIM 1 中，我将确定 α5* nAChR 如何促进胆碱能。使用基于荧光的乙酰胆碱报告器进行体内纤维光度测定的机制。信号传导将用于监测急性可卡因后 NAc 中的胆碱能传输，以确认 α5 基因敲除小鼠缺乏对可卡因引起的胆碱能信号的反馈抑制。化学遗传学方法确定抑制 CIN 如何影响野生型和可卡因自我给药 α5 敲除小鼠将直接测试表达 α5 nAChR 的 NAc 输入对可卡因奖励的作用。首先，切片电生理学与靶向光遗传学操作相结合。评估来自 mPFC 的 α5 nAChR 表达输入如何调节 CIN 的活性以及最后，我将使用 CRISPR/Cas9 方法在表达 α5* nAChR 的 mPFC→NAc 中切割 CHRNA5。输入后进行静脉自我给药范例以评估 CHRNA5 的后果敲除小鼠中的可卡因奖赏将产生重要的新数据。可卡因奖赏的胆碱能机制与易受伤害的遗传机制直接相关此外，量身定制的职业发展计划将为我提供尖端的培训。方法论，使我能够提交一份竞争激烈的 K99/R00 衔接课程申请独立奖。