Role of CB1R expressed in the prefrontal cortex in the control of locomotion

前额皮质表达的 CB1R 在运动控制中的作用

• 批准号: 10590320
• 负责人: Nephi Stella
• 金额: $ 23.33万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-15 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10590320
• 关键词: 2-arachidonylglycerol; Address; Affective; Age; Agonist; Applications Grants; Behavior; Behavioral; Biological; Biosensing Techniques; Brain; CNR1 gene; Calcium; Cannabis; Cells; Clustered Regularly Interspaced Short Palindromic Repeats; Coupled; Data; Disinhibition; Dose; Electrophysiology (science); Endocannabinoids; Equilibrium; Event; Fiber; Foundations; Frequencies; Genetic; Glutamates; Goals; Grant; Human; Immobilization; Impairment; In Situ Hybridization; Individual; Interneurons; Intoxication; Locomotion; Mammals; Maps; Measures; Mediating; Modality; Modernization; Modification; Molecular; Monitor; Motor Activity; Mus; Nature; Neocortex; Neurons; Outcome; Photometry; Physiological; Prefrontal Cortex; Regulation; Resolution; Role; Running; Series; Signal Transduction; Slice; Synaptic Transmission; System; THC exposure; Techniques; Technology; Testing; Tetrahydrocannabinol; Time; Viral; abuse liability; behavioral impairment; cannabimimetics; differential expression; endocannabinoid signaling; excitatory neuron; executive function; gamma-Aminobutyric Acid; hippocampal pyramidal neuron; in vivo; inhibitor; inhibitory neuron; innovation; mRNA Expression; marijuana use; motor impairment; neocortical; neural circuit; novel; optogenetics; pharmacologic; response; sensor

SUMMARY: Cannabis use in the US has alarmingly quadrupled in recent years: from 8.9% in 2016 to 17.5% in 2019 in individuals of age 12+ that used Cannabis in the last year. Thus, it is critical that we better understand the bioactivity of ∆9-tetrahydrocannabinol (THC), the primary intoxicating compound in Cannabis. THC activates cannabinoid receptors 1 (CB1R) and impairs several behaviors, including spontaneous locomotion. While it is known that THC reduces spontaneous locomotion in mice, the neural-circuit basis and involvement of the endocannabinoid (eCB), 2-arachidonoyl glycerol (2-AG), of this response remains largely unexplored. Using a novel 2-AG sensor (eCB2.0) expressed in the prelimbic cortex (PrL) neurons, we discovered a tight correlation between increase in 2-AG levels, calcium transients and initiation of spontaneous locomotion. Remarkably, both 2-AG and GCaMP6f calcium transients were significantly greater in THC-treated mice compared to vehicle- treated mice, and the number of initiated locomotion events greatly reduced as a function of the hypolocomotion response. Our hypothesis is that 2-AG activates CB1R’s on select PrL GABAergic interneuron subpopulations, which disinhibits the glutamatergic activity within the PrL to control the initiation of spontaneous locomotion, and THC potentiates this mechanism. ■ Aim 1 will determine how THC preferentially modulates the physiological activity of select GABAergic subpopulations and glutamatergic neurons in the PrL. We will use RNAscope in situ hybridization to map CB1R expression in GABAergic interneurons (GABA-IN’s) in the PrL. Combining GABA-IN-Cre lines with viral techniques and channel-rhodopsin assisted circuit mapping with slice electrophysiology, we will determine how 2-AG’s action at CB1-R in PFC changes synaptic transmission and how this is impacted by THC. ■ Aim 2 will utilize in vivo fiber photometry and optogenetic manipulation of GABA-IN-Cre and VGLUT-Cre mice expressing cre-dependent GCaMP6f or eCB2.0 in mouse PrL to record changes in neuronal activity and 2-AG levels of inhibitory and excitatory neurons during spontaneous locomotion of mice treated with increasing doses of THC. We will determine the specific 2-AG signaling components involved in this response using selective pharmacological inhibitors. We will also virally express a CRISPR-CB1R construct to eliminate CB1R from GABA- IN’s and glutamatergic neurons to establish the involvement of this target. Based on this premise, we will test the hypothesis that select subpopulations of GABA-IN’s mediate changes in PrL activity associated with increase in 2-AG levels and control of spontaneous locomotion. To determine the necessity and sufficiency of PrL activity in THC treated mice, we will optogenetically stimulated or inhibited GABA-IN’s and glutamatergic neurons and measured changes in spontaneous locomotion in vehicle and THC-treated mice. This exploratory R21 grant proposal provides an ideal mechanisms and scientific foundation for studies aimed at deciphering the mechanisms by which eCBs modulates PrL activity and how THC impairs locomotor behavior.

摘要：近年来，美国的大麻使用量惊人地翻了两番：从 2016 年的 8.9% 上升到 2016 年的 17.5%。 2019 年，去年使用过大麻的 12 岁以上个人的情况因此，我们更好地了解这一点至关重要。 大麻中主要的致醉化合物 Δ9-四氢大麻酚 (THC) 的生物活性被激活。 大麻素受体 1 (CB1R) 并损害多种行为，包括自发运动。 已知 THC 会减少小鼠的自发运动、神经回路基础和参与 内源性大麻素 (eCB)、2-花生四烯酰甘油 (2-AG) 的这种反应在很大程度上尚未被探索。 在前边缘皮层（PrL）神经元中表达的新型 2-AG 传感器（eCB2.0），我们发现了紧密的相关性 值得注意的是，2-AG 水平增加、钙瞬变和自发运动启动之间都有影响。 与载体相比，THC 处理的小鼠中 2-AG 和 GCaMP6f 钙瞬变明显更大 接受治疗的小鼠，由于运动不足，启动运动事件的数量大大减少 我们的假设是 2-AG 激活特定 PrL GABA 能中间神经元亚群上的 CB1R， 它抑制 PrL 内的谷氨酸能活性，以控制自发运动的启动，并且 THC 增强了这种机制。 ■ 目标 1 将确定 THC 如何优先调节所选 GABA 能的生理活性 PrL 中的亚群和谷氨酸能神经元 我们将使用 RNAscope 原位杂交来绘制 CB1R 的图谱。 PrL 中 GABA 能中间神经元 (GABA-IN) 的表达将 GABA-IN-Cre 系与病毒相结合。 技术和通道视紫红质辅助电路映射与切片电生理学，我们将确定如何 2-AG 在 PFC 中 CB1-R 上的作用改变了突触传递以及 THC 对突触传递的影响。 ■ 目标 2 将利用 GABA-IN-Cre 和 VGLUT-Cre 小鼠的体内光纤光度测定和光遗传学操作 在小鼠 PrL 中表达 cre 依赖性 GCaMP6f 或 eCB2.0 以记录神经元活动和 2-AG 的变化 增加剂量治疗的小鼠自发运动期间抑制性和兴奋性神经元的水平 我们将使用选择性确定参与此反应的特定 2-AG 信号成分。 我们还将通过病毒表达 CRISPR-CB1R 构建体，以消除 GABA 中的 CB1R。 基于此前提，我们将测试 IN 和谷氨酸能神经元的参与。 假设 GABA-IN 的选择亚群介导与增加相关的 PrL 活性变化 2-AG 水平和自发运动的控制 确定 PrL 活动的必要性和充分性。 在 THC 治疗的小鼠中，我们将通过光遗传学刺激或抑制 GABA-IN 和谷氨酸能神经元， 测量了媒介物和 THC 处理的小鼠自发运动的变化。 这项探索性的 R21 拨款提案为旨在研究的研究提供了理想的机制和科学基础 破译 eCB 调节 PrL 活性的机制以及 THC 如何损害运动行为。