Desensitization and recovery of D2 autoreceptors

D2 自身受体的脱敏和恢复

• 批准号: 9982928
• 负责人: Alec Condon
• 金额: $ 4.55万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9982928
• 关键词: Acute; Address; Agonist; Autoreceptors; Behavior; Brain; Buffers; Calcium; Cells; Coupled; Data; Dendrites; Dependence; Disease; Dopamine; Dopamine D2 Receptor; Dopamine Receptor; Electrophysiology (science); Future; G-Protein-Coupled Receptors; GTP-Binding Proteins; Individual; Knowledge; Lateral; Lead; Lighting; Location; Measures; Midbrain structure; Modification; Monitor; Neurons; Outcome; Pacemakers; Parkinson Disease; Pathway interactions; Phosphorylation; Physiologic pulse; Play; Potassium; Process; Production; Psychotic Disorders; Receptor Activation; Receptor Signaling; Recovery; Regulation; Rewards; Role; Signal Transduction; Site; Slice; Substantia nigra structure; System; Testing; Therapeutic; Time; Tissues; Ventral Tegmental Area; Work; addiction; desensitization; dopamine system; dopaminergic neuron; experimental study; motor control; mu opioid receptors; neuronal cell body; neuronal excitability; novel therapeutics; pars compacta; receptor; receptor binding; response; reward processing; stem; transmission process; two-photon; voltage clamp

Project Summary/Abstract The dopamine system has a diverse repertoire of functions in the CNS including in voluntary motor control and reward. Mixtures of inhibitory and excitatory inputs overlaid on pacemaker firing of midbrain dopamine neurons controls circuit function. One such controlling factor is lateral inhibition by somatodendritic release of dopamine onto neighboring SNc and VTA dopamine neurons. Dopamine activates inhibitory D2-autoreceptors which couple to GIRK channels. The level of D2 sensitization controls the magnitude of inhibition generated by dopamine release. Little is known about the mechanism of D2 receptor desensitization except that calcium entry has been demonstrated to accelerate desensitization. This is highlighted by recent work which implicates non-canonical mechanisms of G-protein-coupled-receptor (GPCR) desensitization. This proposal seeks to further characterize D2 receptor regulation with two specific aims using whole-cell voltage-clamp recordings of SNc dopamine neurons in acute slice. First, what is the time course and calcium sensitivity of recovery from desensitization? Preliminary findings indicate recovery from desensitization is a rapid process, with signaling returning to baseline in about five minutes. This contrasts with experiments done with similar receptors, such as the µ-opioid receptor, which takes 30 or more minutes to recover. However, additional data are needed to uncover any calcium-dependence of desensitization as well as determine the relationship, if any, between degree of desensitization and time course of recovery. Aim two focuses on the receptor occupancy requirements for D2 receptor signaling; does an individual receptor require a bound agonist for downstream signaling to desensitize? This aim will be pursued by three sub-questions and, once again, how calcium might modify outcomes will be tested throughout these experiments. Does a low concentration of dopamine pre-desensitize the response to a high concentration of dopamine? In preliminary experiments, low concentrations of dopamine (~500 nM) desensitize the response to a high concentration test pulse. Does activation of other GPCRs desensitize D2 receptors? Though the results must be extended to other GPCRs, initial experiments have indicated pronounced heterologous desensitization with by activation of the inhibitory GPCR GABAB receptors. And finally, can desensitization spread from an initial location? The study of D2 signaling in specific compartments has remained inaccessible due to technological limitations. The recent production of CyHQ-O-DA, a photoactivatable caged dopamine with the ability to be photolyzed by 2-photon emission, allows for the spatially defined release of dopamine. This will allow for the induction of desensitization by repeated uncaging on a focal point and the probing to what degree the desensitization is localized by uncaging at various locations along dendrites and soma.

项目总结/摘要 多巴胺系统在中枢神经系统中具有多种功能，包括在自主运动中， 控制和奖励。抑制性和兴奋性混合输入叠加在中脑起搏放电上 多巴胺神经元控制回路功能。一个这样的控制因素是侧抑制， 体树突向邻近的SNc和VTA多巴胺神经元释放多巴胺。多巴胺活化 抑制性D2-自身受体，其与GIRK通道偶联。D2致敏水平控制着 多巴胺释放产生的抑制幅度。D2受体的作用机制尚不清楚 除了钙进入已被证明加速脱敏之外，这是 最近的研究强调了G蛋白偶联受体（GPCR）的非经典机制 脱敏该提案旨在进一步表征D2受体调节，具有两个特定目的， 全细胞电压钳记录的黑质多巴胺神经元在急性切片。 首先，脱敏后恢复的时间过程和钙敏感性如何？初步 研究结果表明，从脱敏中恢复是一个快速的过程，信号在大约 五分钟这与用类似受体（如μ-阿片受体）进行的实验形成对比， 需要30分钟或更长时间才能恢复。然而，需要更多的数据来揭示任何钙依赖性 以及确定脱敏程度与时间之间的关系，如果有的话 恢复的过程。 目的二关注D2受体信号传导的受体占用要求;个体是否 受体需要结合激动剂下游信号脱敏？这一目标将由三个 子问题，并再次，钙如何可能改变结果将在这些测试 实验低浓度的多巴胺是否会使对高浓度多巴胺的反应预先脱敏？ 多巴胺？在初步实验中，低浓度的多巴胺（~500 nM）使反应脱敏。 到高浓度测试脉冲。其他GPCR的激活是否会使D2受体脱敏？尽管结果 必须扩展到其他GPCR，最初的实验表明明显的异源脱敏 通过激活抑制性GPCR GABAB受体。最后，脱敏是否可以从 初始位置？在特定隔室中的D2信号传导的研究仍然是不可访问的， 技术限制。最近生产的CyHQ-O-DA是一种可光活化的笼状多巴胺， 通过双光子发射光解的能力，允许多巴胺的空间限定释放。这将 允许通过在焦点上反复撑开来诱导脱敏，以及探测到什么程度 通过在沿着树突和索马的不同位置撑开来定位脱敏。