Architecture and function of striatal dopamine signaling machinery

总结 多巴胺是一种重要的神经调质，多巴胺信号传导中的病理是脑损伤的标志。 疾病尽管有这些作用，多巴胺信号的组织和调节还不完全清楚。 这个项目的长期目标是剖析轴突多巴胺传递的细胞生物学。 多巴胺信号的时空特征不同于突触传递。在常规 突触，纳米级的突触结构能够以亚毫秒的速度实现强大的受体激活， 将通信限制在选定神经元之间的点对点接触。相比之下，多巴胺是一种体积 胞吐作用后通过细胞外空间扩散的一种递质，可通过 G蛋白偶联受体这些特性表明多巴胺的传递是缓慢和扩散的。最近 然而，来自几个实验室的数据，包括上一个供资周期产生的一些数据， 表明多巴胺的传递是高度动态的，在某些情况下，非常精确。此外，委员会认为， 多巴胺的释放受到纹状体中局部胆碱能中间神经元的有力和快速调节。这些 研究结果表明，多巴胺容量传输的编码比以前认为的更精确。 出现的一个主要问题是多巴胺传输的架构如何支持精确的 信号我们的总体模型是，分子机制已经进化到支持广泛的多巴胺编码 鳞片我们建立在我们以前的发现，轴突多巴胺胞吐执行毫秒精度 由稀疏复杂的蛋白质机器组成，通常存在于突触中。在aim1中，我们放大了强大的 局部调节，并询问胆碱能神经元如何触发多巴胺释放。根据初步数据，我们 假设胆碱能中间神经元的活动诱导多巴胺轴突的异位动作电位放电， 引发多巴胺分泌我们的目标是测试这一假设，并了解潜在的机制。 异位轴突动作电位起始远离多巴胺的内源性机制的鉴定 神经元索马对多巴胺神经元功能具有重要意义。在目标2中，我们剖析了组织 多巴胺受体相对于释放部位的变化。我们建立在最近的工作，确定这些标记 稀疏的分泌部位。我们的初步数据显示多巴胺受体聚集在一到两微米 远离释放位点，并表明D1与D2受体分布的差异。我们将系统地 评估释放受体的组织在超分辨率三维图像的大纹状体体积和将 机械地剖析它是如何建立的。我们认为这种组织不同于纳米级突触， 结构和从扩散组织往往与体积传输，并可能适合于 通过多巴胺神经元放电模式的开关介导不同的通路激活。 我们的工作将剖析专门的多巴胺信号结构的组织和快速的，局部的 脊椎动物纹状体多巴胺释放的触发机制。

Summary Dopamine is an important neuromodulator and pathologies in dopamine signaling are a hallmark of brain disease. Despite these roles, the organization and regulation of dopamine signaling are incompletely understood. The long-term goal of this project is to dissect the cell biology of axonal dopamine transmission. Spatial and temporal features of dopamine signaling are different from synaptic transmission. At conventional synapses, nanometer-scale synaptic structure enables robust receptor activation at sub-millisecond speeds and restricts communication to point-to-point contacts between select neurons. In contrast, dopamine is a volume transmitter that diffuses through the extracellular space after exocytosis and may influence many cells through G-protein coupled receptors. These properties suggest that dopamine transmission is slow and diffuse. Recent data from several laboratories, including some generated during the previous funding cycle, however, have revealed that dopamine transmission is highly dynamic and, in some cases, remarkably precise. Furthermore, dopamine release is powerfully and rapidly regulated by local cholinergic interneurons in the striatum. These findings suggest that the coding of dopamine volume transmission is more precise than previously thought. A major question that arises is how the architecture for dopamine transmission can support precise signaling. Our overarching model is that molecular machinery has evolved to support broad dopamine coding scales. We build on our previous findings that axonal dopamine exocytosis is executed with millisecond precision by sparse, sophisticated protein machinery typically present at synapses. In aim 1, we zoom in on the powerful local regulation and ask how cholinergic neurons trigger dopamine release. Based on preliminary data, we hypothesize that activity in cholinergic interneurons induces ectopic action potential firing in dopamine axons to trigger dopamine secretion. Our goal is to test this hypothesis and to understand the underlying mechanisms. Identification of an endogenous mechanism for ectopic axonal action potential initiation away from the dopamine neuron soma has important implications for dopamine neuron function. In aim 2, we dissect the organization of dopamine receptors relative to release sites. We build on recent work that identified markers for these sparse secretory sites. Our preliminary data reveal that dopamine receptors are clustered one to two micrometers away from release sites and suggest differences in D1 vs. D2 receptor distributions. We will systematically assess release-receptor organization in super-resolved 3D-images of large striatal volumes and will mechanistically dissect how it is set up. We propose that the organization is different from nanoscale synaptic structure and from the diffuse organization often associated with volume transmission, and may be suited to mediate distinct pathway activation by switches in dopamine neuron firing modes. Our work will dissect the organization of specialized dopamine signaling architecture and rapid, local triggering mechanisms of dopamine release in the vertebrate striatum.