Multiplex interrogation of neuromodulatory signaling in behaving animals with enhanced depth and resolution

项目摘要 哺乳动物大脑对环境变化的动态适应性是显著的，因为它的复杂性 在允许这种适应的操作基础上的神经元网络。虽然我们有一些 对这一现象的解剖学和功能基础的了解，我们仍然缺乏详细的图像如何 神经元活动的调节起作用。这些神经调节剂释放的时间和位置是什么？ 与兴奋/抑制回路的关系？神经调节剂回路是如何完成这一调节的 靶向神经元的放电和突触特性？填补这些知识空白将推动我们的 了解神经调节剂生物学的各个方面，并有助于发现新的治疗策略。至 帮助缩小这一差距，我们已经使用了创造性的方法来开发基因编码的直接 报道行为触发和调节的神经调节剂释放，包括5-羟色胺(5-羟色胺)和多巴胺(DA) 去甲肾上腺素(NE)。我们已经向神经科学界传播了这些指标，并鼓励 调节神经调节剂释放、潜在动机和成瘾的新机制的重大发现。 在这一初步成功的基础上，我们建议进一步扩展此NM传感器工具箱的有效性，以 启用深度和亚细胞分辨率的成像稀疏释放。我们的具体目标是：(1)提高 我们目前传感器的敏感度使稀疏神经调节剂释放能够进行稳健的成像，推动其空间 分辨率达到亚细胞水平，并在较低浓度下增加响应的线性；(2)扩展其 光谱范围到红色/远红，以增强成像深度、SNR和活体多路测量 使用两种或三种不同的颜色操作多个电路组件，以及(3)描述可能的 电流传感器对内源信号的干扰，并系统地验证新出现的传感器 大范围的显微镜接近活体。我们的策略依赖于传感器之间的动态协作 设计团队和最终用户获得持续反馈，以实施对传感器的有效改进。 我们的目标是迅速传播一系列具有良好特点的、高度敏感的指标，以 神经科学界将被用来研究老鼠、鱼、苍蝇和蠕虫的行为，以丰富我们的知识 关于神经调节剂在大脑回路中的功能作用。

Project Summary The dynamic adaptability of the mammalian brain to environmental changes is remarkable, as it is the complexity of the networks of neurons underlying the operations that allow for such adaptations. Although we have some understanding of the anatomical and functional basis of this, we are still lacking a detailed picture of how the modulation of neuronal activity works. What is the timing and locations of these neuromodulator release and relationship with excitatory/inhibitory circuits? How does the neuromodulators circuitry accomplish the regulation of firing and synaptic properties of targeted neurons? Filling these gaps in knowledge would advance our understanding of all aspects of neuromodulator biology and allow discovery of new therapeutic strategies. To help close this gap, we have used creative approaches to the development of genetically encoded to directly report behaviorally triggered and modulated neuromodulator release including serotonin (5-HT), dopamine (DA) and norepinephrine (NE). We have disseminated these indicators to the neuroscience community and spurred major discoveries of novel mechanisms regulating neuromodulator release underlying motivation and addiction. Build on this initial success, we propose to further expand the effectiveness of this toolbox of NM sensors to enable imaging sparse release at depth and subcellular resolution. Our specific goals are to (1) improve the sensitivity of our current sensors to enable robust imaging of sparse neuromodulator release, push their spatial resolution to the subcellular level and increase linearity of response at lower concentrations; (2) expand their spectral range to red/far-red to enhance imaging depth, SNR and in vivo multiplex measurement and manipulation of multiple circuit components using two or three distinct colors, and (3) characterize the possible interference of current sensors with endogenous signaling and systematically validate emerging sensors with a wide-ranging microscopy approaches in vivo. Our strategy relies on a dynamic collaboration between the sensor design team and end users to obtain continuous feedback to implement efficient improvements to the sensors. It is our goal to rapidly disseminate a wide range of well-characterized, highly sensitive indicators for the neuroscience community to be employed to study behaving mice, fish, flies and worms, to enrich our knowledge on the functional roles of neuromodulators in the brain circuitry.