Molecular and Neural Mechanisms regulating Foraging and Food Intake

项目摘要 在我的实验室里，NIGMS资助的研究的总体目标是了解 大脑将食物的感觉感受与饥饿感整合在一起，以调节食物摄入水平。 分子、细胞和电路。我们使用的是遗传学上易于处理的模式生物--苍蝇（果蝇 研究食物摄入回路如何在大脑中发挥作用和调节。的主要目标之一 这个项目是捕捉神经活动的电路，调节食物的摄入量时，苍蝇正在改变他们的 代谢状态。这些实验需要在长时间尺度（> 6小时）下对苍蝇大脑进行非侵入性成像。 目前用于果蝇光学神经生理学的方法不允许这样的实验，因为表皮开放 通常用于苍蝇神经科学研究的成像制剂显示出对 最多约3 - 4小时后，苍蝇的大脑开始退化。最近，我们开发了一种非侵入性的， 慢性功能成像方法。我们首先表明，与该领域的误解相反， 蝇头角质层在> 900 nm的波长下具有令人惊讶的高透射率，并且穿透角质层的困难 成像是由于头部角质层下面的气囊和/或脂肪组织。完全去除气囊或 通过非侵入性压缩飞头将它们重新定位到成像窗口之外允许光学访问 而不解剖头部角质层。使用我们的穿透表皮成像方法， 通过表皮观察了果蝇脑内的蘑菇体Kenyon细胞和中央复合环神经元 2P和3P显微镜。我们的测量结果表明，2P和3P激发在浅层中表现相似， 区域的苍蝇大脑，但在1320 nm的3P激发是上级更深的大脑结构。我们证明 通过捕获气味诱发的钙离子的透表皮多光子成像的功能成像性能 表达GCaMP6s的蘑菇体Kenyon细胞的反应在短期和长期，如苍蝇， 被剥夺食物。我们计划使用穿透表皮的功能成像来捕捉神经回路的活动， 调节果蝇的味觉和食物摄入。这些神经回路位于大脑的深埋区域， 食管下的苍蝇大脑（食管下区），因此无法用2P激发（920 nm）进入 穿过表皮因此，我们的计划是使用3P激发（1320nm）来对这些深层神经回路进行成像， 完整的苍蝇大脑穿透表皮3P显微镜将使味觉敏感神经元的慢性成像成为可能， 食物摄入回路，因为苍蝇正在改变它们的代谢状态，这将使我们能够回答重要的问题， 在神经科学中，例如：当动物改变其生理状态时，味觉回路如何改变活动 如饥饿或口渴。神经回路是如何编码饥饿/口渴等行为状态的？这个项目 不仅将突破苍蝇体内功能成像的极限，还将回答果蝇体内功能成像的基本问题。 神经科学

PROJECT SUMMARY The overall goal of NIGMS-funded research in my lab is to understand the fundamental principles of how the brain integrates the sensory percept of food with the sensation of hunger to regulate food intake on the level of molecules, cells and circuits. We are using the genetically tractable model organism, the fly (Drosophila melanogaster) to study how food intake circuits function and are regulated in the brain. One of the main aims of this project is to capture neural activity from circuits that regulate food intake when flies are changing their metabolic states. These experiments require non-invasive imaging of the fly brain at long time scales (>6 hours). Current methods used in fly optical neurophysiology do not allow such experiments because the cuticle-open imaging preparations that are commonly used in fly neuroscience research show reliable calcium responses for a maximum of ~3-4 hours before the fly brain starts degenerating. Recently, we have developed a non-invasive, chronic functional imaging method in flies. We first showed that, in contrast to a misconception in the field, the fly head cuticle has surprisingly high transmission at wavelengths > 900 nm, and the difficulty of through-cuticle imaging is due to the air sacs and/or fat tissue underneath the head cuticle. Removing air sacs completely or relocating them out of the imaging window by non-invasive compression of the fly head allows optical access to the fly brain without dissecting away the head cuticle. Using our through-cuticle imaging method, we first imaged the mushroom body Kenyon cells and the central complex ring neurons in the fly brain through the cuticle using 2P and 3P microscopy. Our measurements showed that 2P and 3P excitation performed similarly in shallow regions of the fly brain, but 3P excitation at 1320 nm is superior for deeper brain structures. We demonstrated the functional imaging performance of through-cuticle multiphoton imaging by capturing odor evoked calcium responses from mushroom body Kenyon cells expressing GCaMP6s in short-term and in long-term as flies are being food deprived. We plan to use through-cuticle functional imaging to capture the activity of neural circuits that regulate taste perception and food intake in flies. These neural circuits are located in deeply buried regions of the fly brain below the esophagus (subesophageal zone), therefore cannot be accessed with 2P excitation (920nm) through the cuticle. Therefore, our plan is to use 3P excitation (1320nm) to image these deep neural circuits in the intact fly brain through-cuticle. 3P-microscopy will enable chronic imaging of the taste sensitive neurons and food intake circuits as flies are changing their metabolic states and will allow us to answer important questions in neuroscience, such as: How do the taste circuits change activity when animals are changing their physiological states such as hunger/thirst? How do neural circuits encode behavioral states such as hunger/thirst? This project will not only push the limits of in vivo functional imaging in flies but also answer fundamental questions in neuroscience.