Imaging Molecules and Circuits in Peripheral Taste Pathways

描述（由申请人提供）：我们开发了一种创新的方法来记录味觉传入神经元的味觉诱发活动，具有良好的细胞和时间分辨率。该方法使用共聚焦功能钙成像和表达GCaMP3的转基因小鼠在感觉神经元。通过仔细地暴露活体麻醉gcamp3小鼠的腿状神经节，并对其口腔施加味觉刺激，我们现在可以记录稳健可靠的反应，从而辨别味觉诱发信号从味蕾到后脑传递背后的原理。这些数据将大大扩展鼓室索和大浅表岩神经或膝状神经节细胞的单单位电记录。我们能够同时记录大量的神经元。我们还设计了一种强大的策略，将单个膝状神经节神经元的功能反应谱与其基因表达模式联系起来，并为神经元的不同功能类别建立了强大的分子标记。在两个成熟的实验室的共同努力下，我们现在提出了一个多pi项目来开发这种新颖的制备方法，并回答有关味道的关键问题。我们的新方法将极大地扩展我们对味觉外周感觉处理的知识。我们的初步数据证明了所有4个紧密聚焦的具体目标的可行性：目标1：味觉感觉传入细胞如何对甜、咸、苦、酸、鲜味和脂肪做出反应？目标2：混合味觉刺激是否能增强味觉感觉传入神经元的反应？目标3：表达某些递质受体的神经节神经元是否支配特定的味觉细胞？目标4：是否存在检测每种味觉刺激的专用神经元，如果存在，是否可以识别与每种味觉质量的味觉传入神经元反应相关的特定分子标记？

DESCRIPTION (provided by applicant): We have developed an innovative method to record taste-evoked activity in gustatory afferent neurons with good cellular and temporal resolution. The method uses confocal functional calcium imaging and transgenic mice that express GCaMP3 in sensory neurons. By carefully exposing geniculate ganglia in living, anesthetized GCaMP3-mice and applying taste stimuli to the oral cavity, we can now record robust and reliable responses to discern the principles behind the transmission of gustatory evoked signals from taste buds to the hindbrain. These data will considerably extend single unit electrical recordings from the chorda tympani and greater superficial petrosal nerves or from geniculate ganglion cells. We are able to recording simultaneously from large ensembles of neurons. We have also designed a powerful strategy to relate the functional response profiles of individual geniculate ganglion neurons to their patterns of gene expression and establish robust molecular markers for separate functional classes of neurons. In a concerted effort from two well-established laboratories, we now propose a multi-PI project to exploit this novel preparation and to answer key questions regarding taste. Our new approach will tremendously expand our knowledge of peripheral sensory processing in taste. Our preliminary data demonstrate the feasibility of all 4 tightly-focused Specific Aims: Aim 1: How do gustatory sensory afferent cells respond to sweet, salty, bitter, sour, umami tastes and fats? Aim 2: Do mixtures of taste stimuli enhance responses from gustatory sensory afferent neurons? Aim 3: Do ganglion neurons that express certain transmitter receptors innervate specific taste cells? Aim 4: Are there dedicated neurons that detect each taste stimulus, and if so, can specific molecular markers be identified that associate with gustatory afferent neuron responses for each taste quality?