Neurotransmission in tastebuds

味蕾的神经传递

• 批准号: 9397936
• 负责人: Sue C. Kinnamon
• 金额: $ 44.38万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9397936
• 关键词: Acids; Action Potentials; Address; Affect; Altered Taste; Behavior; Behavioral Paradigm; Brain Stem; Calcium; Cell Nucleus; Cells; Chemicals; Citric Acid; Data; Detection; Development; Dose; Eating; Enterobacteria phage P1 Cre recombinase; Event; Fiber; Gene Targeting; Glossopharyngeal nerve structure; Image; Knock-out; Lead; Light; LoxP-flanked allele; Molecular; Morphology; Mus; NTPDase2; Nerve; Nerve Fibers; Nervous system structure; Neurons; Neurotransmitters; Norepinephrine; Nucleus solitarius; Nutritional; Obesity; Oral; Oral cavity; P2X-receptor; Pattern; Pharmacologic Substance; Pharmacology; Physiological; Play; Poison; Purinoceptor; Receptor Cell; Role; Salts; Serotonin; Serotonin Receptors 5-HT-3; Signal Transduction; Sodium Chloride; Stimulus; Structure of geniculate ganglion; Supporting Cell; Synapses; System; Taste Buds; Taste Perception; Taste aversion; Testing; Tongue; Type II Epithelial Receptor Cell; Type III Epithelial Receptor Cell; Vesicle; afferent nerve; awake; cell type; chorda tympani; ectoATPase; gamma-Aminobutyric Acid; ganglion cell; immunoreactivity; in vivo; mouse model; neurotransmission; neurotransmitter release; optogenetics; patch clamp; presynaptic; receptor; relating to nervous system; response; taste stimuli; taste system; transmission process; uptake

PROJECT SUMMARY Type III “presynaptic” taste cells respond to sour and some salty stimuli with action potentials and release neurotransmitter at conventional synapses with afferent nerve fibers. The identity of the transmitters involved, however, is still unclear. We have shown previously that all taste stimuli (including sour and salty) require ATP signaling to neural P2X receptors for transmission to afferent fibers. Nonetheless, ATP release from Type III cells has not been detected. Conversely, Type III cells do release serotonin (5-HT) which activates 5-HT3 receptors on afferent nerve fibers but neither knockout nor pharmacological inhibition of 5-HT3 completely eliminates responses to acids or salts, suggesting other neurotransmitters are involved. One problem in studying transmission from Type III cells is that the conventional stimuli used to activate these cells -- acids, salts, and KCl -- all have non-specific effects on other cell types in the taste bud. To circumvent this problem, we employ an optogenetic strategy permitting direct stimulation of Type III cells with light rather than chemicals. We developed a mouse that expresses Cre recombinase selectively in type III taste cells relying on Pkd2l1 as a Cre driver. When these Pkd2l1-Cre mice are crossed with “floxed” channelrhodopsin (ChR2) mice ChR2 is faithfully expressed in taste cells immunoreactive for PKD2L1 with no off-target expression in other taste cell types. Flashing 470 nm light onto the tongue elicits responses in the chorda tympani and glossopharyngeal nerves that are robust and repeatable, resembling responses to sour and salty stimuli. In this proposal we will utilize these Pkd2l1-ChR2 mice plus other gene-targeted mice to investigate the neurotransmitters that are used by these cells to communicate with the nervous system, and the perceptual qualities evoked by selective stimulation of PKD2l1 Type III cells. Aim 1 will investigate the role of ATP and other neurotransmitters (i.e., 5-HT, GABA, and NE) in activating geniculate ganglion neurons that selectively innervate Type III cells. Calcium imaging of isolated geniculate ganglion cells will identify transmitters that activate these neurons, and chorda tympani and glossopharyngeal nerve responses to light in the Pkd2l1- ChR2 mice will allow us to test the role of their cognate receptors in vivo. Aim 2 will address the perceptual quality elicited by stimulation of Pkd2l1-ChR2 cells, using behavioral paradigms, and cFos activation will be used to address the central representation of PKD2L1-expressing Type III cells. In Aim 3 we will test the hypothesis that PKD2L1-Type III cells participate in an intragemmal (intra taste bud) circuit, resulting in modulation of other taste qualities, particularly those transduced by Type II cells. Results from these studies will provide important new information about transduction, afferent neurotransmission, and intragemmal signaling in Type III taste cells.

项目概要 III 型“突触前”味觉细胞通过动作电位响应酸味和一些咸味刺激并释放 具有传入神经纤维的传统突触处的神经递质。涉及的发射机的身份， 然而，目前还不清楚。我们之前已经证明，所有味觉刺激（包括酸味和咸味）都需要 ATP 向神经 P2X 受体发送信号以传输至传入纤维。尽管如此，III 型中的 ATP 释放 尚未检测到细胞。相反，III 型细胞会释放血清素 (5-HT)，从而激活 5-HT3 传入神经纤维上的受体，但既没有完全敲除也没有完全药物抑制 5-HT3 消除了对酸或盐的反应，表明其他神经递质也参与其中。一个问题在 研究 III 型细胞的传输的方法是，用于激活这些细胞的传统刺激物——酸、 盐和氯化钾——都对味蕾中的其他细胞类型产生非特异性影响。为了规避这个问题， 我们采用光遗传学策略，允许用光直接刺激 III 型细胞，而不是 化学品。我们开发了一种在 III 型味觉细胞中选择性表达 Cre 重组酶的小鼠，依赖于 Pkd2l1 作为 Cre 驱动程序。当这些 Pkd2l1-Cre 小鼠与“floxed”视紫红质通道 (ChR2) 小鼠杂交时 ChR2 在对 PKD2L1 具有免疫反应性的味觉细胞中忠实表达，在其他细胞中没有脱靶表达 味觉细胞类型。将 470 nm 的光闪烁到舌头上会引起鼓索和 舌咽神经强健且可重复，类似于对酸和咸刺激的反应。在 根据该提议，我们将利用这些 Pkd2l1-ChR2 小鼠和其他基因靶向小鼠来研究 这些细胞使用神经递质与神经系统和感知系统进行通信 通过选择性刺激 PKD2l1 III 型细胞而引发的品质。目标 1 将研究 ATP 的作用和 其他神经递质（即 5-HT、GABA 和 NE）选择性地激活膝状神经节神经元 支配 III 型细胞。分离的膝状神经节细胞的钙成像将识别发射器 激活这些神经元，鼓索和舌咽神经对 Pkd2l1- 中的光做出反应 ChR2 小鼠将使我们能够测试其同源受体在体内的作用。目标 2 将解决感知问题 使用行为范例刺激 Pkd2l1-ChR2 细胞而引发的质量，以及 cFos 激活将 用于解决表达 PKD2L1 的 III 型细胞的中心表达。在目标 3 中，我们将测试 假设 PKD2L1-III 型细胞参与胞内（味蕾内）回路，导致 其他味觉品质的调节，特别是由 II 型细胞转导的味觉品质。这些研究的结果 将提供有关转导、传入神经传递和胞内神经传递的重要新信息 III 型味觉细胞中的信号传导。