The Action Potential as a Modulator of Synaptic Transmission

作为突触传递调节剂的动作电位

• 批准号: 9911037
• 负责人: Lauren C Panzera
• 金额: $ 4.5万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-12 至 2021-09-11
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9911037
• 关键词: Action Potentials; Affect; Affinity; Arrhythmia; Ataxia; Axon; Cells; Chelating Agents; Chemicals; Clustered Regularly Interspaced Short Palindromic Repeats; Coupling; Critical Thinking; Data; Development; Disease; Electrodes; Epilepsy; Exocytosis; Family; Frequencies; Gene Silencing; Goals; Heterogeneity; Hippocampus (Brain); Image; Impairment; Individual; Ion Channel; Kinetics; Measurement; Measures; Mediating; Membrane; Memory; Monitor; Mutation; National Research Service Awards; Nature; Nerve; Neuraxis; Neurons; Optics; Pharmacology; Phenotype; Physiological; Play; Potassium Channel; Presynaptic Terminals; Probability; Process; Protein Isoforms; Proteins; Radial; Resolution; Role; Scientist; Shapes; Signal Transduction; Structure; Synapses; Synaptic Transmission; Synaptic Vesicles; Synaptic plasticity; System; Technical Expertise; Testing; Time; Toxin; Training; Variant; Vesicle; Voltage-Gated Potassium Channel; Width; Work; chemical release; dendrotoxin; excitatory neuron; experience; experimental study; inhibitor/antagonist; insight; knock-down; mossy fiber; neuronal cell body; neuroregulation; neurotransmission; neurotransmitter release; new therapeutic target; optogenetics; patch clamp; presynaptic; response; sensor; submicron; synaptic function; synaptotagmin; synaptotagmin I; tool; voltage; voltage gated channel

Project Summary The action potential (AP) is a command signal that sharply controls the activity of voltage-gated Ca2+ channels (Cavs) and neurotransmission. The AP waveform has traditionally been considered to be a uniform, binary signal as it propagates across an axonal arbor, however recent work has suggested there is a surprising amount of variability in the width of the waveform arising from a heterogeneous distribution of voltage-gated Na+ and K+ channels across synapses. In the hippocampus, neurons burst in high frequency trains that undergo waveform broadening though it is unclear what the ramifications of this broadening are. A fundamental gap exists in understanding how variations in the AP waveform mechanistically affect neurotransmission as an experimental approach is required with subcellular resolution that can integrate at the microsecond time scale. The development of optogenetics has provided opportunities for manipulating and imaging activity within the small en passant synapses of the central nervous system such as in the hippocampus. These experiments will provide the first optical measurements of presynaptic APs, Ca2+ influx and exocytosis in single hippocampal neurons. Modulation of the AP waveform is achieved pharmacologically by inhibiting a family of voltage-gated K+ channels which results in a predictable broadening of the AP. An unexpected phenotype occurs with this treatment in excitatory neurons: a dramatic increase in exocytosis corresponds with a minimal increase in Ca2+, suggesting an uncoupling of the clearly defined Ca2+ and vesicle fusion relationship and an enhancement of synaptic efficacy. The central hypotheses of this proposal are that a broadened AP waveform alters the Ca2+ contribution of a specific Cav isoform, changes the radius of Ca2+ microdomains and/or differentially activates vesicle fusion machinery. The overall objective of this proposal is to investigate the mechanisms behind the transduction of an electrical action potential signal to the chemical release of neurotransmitter. The long-term goals are to determine how heterogeneity of the AP waveform informs synaptic strength and plasticity. Specific Aim 1 will determine how waveform broadening modulates presynaptic Cav isoform contribution using selective toxins to isolate specific isoforms with and without a broadened AP. Specific Aim 2 will determine how AP shape alters Ca2+ microdomains and vesicle exocytosis. Using optogenetics, gene silencing and pharmacology these experiments will demonstrate if AP broadening influences the radius of microdomains or differentially activates the protein Ca2+ sensors that mediate exocytosis. Given the complexity of this system as well as the essential nature of electrical signaling in excitable cells it is unsurprising that mutations in the voltage- gated channels which control the shape of the AP are implicated in several diseases including epilepsy, ataxia and arrhythmias. Due to the role the AP plays as an information carrier by modulating intracellular Ca2+ and subsequently neurotransmission, there is a critical need for a better understanding of this level of neural regulation.

项目摘要 动作电位（AP）是一种指令信号，它急剧控制电压门控性Ca 2+的活动 通道（Cavs）和神经传递。AP波形传统上被认为是均匀的， 二元信号，因为它传播通过轴突乔木，但最近的工作表明，有一个令人惊讶的 由电压门控Na+的不均匀分布引起的波形宽度的变化量 和跨突触的K+通道。在海马体中，神经元以高频率的序列爆发， 波形展宽，尽管不清楚这种展宽的后果是什么。一个根本的差距存在 在理解AP波形的变化如何机械地影响神经传递方面， 这种方法需要具有亚细胞分辨率，可以在微秒时间尺度上积分。的 光遗传学的发展提供了在小范围内操纵和成像活动的机会。 中枢神经系统的传递突触，例如海马体中的突触。这些实验将提供 首次光学测量单个海马神经元突触前AP、Ca 2+内流和胞吐。 AP波形的调制是通过抑制一个电压门控K+通道家族而间接实现的 这导致AP的可预测的加宽。这种治疗会出现意想不到的表型， 兴奋性神经元：胞吐的急剧增加对应于Ca 2+的最小增加，表明 明确定义的Ca 2+和囊泡融合关系的解偶联和突触的增强， 功效该建议的中心假设是，加宽的AP波形改变了Ca 2+的贡献 改变Ca 2+微区的半径和/或差异激活囊泡融合 机械.这项建议的总体目标是调查转导背后的机制， 电动作电位信号的神经递质的化学释放。长期目标是 确定AP波形的异质性如何通知突触强度和可塑性。 具体目标1将确定波形加宽如何使用以下方法调制突触前Cav亚型贡献： 选择性毒素以分离具有和不具有加宽AP的特定同种型。具体目标2将决定如何 AP形状改变Ca 2+微区和囊泡胞吐。利用光遗传学，基因沉默和 这些实验将证明AP加宽是否影响微区的半径， 差异激活介导胞吐作用的蛋白质Ca 2+传感器。鉴于该系统的复杂性， 以及可兴奋细胞中电信号的基本性质，电压- 控制AP形状的门控通道与几种疾病有关， 和心律失常。由于AP通过调节细胞内Ca 2+和Ca ~（2+）而作为信息载体的作用， 随后是神经传递，迫切需要更好地了解这一水平的神经传递 调控