Neuronal mechanisms controlling number and function of presynaptic mitochondria

描述（由申请人提供）：我们的总体目标是阐明控制线粒体以满足神经末梢能量需求的神经元机制。线粒体积聚在神经末梢，在那里它们产生释放和循环神经递质所需的大部分ATP。因此，神经功能依赖于线粒体产生足够的ATP来维持神经递质释放。类似地，线粒体促进突触前Ca2+稳态。神经元Ca2+稳态的失败具有灾难性的后果，是许多神经退行性疾病的标志。令人惊讶的是，我们对协调线粒体数量和功能与突触前能量需求的机制知之甚少，但了解这些机制对于理解神经退行性疾病的进展至关重要。我们的中心假设是神经元机制控制线粒体的数量和功能以适应突触前的能量需求，并且这些机制是突触特异性的。我们建议阐明果蝇幼虫肌肉骨骼系统中的这些机制，其中每个运动神经元终端具有不同的工作速率，我们可以使用电生理和Ca2+成像技术进行量化。突触前能量需求的多样性，遗传易感性和神经生理技术的可及性，使其成为研究控制线粒体以适应突触前能量需求的神经元机制的理想系统。在目的1中，我们将确定线粒体是否以与其工作速率成正比的数量供应给运动神经末梢。3D-EM重建将用于确定线粒体数量。我们还将探讨线粒体数量与功能之间的关系。在Aim 2中，我们将验证线粒体体积在同一轴突上不同末端的水平上受到控制的假设。将在单个终端确定线粒体功能参数，以测试同一轴突上不同终端的线粒体功能是否可能不同。在Aim 3中，我们将测试假设，在发育过程中，活动区间距和钮扣直径调整到运动神经元的放电速率，将突触前Ca2+水平带入突触前活动期间刺激线粒体能量代谢最有效的范围。到目前为止，Ca2+调节是突触前ATP的大量消耗者，这种突触前生物能量的原位模型将为更好地理解神经退行性疾病中涉及线粒体功能障碍和Ca2+失调的早期事件提供必要的背景。

DESCRIPTION (provided by applicant): Our overall goal is to elucidate the neuronal mechanisms that control mitochondria to satisfy the energy demands of nerve terminals. Mitochondria accumulate within nerve terminals where they generate most of the ATP required for the release and recycling of neurotransmitters. Neural function, therefore, relies on mitochondria generating sufficient ATP to sustain neurotransmitter release. Similarly, mitochondria power presynaptic Ca2+ homeostasis. A failure in neuronal Ca2+ homeostasis has catastrophic consequences and is a hallmark of many neurodegenerative diseases. Surprisingly, we know very little about the mechanisms that coordinate mitochondrial number and function with presynaptic energy requirements, yet understanding these mechanisms will be critical to understanding the progression of neurodegenerative disease. Our central hypothesis is that neuronal mechanisms control the number and function of mitochondria to accommodate presynaptic energy requirements, and that these mechanisms are synapse specific. We propose to elucidate these mechanisms in the musculoskeletal system of the fruit fly larva, where each motor neuron terminal has a different work rate which we can quantify using electrophysiological and Ca2+-imaging techniques. Diversity in presynaptic energy requirements, genetic tractability and accessibility to neurophysiological techniques, make this an ideal system in which to investigate neuronal mechanisms that control mitochondria to accommodate presynaptic energy requirements. In Aim 1, we will determine whether mitochondria are supplied to motor nerve terminals in numbers that are proportional to their work rate. 3D-EM reconstruction will be used to determine mitochondrial number. We will also probe the relationship between mitochondrial number and function. In Aim 2, we will test the hypothesis that mitochondrial volume is controlled at the level of different terminals on the same axon. Mitochondrial functional parameters will be determined at individual terminals to test whether mitochondrial function may be different between terminals on the same axon. In Aim 3, we will test the hypothesis that, over the course of development, active zone spacing and bouton diameter adjust to the firing rate of the motor neuron to bring presynaptic Ca2+ levels into a range most effective at stimulating mitochondrial energy metabolism during presynaptic activity. In so far as Ca2+ regulation is a heavy consumer of presynaptic ATP, this in situ model of presynaptic bioenergetics will provide an essential context for a better understanding of the early events involving mitochondrial dysfunction and Ca2+ dysregulation in neurodegenerative disease.