Chronic imaging of odor-evoked activity in olfactory bulb circuits of awake mice

描述（申请人提供）：嗅觉在我们的生活质量中起着重要的作用。然而，控制大脑中嗅觉信息处理和表征的机制尚不清楚。在哺乳动物中，嗅球是大脑中负责嗅觉信息初始处理的关键区域。我们目前对嗅球功能的大部分理解是基于先前使用体外脑切片或麻醉动物的急性制剂的研究。虽然这些方法提供了有价值的见解，但对清醒、行为正常的动物的气味编码知之甚少。例如，在长时间尺度（即几天、几周、几个月）内，同一动物的气味表征的动态是什么？在清醒、有行为的动物中，嗅球中的气味编码是如何通过经验和学习形成的？为了解决这些问题，我们提出了一种实验策略，使用慢性双光子钙成像来研究清醒小鼠嗅球的气味诱发活动。特异性目标1提出了一种利用cre依赖的腺相关病毒系统，在主要（二尖瓣/簇状）细胞或局部中间神经元中选择性表达遗传编码钙指示剂GCaMP3的方法。我们假设，这种方法将提供单细胞分辨率的动作电位依赖钙信号，在大量的神经系统中，可以长期成像的清醒，头部固定的小鼠。特异性目标2提出了成像实验，以确定二尖瓣细胞和中间神经元（颗粒细胞）在清醒和麻醉状态下气味诱发反应的差异。我们还将在清醒的动物身上使用慢性成像技术来确定气味诱发的神经系统活动模式是稳定的还是动态的。这些实验将对清醒大脑中气味编码的本质提供新的见解。

DESCRIPTION (provided by applicant): The sense of smell plays an important role in our quality of life. However, the mechanisms governing the processing and representation of olfactory information in the brain are not well understood. In mammals, the olfactory bulb is a critical brain region responsible for the initial processing of olfactory information. Much of our current understanding of olfactory bulb function is based on previous studies using in vitro brain slices or acute preparations of anesthetized animals. While these approaches have provided valuable insight, much less is known about odor coding in awake, behaving animals. For example, what are the dynamics of odor representations in the same animal over long time scales (i.e. days, weeks, months) and how is odor coding in the olfactory bulb shaped by experience and learning in awake, behaving animals? To address these questions, we propose an experimental strategy using chronic 2-photon calcium imaging to study odor-evoked activity in the olfactory bulbs of awake mice. Specific Aim 1 proposes the development of an approach for the selective expression of the genetically-encoded calcium indicator GCaMP3 in principal (mitral/tufted) cells or local interneurons using a Cre-dependent, adeno-associated virus system. We hypothesize that this approach will provide single cell resolution of action potential-dependent calcium signals in large populations of neural ensembles that can be imaged chronically in awake, head fixed mice. Specific Aim 2 proposes imaging experiments to determine how odor-evoked responses of mitral cells and interneurons (granule cells) differ between the awake and anesthetized state. We will also use chronic imaging in awake animals to determine whether patterns of odor-evoked activity in neural ensembles are stable or dynamic over days of repeated testing. These experiments will provide new insight into the nature of odor coding in the awake brain.