Cell type-specific calcium imaging during hippocampus-dependent memory

描述(由申请人提供)：来自人类和动物的证据表明，海马体在形成长期情景记忆中起着关键作用。海马体由解剖学上分离的亚区组成，假设有多种神经元类型来编码一种体验的不同方面。尽管海马神经元活动依赖可塑性的分子机制已经被仔细研究，但我们对特定神经元群体在记忆编码、巩固和回忆过程中所表现出的活动模式的了解有限。因此，人们强烈需要开发新的成像方法，能够根据海马体电路被认为支持的行为来询问它们的活动。由于神经元尖峰与大的细胞内钙电导有关，钙指示剂可以作为神经元激活的替代指标。在这项拟议的研究中，我们将使用光纤荧光内窥镜(FFE)来长期监测接受海马区依赖任务的小鼠海马神经元的激活情况。在CA1锥体细胞、齿状颗粒细胞和CA3锥体细胞中选择性表达Cre重组酶的转基因小鼠中，将有条件地表达遗传编码的钙指示剂。我们最初在完全清醒的、不受限制的小鼠身上进行的原理验证实验表明，这种方法对于单个神经元活动的大群体成像是可行的。在我们的第一个目标中，我们将优化FFE的参数，以实现对不同神经元群体的稳定、长期成像。在我们的第二个目标中，我们将使用这种方法来定义编码背景恐惧和痕迹恐惧条件反射的神经元集合，这是两种海马体依赖的联想记忆变体，假设它们参与不同的电路。延时实验将可以比较在训练、巩固和回忆过程中观察到的激活模式。通过对这些记忆阶段的每个阶段进行遗传或药物干预，我们将把选定神经元的活动与认知联系起来。因此，这项拟议的研究将通过引入FFE作为一种新的体内方法来研究记忆机制，并将利用这一技术开始解决以前在细胞基础上无法解决的记忆问题，从而推动该领域的发展。 公共卫生相关性：记忆障碍与衰老和精神疾病有关。该项目使用先进的钙成像在行为小鼠，以揭示在记忆形成和提取过程中不同类型的海马细胞中神经元激活的特性。因此，这项研究可能会确定特定的细胞靶点或活动依赖的网络模式，可以利用这些模式来防止记忆损伤。

DESCRIPTION (provided by applicant): Evidence from humans and animals indicate a critical role for the hippocampus in the formation of long- term episodic memories. The hippocampus consists of anatomically segregated subregions with a diversity of neuronal types hypothesized to encode different aspects of an experience. Although the molecular mechanisms of activity-dependent plasticity in hippocampal neurons have been intensely scrutinized, we have limited understanding of the activity patterns shown by select neuronal populations during memory encoding, consolidation and recall. Therefore, there is a strong demand to develop new imaging approaches that can interrogate the activity of hippocampal circuits in the context of the behavior that they are thought to support. Because neuronal spikes are associated with large intracellular calcium conductances, calcium indicators can be used as a proxy for neuronal activation. In the proposed research, we will implement the use of fiber optic fluorescence endomicroscopy (FFE) to chronically monitor the activation of defined hippocampal neurons in mice subjected to hippocampus-dependent tasks. A genetically-encoded calcium indicator will be conditionally expressed in transgenic mice selectively expressing Cre recombinase in CA1 pyramidal cells, or in dentate granule cells and CA3 pyramidal cells. Our initial proof-of-principl experiments in fully awake, unrestricted mice advocate the feasibility of this approach for large-population imaging of single-neuron activity. In our first aim, we will optimize the parameters of FFE required to achieve stable, long-term imaging of the different neuronal populations. In our second aim, we will use this method to define the neuronal ensembles that encode contextual- and trace-fear conditioning, two hippocampus-dependent variants of associative memory hypothesized to engage different circuits. Time-lapse experiments will allow comparisons of the activation patterns observed during training, consolidation and recall. Through genetic or pharmacological interventions against each one of these memory phases, we will link the activity of select neurons to cognition. Thus, the proposed research will advance the field by introducing FFE as a novel in vivo approach to study memory mechanisms, and will use this technique to begin to address previously inaccessible questions on the cellular basis of memory. PUBLIC HEALTH RELEVANCE: Memory impairments are associated with aging and psychiatric diseases. This project uses advanced calcium imaging in behaving mice to reveal the properties of neuronal activation in different types of hippocampal cells during memory formation and retrieval. Consequently, this research may identify specific cellular targets or activity-dependent network patterns that can be exploited to prevent memory impairments.