MEMORY CONSOLIDATION: HIPPOCAMPUS & GENE EXPRESSION

记忆是分子、细胞和大脑系统相互作用的结果。为了更好地治疗和诊断影响记忆和认知的心理健康疾病，我们必须了解大脑中这些不同水平发生的复杂相互作用。这项研究计划的长期目标是增加对经验依赖型神经元基因表达在长期记忆(LTM)形成中的作用的理解。在这个广泛的框架内，这一提议的实验将有助于确定：(1)感觉信息如何影响海马结构(HF)中的即刻早期基因(IEG)转录，以及(2)任务需求如何影响HF中特定神经元群体的IEG转录。虽然这项研究的重点是在系统神经生物学和行为学的水平上，但拟议研究的结果将为未来重点研究不同生理条件下控制IEG表达的分子机制提供重要的初步信息。我们将使用我们最近开发的一种改进的荧光原位杂交(FISH)技术来可视化脑组织中的多个不同的IEG RNA。这种FISH分析非常敏感，能够检测脑神经元中活跃的IEG转录(转录焦点，或TF)的假定位置。我们已经在大鼠身上进行了一系列实验，以表明在电刺激或探索新环境的2分钟内，可以看到许多IEG的Tf。此外，在行为和电刺激后30分钟，这种诱导转录是瞬时的，IEG TF处于基线水平，在这个时间点，新生的mRNA分布在细胞质中。由于核内和胞浆内RNA积累的时间进程不同，通过共聚焦显微镜测定的RNA在亚细胞内的分布可以用来推断单个神经元在两个不同时间的活动历史。我们将这种方法命名为功能性脑成像细胞隔间分析鱼的时间活动，或称鲶鱼。我们已经开始使用IEG鲶鱼来检查离散的感觉信息对HF中IEG转录激活模式的作用。在许多这样的研究中，我们利用鲶鱼的时间分辨率特性来了解离散的行为经验是如何影响不同神经元群体的转录模式的。我们的早期数据表明，两组CA1神经元在探索两种环境后诱导IEG Arc的转录，而两次探索同一环境只能激活一组神经元中Arc RNA的转录。这提供了令人信服的证据，证明CA1神经元中的IEG转录与信息处理有关，而不是受到普遍或非特定影响的驱动。来自拟议研究的数据将为经验依赖型IEG表达在记忆形成过程中的作用，以及在动物处理不同类型信息时发生在HF内的神经元群体相互作用提供新的见解。

Memory is the result of molecular, cellular, and brain system interactions. To better treat and diagnose mental health disorders that affect memory and cognition, we must understand the complex interactions that occur at these different levels in the brain. The long-term objective of this research program is to increase understanding on the role of experience-dependent neuronal gene expression in the formation of long-term memory (LTM). Within this broad framework, the experiments of this proposal will help determine: (1) how sensory information affects immediate-early gene (IEG) transcription in the hippocampal formation (HF) and (2) how task demands influence IEG transcription in defined neuronal populations in the HF. While the central focus of this research is at the level of systems neurobiology and behavior, the findings from the proposed studies will provide important primary information for future focused studies on the molecular mechanisms that control IEG expression under different physiologic conditions. We will use a modified fluorescent in situ hybridization (FISH) protocol that we developed recently to visualize multiple different IEG RNAs in brain tissue. This FISH assay is extremely sensitive and is capable of detecting the putative sites of active IEG transcription (transcription foci, or TF) in brain neurons. We have performed a series of experiments in rats to show that TF for a number of IEGs can be visualized within 2 minutes of either an electrical stimulus or by exploration of a new environment. Moreover, this induced transcription is transient with IEG TF at baseline levels by 30 minutes after behavioral and electrical stimulus, and the nascent mRNA found distributed in the cytoplasm at this time point. Because the time course of nuclear versus cytoplasmic RNA accumulation is distinct, the subcellular distribution of RNA, determined by confocal microscopy, can be used to infer the activity history of individual neurons at two different times. We have termed this approach to functional brain imaging cellular compartment analysis of temporal activity by FISH, or catFISH. We have begun to use IEG catFISH to examine the role that discrete sensory information has on patterns of IEG transcriptional activation in the HF. In many of these studies, we exploit the temporal resolution properties of catFISH to see how discrete behavioral experiences, separated in time by 20 minutes, influence transcriptional patterns in different neuronal populations. Our early data show that two populations of CA1 neurons induce transcription of the IEG Arc following exploration of two environments, whereas exploration of the same environment twice only activates transcription of Arc RNA in one population of neurons. This provides compelling evidence that IEG transcription in CA1 neurons is linked to information processing, and is not driven by generalized or nonspecific influences. Data from the proposed studies will provide new insights into the role of experience-dependent IEG expression during memory formation, and in the neuronal population interactions that occur within the HF as an animal processes different types of information.