Molecular and cellular mechanisms underlying activity dependent gene regulation in neurons

项目摘要/摘要 在神经元中，膜去极化导致即刻早期基因的表达 转录因子(IEG-TF)，包括NPAS4，调节基因表达程序 与可塑性有关。IEG-TF被广泛用作识别脑内任务相关神经元的工具 然而，目前尚不清楚这些蛋白质是否因动作电位的变化而被诱导 (AP)神经元的输出或突触输入(EPSP)。更不知道AP是否 EPSPS可以导致不同的基因调控模式和细胞表型。这 信息传递函数是神经元监控和调节的重要组成部分 他们自己的活动。在NPAS4的特定情况下，调节兴奋性抑制的IEG-TF (E-I)平衡，研究这一传递函数将对机制提供有价值的见解 由E-I调节失调引起的潜在神经发育和精神障碍 平衡。我们已经从小鼠身上开发出一种急性海马片制剂 允许我们从定义的输入群体中独立地调用AP或EPSP 完整电路的上下文。我们建议调查NPAS4的活动需求 每种类型的表达及其不同的基因组和突触调节 刺激。我们已经使用此方法来演示AP和EPSP会导致NPAS4 表达具有不同的时空分布，并有广泛的初步结果 描述了意想不到的潜在机制。使用为此开发的方法 建议，结合电生理学、光学和测序技术，我们正在 准备确定AP和EPSP如何不同地影响活动依赖基因 调节和突触功能。这一提议与IEG-TF的方式有很大的不同 通常被研究过。这些目标的实现将产生对 神经元中活性依赖的基因调控机制以及这种生物学是如何在 脑部疾病，如自闭症、谱系障碍和精神分裂症。

Project Abstract/Summary In neurons, membrane depolarization leads to the expression of immediate early gene transcription factors (IEG-TFs), including NPAS4, that regulate programs of gene expression associated with plasticity. IEG-TFs are widely used as tools to identify task-relevant neurons in vivo, yet it is unclear if these proteins are induced in response to changes in the action potential (AP) output or synaptic inputs (EPSPs) to the neuron. Even less is known about whether APs and EPSPs can lead to distinct patterns of gene regulation and cellular phenotypes. This information “transfer function” is an essential component of how neurons monitor and regulate their own activity. In the specific case of NPAS4, an IEG-TF that regulates excitatory-inhibitory (E-I) balance, studying this transfer function will provide valuable insight into the mechanisms underlying neurodevelopmental and psychiatric disorders that stem from dysregulation of E-I balance. We have developed an acute hippocampal slice preparation from the mouse that allows us to independently evoke APs or EPSPs, from defined populations of inputs, within the context of an intact circuit. We propose investigating the activity requirements for NPAS4 expression and the divergent genomic and synaptic regulation that follows from each type of stimulus. We have used this approach to demonstrate that APs and EPSPs lead to NPAS4 expression with distinct spatio-temporal profiles and have extensive preliminary results characterizing the unexpected underlying mechanisms. Using the methods developed for this proposal, in combination with electrophysiology, optical, and sequencing techniques, we are poised to determine how APs and EPSPs differentially impact activity-dependent gene regulation and synapse function. This proposal is a significant departure from how IEG-TFs are typically studied. The execution of these aims will yield important new insights into the mechanics of activity-dependent gene regulation in neurons and how this biology is disrupted in disorders of the brain such as Autism Spectrum Disorders and schizophrenia.