Molecular and cellular mechanisms underlying activity dependent gene regulation in neurons

在神经元中，膜去极化导致即时早期基因转录因子(IEG-TF)的表达，包括NPAS4，它调节与可塑性相关的基因表达程序。IEG-TF在体内被广泛用作识别任务相关神经元的工具，但目前尚不清楚这些蛋白是在动作电位(AP)输出或突触输入(EPSP)发生变化时诱导的。AP和EPSP是否能导致不同的基因调控模式和细胞表型更是鲜为人知。这种信息“传递函数”是神经元如何监控和调节自身活动的重要组成部分。在NPAS4这一调节兴奋性-抑制性(E-I)平衡的IEG-TF的具体案例中，研究这一传递函数将为揭示E-I平衡失调引起的神经发育和精神障碍的机制提供有价值的见解。我们已经开发出一种从小鼠身上制备急性海马片的方法，它允许我们在一个完整的电路环境中，从确定的输入群体中独立地诱发AP或EPSP。我们建议研究NPAS4表达的活性要求以及每种类型的神经细胞所遵循的不同的基因组和突触调节。 刺激。我们已经用这种方法证明了AP和EPSP导致NPAS4表达具有不同的时空分布，并有大量的初步结果表征了意想不到的潜在机制。使用为这一提议开发的方法，结合电生理学、光学和测序技术，我们准备确定AP和EPSP如何不同地影响活性依赖的基因调控和突触功能。这一提议与通常研究IEG-TF的方式有很大不同。这些目标的实现将对神经元中依赖活动的基因调控机制以及这种生物学如何在自闭症、谱系障碍和精神分裂症等大脑疾病中受到干扰产生重要的新见解。

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.