Role of Gadd45b in Cocaine-driven Epigenetic and Behavioral Dynamics

精神兴奋剂滥用是一场公共卫生危机，影响着美国数百万人， 造成深刻的经济、社会和个人伤害。然而，尽管吸毒过量的人数迅速增加， 与可卡因等兴奋剂药物有关的死亡，仍然没有批准的兴奋剂治疗方案 滥用障碍精神兴奋剂药物通过明确的信号机制发挥作用，以提高 多巴胺能神经传递的神经核（NAc），一个关键的奖励相关的大脑结构， 整合来自不同大脑区域的信息，直接影响动机行为。此外，可卡因 引起NAc中中型棘神经元（MSN）的表观遗传和转录重组， 细胞信号传导和突触功能的不适应转变。我们的初步数据表明， Gadd45 b（生长、停滞和DNA损伤诱导蛋白45 b）mRNA在MSNs中上调， 可卡因和多巴胺受体刺激，Gadd45b是可卡因相关记忆所必需的， 阵然而，尽管Gadd45 b在表观遗传重编程和记忆中起着关键作用， Gadd45 b在可卡因相关的表观遗传、分子和行为中的作用 适应性并不明确。在这项提案中，我们将测试Gadd45b调节药物的总体假设， 通过控制NAc中的活动依赖性DNA去甲基化来诱导行为可塑性。具体目标1 该提案将结合联合收割机基于双向CRISPR的操作和单核RNA测序， 确定Gadd45b信号如何影响可卡因的转录反应。《Specific Aim 2》将使用新的 Gadd45b工具和全基因组DNA甲基化谱分析，以确定调节 NAc中Gadd45b依赖的表观遗传编程。最后，具体目标3将使用细胞特异性体内 Gadd45b操作与可卡因和自然奖赏的行为测定组合以测试可卡因的释放。 Gadd45b增强可卡因的行为效应的假说。总之，这些实验将确定 Gadd45b靶基因在NAc，剖析表观遗传机制，其中Gadd45b有助于可卡因的 表观遗传效应，并定义了可卡因相关的行为可塑性的Gadd45 b的作用。这些研究将 揭示了Gadd45b有助于精神兴奋反应的基本机制，并将为 未来的实验将探索滥用药物如何利用表观基因组来改变动机行为。

Psychostimulant abuse is a public health crisis that affects millions of individuals in the United States and results in profound economic, social, and individual harm. However, despite rapid increases in overdose deaths linked to stimulant drugs like cocaine, there are still no approved therapeutic options for stimulant abuse disorders. Psychostimulant drugs act through well-defined signaling mechanisms to elevate dopaminergic neurotransmission in the nucleus accumbens (NAc), a key reward-linked brain structure that integrates information from diverse brain regions to directly influence motivated behavior. Further, cocaine causes epigenetic and transcriptional reorganization in medium spiny neurons (MSNs) in the NAc, promoting maladaptive shifts in cell signaling and synaptic function. Our preliminary data indicates that expression of Gadd45b (Growth, arrest, and DNA-damage inducible protein 45b) mRNA is upregulated in the MSNs after both cocaine and dopamine receptor stimulation, and that Gadd45b is required for cocaine-related memory formation. However, although Gadd45b plays a critical role in epigenetic reprogramming and memory formation in other brain regions, the role of Gadd45b in cocaine-related epigenetic, molecular, and behavioral adaptations is not clear. In this proposal, we will test the overarching hypothesis that Gadd45b regulates drug- induced behavioral plasticity by control of activity-dependent DNA demethylation in the NAc. Specific Aim 1 of this proposal will combine bidirectional CRISPR-based manipulations and single-nucleus RNA sequencing to determine how Gadd45b signaling impacts transcriptional responses to cocaine. Specific Aim 2 will use novel Gadd45b tools and genome-wide DNA methylation profiling to identify the molecular interactions that regulate Gadd45b-dependent epigenetic programming in the NAc. Finally, Specific Aim 3 will use cell-specific in vivo Gadd45b manipulations in combination with behavioral assays of cocaine and natural reward to test the hypothesis that Gadd45b enhances the behavioral effects of cocaine. Together, these experiments will identify Gadd45b target genes in the NAc, dissect epigenetic mechanisms by which Gadd45b contributes to cocaine’s epigenetic effects, and define a role for Gadd45b in cocaine-related behavioral plasticity. These studies will reveal fundamental mechanisms by which Gadd45b contributes to psychostimulant response, and will pave the way for future experiments to explore how drugs of abuse engage the epigenome to alter motivated behavior.