Alcohol-Induced Alternative Splicing in Drosophila Memory Circuits

反复的酒精体验会对与醉酒有关的感觉线索产生持久的记忆。 这些记忆可能会在从酒精使用障碍(AUD)中恢复的个人中引发复发。 酒精改变记忆变得持久和僵硬的分子机制仍然存在 不清楚。我们最近证明，这些记忆的形成会导致另一种选择的表达 黑腹果蝇记忆编码神经元中的转录异构体。果蝇靠蘑菇身体为生 (MB)神经元产生联想记忆，包括与酒精相关的感觉线索的记忆。 在成年MB神经元中，发挥剪接作用的基因表达减少，减少了这些基因的形成 记忆，论证了RNA加工在乙醇记忆形成中的必要性。此外，减少 多巴胺-2样受体等选择性剪接基因在成人MB中的表达 神经元减少酒精记忆的形成。这表明这些基因的剪接变化已经 对未来记忆形成的功能暗示。这一提议的中心假设是，动态 酒精暴露反应中DOP2R的选择性剪接通过改变活动来指导奖励行为 神经回路。 我们产生了突变的果蝇，它们强迫表达幼稚的或训练有素的多巴2R亚型，这 使我们能够询问这些剪接变体如何在高度机械化的情况下影响神经动力学和行为 水平。申请人将揭示DOP2R蛋白在细胞内的定位(目标1，如何进行选择性剪接 多巴胺2R影响奖赏记忆(目标2)和记忆回路内的可塑性(目标3)。通过完成这些目标， 申请者将获得细胞神经生物学和神经遗传学、行为学方面的科学和技术专长。 评估果蝇，超分辨率成像，和双光子钙成像。通过全面的 培训计划，该奖学金将促进申请者在稳健实验中的专业发展 设计和数据分析；书面和口头科学交流；有效和包容的指导。 项目和培训目标的顺利完成得到了互动和支持的充分支持 布朗大学和神经科学研究生项目的机构环境，并将准备 申请者下一步迈向独立的科学事业。

Repeated alcohol experiences can produce long-lasting memories for sensory cues associated with intoxication. These memories can problematically trigger relapse in individuals recovering from alcohol use disorder (AUD). The molecular mechanisms by which ethanol changes memories to become long-lasting and inflexible remain unclear. We recently demonstrated that formation of these memories results in expression of alternative transcript isoforms in memory-encoding neurons in Drosophila melanogaster. Drosophila rely on mushroom body (MB) neurons to make associative memories, including memories of ethanol-associated sensory cues. Decreasing expression of genes that play a role in splicing in adult MB neurons reduces formation of these memories, demonstrating the necessity of RNA processing in ethanol memory formation. Moreover, decreasing expression of genes that are alternatively spliced like the Dopamine-2-like Receptor (Dop2R) in adult MB neurons reduces ethanol memory formation. This suggests that the splicing changes in these genes has functional implications for future memory formation. The central hypothesis of this proposal is that the dynamic alternative splicing of Dop2R in response to alcohol exposure directs reward behavior by changing the activity of neural circuits. We generated mutant Drosophila that have forced expression of the naïve or trained Dop2R isoform, which allows us to interrogate how these splice variants affect neural dynamics and behavior at a highly mechanistic level. The applicant will reveal intracellular localization of DOP2R protein (Aim 1, how alternative splicing of Dop2R affects reward memory (Aim 2), and plasticity within memory circuits (Aim 3). By completing these aims, the applicant will gain scientific and technical expertise in cellular neurobiology and neurogenetics, behavioral assessment of Drosophila, super-resolution imaging, and 2-photon calcium imaging. Through a comprehensive training plan, this fellowship will facilitate the professional development of the applicant in robust experimental design and data analysis; written and oral scientific communication; and effective and inclusive mentoring. Successful completion of the project and training goals are fully supported by the interactive and supportive institutional environment of Brown University and in the Neuroscience Graduate Program, and will prepare the applicant for the next steps towards an independent scientific career.