Circuit and molecular architecture of a ventral hippocampal network

The ventral hippocampus (vHPC) is a critical hub in networks that process emotional information. While recent studies have indicated that vHPC projection neurons are functionally dissociable, the basic principles of how the inputs and outputs of the vHPC are organized remain unclear. Here we used viral and sequencing approaches to define the logic of the extended vHPC circuit. Through molecular profiling of vHPC projection neurons, we show that vHPC neurons with distinct projection targets differ in their transcriptional signatures. Overall design: In this study we determined whether ventral hippocampal (vHPC) neurons that project to one of four target areas, lateral hypothalamus (LH) basomedial amygdala (BMA), medial prefrontal cortex (mPFC), and nucleus accumbens (NAc), differ in terms of the genes they express. To do so, we used transcriptional profiling methods that accessed translating mRNAs in ventral hippocampal neurons defined by their projection to one of these four targets. We labeled ventral hippocampal neurons that projected to ateral hypothalamus (LH) basomedial amygdala (BMA), medial prefrontal cortex (mPFC), and nucleus accumbens (NAc), in different cohorts of B6.129S4-Gt(ROSA)26Sortm1(CAG-EGFP/Rpl10a,-birA)Wtp/J (Jackson labs stock #022367) by injection of a retrograde traveling adenoassociated virus expressing Cre recombinase in each of the target areas, then used a translating ribosome affinity purification (TRAP) approach to profile translation in each of these populations . RNA sequencing (RNAseq) was then performed to identify differentially enriched mRNAs in the distinct vHPC projection neurons . Each sample correspond to 6 hippocampi from each projection pathway, perfomed in duplicate or triplicate.

腹侧海马体（ventral hippocampus, vHPC）是参与情绪信息处理的神经网络关键枢纽。尽管近期研究表明vHPC投射神经元存在功能分化，但vHPC的输入与输出组织的基本原理仍未明确。
本研究借助病毒介导技术与测序手段，阐明了延伸性vHPC环路的运作逻辑。通过对vHPC投射神经元的分子谱分析，我们发现具有不同投射靶点的vHPC神经元，其转录特征存在显著差异。

整体实验设计：本研究旨在明确，投射至四个靶脑区——外侧下丘脑（lateral hypothalamus, LH）、基底内侧杏仁核（basomedial amygdala, BMA）、内侧前额叶皮层（medial prefrontal cortex, mPFC）以及伏隔核（nucleus accumbens, NAc）的vHPC神经元，在基因表达层面是否存在差异。为达成该目标，我们采用靶向鉴定投射至上述四个靶区之一的vHPC神经元的转录谱分析技术，以检测其翻译中的mRNA。

我们在不同批次的B6.129S4-Gt(ROSA)26Sortm1(CAG-EGFP/Rpl10a,-birA)Wtp/J小鼠（杰克逊实验室货号：022367）中，通过向四个靶脑区分别注射表达Cre重组酶的逆行腺相关病毒，标记投射至LH、BMA、mPFC以及NAc的vHPC神经元；随后采用翻译核糖体亲和纯化（translating ribosome affinity purification, TRAP）技术，对上述各神经元群体的翻译状态进行谱分析。随后通过RNA测序（RNA sequencing, RNA-seq），鉴定不同vHPC投射神经元群体中差异富集的mRNA。每个样本对应来自每条投射通路的6个海马体，实验设置了2~3次生物学重复。