Functional and molecular profiling of cell diversity and identity in the central auditory system: Patch-seq in a sound localization nucleus

大脑就像万花筒。它由无数在结构和功能上不同的细胞组成。对细胞类型的全面了解对于了解大脑功能是必不可少的。然而，我们对细胞多样性和特性的了解还远远不完整。本项目将揭示外侧上橄榄(LSO)神经元异质性的生物物理、解剖学和遗传学决定因素。该核团是哺乳动物听觉脑干的重要整合中心，参与声音定位。通过Patch-Seq方法的单细胞RNA测序，我们的目标是识别细胞类型特定的分子特征，并生成具有全基因组表达数据的全面的神经元类型图谱。我们推断，功能和解剖细胞间的差异是由转录组的差异因果反映出来的。我们的单细胞分析将获得对特定神经元中存在的信使核糖核酸的公正看法。因此，无人监督的探索性数据分析将是我们基于生物信息学的方法的主要部分。我们希望通过寻找迄今未检测到的亚群来扩大LSO中神经元类型的光谱。我们的项目还将包括有针对性的假设驱动的分析。我们假设生物物理参数与标记基因相关(例如，低输入阻力)。另一个例子是膜时间常数，它对突触整合至关重要，突触整合是声音定位的一个非常重要的方面。我们进一步假设，与神经生理学相关的基因沿着LSO的紧张轴在空间上存在差异表达，其中几个基因以相反的梯度表达。转录组分析将伴随着组织学和电生理学的研究。我们的提案是以“一揽子提案”的形式提交的，与Felix Felmy的提案一起提交。这种交互作用使我们能够将我们关于LSO神经元的数据集与Felmy关于另一个听觉脑干核团(INLL)的数据集结合起来，以便在荟萃分析中进行联合比较和评估。总之，通过将全基因组表达谱与单细胞分辨率下的功能表型相关联，我们的项目将大大有助于更全面地理解中枢听觉系统的功能组织。

The brain is like a kaleidoscope. It consists of countless cells that vary structurally and functionally. A comprehensive knowledge about the cell types is imperative to understand brain function. However, our knowledge of cellular diversity and identity is far from being complete. The present project will unravel the biophysical, anatomical, and genetic determinants of neuronal heterogeneity in the lateral superior olive (LSO). This nucleus is a prominent integration center in the mammalian auditory brainstem and involved in sound localization. By means of single-cell RNA sequencing via the Patch-seq methodology, we aim to identify cell type-specific molecular signatures and to generate a comprehensive neuron type atlas with genome-wide expression data. We reason that functional and anatomical cell-to-cell variability is causally reflected by differences in the transcriptome profile. Our single-cell analyses will obtain an unbiased view of the mRNA present in a given neuron. Unsupervised explorative data analysis will therefore be a major part of our bioinformatics-based approach. We expect to expand the spectrum of neuron types in the LSO by finding hitherto undetected subpopulations. Our project will also include targeted hypothesis-driven analyses. We hypothesize that biophysical parameters correlate with marker genes (e.g. a low input resistance). Another example are membrane time constants, which are crucial for synaptic integration, an utterly important aspect in sound localization. We further hypothesize that genes relevant to neuronal physiology are spatially differentially expressed along the tonotopic axis of the LSO, with several of them in opposing gradients. The transcriptome analyses will be flanked by histological and electrophysiological investigations. Our proposal is submitted in the format of a ‘package proposal’, together with a proposal by Felix Felmy. The interaction enables us to combine our dataset on LSO neurons with Felmy’s dataset on another auditory brainstem nucleus (the INLL) for joint comparison and assessment within a meta-analysis. In summary, by correlating genome-wide expression profiles with the functional phenotype at single-cell resolution, our project will contribute substantially to a more comprehensive understanding of the functional organization of the central auditory system.