A Multimodal Developmental Atlas for Cerebellar Molecular Layer Interneurons

Student: Wendy Wang

Supervisor: Dr. Julie Lefebvre

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Abstract

The central nervous system is comprised of an extraordinary diversity of neuronal cell-types. Classification of neurons into types and subtypes enables systematic studies of nervous system organization and function in health and disease. Defining cell types is complicated by the need to consider multiple modalities of classification including morphology, gene expression and function. Additionally, neuronal cell types diverge from common progenitors during development through protracted and dynamic processes. Studies at maturity have shown that multimodal approaches are needed to delineate the repertoire of neuronal cell-type diversity. However, similar methods have not been extended to interrogate the steps by which neuronal progenitors are programmed to acquire their adult forms and functions.

In this thesis, I present a multimodal diversification map of the cerebellar molecular layer interneurons (MLIs), a heterogeneous inhibitory interneuron population that derives from a common progenitor pool. Using genetic, histological and computational approaches, I first define the diversity of mature MLIs along morphological and transcriptional axes. My analyses reveal examples of both discrete and continuous heterogeneity within each modality. Interestingly, I did not find direct correlation between MLI morphological and transcriptional identities. Instead, I identify graded transitions in gene expression profiles between morphologically discrete neurons, as well as graded transitions in neuronal morphology within transcriptionally discrete neurons. I demonstrate that neither morphology nor gene expression alone can be used to fully annotate interneuron diversity.

I further define MLI subtypes by tracking the emergence of distinct identities during development using a novel application of pseudo time trajectory inference to morphological reconstructions. MLI subtype-specific morphological phenotypes emerge early during neuronal migration within the molecular layer, challenging prevailing views that MLI subtype specification is dependent on positional cues. In a complementary approach, I use in situ single molecule RNA imaging to visualize changes in marker gene expression during the timeframe of MLI migration and maturation. I find that MLIs do not establish the full repertoire of subtype-specific expression signatures until days after the completion of neuronal migration. Altogether, these studies present a multimodal map of MLI diversification and offer a framework that is broadly applicable for defining the developmental trajectory of interneuron populations.