Cascade Diversification Directs the Generation of Neuronal Diversity in Hypothalamus

Abstract

The hypothalamus contains an astounding heterogeneity of neurons to achieve its role in regulating endocrine, autonomic and behavioral functions. Despite previous progress in deciphering the gene regulatory programs linked to hypothalamus development, its molecular developmental trajectory and origin of neuronal diversity remain largely unknown. Here we combine transcriptomic profiling of 43,261 cells derived from Rax + hypothalamic neuroepithelium with lineage tracing to map a developmental landscape of mouse hypothalamus and delineate the developmental trajectory of radial glial cells (RGCs), intermediate progenitor cells (IPCs), nascent neurons and peptidergic neurons in the lineage hierarchy. We show that RGCs adopt a conserved strategy for multipotential differentiation but generate both Ascl1 + and Neurog2 + IPCs, which display regionally differential origins in telencephalon. As transit-amplifying cells, Ascl1 + IPCs differ from their telencephalic counterpart by displaying fate bifurcation to produce both glutamatergic and GABAergic neurons.After classifying the developing neurons into 29 subtypes coded by diverse transcription factors, neurotransmitters and neuropeptides, we identified their molecular determinants via regulon analysis and further found that postmitotic neurons at nascent state possess the potential to resolve into more diverse subtypes of peptidergic neurons. Together, our study offers a single-cell framework for hypothalamus development and reveals that multiple cell types along the order of lineage hierarchy contribute to the fate diversification of hypothalamic neurons in a stepwise fashion, suggesting that a cascade diversifying model can deconstruct the origin of neuronal diversity.

The cellular and molecular landscape of hypothalamic patterning and differentiation from embryonic to late postnatal development

“…To identify region-specific differences between mature oligodendrocytes or astrocytes of the hypothalamus and other brain regions, previously published cortical scRNA-seq 12,13 were used to identify differential gene expression, using age as the variance with default parameters (first-pass gene lists) 14 . Published hypothalamic datasets 13,[15][16][17] were then used to compare to these cortical scRNAseq datasets using the identified genes (second-pass gene lists). Identified differential genes were then validated by matching the Allen Brain in situ atlas data using co-coframer 11 to validate spatial expression of these differential genes, as some observed differences could reflect batch effects resulting from scRNA-seq library preparations from different laboratories (third-pass gene lists).…”

“…Progress in this area has been hampered, however, by the fact that hypothalamic cell types thus far have remained quite poorly characterized, despite recent efforts aimed at using scRNA-seq to classify cells in different regions of the adult hypothalamus [11][12][13][14][15] , and more recently in limited embryonic and early postnatal periods 16,17 . Still less is known about how hypothalamic cell types acquire their identities during development.…”

Selective Requirement for Polycomb Repressor Complex 2 in the Generation of Specific Hypothalamic Neuronal Sub-types

“…To address the effects of Eed mutation upon hypothalamus development in more detail we performed scRNA-seq analysis. We focused first upon a late embryonic stage, E18.5, because the majority of distinct neuronal sub-types -such as those specifically expressing neuropeptides and neurotransmitters -have been generated by this stage (Kim et al, 2020;Romanov et al, 2020;Zhang et al, 2021). We dissected the hypothalamus from 6 control (Eed fl/fl ) and 9 Eed-cKO embryos, at E18.5.…”

“…These early TFs in turn activate panels of late TFs within subdomains of the developing hypothalamus, which act with more restrictive mandates to specify diverse subsets of hypothalamic cell fates (Alvarez-Bolado, 2019;Bedont et al, 2015;Blackshaw et al, 2010;Burbridge et al, 2016;Ferran et al, 2015;Nesan and Kurrasch, 2016;Puelles and Rubenstein, 2015;Xie and Dorsky, 2017). More recently, single cell transcriptomic analysis of the embryonic hypothalamus has greatly increased our understanding of its developmental process (Huisman et al, 2019;Kim et al, 2020;Romanov et al, 2020;Zhang et al, 2021;Zhou et al, 2020).…”

“…Yaghmaeian Salmani et al, PRC2 affects hypothalamus development the majority of distinct neuronal sub-types -such as those specifically expressing neuropeptides and neurotransmitters -have been generated by this stage (Kim et al, 2020;Romanov et al, 2020;Zhang et al, 2021). We dissected the hypothalamus from 6 control (Eed fl/fl ) and 9 Eed-cKO embryos, at E18.5.…”