A neuroepithelial wave of BMP signalling drives anteroposterior specification of the tuberal hypothalamus

Abstract

The tuberal hypothalamus controls life-supporting homeostatic processes, but despite its fundamental role, the cells and signalling pathways that specify this unique region of the CNS in embryogenesis are poorly characterised. Here we combine experimental and bioinformatic approaches in the embryonic chick to show that the tuberal hypothalamus is progressively generated from hypothalamic floor plate-like cells. Fate-mapping studies show that a stream of tuberal progenitors develops in the anterior-ventral neural tube as a wave of neuroepithelial-derived BMP signalling sweeps from anterior to posterior through the hypothalamic floor plate. As later-specified posterior tuberal progenitors are generated, early-specified anterior tuberal progenitors become progressively more distant from these BMP signals and differentiate into tuberal neurogenic cells. Gain- and loss-of-function experiments in vivo and ex vivo show that BMP signalling initiates tuberal progenitor specification, but must be eliminated for these to progress to anterior neurogenic progenitors. ScRNA-Seq profiling shows that tuberal progenitors that are specified after the major period of anterior tuberal specification begin to upregulate genes that characterise radial glial cells. This study provides an integrated account of the development of the tuberal hypothalamus.

A Methodology for the Enzymatic Isolation of Embryonic Hypothalamus Tissue and Its Acute or Post-Culture Analysis by Multiplex Hybridisation Chain Reaction

“…Here, we extend these reports to provide details on how to take embryos and explants through repeated rounds of multiplex HCR. Such stripping and re-probing enables samples to be routinely analysed with 10–12 genes per sample ( Chinnaiya et al, 2023 ) ( Figure 5 ).…”

“…Nonetheless, where anterior neural tube explants have been used, they, too, have provided new insights into the tissue, cell, and molecular interactions that mediate brain patterning ( Ericson et al, 1995 ; Guinazu et al, 2007 ; Placzek and Briscoe, 2018 ; Shirahama et al, 2019 ). In particular, their use has significantly improved our understanding of the development of the hypothalamus ( Dale et al, 1997 ; Manning et al, 2006 ; Kim et al, 2022 ; Chinnaiya et al, 2023 ). This intricate region of the ventral forebrain regulates physiological processes that maintain homeostasis and orchestrate complex behaviours.…”

“…The enzymatic isolation of entire embryonic chick hypothalamic tissues at different patterning stages, and their acute analysis through scRNA-seq, has generated a comprehensive roadmap of hypothalamic development, from induction to patterning and to neurogenesis, and revealed many previously uncharacterized candidate regulators of hypothalamic patterning ( Kim et al, 2022 ). A powerful new development has been the use of multiplex hybridisation chain reaction (HCR) ( Choi et al, 2018 ) in the analysis of wholemount and isolated hypothalamic tissues or hypothalamic tissue explants ( Kim et al, 2022 ; Chinnaiya et al, 2023 ), for detecting and visualising specific DNA or RNA sequences in a sample. In the protocol we describe here, RNA probes specific to a target gene trigger chain reaction events between two sets of hairpin molecules, allowing for the rapid detection of mRNA ( Choi et al, 2014 ).…”

“…In the complex environment of the developing brain, multiplex HCR has provided an incisive tool to define distinct progenitor subtypes and examine dynamic gene expression patterns. Multiplex HCR can similarly be applied to cultured hypothalamic tissue explants to reveal how naïve or pre-patterned tissues respond to extrinsic signals and/or self-organise ( Kim et al, 2022 ; Chinnaiya et al, 2023 ). To date, these techniques have provided particularly powerful insights into the development of the tuberal hypothalamus ( Chinnaiya et al, 2023 ).…”

“…Multiplex HCR can similarly be applied to cultured hypothalamic tissue explants to reveal how naïve or pre-patterned tissues respond to extrinsic signals and/or self-organise ( Kim et al, 2022 ; Chinnaiya et al, 2023 ). To date, these techniques have provided particularly powerful insights into the development of the tuberal hypothalamus ( Chinnaiya et al, 2023 ).…”