Single-Cell Profiling Uncovers Evolutionary Divergence of Hypocretin/Orexin Neuronal Subpopulations

Area of Science:

• Neuroscience
• Genomics
• Comparative Biology

Background:

• The hypothalamus regulates vital behaviors like sleep-wake cycles via discrete neuronal populations.
• Hypocretin/orexin (Hcrt) neurons are known to be functionally diverse.
• Understanding hypothalamic cell-type heterogeneity is crucial for deciphering brain function.

Purpose of the Study:

• To comprehensively identify and annotate hypothalamic cell types in zebrafish using single-cell RNA sequencing (scRNA-seq).
• To compare zebrafish and mouse hypothalamic cell types to identify conserved and divergent populations.
• To investigate the functional heterogeneity of hypocretin/orexin (Hcrt) neurons, particularly those expressing Npvf.

Main Methods:

• Single-cell RNA sequencing (scRNA-seq) of adult male and female zebrafish hypothalamic periventricular zone.
• High-resolution imaging of zebrafish brain tissue.
• Integration of zebrafish and mouse scRNA-seq datasets for cross-species comparison.

Main Results:

• Identification of 21 glutamatergic and 28 GABAergic cell types in the zebrafish hypothalamus.
• Discovery of evolutionarily conserved and divergent hypothalamic cell types between zebrafish and mice.
• Enrichment of the sleep-regulating neuropeptide Npvf gene in a subset of glutamatergic Hcrt neurons (Hcrt+Npvf+).
• Demonstration that Hcrt+Npvf+ neurons exhibit distinct genetic profiles, activity, and neurite morphology compared to other Hcrt neurons.

Conclusions:

• Provides a unified annotation of hypothalamic cell types across species.
• Suggests that Hcrt neuron heterogeneity allows for multifunctionality in regulating behavior.
• Highlights the Hcrt- and Npvf-releasing neuronal subpopulation's role in consolidating both wake and sleep states.