Unveiling Hypothalamic Molecular Signatures via Retrograde Viral Tracing and Single-Cell Transcriptomics

Area of Science:

• Neuroscience
• Molecular Biology
• Genomics

Background:

• Hypothalamic neurocircuits regulate vital homeostatic and survival behaviors.
• Limited understanding of the molecular identities of neurons in complex hypothalamic circuits.

Purpose of the Study:

• To develop a novel method for mapping hypothalamic neurocircuits.
• To generate a high-resolution transcriptomic dataset of hypothalamic cells.
• To identify molecular identities of upstream neurons in hypothalamic circuits.

Main Methods:

• Constructed a Cre recombinase-dependent pseudorabies virus (PRVs) for retrograde tracing.
• Utilized retrograde nuclear Connect-seq (nuConnect-seq) for transcriptome analysis of single upstream neurons.
• Generated a single nuclei RNA-seq (snRNA-seq) dataset from CRH-IRES-Cre (CRH-Cre) mice hypothalamus.
• Validated the dataset using label transfer against a large integrated reference dataset.

Main Results:

• Generated a snRNA-seq dataset of 1,533 hypothalamic cells.
• Successfully mapped single upstream neurons using PRVs and nuConnect-seq.
• Integrated diverse datasets for enhanced technical validity and cell type diversity.
• Provided a validated transcriptomic resource for hypothalamic neurocircuit research.

Conclusions:

• The nuConnect-seq approach offers a powerful tool for dissecting hypothalamic neurocircuits.
• The validated dataset enhances understanding of hypothalamic cell type diversity and function.
• Integrated transcriptomic data is essential for robust neurobiological research.