Single-nuclei RNA sequencing reveals heterogeneity within developing GnRH3 neurons in zebrafish

Area of Science:

• Neuroscience
• Developmental Biology
• Genomics

Background:

• Gonadotropin-releasing hormone (GnRH) regulates reproduction by controlling pituitary gonadotropins.
• GnRH3 neurons in zebrafish migrate from the nasal placode to the hypothalamus, forming distinct subpopulations.
• The molecular basis of GnRH3 neuron migration heterogeneity is not fully understood.

Purpose of the Study:

• To delineate the molecular and spatial heterogeneity of GnRH3 neurons during development.
• To identify distinct subpopulations of GnRH3 neurons and their molecular markers.
• To investigate the migratory pathways and transcriptomic continuum of GnRH3 neurons.

Main Methods:

• Single-nuclei RNA sequencing of EGFP-labeled GnRH3 nuclei from zebrafish.
• Clustering analysis, differential gene expression, and trajectory inference.
• In situ hybridization and immunohistochemistry for marker gene validation and cell distribution analysis.

Main Results:

• Three main GnRH3 subpopulations were identified: Gnrh3^cyp1a, Gnrh3^nrg1, and Gnrh3^npffl/prl2.
• Gnrh3^cyp1a and Gnrh3^nrg1 were located in the olfactory epithelium; Gnrh3^npffl/prl2 subpopulations were in the olfactory bulbs/terminal nerve and telencephalon.
• Trajectory inference suggested a transcriptomic continuum from Gnrh3^cyp1a to Gnrh3^npffl/prl2, with the latter lacking canonical migratory gene signatures.

Conclusions:

• At least three distinct GnRH3 neuron subpopulations exist with unique transcriptomes and characteristics.
• These subpopulations exhibit specific spatial distributions related to their developmental origins.
• The findings provide a foundation for understanding the diverse roles of GnRH3 in neuronal migration and reproductive regulation.