EASI-FISH for thick tissue defines lateral hypothalamus spatio-molecular organization

Area of Science:

• Neuroscience
• Molecular Biology
• Computational Biology

Background:

• Characterizing brain architecture requires understanding the spatial organization and morphology of molecularly defined cell types.
• Current methods face bottlenecks in surveying gene expression across large tissue volumes.
• The lateral hypothalamic area (LHA) has a poorly defined anatomical organization.

Purpose of the Study:

• To develop a novel method for surveying gene expression in thick brain sections.
• To create a computational pipeline for processing large image datasets generated by the new method.
• To investigate the spatial distribution of cell types within the LHA and identify its subregions.

Main Methods:

• Developed Expansion-Assisted Iterative Fluorescence In Situ Hybridization (EASI-FISH) for gene expression analysis in 300 μm brain sections.
• Created a computational pipeline for rapid processing of EASI-FISH image data.
• Applied EASI-FISH to map dozens of molecularly defined cell types in the lateral hypothalamic area.

Main Results:

• EASI-FISH enabled the reconstruction of spatio-molecular domains that generalize across brains.
• Mapping cell types in the LHA revealed nine spatially and molecularly defined subregions.
• Iterative reanalysis of scRNA-seq datasets identified marker genes that resolved spatial and morphological heterogeneity.

Conclusions:

• EASI-FISH provides a scalable approach to map molecularly defined cell types in brain tissue.
• The developed computational pipeline democratizes cell type mapping and facilitates discoveries in brain organization.
• This technique enhances the understanding of complex brain regions like the LHA.