Connectome-seq: High-throughput Mapping of Neuronal Connectivity at Single-Synapse Resolution via Barcode Sequencing

Area of Science:

• Neuroscience
• Molecular Biology
• Genomics

Background:

• Mapping neuronal connectivity is crucial for understanding brain function.
• Current methods struggle with long-distance circuits and preserving cell type information.
• Single-cell resolution is needed for detailed circuit analysis.

Purpose of the Study:

• To develop a high-throughput method for mapping neuronal connectivity at single-synapse resolution.
• To simultaneously capture synaptic connections and molecular identities of connected neurons.
• To identify molecular determinants of neuronal circuit organization.

Main Methods:

• Connectome-seq combines engineered synaptic proteins, RNA barcoding, and parallel single-nucleus and single-synaptosome sequencing.
• An adeno-associated virus (AAV)-based approach was utilized.
• The method was validated in the mouse pontocerebellar circuit.

Main Results:

• Connectome-seq successfully mapped neuronal connectivity at single-synapse resolution.
• Established projections and novel synaptic partnerships were identified in the mouse pontocerebellar circuit.
• Integrated analysis revealed molecular markers enriched in connected neurons, suggesting circuit organization determinants.

Conclusions:

• Connectome-seq offers a scalable platform for comprehensive circuit analysis with single-cell precision and gene expression data.
• This method enables systematic mapping of neuronal connectivity across brain regions.
• Connectome-seq advances the understanding of complex mammalian brain circuit organization.