Integrative analysis of single-neuron projectomes links connectome, transcriptome, and function in the mouse cortex

The mammalian cortex orchestrates sensory processing, motor control, and cognition through a complex neural network, yet the organization principles governing single-neuron connectivity and its relationship with transcriptome and neuron activity remain elusive. Here, we reconstructed the projectomes of nearly 20,000 neurons in the whole mouse cortex, identifying 346 projection-defined subtypes with region- and layer-specific distributions. Our analysis uncovered seven cortico-cortical modules with submodular organization, high-resolution cortical hierarchy, and diverse terminal arborization patterns of single neurons refining the traditional feedforward/feedback model. Furthermore, we revealed cortico-subcortical projections, including multiple hyper-direct cortico-basal ganglia pathways, and constructed the detailed topographic maps between cortical neurons and their subcortical target regions. Finally, we showed that our fine-grained cortico-cortical connectivity map is closely related to the cortex-wide Ca²⁺ activity, electrical neuronal activity, and gene expression. Together, our multimodal analysis provides a foundational framework for understanding the cortical neural network in the mouse brain.