Axonal ensembles repeatedly cluster and order synapses along dendrites in mouse cortex

Neuronal ensembles—groups of neurons that exhibit coordinated activity during behavior—are a fundamental feature of cortical computation. Dendritic branches amplify clustered synaptic inputs through local nonlinearities, suggesting that presynaptic groups might organize their connections in specific spatial patterns to engage these mechanisms. Whether the same axon groups form synaptic clusters with consistent spatial arrangements across different target neurons remains unknown, but nanoscale connectomes would resolve such anatomical motifs if they exist. We analyzed millions of synaptic connections in a connectome of mouse visual cortex and found over 700,000 axon groups that repeatedly cluster their synapses onto dendritic branches of multiple pyramidal cells, with over 500,000 maintaining consistent distal-to-proximal arrangements. These repeated patterns occur far more frequently than expected from spatial proximity or layer-based connectivity rules. Axon groups preferentially target specific dendritic branches and position their synapses in stereotyped spatial configurations across multiple postsynaptic partners, revealing that functional ensembles leave characteristic anatomical signatures in cortical microarchitecture.