The organization of intracortical connections by layer and cell class in the mouse brain

The mammalian cortex is a laminar structure composed of many cell types densely interconnected in complex ways. Recent systematic efforts to map the mouse mesoscale connectome provide comprehensive projection data on inter-areal connections, but not at the level of specific cell classes or layers within cortical areas. We present here a significant expansion of the Allen Mouse Brain Connectivity Atlas, with ~1000 new axonal projection mapping experiments across nearly all isocortical areas in 50 Cre driver lines. Using 13 lines most selective for cortical layer and/or projection neuron class we identify the differential contribution of each layer/class to the overall intracortical connectivity patterns. We find that layer 5 (L5) projection neurons account for essentially all intracortical outputs. L2/3, L4, and L6 neurons contact a subset of the L5 cortical targets. We describe the most common axon lamination patterns in target regions, and their relationships to source layer/class. Most patterns were consistent with previous anatomical rules used to determine hierarchical position between cortical areas (feedforward, feedback), with notable exceptions. We observe a diversity of target patterns arising from every source layer/class, but supragranular (L2/3 and upper L4) neurons are most associated with feedforward type patterns, whereas infragranular (L5 and L6) neurons have both feedforward and feedback. Network analyses revealed a modular organization of the intracortical connectome. Using the cell class-based target lamination patterns, we labeled all connections and intermodule connections as feed-forward or -back, and finally present an integrated view of the intracortical connectome as a hierarchical network.