Spatial clusters of constitutively active neurons in mouse visual cortex

The networks of neocortical neurons are coordinated by spontaneous activity, the level of which exhibits high heterogeneity among neurons, ranging from low activity levels to very high activity levels, even in the same network. Highly active neurons represent a small proportion in the cerebral cortex and are mingled in a web of the vast majority of neurons with low firing rates. However, little is known about the spatial arrangement of these highly active cells within the cerebral cortex. Here, we visualized their spatial distribution by labeling them with c-Fos, a neuronal activity marker, in the mouse primary visual cortex. By introducing energy-like and entropy-like parameters that did not require arbitrary thresholds for c-Fos positivity, we found that strongly c-Fos-expressing neurons were clustered in the vicinity. The cluster size measured approximately 100 μm in diameter and was smaller in layer 2/3 than in layers 5 and 6. Layer 1 neurons did not exhibit a clustered pattern of c-Fos-expressing neurons. Our novel statistical approaches are not subject to human bias and are thus widely applicable to evaluate the spatial bias of any particles.