The mouse midbrain periaqueductal gray is organized into target-specific output populations

How brains select between behaviours and coordinate their execution is a central question in neuroscience. The vertebrate periaqueductal gray (PAG) controls a remarkable range of instinctive behaviours, from escape and freezing to hunting, vocalization, and micturition. Its columnar organization, the classical framework for mapping these functions, cannot explain how one region controls so many, often mutually exclusive, behaviours. To determine how PAG output is organized, we generated a whole-brain, column- and cell-type-specific synaptic projection atlas of the mouse PAG, showing distinct dorsal and ventral outputs and dense within-PAG connectivity. Retrograde tracing from four motor-related target regions identified segregated PAG populations, biased towards, but not confined to, individual columns. Channelrhodopsin-2-assisted circuit mapping showed that one such population, projecting to the cuneiform nucleus, provides substantial local collaterals, positioned to shape ongoing PAG activity. These findings uncover a modular output architecture that extends the columnar framework, offering a circuit logic for how one brain region produces diverse behaviours.