Spatial Interrogation of Periaqueductal Gray and Hypothalamic Cell Types Involved in Instinctive Behavior

Abstract

The midbrain periaqueductal gray (PAG) and hypothalamic preoptic area (POA) are essential for orchestrating instinctive behaviors. The current organization of the PAG into four main radial columns and the coarse nuclear organization of the POA lack the resolution needed to account for the vast range of functionalities displayed by these two large neural structures. Using single nuclear sequencing and spatially resolved single-cell transcriptomic measurements, we uncovered widespread transcriptional heterogeneity in the mouse PAG and POA with ~140 neuronal populations defined in the PAG and ~70 neuronal populations in the POA. Within the PAG, we used each population’s neighborhood properties to further assemble them into 19 discrete groups that share the same three-dimensional spatial motifs. We explored the transcriptional identity of PAG and POA cell types activated during various instinctive behaviors, and examined the spatial logic of PAG function, demonstrating the regional, yet selective recruitment of PAG cell types for distinct behaviors. Remarkably, certain behaviors trigger differential spatial activation patterns within a given cell type, illustrating the complexity of PAG molecular and functional 3D organization. Altogether, this work identifies novel neural spatial motifs and establishes new spatially informed functional and molecular maps of the POA and PAG during instinctive behavior.