Organ scale Three-dimensional atlas of developing pelvic innervation for the mouse lower urinary tract.

Pelvic autonomic neurons are responsible for coordinating pelvic organ function, including storage and emptying of the urinary bladder. These neurons arise from sacral neural crest-derived progenitors which migrate to the developing urogenital sinus and undergo differentiation alongside pelvic organs, ultimately organizing into paired pelvic ganglia. To circumvent the complex morphological changes that occur during genitourinary organ development, we have optimized light sheet fluorescence microscopy imaging for three-dimensional mapping of the development and spatial distribution of pelvic autonomic neurons. To track neural crest progenitor migration, neurogenesis, and formation of mature pelvic ganglia morphology in mice, we imaged intact lower urinary tract tissue from mid-gestational to post-weaning stages at single-cell resolution. The resulting atlas identifies the precise timing and initial routes of neural crest progenitor migration, defines timing for pelvic ganglion neurogenesis, and reveals previously unrecognized neuronal cell bodies outside of the centralized pelvic ganglia structures. The analysis further highlights morphological changes to pelvic ganglion structure that occur in parallel with genitourinary maturation, including the divergence of the major pelvic ganglia and bladder-proximal accessory ganglia. This work acts as the first wholemount imaging atlas of pelvic innervation in the mouse spanning fetal and postnatal timepoints, and thus we present an informative tool for future studies of developmental neurourology and neurogenic bladder dysfunction.