Nonbinary 2D Distribution Tool Maps Autonomic Nerve Fiber Clustering in Lumbosacral Ventral Roots of Rhesus Macaques

Neuromodulation of the peripheral nervous system (PNS) by electrical stimulation may augment autonomic function after injury or in neurodegenerative disorders. Nerve fiber size, myelination, and distance between individual fibers and the stimulation electrode may influence response thresholds to electrical stimulation. However, information on the spatial distribution of nerve fibers within the PNS is sparse. We developed a new two-dimensional (2D) morphological mapping tool to assess spatial heterogeneity and clustering of nerve fibers. The L6-S3 ventral roots (VRs) in rhesus macaques were used as a model system to map preganglionic parasympathetic, γ-motor, and α-motor fibers. Random and ground truth distributions of nerve fiber centroids were determined for each VR by light microscopy. The proposed tool allows for nonbinary determinations of fiber heterogeneity by defining the minimum distance between nerve fibers for cluster inclusion and comparisons with random fiber distributions for each VR. There was extensive variability in the relative composition of nerve fiber types and degree of 2D fiber heterogeneity between different L6-S3 VR levels within and across different animals. There was a positive correlation between the proportion of autonomic fibers and the degree of nerve fiber clustering. Nerve fiber cluster heterogeneity between VRs may contribute to varied functional outcomes from neuromodulation.