Spatial clustering analysis in neuroanatomy: Applications of different approaches to motor nerve fiber distribution

Spatial organization of the nerve fibers in the peripheral nerves may be important for the studies of axonal regeneration, the degenerative nerve diseases and the construction of interfaces with peripheral nerves, such as nerve prostheses. Functional topography of motor axons related to the gastrocnemius muscle was revealed in the ventral spinal roots by retrograde tracing. Gastrocnemius muscles of adult rats were injected with the tracer Fluoro-Gold. After 3 days of survival the animals were sacrificed and their ventral roots were harvested, sectioned, and imaged on a fluorescence microscope. Maps of the traced fibers were automatically analyzed using a novel approach, local spatial clustering statistics, that tested for occurrences of clusters of motor fibers and visualized them. Thresholds indicating the presence of clustering at various scales of observation were computed based on series of Monte Carlo simulations of random spatial point patterns. Clusters were visualized by kernel interpolation. The approach was tested on simulated data and subsequently applied to the motor fiber maps in the ventral roots. Results revealed clustering of the motor fibers innervating the gastrocnemius muscle at the level of the L6 ventral spinal root. The analysis was validated using Voronoi tessellation and nearest neighbor analysis.