A130 RETROGRADE TRACING ANALYSIS OF THE SENSORY INNERVATION OF THE MOUSE COLON

Abstract

Abstract Background Visceral pain is a primary symptom of many gastrointestinal diseases. One feature of visceral pain is its vague localization. We hypothesized that overlap in the receptive fields of spinal primary afferent neurons that innervate the gut may contribute to this vague localization. Many studies have confirmed that the proximal and distal colon are mainly innervated by spinal afferent neurons with cell bodies in thoraco-lumbar and lumbo-sacral dorsal root ganglia (DRG), respectively. However, no murine studies have examined whether individual DRG neurons simultaneously innervate both proximal and distal colon. Aims To determine the extent of overlap in receptive fields of colon-projecting DRG neurons. Methods C57BL/6 mice (n=8) were anesthetized, and two retrograde neuronal tracers with distinct fluorescence emission spectra (Fast blue and DiI) were injected separately into the smooth muscle layers of proximal and distal colon. Mice were left for 10–13 days for dye transport, before being euthanized. Thoraco, lumbar and lumbo-sacral DRGs (T8-13, L1-4, L5-S2) were dissected and fixed in 4% paraformaldehyde overnight. 12μm cryostat sections were obtained and analyzed using a fluorescent microscope equipped with filter cubes that detect Fast blue and DiI. Results When DiI was injected into the proximal colon, we observed labelling to be highest in T8-13 DRG with 12.6 +/- 4.5% of cell bodies labelled, followed by L1-4 was (8.2 +/- 1.4%) and in L5-S2 (6.5 +/- 0.8%). DiI injections into the distal colon resulted in labelling of similar numbers of neurons labelled in T8-13 and L1-4 ganglia, whereas half as many neurons were labelled in L5-S2 ganglia. This data shows that the majority of spinal afferent innervation of the colon originates in thoracolumbar DRG. Most importantly, 26.4% and 17.6% of thoracolumbar and lumbo-sacral DRG neurons labelled by Fast blue injection into the proximal colon were also double-labelled by DiI injected into the distal colon. Similarly, 16.6% and 13.8% of neurons in thoracolumbar and lumbosacral DRG labelled by Fast blue injection into the distal colon were double-labelled by DiI injected into the proximal colon. Conclusions These data reveal a surprisingly large number of DRG neurons that innervate the colon have receptive fields that cover both the proximal and distal colon, which may contribute to the poor spatial localization of pain emanating from the colon. Funding Agencies CCC, CIHR