Resiniferatoxin induces paradoxical changes in thermal and mechanical sensitivities in rats: mechanism of action

Abstract

Resiniferatoxin (RTX), an ultrapotent analog of capsaicin, has been used as a tool to study the role of capsaicin-sensitive C fibers in pain. Recently, we found that RTX diminished the thermal sensitivity but unexpectedly increased the sensitivity to tactile stimulation in adult rats. In this study, we explored the potential mechanisms involved in RTX-induced changes in somatosensory function. An intraperitoneal injection of 200 microg/kg RTX, but not its vehicle, rapidly produced an increase in the paw withdrawal latency to a heat stimulus. Also, profound tactile allodynia developed in all the RTX-treated rats in 3 weeks. This paradoxical change in thermal and mechanical sensitivities lasted for at least 6 weeks. Electron microscopic examination of the sciatic nerve revealed a loss of unmyelinated fibers and extensive ultrastructural damage of myelinated fibers in RTX-treated rats. Immunofluorescence labeling showed a diminished vanilloid receptor 1 immunoreactivity in dorsal root ganglia neurons and the spinal dorsal horn of RTX-treated rats. Furthermore, two transganglionic tracers, horseradish peroxidase conjugates of cholera toxin B subunit (CTB) and isolectin-B(4) of Bandeiraea simplicifolia (IB(4)), were injected into the opposite sides of the sciatic nerve to trace myelinated and unmyelinated afferent terminations, respectively, in the spinal dorsal horn. In RTX-treated rats, IB(4)-labeled terminals in the dorsal horn were significantly reduced, and CTB-labeled terminals appeared to sprout into lamina II of the spinal dorsal horn. Thus, this study demonstrates that systemic RTX diminishes the thermal pain sensitivity by depletion of unmyelinated afferent neurons. The delayed tactile allodynia induced by RTX is likely attributable to damage to myelinated afferent fibers and their abnormal sprouting in lamina II of the spinal dorsal horn. These data provide new insights into the potential mechanisms of postherpetic neuralgia.

Fig. 1.

Time course of the paw withdrawal latency to a noxious heat stimulus in 6 vehicle- and 10 RTX-treated rats. *p < 0.05 compared with the pretreatment control. The paw withdrawal latency was determined by a radiant heat stimulus.

Fig. 2.

Time course of the development of tactile allodynia in 10 rats treated with RTX and the mechanical withdrawal threshold in 6 vehicle-treated rats. *p < 0.05 compared with the pretreatment control. The paw withdrawal thresholds were determined using von Frey filaments.

Fig. 3.

Representative electron photomicrographs showing ultrastructural changes of myelinated fibers in the sciatic nerve in a vehicle (A)- and an RTX (B–D)-treated rats. Note that the ultrastructural changes induced by RTX include the loss of unmyelinated fibers and noticeable swelling of the myelinated fibers. Magnification, ×4439.

Fig. 4.

VR1 receptor immunoreactivity in lumbar DRG and spinal dorsal horn in a vehicle- and an RTX-treated rat. Densely stained VR1 receptor immunoreactivity is present in the DRG and superficial dorsal horn in the vehicle- but not in the RTX-treated rat. Scale bar, 100 μm.

Fig. 5.

Photomicrographs showing reconstructed plots (A) and the original staining photomicrographs (B) of CTB- and IB₄-labeled afferent terminals in the spinal dorsal horn of a vehicle control and an RTX-treated rat. In A and B, the left side of the dorsal horn contains CTB-labeled terminals and the right side contains IB₄-labeled afferent terminals.C, Magnification of the inset inB. Note that the scattered CTB-labeled terminals are present in lamina II of the RTX-treated rat (C).

Fig. 6.

Mean amount of CTB- and IB₄-labeled areas in the dorsal horn of lumbar spinal sections in two vehicle- (26 sections) and three RTX-treated (40 sections) rats. The mean area was calculated from reconstructed sections as shown in Figure5A. Data are presented as means ± SEM. *p < 0.05 compared with the control in the IB₄ group (Student's t test).