REMOTE MICROGLIAL ACTIVATION AND THE PRODUCTION OF PROINFLAMMATORY CYTOKINES PREDICTS THE DEVELOPMENT OF NEUROPATHIC PAIN AFTER SPINAL CORD INJURY

Abstract

Purpose/Hypothesis: Spinal cord injury (SCI) often impairs sensory systems causing chronic allodynia. The mechanism of neuropathic pain has been studied extensively following peripheral nerve injury, while little is known about the mechanism underlying central trauma. Microglial activation via P2X4, an ionotropic ATP receptor and the production of proinfammatory cytokines including TNFα, IL-1β and IL-6 in the L5 dorsal horn is involved in the development of mechanical allodynia, following peripheral nerve injury. Terefore, we investigated whether below-level allodynia that develops after mid-thoracic SCI is associated with ATP-mediated microglial activation and proinfammatory cytokine production in the L5 dorsal horn. Number of Subjects: Sixty-one adult, female Sprague-Dawley rats were used. Materials/Methods: Rats were randomly assigned to naïve, laminectomy control, spinal nerve ligation (SNL), mild SCI (0.5 mm cord displacement) or moderate SCI (1.1 mm cord displacement) and were sacrifced at 7, 14, 21 or 35 dpo. SNL, a peripheral nerve injury pain model, served as allodynia controls. We tested weekly for tactile allodynia and thermal hyperalgesia using von Frey hair and plantar heat tests, respectively. Microglia (OX42), astrocytes (GFAP), and P2X4 ATP receptors were quantified at L5. Changes in gene expression (mRNA) were measured from superfcial dorsal horn obtained by laser capture microdissection. We assessed gene changes for microglial activators including p38, P2X4 and fractalkine as well as microglia and astrocytes. Using ELISA and Western blotting, we examined the secretory profile of activated microglial within the L5 dorsal horn. Results: Significant allodynia developed for moderate SCI and SNL groups (p<;.01). These groups had more activated microglia but not astrocytes in the L5 dorsal horn. Chronic allodynia correlates to microglial activation with sensitivity and specificity values at 86% and 92%, respectively. A cut-of of 200% microglial activation accurately predicted allodynia in 86% of animals. Surprisingly, there was no upregulation of P2X4 ATP receptor mRNA or protein in the dorsal horn of rats exhibiting neuropathic pain. After moderate SCI, TNF-α and IL-1β levels significantly increase in the L5 dorsal horn by 7 dpo and return to normal levels by 35 dpo. Interestingly, IL-6 remains at normal levels early after SCI and increases at chronic timepoints. Conclusions: These data suggest that remote microglia through pro-infammatory pathways may be key mediators of the development and maintenance of below-level allodynia after moderate SCI. TNF-α and IL-1β may induce allodynia while IL-6 may mediate the maintenance of pain. Clinical Relevance: Modulating the microglial response after SCI by either preventing microglial activation itself or blocking subsequent pro-infammatory cascades may prove to be effective at limiting or preventing neuropathic pain after SCI in humans. Support Contributed By: NIH NINDS #NS43798-01 (DMB) and NINDS #NS037846 (PGP).