A Study Of Mitochondrial Morphology And Function At Spinal Level Involved In Inflammatory And Neuropathic Pain In Mice

1Significant changes in mitochondrial distribution and subcellularmorphology in models of inflammatory and neuropathic pain of miceMitochondria play an important role in pathophysiology of inflammatoryand neuropathic pain, but the mechanism is unclear. So far no comprehensivestudy exists that evaluates the changes of mitochondrial dynamics following thepain. In this study, we detected the mitochondrial distribution and subcellularmorphology by using intrathecal injection of mitochondrial tracer, MitotrackerRed CMXRox probe (Mito-Red) and confocal microscopic analysis in modelsof formalin-induced acute inflammatory pain, Complete Freund’sAdjuvant(CFA)-induced persistent inflammatory pain and spared nerve injury(SNI)-induced neuropathic pain. The results demonstrated that subcutaneousformalin injection did not affect the number of Mito-Red positive cells withinthe spinal dorsal horn at both acute and tonic phases, but significantly increasedthe number of cluster type mitochondria in superficial spinal dorsal horn (laminaI–II) at tonic phase. Differently, the number of Mito-Red cells significantly increased in superficial and deep spinal dorsal horn (lamina III–V) followingpersistent CFA and SNI neuropathic pain. Moreover, both CFA and SNIremarkably increased the number of cluster type mitochondria and decreased thenumber of granule type mitochondria, in both superficial and deep spinal dorsalhorn. Based on these data, we concluded that abnormal mitochondrialdistribution and subcellular morphology that results in mitochondrial andneuronal dysfunction contribute to neuropathic and persistent inflammatorypain.2Different functions of mitochondria in the model of formalin-inducedacute and tonic inflammatory pain in spinal cordPrevious studies suggested that reactive oxygen species (ROS) are involvedin the central sensitization of formalin-induced tonic phase of pain at spinal cordlevel. Mitochondria are the main sources of ROS, but its functions in pain arenot clear. In this study, firstly we extracted homogenate and mitochondria fromthe spinal cord of formalin inflammatory pain model mice, and detectedmalondialdehyde (MDA), ROS and hydroperoxide (HP) to reflect oxidativestress in spinal tissues and mitochondria. Then we detected the activities ofmitochondrial ATPase, changes of mitochondrial membrane potential (m)and expression changes of mitochondrial uncoupling protein UCP2and UCP4protein in spinal cord to assess mitochondrial function changes and possibleinvolvement of mitochondrial protein system. At last we observed the painbehavior changes induced by formalin after intrathecal injection of antioxidantTEMPOL (4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl) and/or UCP4RNAi.The present results showed that the levels of oxidative stress in spinal cordexaggerated in both acute and tonic phases of formalin pain model. Moreover,mitochondria function in acute phase was impaired significantly, whereas recovered and enhanced in tonic phase. The UCP4-related mitochondrialprotection system was trigged in tonic phase and contributed to the centralsensitization of formalin pain. In conclusion, spinal mitochondria may playdifferent functions in biphasic phases of formalin pain model: damaged in theacute phase, but recoverd in tonic phase to resist the imbalance of mitochondrialhomeostasis. Therefore, we suggest that the mitochondria at the spinal cord levelmay play different roles during generation and maintenance of neuropathic pain;the understanding of mitochondrial function in pain could help us treat thechronic pain.