Neuronal-Glial Interactions Maintain Chronic Neuropathic Pain after Spinal Cord Injury.

The hyperactive state of sensory neurons in the spinal cord enhances pain transmission. Spinal glial cells have also been implicated in enhanced excitability of spinal dorsal horn neurons, resulting in pain amplification and distortions. Traumatic injuries of the neural system such as spinal cord injury (SCI) induce neuronal hyperactivity and glial activation, causing maladaptive synaptic plasticity in the spinal cord. Recent studies demonstrate that SCI causes persistent glial activation with concomitant neuronal hyperactivity, thus providing the substrate for central neuropathic pain. Hyperactive sensory neurons and activated glial cells increase intracellular and extracellular glutamate, neuropeptides, adenosine triphosphates, proinflammatory cytokines, and reactive oxygen species concentrations, all of which enhance pain transmission. In addition, hyperactive sensory neurons and glial cells overexpress receptors and ion channels that maintain this enhanced pain transmission. Therefore, post-SCI neuronal-glial interactions create maladaptive synaptic circuits and activate intracellular signaling events that permanently contribute to enhanced neuropathic pain. In this review, we describe how hyperactivity of sensory neurons contributes to the maintenance of chronic neuropathic pain via neuronal-glial interactions following SCI.

Comparison of Mechanical Allodynia and Recovery of Locomotion and Bladder Function by Different Parameters of Low Thoracic Spinal Contusion Injury in Rats.

BACKGROUND: The present study was designed to examine the functional recovery following spinal cord injury (SCI) by adjusting the parameters of impact force and dwell-time using the Infinite Horizon (IH) impactor device. METHODS: Sprague-Dawley rats (225-240 g) were divided into eight injury groups based on force of injury (Kdyn) and dwell time (seconds), indicated as Force-Dwell time: 150-4, 150-3, 150-2, 150-1, 150-0, 200-0, 90-2 and sham controls, respectively. RESULTS: After T10 SCI, higher injury force produced greater spinal cord displacement (P < 0.05) and showed a significant correlation (r = 0.813) between the displacement and the force (P < 0.05). In neuropathic pain-like behavior, the percent of paw withdrawals scores in the hindpaw for the 150-4, 150-3, 150-2, 150-1 and the 200-0 injury groups were significantly lowered compared with sham controls (P < 0.05). The recovery of locomotion had a significant within-subjects effect of time (P < 0.05) and the 150-0 group had increased recovery compared to other groups (P < 0.05). In addition, the 200-0 and the 90-2 recovered significantly better than all the 150 kdyn impact groups that included a dwell-time (P < 0.05). In recovery of spontaneous bladder function, the 150-4 injury group took significantly longer recovery time whereas the 150-0 and the 90-2 groups had the shortest recovery times. CONCLUSIONS: The present study demonstrates SCI parameters optimize development of mechanical allodynia and other pathological outcomes.

Reactive oxygen species and lipid peroxidation inhibitors reduce mechanical sensitivity in a chronic neuropathic pain model of spinal cord injury in rats.

Chronic neuropathic pain is a common consequence of spinal cord injury (SCI), develops over time and negatively impacts quality of life, often leading to substance abuse and suicide. Recent evidence has demonstrated that reactive oxygen species (ROS) play a role in contributing to neuropathic pain in SCI animal models. This investigation examines four compounds that reduce ROS and the downstream lipid peroxidation products, apocynin, 4-oxo-tempo, U-83836E, and tirilazad, and tests if these compounds can reduce nocioceptive behaviors in chronic SCI animals. Apocynin and 4-oxo-tempo significantly reduced abnormal mechanical hypersensitivity measured in forelimbs and hindlimbs in a model of chronic SCI-induced neuropathic pain. Thus, compounds that inhibit ROS or lipid peroxidation products can be used to ameliorate chronic neuropathic pain. We propose that the application of compounds that inhibit reactive oxygen species (ROS) and related downstream molecules will also reduce the behavioral measures of chronic neuropathic pain. Injury or trauma to nervous tissue leads to increased concentrations of ROS in the surviving tissue. Further damage from ROS molecules to dorsal lamina neurons leads to membrane excitability, the physiological correlate of chronic pain. Chronic pain is difficult to treat with current analgesics and this research will provide a novel therapy for this disease.

Reactive oxygen species contribute to neuropathic pain and locomotor dysfunction via activation of CamKII in remote segments following spinal cord contusion injury in rats.

In this study, we examined whether blocking spinal cord injury (SCI)-induced increases in reactive oxygen species (ROS) by a ROS scavenger would attenuate below-level central neuropathic pain and promote recovery of locomotion. Rats with T10 SCI developed mechanical allodynia in both hind paws and overproduction of ROS, as assayed by Dhet intensity, in neurons in the lumbar 4/5 dorsal horn ((∗)P<0.05). To scavenge ROS, phenyl-N-tert-butylnitrone (PBN, a ROS scavenger) was administered immediately after SCI and for 7 consecutive days (early treatment) by either intrathecal (it; 1 and 3mg) or systemic (ip; 10, 50 and 100mg) injections. In addition, the high doses of it (3mg) or ip (100mg) injections were performed at 35 days (delayed treatment) after SCI. High doses of PBN (ip, 100mg, and it, 3mg) significantly attenuated mechanical allodynia in both hind paws at both early and delayed treatments, respectively ((∗)P<0.05). The abnormal hyperexcitability of wide dynamic range neurons after SCI was significantly attenuated by both early and delayed PBN treatment ((∗)P<0.05). Early PBN treatment (100mg, ip, and 3mg, it) attenuated overproduction of ROS in neurons in the lumbar 4/5 dorsal horn. In addition, it and ip t-BOOH (ROS donor) treatment dose-dependently produced mechanical allodynia in both hind paws ((∗)P<0.05). Both SCI and t-BOOH treatment groups showed significantly increased phospho-CamKII (pCamKII) expression in neurons and KN-93 (an inhibitor of pCamKII) significantly attenuated mechanical allodynia ((∗)P<0.05). In addition, high doses of PBN significantly promoted the recovery of locomotion ((∗)P<0.05). In conclusion, the present data suggest that overproduction of ROS contribute to sensory and motor abnormalities in remote segments below the lesion after thoracic SCI.

Inflammatory consequences in a rodent model of mild traumatic brain injury.

Mild traumatic brain injury (mTBI), particularly mild "blast type" injuries resulting from improvised exploding devices and many sport-caused injuries to the brain, result in long-term impairment of cognition and behavior. Our central hypothesis is that there are inflammatory consequences to mTBI that persist over time and, in part, are responsible for resultant pathogenesis and clinical outcomes. We used an adaptation (1 atmosphere pressure) of a well-characterized moderate-to-severe brain lateral fluid percussion (LFP) brain injury rat model. Our mild LFP injury resulted in acute increases in interleukin-1α/ß and tumor necrosis factor alpha levels, macrophage/microglial and astrocytic activation, evidence of heightened cellular stress, and blood-brain barrier (BBB) dysfunction that were evident as early as 3-6 h postinjury. Both glial activation and BBB dysfunction persisted for 18 days postinjury.

Challenges in the development of rodent models of mild traumatic brain injury.

Approximately 75% of traumatic brain injuries (TBI) are classified mild (mTBI). Despite the high frequency of mTBI, it is the least well studied. The prevalence of mTBI among service personnel returning from Operations Iraqi Freedom (OIF) and Enduring Freedom (OEF) and the recent reports of an association between repeated mTBI and the early onset of Alzheimer's and other types of dementias in retired athletes has focused much attention on mTBI. The study of mTBI requires the development and validation of experimental models and one of the most basic requirements for an experimental model is that it replicates important features of the injury or disease in humans. mTBI in humans is associated with acute symptoms such as loss of consciousness and pre- and/or posttraumatic amnesia. In addition, many mTBI patients experience long-term effects of mTBI, including deficits in speed of information processing, attention and concentration, memory acquisition, retention and retrieval, and reasoning and decision-making. Although methods for the diagnosis and evaluation of the acute and chronic effects of mTBI in humans are well established, the same is not the case for rodents, the most widely used animal for TBI studies. Despite the magnitude of the difficulties associated with adapting these methods for experimental mTBI research, they must be surmounted. The identification and testing of treatments for mTBI depends of the development, characterization and validation of reproducible, clinically relevant models of mTBI.

Calcium/calmodulin dependent kinase II contributes to persistent central neuropathic pain following spinal cord injury.

Chronic central neuropathic pain after central nervous system injuries remains refractory to therapeutic interventions. A novel approach would be to target key intracellular signaling proteins that are known to contribute to continued activation by phosphorylation of kinases, transcription factors, and/or receptors that contribute to changes in membrane excitability. We demonstrate that one signaling kinase, calcium/calmodulin-dependent kinase II (CaMKII), is critical in maintaining aberrant dorsal horn neuron hyperexcitability in the neuropathic pain condition after spinal cord injury (SCI). After contusion SCI at spinal level T10, activated CaMKII (phosphorylated, pCaMKII) expression is significantly upregulated in the T7/8 spinal dorsal horn in neurons, but not glial cells, and in oligodendrocytes in the dorsal column in the same rats that displayed at-level mechanical allodynia. Furthermore, identified spinothalamic neurons demonstrated significant increases of pCaMKII after SCI compared to sham-treated control animals. However, neither astrocytes nor microglia showed pCaMKII expression in either sham-treated or SCI rats. To demonstrate causality, treatment of SCI rats with KN-93, which prevents CaMKII activation, significantly attenuated at-level mechanical allodynia and aberrant wide dynamic range neuronal activity evoked by brush, pressure, and pinch stimuli and a graded series of von Frey stimuli, respectively. Persistent CaMKII activation contributes to chronic central neuropathic pain by mechanisms that involve maintained hyperexcitability of wide dynamic range dorsal horn neurons. Furthermore, targeting key signaling proteins is a novel, useful therapeutic strategy for treating chronic central neuropathic pain.

Special issue on microglia and chronic pain.

This special issue of Experimental Neurology is devoted to the role of Microglia and Chronic Pain. Chronic pain affects 116 million people per year in the United States, which is more than heart disease, cancer, and diabetes combined. Nervous system trauma and disease are principal contributors to the establishment of chronic pain in people and in animal models. Central nervous system (CNS) injury or tumor development, peripheral nerve injury, multiple sclerosis, diabetes and many other neurological disruptions can serve as the instigating pathophysiolgical conditions that lead to chronic pain. Once considered to function solely as the phagocytotic cells of the CNS, more recent work has demonstrated that persistent activation of the microglial population may contribute to continued dysfunction including chronic pain. In the invited articles for this special issue on Microglia and Chronic Pain, we present evidence for the role of persistent microglial activation in chronic pain after peripheral and central nervous system injury, as well as in diabetic pain, post-herpetic neuralgia pain and related diseases. Collectively, the body of work indicates the importance of understanding the roles of microglial cells in chronic pain which will lead to targeted treatment to attenuate or alleviate chronic neuropathic pain syndromes.

Spatial and temporal activation of spinal glial cells: role of gliopathy in central neuropathic pain following spinal cord injury in rats.

In the spinal cord, neuron and glial cells actively interact and contribute to neurofunction. Surprisingly, both cell types have similar receptors, transporters and ion channels and also produce similar neurotransmitters and cytokines. The neuroanatomical and neurochemical similarities work synergistically to maintain physiological homeostasis in the normal spinal cord. However, in trauma or disease states, spinal glia become activated, dorsal horn neurons become hyperexcitable contributing to sensitized neuronal-glial circuits. The maladaptive spinal circuits directly affect synaptic excitability, including activation of intracellular downstream cascades that result in enhanced evoked and spontaneous activity in dorsal horn neurons with the result that abnormal pain syndromes develop. Recent literature reported that spinal cord injury produces glial activation in the dorsal horn; however, the majority of glial activation studies after SCI have focused on transient and/or acute time points, from a few hours to 1 month, and peri-lesion sites, a few millimeters rostral and caudal to the lesion site. In addition, thoracic spinal cord injury produces activation of astrocytes and microglia that contributes to dorsal horn neuronal hyperexcitability and central neuropathic pain in above-level, at-level and below-level segments remote from the lesion in the spinal cord. The cellular and molecular events of glial activation are not simple events, rather they are the consequence of a combination of several neurochemical and neurophysiological changes following SCI. The ionic imbalances, neuroinflammation and alterations of cell cycle proteins after SCI are predominant components for neuroanatomical and neurochemical changes that result in glial activation. More importantly, SCI induced release of glutamate, proinflammatory cytokines, ATP, reactive oxygen species (ROS) and neurotrophic factors trigger activation of postsynaptic neuron and glial cells via their own receptors and channels that, in turn, contribute to neuronal-neuronal and neuronal-glial interaction as well as microglia-astrocytic interactions. However, a systematic review of temporal and spatial glial activation following SCI has not been done. In this review, we describe time and regional dependence of glial activation and describe activation mechanisms in various SCI models in rats. These data are placed in the broader context of glial activation mechanisms and chronic pain states. Our work in the context of work by others in SCI models demonstrates that dysfunctional glia, a condition called "gliopathy", is a key contributor in the underlying cellular mechanisms contributing to neuropathic pain.

Neuronal hyperexcitability: a substrate for central neuropathic pain after spinal cord injury.

Neuronal hyperexcitability produces enhanced pain transmission in the spinal dorsal horn after spinal cord injury (SCI). Spontaneous and evoked neuronal excitability normally are well controlled by neural circuits. However, SCI produces maladaptive synaptic circuits in the spinal dorsal horn that result in neuronal hyperexcitability. After SCI, activated primary afferent neurons produce enhanced release of glutamate, neuropeptides, adenosine triphosphate, and proinflammatory cytokines, which are known to be major components for pain transmission in the spinal dorsal horn. Enhanced neurochemical events contribute to neuronal hyperexcitability, and neuroanatomical changes also contribute to maladaptive synaptic circuits and neuronal hyperexcitability. These neurochemical and neuroanatomical changes produce enhanced cellular signaling cascades that ensure persistently enhanced pain transmission. This review describes altered neurochemical and neuroanatomical contributions on neuronal hyperexcitability in the spinal dorsal horn, which serve as substrates for central neuropathic pain after SCI.

GABA and central neuropathic pain following spinal cord injury.

Spinal cord injury induces maladaptive synaptic transmission in the somatosensory system that results in chronic central neuropathic pain. Recent literature suggests that glial-neuronal interactions are important modulators in synaptic transmission following spinal cord injury. Neuronal hyperexcitability is one of the predominant phenomenon caused by maladaptive synaptic transmission via altered glial-neuronal interactions after spinal cord injury. In the somatosensory system, spinal inhibitory neurons counter balance the enhanced synaptic transmission from peripheral input. For a decade, the literature suggests that hypofunction of GABAergic inhibitory tone is an important factor in the enhanced synaptic transmission that often results in neuronal hyperexcitability in dorsal horn neurons following spinal cord injury. Neurons and glial cells synergistically control intracellular chloride ion gradients via modulation of chloride transporters, extracellular glutamate and GABA concentrations via uptake mechanisms. Thus, the intracellular "GABA-glutamate-glutamine cycle" is maintained for normal physiological homeostasis. However, hyperexcitable neurons and glial activation after spinal cord injury disrupts the balance of chloride ions, glutamate and GABA distribution in the spinal dorsal horn and results in chronic neuropathic pain. In this review, we address spinal cord injury induced mechanisms in hypofunction of GABAergic tone that results in chronic central neuropathic pain. This article is part of a Special Issue entitled 'Synaptic Plasticity & Interneurons'.

Ionotropic glutamate receptors contribute to maintained neuronal hyperexcitability following spinal cord injury in rats.

In this study, we examined whether topical treatment of glutamate receptor antagonists attenuate hyperexcitability of lumbar spinal dorsal horn neurons following low thoracic hemisection spinal cord injury in rats. Four weeks after spinal hemisection, neuronal activity in response to mechanical stimuli applied on the peripheral receptive field was significantly increased in three different phenotypes of lumbar spinal dorsal horn neurons: wide dynamic range (WDR), low threshold (LT) and high threshold (HT). Topical application of MK-801 (NMDA receptor antagonist, 50 microg) significantly attenuated the activity of WDR, but not LT and HT neurons; whereas, NBQX (AMPA receptor antagonist, 0.5 and 1 microg) significantly attenuated neuronal activity in all three phenotypes of neurons (*p<0.05). However, MCPG (group I/II metabotropic glutamate receptor antagonist, 100 microg) had no effect. The present study, in the context of previous work, suggests that ionotropic glutamate receptor activation play critical roles in the maintenance of neuronal hyperexcitability and neuropathic "below-level" pain behavior following spinal hemisection injury.

Peripheral and central sensitization in remote spinal cord regions contribute to central neuropathic pain after spinal cord injury.

Central neuropathic pain (CNP) developing after spinal cord injury (SCI) is described by the region affected: above-level, at-level and below-level pain occurs in dermatomes rostral, at/near, or below the SCI level, respectively. People with SCI and rodent models of SCI develop above-level pain characterized by mechanical allodynia and thermal hyperalgesia. Mechanisms underlying this pain are unknown and the goals of this study were to elucidate components contributing to the generation of above-level CNP. Following a thoracic (T10) contusion, forelimb nociceptors had enhanced spontaneous activity and were sensitized to mechanical and thermal stimulation of the forepaws 35 days post-injury. Cervical dorsal horn neurons showed enhanced responses to non-noxious and noxious mechanical stimulation as well as thermal stimulation of receptive fields. Immunostaining dorsal root ganglion (DRG) cells and cord segments with activating transcription factor 3 (ATF3, a marker for neuronal injury) ruled out neuronal damage as a cause for above-level sensitization since few C8 DRG cells expressed AFT3 and cervical cord segments had few to no ATF3-labeled cells. Finally, activated microglia and astrocytes were present in thoracic and cervical cord at 35 days post-SCI, indicating a rostral spread of glial activation from the injury site. Based on these data, we conclude that peripheral and central sensitization as well as reactive glia in the uninjured cervical cord contribute to CNP. We hypothesize that reactive glia in the cervical cord release pro-inflammatory substances which drive chronic CNP. Thus a complex cascade of events spanning many cord segments underlies above-level CNP.

Activation of p-38alpha MAPK contributes to neuronal hyperexcitability in caudal regions remote from spinal cord injury.

In the present study, we examined whether activation of p-38alpha MAPK modulates mechanical allodynia and neuronal hyperexcitability, and if propentofylline (PPF, a glial modulator) modulates specifically localized activated p-38alpha MAPK expression in caudal regions remote from a low thoracic hemisection injury in rats. T13 spinal hemisection produces bilateral mechanical allodynia in hindpaws with evoked (in response to mechanical stimuli) neuronal hyperexcitability in lumbar spinal wide dynamic range (WDR) neurons compared to sham controls. The mechanical allodynia and the evoked activity of WDR neurons is attenuated by intrathecal and topical administration of SB203580, an inhibitor of p-38 MAPK activation, dose dependently (p<0.05); however, the spontaneous activity showed no significant differences compared to sham controls. After T13 spinal hemisection, significantly increased phosphorylated (activated form) p-38alpha MAPK expression was present in both superficial and deep dorsal horn neurons as well as in microglia, but not in astrocytes, in the lumbar spinal cord compared to sham controls (p<0.05). Intrathecal application of PPF significantly attenuated the expression of phosphorylated p-38alpha MAPK in superficial dorsal horn neurons (10 mM) and in microglia (1 and 10 mM) in the lumbar spinal cord compared to the hemisection group (p<0.05). In conclusion, our present data demonstrate that activated neuronal and microglial, but not astrocytic, p-38alpha MAPK contributes to the maintenance of neuronal hyperexcitability in caudal regions following spinal cord injury.

Mechanisms of chronic central neuropathic pain after spinal cord injury.

Not all spinal contusions result in mechanical allodynia, in which non-noxious stimuli become noxious. The studies presented use the NYU impactor at 12.5 mm drop or the Infinite Horizons Impactor (150 kdyn, 1 s dwell) devices to model spinal cord injury (SCI). Both of these devices and injury parameters, if done correctly, will result in animals with above level (forelimb), at level (trunk) and below level (hindlimb) mechanical allodynia that model the changes in evoked somatosensation experienced by the majority of people with SCI. The sections are as follows: 1) Mechanisms of remote microglial activation and pain signaling in "below-level" central pain 2) Intracellular signaling mechanisms in central sensitization in "at-level" pain 3) Peripheral sensitization contributes to "above level" injury pain following spinal cord injury and 4) Role of reactive oxygen species in central sensitization in regional neuropathic pain following SCI. To summarize, differential regional mechanisms contribute to the regional chronic pain states. We propose the importance of understanding the mechanisms in the differential regional pain syndromes after SCI in the chronic condition. Targeting regional mechanisms will be of enormous benefit to the SCI population that suffer chronic pain, and will contribute to better treatment strategies for other chronic pain syndromes.

Gliopathy ensures persistent inflammation and chronic pain after spinal cord injury.

Research focused on improving recovery of function, including the reduction of central neuropathic pain (CNP) after spinal cord injury (SCI) is essential. After SCI, regional neuropathic pain syndromes above, at and below the level or spinal injury develop and are thought to have different mechanisms, but may share common dysfunctional glial mechanisms. Detloff et al., [Detloff, M.R., Fisher, L.C., McGaughy, V., Longbrake, E.E., Popovich, P.G., Basso, D.M., Remote activation of microglia and pro-inflammatory cytokines predict the onset and severity of below-level neuropathic pain after spinal cord injury in rats. Exp. Neurol. (2008), doi: 10.1016/j.expneurol.2008.04.009.] describe events in the lumbar region of the spinal cord after a midthoracic SCI injury, the so called "below-level" pain and compares the findings to peripheral nerve lesion findings. This commentary briefly reviews glial contributions and intracellular signaling mechanisms, both neuronal and glial, that provide the substrate for CNP after SCI, including the persistent glial production of factors that can maintain sensitization of dorsal horn neurons in segments remote from the spinal injury; ie. dorsal horn hyperexcitability to formerly non noxious stimuli that become noxious after SCI resulting in allodynia. The term "gliopathy" is proposed to describe the dysfunctional and maladaptive response of glial cells, specifically astrocytes and microglia, to neural injury that is initiated by the sudden injury induced increase in extracellular concentrations of glutamate and concomitant production of several proinflammatory molecules. It is important to understand the roles that different glia play in "gliopathy", a condition that appears to persist after SCI. Furthermore, targeted treatment of gliopathy will attenuate mechanical allodynia in both central and peripheral neuropathic pain syndromes.

Regulation of interleukin-1beta by the interleukin-1 receptor antagonist in the glutamate-injured spinal cord: endogenous neuroprotection.

Elevation of extracellular glutamate contributes to cell death and functional impairments generated by spinal cord injury (SCI), in part through the activation of the neurotoxic cytokine interleukin-1beta (IL-1beta). This study examines the participation of IL-1beta and its regulation by the endogenous interleukin-1 receptor antagonist (IL-1ra) in glutamate toxicity following SCI. Glutamate, glutamatergic agonists and SCI had similar effects on levels of IL-1beta and IL-1ra. Following spinal cord contusion or exposure to elevated glutamate, concentrations of IL-1beta first increased as IL-1ra decreased, and both then changed in the opposite directions. Applying the glutamate agonists NMDA and S-AMPA to the spinal cord caused changes in IL-1beta and IL-1ra levels very similar to those produced by contusion and glutamate. The glutamate antagonists MK801 and NBQX blocked the glutamate-induced changes in IL-1beta and IL-1ra levels. Administering IL-1beta elevated IL-1ra, and administering IL-1ra depressed IL-1beta levels. Infusing IL-beta into the spinal cord impaired locomotion, and infusing IL-1ra improved recovery from glutamate-induced motor impairments. We hypothesize that elevating IL-1ra opposes the damage caused by IL-1beta in SCI by reducing IL-1beta levels as well as by blocking binding of IL-1beta to its receptor. Our results demonstrate that IL-1beta contributes to glutamate damage following SCI; blocking IL-1beta may usefully counteract glutamate toxicity.

Activation of p38 MAP kinase is involved in central neuropathic pain following spinal cord injury.

Recent work regarding chronic central neuropathic pain (CNP) following spinal cord injury (SCI) suggests that activation of key signaling molecules such as members of the mitogen activated protein kinase (MAPK) family play a role in the expression of at-level mechanical allodynia. Previously, we have shown that the development of at-level CNP following moderate spinal cord injury is correlated with increased expression of the activated (and thus phosphorylated) forms of the MAPKs extracellular signal related kinase and p38 MAPK. The current study extends this work by directly examining the role of p38 MAPK in the maintenance of at-level CNP following spinal cord injury. Using a combination of behavioral, immunocytochemical, and electrophysiological measures we demonstrate that increased activation of p38 MAPK occurs in the spinal cord just rostral to the site of injury in rats that develop at-level mechanical allodynia after moderate SCI. Immunocytochemical analyses indicate that the increases in p38 MAPK activation occurred in astrocytes, microglia, and dorsal horn neurons in the spinal cord rostral to the site of injury. Inhibiting the enzymatic activity of p38 MAPK dose dependently reverses the behavioral expression of at-level mechanical allodynia and also decreases the hyperexcitability seen in thoracic dorsal horn neurons after moderate SCI. Taken together, these novel data are the first to demonstrate causality that increased activation of p38 MAPK in multiple cell types play an important role in the maintenance of at-level CNP following spinal cord injury.