D1/D5 Dopamine Receptors and mGluR5 Jointly Enable Non-Hebbian Long-Term Potentiation at Sensory Synapses onto Lamina I Spinoparabrachial Neurons.

Highly correlated firing of primary afferent inputs and lamina I projection neurons evokes synaptic long-term potentiation (LTP), a mechanism by which ascending nociceptive transmission can be amplified at the level of the spinal dorsal horn. However, the degree to which neuromodulatory signaling shapes the temporal window governing spike-timing-dependent plasticity (STDP) at sensory synapses onto projection neurons remains unclear. The present study demonstrates that activation of spinal D1/D5 dopamine receptors (D1/D5Rs) creates a highly permissive environment for the production of LTP in male and female adult mouse spinoparabrachial neurons by promoting non-Hebbian plasticity. Bath application of the mixed D1/D5R agonist SKF82958 unmasked LTP at STDP pairing intervals that normally fail to alter synaptic efficacy. Furthermore, during D1/D5R signaling, action potential discharge in projection neurons became dispensable for LTP generation, and primary afferent stimulation alone was sufficient to induce strengthening of sensory synapses. This non-Hebbian LTP was blocked by the D1/D5R antagonist SCH 39166 or genetic deletion of D5R, and required activation of mGluR5 and intracellular Ca2+ release but was independent of NMDAR activation. D1/D5R-enabled non-Hebbian plasticity was observed across multiple neuronal subpopulations in the superficial dorsal horn but was more prevalent in spinoparabrachial neurons than interneurons. Interestingly, the ability of neonatal tissue damage to promote non-Hebbian LTP in adult projection neurons was not observed in D5R knock-out mice. Collectively, these findings suggest that joint spinal D1/D5R and mGluR5 activation can allow unfettered potentiation of sensory synapses onto the output neurons responsible for conveying pain and itch information to the brain.SIGNIFICANCE STATEMENT Synaptic LTP in spinal projection neurons has been implicated in the generation of chronic pain. Under normal conditions, plasticity at sensory synapses onto adult mouse spinoparabrachial neurons follows strict Hebbian learning rules, requiring coincident presynaptic and postsynaptic firing. Here, we demonstrate that the activation of spinal D1/D5Rs promotes a switch from Hebbian to non-Hebbian LTP so that primary afferent stimulation alone is sufficient to evoke LTP in the absence of action potential discharge in projection neurons, which required joint activation of mGluR5 and intracellular Ca2+ release but not NMDARs. These results suggest that D1/D5Rs cooperate with mGluR5 receptors in the spinal dorsal horn to powerfully influence the amplification of ascending nociceptive transmission to the brain.

Chemokine CCL2 prevents opioid-induced inhibition of nociceptive synaptic transmission in spinal cord dorsal horn.

BACKGROUND: Opioid analgesics remain widely used for pain treatment despite the related serious side effects. Some of those, such as opioid tolerance and opioid-induced hyperalgesia may be at least partially due to modulation of opioid receptors (OR) function at nociceptive synapses in the spinal cord dorsal horn. It was suggested that increased release of different chemokines under pathological conditions may play a role in this process. The goal of this study was to investigate the crosstalk between the µOR, transient receptor potential vanilloid 1 (TRPV1) receptor and C-C motif ligand 2 (CCL2) chemokine and the involvement of spinal microglia in the modulation of opioid analgesia. METHODS: Patch-clamp recordings of miniature excitatory postsynaptic currents (mEPSCs) and dorsal root evoked currents (eEPSC) in spinal cord slices superficial dorsal horn neurons were used to evaluate the effect of µOR agonist [D-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin (DAMGO), CCL2, TRPV1 antagonist SB366791 and minocycline. Paw withdrawal test to thermal stimuli was combined with intrathecal (i.t.) delivery of CCL2 and DAMGO to investigate the modulation in vivo. RESULTS: Application of DAMGO induced a rapid decrease of mEPSC frequency and eEPSC amplitude, followed by a delayed increase of the eESPC amplitude, which was prevented by SB366791. Chemokine CCL2 treatment significantly diminished all the DAMGO-induced changes. Minocycline treatment prevented the CCL2 effects on the DAMGO-induced eEPSC depression, while mEPSC changes were unaffected. In behavioral experiments, i.t. injection of CCL2 completely blocked DAMGO-induced thermal hypoalgesia and intraperitoneal pre-treatment with minocycline prevented the CCL2 effect. CONCLUSIONS: Our results indicate that opioid-induced inhibition of the excitatory synaptic transmission could be severely attenuated by increased CCL2 levels most likely through a microglia activation-dependent mechanism. Delayed potentiation of neurotransmission after µOR activation is dependent on TRPV1 receptors activation. Targeting CCL2 and its receptors and TRPV1 receptors in combination with opioid therapy could significantly improve the analgesic properties of opioids, especially during pathological states.

Propofol Alleviates Neuropathic Pain Induced by Chronic Contractile Injury by Regulating the Spinal glun2b-p38mapkepac1 Pathway.

BACKGROUND: Propofol acts as an intravenous anesthetic cure which is widely used as a therapy for the craniocerebral injury that comprised surgical anesthesia as well as the sedation done in the intensive care units. Propofol is one of the most commonly used and efficient anesthetics where the painful effects are followed by an injection of propofol. In many cases, patients experience pain followed by anxiety, boredom, fear, and even myocardial ischemia. OBJECTIVE: This study was performed to investigate the underlying mechanism of propofol and its effect on regulating spinal glun2b-p38mapkepac1 pathways in chronic contractile injury. Material and Methods. Contractile injury was performed by ligation around the nerve of the thigh region postanesthesia. Rats were divided into three groups to analyze the changes like mechanical allodynia by the paw withdrawal threshold and histopathological analysis for assessing cellular degradation. L4-L6 from the spinal dorsal horns were isolated and harvested for studying protein expression, by the method of western blotting and immunofluorescence analysis. RESULTS: The pain caused due to mechanical allodynia in the paw region was highest at 1 hour postinduction and lasted for three days postinjury. Pain was significantly less in the group receiving propofol when compared with the isoflurane group for the first two hours of injury. In the propofol group, EPAC1, GluN2B, and p38 MAP K were significantly lower. CONCLUSION: In the rat model of induced chronic contractile injury, postsurgery there was a suppression of the GluN2B-p38MAPK/EPAC1 signaling pathway in the propofol group. As the p38MAPK/EPAC pathway has a significant role in the postoperative hyperalgesia, thus our experiment suggests that propofol has analgesic effects.

Exercise Reduces Morphine-Induced Hyperalgesia and Antinociceptive Tolerance.

Chronic morphine intake for treating various pain is frequently concomitant with morphine-induced hyperalgesia and tolerance. The mechanisms can be explained by the activation of p38-MAPK proteins in microglia in the spinal cord horn. Exercise has been shown to prevent the development of microglia overactivation. Thus, we designed to test whether exercise prevents the morphine-induced hyperalgesia and tolerance as well as suppression of p38 phosphorylation. A p38 inhibitor SB203580, exercise, and exercise preconditioning were used for treating morphine-induced hyperalgesia and tolerance development in the present study. The behavior tests for hyperalgesia and tolerance were performed in male Wistar rats before and after morphine administration. Western blotting and immunostaining for examining phosphorylated-p38 expression were performed after the behavior tests. Our results showed that SB203580 and exercise, but not exercise preconditioning, prevented the occurrence of morphine-induced hyperalgesia and tolerance. Meanwhile, exercise decreased morphine-induced phosphorylated-p38 overexpression. In summary, exercise prevented the development of morphine-induced hyperalgesia and tolerance. The mechanism may be related to inhibition of p38 phosphorylation.

Repurposing cancer drugs identifies kenpaullone which ameliorates pathologic pain in preclinical models via normalization of inhibitory neurotransmission.

Inhibitory GABA-ergic neurotransmission is fundamental for the adult vertebrate central nervous system and requires low chloride concentration in neurons, maintained by KCC2, a neuroprotective ion transporter that extrudes intracellular neuronal chloride. To identify Kcc2 gene expression­enhancing compounds, we screened 1057 cell growth-regulating compounds in cultured primary cortical neurons. We identified kenpaullone (KP), which enhanced Kcc2/KCC2 expression and function in cultured rodent and human neurons by inhibiting GSK3ß. KP effectively reduced pathologic pain-like behavior in mouse models of nerve injury and bone cancer. In a nerve-injury pain model, KP restored Kcc2 expression and GABA-evoked chloride reversal potential in the spinal cord dorsal horn. Delta-catenin, a phosphorylation-target of GSK3ß in neurons, activated the Kcc2 promoter via KAISO transcription factor. Transient spinal over-expression of delta-catenin mimicked KP analgesia. Our findings of a newly repurposed compound and a novel, genetically-encoded mechanism that each enhance Kcc2 gene expression enable us to re-normalize disrupted inhibitory neurotransmission through genetic re-programming.

AKAP150 and its Palmitoylation Contributed to Pain Hypersensitivity Via Facilitating Synaptic Incorporation of GluA1-Containing AMPA Receptor in Spinal Dorsal Horn.

The A-kinase anchoring protein 150 (AKAP150) organizes kinases and phosphatases to regulate AMPA receptors (AMPARs) that are pivotal for synaptic plasticity. AKAP150 itself undergoes S-palmitoylation. However, the roles of AKAP150 and its palmitoylation in spinal nociceptive processing remain unknown. In this study, we found that intraplantar injection of complete Freund's adjuvant (CFA) significantly increased the synaptic expression of AKAP150 and caused a reorganization of AKAP150 signaling complex in spinal dorsal horn. Knockdown of AKAP150 or interruption of its interactions with kinases effectively suppressed the CFA-induced synaptic expression of GluA1 subunit of AMPARs. Our data also showed that an upregulation of AKAP150 palmitoylation was involved in the synaptic redistribution of AKAP150. Inhibition of AKAP150 palmitoylation by expression of palmitoylation-defective mutant AKAP150 (C36, 123S) effectively repressed the CFA-induced phosphorylation and synaptic expression of GluA1 subunit, meanwhile, attenuated the development of mechanical allodynia and thermal hyperalgesia. Furthermore, we found that an increased expression of palmitoyl acyltransferase ZDHHC2 contributed to the upregulation of AKAP150 palmitoylation and GluA1 accumulation in inflamed mouse. These data indicated that AKAP150 and its palmitoylation were involved in AMPA receptor-dependent modification of nociceptive transmission, and the manipulations of AKAP150 signaling complex and palmitoylation might serve as potential therapeutic strategies for persistent pain after inflammation.

Sulforaphane alleviates hyperalgesia and enhances analgesic potency of morphine in rats with cancer-induced bone pain.

Due to the efficacy and tolerability of the available drugs, the current treatment for cancer-induced bone pain (CIBP) is not considered ideal, and new drugs are required for better treatment results. This study investigated whether intrathecal injection of sulforaphane (SFN) can modulates the noxious behavior associated with CIBP and enhances the analgesic effects of morphine and the possible mechanisms related to these effects were investigated. Walker256 breast cancer cells were injected into the bone marrow cavity of rats to establish the CIBP model. When CIBP rats began to exhibit painful behavior (CIBP 6 days), SFN was injected intrathecally for 7 days. The results showed that SFN alleviated the painful behavioral hypersensitivity caused by cancer, accompanied by nuclear factor, erythroid 2 like 2 (Nrf2), Haem oxygenase 1 (HO-1) activation, nuclear factor kappa B (NF-κB) inhibition and inflammation-related factors (tumor necrosis factor-alpha (TNF-α), interleukin-1ß (IL-ß), interleukin-6 (IL-6), and inducible nitric oxide synthase (iNOS) reduction. In addition, SFN treatment inhibited the proliferation of Walker 256 cells in a dose-dependent manner, promoted mu-opioid receptor (MOR) expression in SH-SY5Y cells and enhanced the antihyperalgesic effects of morphine on CIBP rats by restoring the downregulation of MOR expression in the spinal cord. Interestingly, the antihyperalgesic effects of SFN were partially blocked by opioid receptor antagonists. This study showed that SFN combined with morphine might be a new way to treat CIBP.

Glycine Receptors in Spinal Nociceptive Control-An Update.

Diminished inhibitory control of spinal nociception is one of the major culprits of chronic pain states. Restoring proper synaptic inhibition is a well-established rational therapeutic approach explored by several pharmaceutical companies. A particular challenge arises from the need for site-specific intervention to avoid deleterious side effects such as sedation, addiction, or impaired motor control, which would arise from wide-range facilitation of inhibition. Specific targeting of glycinergic inhibition, which dominates in the spinal cord and parts of the hindbrain, may help reduce these side effects. Selective targeting of the α3 subtype of glycine receptors (GlyRs), which is highly enriched in the superficial layers of the spinal dorsal horn, a key site of nociceptive processing, may help to further narrow down pharmacological intervention on the nociceptive system and increase tolerability. This review provides an update on the physiological properties and functions of α3 subtype GlyRs and on the present state of related drug discovery programs.

Pain and itch processing by subpopulations of molecularly diverse spinal and trigeminal projection neurons.

A remarkable molecular and functional heterogeneity of the primary sensory neurons and dorsal horn interneurons transmits pain- and or itch-relevant information, but the molecular signature of the projection neurons that convey the messages to the brain is unclear. Here, using retro-TRAP (translating ribosome affinity purification) and RNA sequencing, we reveal extensive molecular diversity of spino- and trigeminoparabrachial projection neurons. Among the many genes identified, we highlight distinct subsets of Cck+ -, Nptx2+ -, Nmb+ -, and Crh+ -expressing projection neurons. By combining in situ hybridization of retrogradely labeled neurons with Fos-based assays, we also demonstrate significant functional heterogeneity, including both convergence and segregation of pain- and itch-provoking inputs into molecularly diverse subsets of NK1R- and non-NK1R-expressing projection neurons.

Mammalian target of rapamycin signaling pathway is involved in synaptic plasticity of the spinal dorsal horn and neuropathic pain in rats by regulating autophagy.

Unveiling the etiology and the underlying mechanism of neuropathic pain, a poorly treated disease, is essential for the development of effective therapies. This study aimed to explore the role of mammalian target of rapamycin (mTOR) signaling in autophagy-mediated neuropathic pain. We established a spared nerve injury (SNI) model in adult male SD rats by ligating the common peroneal nerve and tibial, with the distal end cutoff. The paw withdrawal threshold (PWT) and C/A-fiber evoked field potentials were determined by electrophysiologic tests at day 0 (before operation), day 7 and day 14 postoperation, and SNI significantly increased field potentials (P < 0.05). Immunohistochemistry and western blots using spinal cord tissues showed that the expressions of GluR1, GluR2, Beclin-1, p62, mTOR and 4EBP1 were significantly increased after SNI (all P < 0.05), whereas the expressions of LC3 and LAMP2 were significantly decreased after SNI (all P < 0.05). Rapamycin efficiently counteracted the effect of SNI and restored the phenotypes to the level comparable to the sham control. In conclusion, rapamycin inhibits C/A-fiber-mediated long-term potentiation in the SNI rat model of neuropathic pain, which might be mediated by activation of autophagy signaling and downregulation of GluRs expression.

ZEB1 Induces Ddr1 Promoter Hypermethylation and Contributes to the Chronic Pain in Spinal Cord in Rats Following Oxaliplatin Treatment.

Application of chemotherapeutic oxaliplatin represses gene transcription through induction of DNA methylation, which may contribute to oxaliplatin-induced chronic pain. Here, Ddr1, which showed an increased methylation in the promoter, was screened from the SRA methylation database (PRJNA587622) after oxaliplatin treatment. qPCR and MeDIP assays verified that oxaliplatin treatment increased the methylation in Ddr1 promoter region and decreased the expression of DDR1 in the neurons of spinal dorsal horn. In addition, overexpression of DDR1 by intraspinal injection of AAV-hSyn-Ddr1 significantly alleviated the mechanical allodynia induced by oxaliplatin. Furthermore, we found that oxaliplatin treatment increased the expression of DNMT3b and ZEB1 in dorsal horn neurons, and promoted the interaction between DNMT3b and ZEB1. Intrathecal injection of ZEB1 siRNA inhibited the enhanced recruitment of DNMT3b and the hypermethylation in Ddr1 promoter induced by oxaliplatin. Finally, ZEB1 siRNA rescued the DDR1 downregulation and mechanical allodynia induced by oxaliplatin. In conclusion, these results suggested that the ZEB1 recruited DNMT3b to the Ddr1 promoter, which induced the DDR1 downregulation and contributed to the oxaliplatin-induced chronic pain.

Role of innate immunity in chemotherapy-induced peripheral neuropathy.

It has become increasingly clear that the innate immune system plays an essential role in the generation of many types of neuropathic pain including that which accompanies cancer treatment. In this article we review current findings of the role of the innate immune system in contributing to cancer treatment pain at the distal endings of peripheral nerve, in the nerve trunk, in the dorsal root ganglion and in the spinal dorsal horn.

Involvement of nuclear factor κB and descending pain pathways in the anti-hyperalgesic effect of ß-citronellol, a food ingredient, complexed in ß-cyclodextrin in a model of complex regional pain syndrome - Type 1.

Complex regional pain syndrome type 1 (CRPS-1) is a painful syndrome without effective treatment. In order to explore possible new treatments, we used an animal model of CRPS-1 to examine the effects of ß-Citronellol (ßCT), a monoterpene found in a variety of plants that has been shown to have analgesic effects. We aimed to assess its effects alone, and complexed with ß-cyclodextrin (ßCD), which has been previously used to enhance the effects of a number of medicines. The ßCT-ßCD was characterized physiochemically using high performance liquid chromatography (HPLC) and differential scanning calorimetry (DSC) and shown to have 80% efficiency. In the animal model, Swiss mice were treated with ßCT, ßCT-ßCD, vehicle, pregabalin or sham and evaluated for hyperalgesia and motor coordination. Inflammatory mediators were measured by Western blot or ELISA and the descending pain pathway by immunofluorescence. ßCT was shown to have an anti-hyperalgesic effect (without affecting motor coordination) that reduced inflammatory mediators and activated the descending pain pathway, and these effects were increased with complexation in ßCD. Our results showed ßCT-ßCD to be a promising treatment for CRPS-1.

Calcium-independent phospholipase A2 inhibitor produces an analgesic effect in a rat model of neuropathic pain by reducing central sensitization in the dorsal horn.

OBJECTIVE: Phospholipase A2 (PLA2) plays an important role in regulating the production of arachidonic acid and various eicosanoids. The aim of our study was to investigate the analgesic mechanisms of calcium-dependent cytosolic phospholipase A2 and calcium-independent PLA2 (iPLA2) inhibitors in the spinal cord in a rat model of neuropathic pain. METHODS: Lumbar 5 spinal nerve ligation was performed in male Sprague-Dawley rats to develop a peripheral neuropathic pain model. Paw withdrawal thresholds in response to von Frey filaments, brush, pressure, and pinch were measured. Lumbar wide dynamic range neuronal firing rates and iPLA2 subtype expression were measured by in vivo extracellular recording and double immunofluorescence staining, respectively. RESULTS: In our rat models, oral administration of prednisolone, a non-selective PLA2 inhibitor, and intrathecal injection of bromoenolactone, a iPLA2 inhibitor, significantly increased the ipsilateral hindpaw withdrawal thresholds in response to von Frey filament stimulation, but intrathecal injection of arachidonyl trifluoromethyl ketone, a selective cytosolic PLA2 inhibitor, did not show significant changes. In spinal dorsal horn neurons, bromoenolactone reduced neuronal firing rates in response to withdrawal stimulation and spontaneous firing rates in the ipsilateral side of the spinal dorsal horn. In addition, the expression of iPLA2 was co-localized with astrocytes and neurons on the ipsilateral side of the dorsal horn in rats that underwent spinal nerve ligation. DISCUSSION: These data suggest that selective iPLA2 inhibitor produce analgesia in neuropathic rats by reducing central sensitization in the dorsal horn.

Blockade of Erythropoietin-Producing Human Hepatocellular Carcinoma Receptor B1 in Spinal Dorsal Horn Alleviates Visceral Pain in Rats.

Objective: This experiment was designed to determine whether erythropoietin-producing human hepatocellular carcinoma (Eph) receptors were involved in the development of visceral pain. Methods: Adult male Sprague-Dawley rats were randomly divided into three groups receiving different treatments (n = 16 per group): intracolonic vehicle (control group), intracolonic 2, 4, 6-trinitrobenzene sulfonic acid (TNBS) (TNBS group), and intracolonic TNBS and intrathecal EphB1 receptor blocking reagent (TNBS + EphB2-Fc group). Visceral hyperalgesia was evaluated with quantification of visceral pain threshold induced by colorectal distention. The spinal expressions of EphB1 and ephrinB2 and levels of their phosphorylated forms (p-EphB1 and p-ephrinB2) were assessed by Western blotting and immunohistochemistry. Results: The TNBS-treated rats developed significant visceral hyperalgesia. The spinal expressions of EphB1, p-EphB1, ephrinB2, and p-ephrinB2 were significantly increased in the TNBS group compared with the control group, but visceral hyperalgesia and elevation of spinal EphB1 and p-EphB1 expressions were evidently alleviated by intrathecal administration of EphB2-Fc in the TNBS + EphB2-Fc group. The number of EphB1- and p-EphB1-immunopositive cells, the average optical (AO) value of EphB1, and its phosphorylated form in the spinal dorsal horn were significantly increased in the TNBS group than in the control group, but they were obviously reduced by intrathecal administration of EphB2-Fc. There were no significant differences in the number of ephrinB2- and p-ephrinB2-immunopositive cells and the AO value of ephrinB2 and its phosphorylated form between the TNBS and TNBS + EphB2-Fc groups. Conclusion: EphB1 receptors in the spinal dorsal horn play a pivotal role in the development of visceral pain and may be considered as a potential target for the treatment of visceral pain.

Methylmercury induces hyperalgesia/allodynia through spinal cord dorsal horn neuronal activation and subsequent somatosensory cortical circuit formation in rats.

Methylmercury (MeHg) is known to cause serious neurological deficits in humans. In this study, we investigated the occurrence of MeHg-mediated neuropathic pain and identified the underlying pathophysiological mechanism in a rat model of MeHg exposure. Rats were exposed to MeHg (20 ppm in drinking water) for 3 weeks. Neurological damage was observed in the primary afferent neuronal system, including the dorsal root nerve and the dorsal column of the spinal cord. The MeHg-exposed rats showed hyperalgesia/allodynia, compared to controls, as evidenced by a significant decrease in the threshold of mechanical pain evaluated using an algometer with calibrated forceps. Immunohistochemistry revealed the accumulation of activated microglia in the dorsal root nerve, dorsal column, and dorsal horn of the spinal cord. Western blot analyses of the dorsal part of the spinal cord demonstrated an increase in inflammotoxic and inflammatory cytokines and a neuronal activation related protein, phospho-CRE bunding protein (CREB). The results suggest that dorsal horn neuronal activation was mediated by inflammatory factors excreted by accumulated microglia. Furthermore, analyses of the cerebral cortex demonstrated increased expression of phospho-CREB and thrombospondin-1, which is known to be an important factor for excitatory synapse formation, specifically in the somatosensory cortical area. In addition, the expression of pre- and post-synaptic markers was increased in this cortex area. These results suggested that the new cortical circuit was wired specifically in the somatosensory cortex. In conclusion, MeHg-mediated dorsal horn neuronal activation with inflammatory microglia might induce somatosensory cortical rewiring, leading to hyperalgesia/allodynia.

The Roles of Superoxide on At-Level Spinal Cord Injury Pain in Rats.

BACKGROUND: In the present study, we examined superoxide-mediated excitatory nociceptive transmission on at-level neuropathic pain following spinal thoracic 10 contusion injury (SCI) in male Sprague Dawley rats. METHODS: Mechanical sensitivity at body trunk, neuronal firing activity, and expression of superoxide marker/ionotropic glutamate receptors (iGluRs)/CamKII were measured in the T7/8 dorsal horn, respectively. RESULTS: Topical treatment of superoxide donor t-BOOH (0.4 mg/kg) increased neuronal firing rates and pCamKII expression in the naïve group, whereas superoxide scavenger Tempol (1 mg/kg) and non-specific ROS scavenger PBN (3 mg/kg) decreased firing rates in the SCI group (* p < 0.05). SCI showed increases of iGluRs-mediated neuronal firing rates and pCamKII expression (* p < 0.05); however, t-BOOH treatment did not show significant changes in the naïve group. The mechanical sensitivity at the body trunk in the SCI group (6.2 ± 0.5) was attenuated by CamKII inhibitor KN-93 (50 µg, 3.9 ± 0.4) or Tempol (1 mg, 4 ± 0.4) treatment (* p < 0.05). In addition, the level of superoxide marker Dhet showed significant increase in SCI rats compared to the sham group (11.7 ± 1.7 vs. 6.6 ± 1.5, * p < 0.05). CONCLUSIONS: Superoxide and the pCamKII pathway contribute to chronic at-level neuropathic pain without involvement of iGluRs following SCI.

Perineural high-mobility group box 1 induces mechanical hypersensitivity through activation of spinal microglia: Involvement of glutamate-NMDA receptor dependent mechanism in spinal dorsal horn.

High mobility box 1 (HMGB1), a damage-associated molecular pattern, has crucial roles in induction of neuropathic pain. Upregulation of HMGB1 around the injured sciatic nerve contributes to mechanical hypersensitivity following partial sciatic nerve ligation (PSNL) of mice. However, central mechanisms mediating perineural HMGB1-induced nociceptive hypersensitivity, especially within the spinal dorsal horn, have not been determined. The current study shows that perineural treatment of naïve mice with recombinant HMGB1, which mimics increased HMGB1 around the injured sciatic nerve of PSNL mice, significantly induced activation of microglia, but not astrocytes, in the spinal dorsal horn. Intraperitoneal injection of minocycline, a microglial inhibitor, ameliorated perineural rHMGB1-induced mechanical hypersensitivity. In addition, blockade of spinal N-methyl-D-aspartate (NMDA) receptors significantly prevented perineural rHMGB1-induced mechanical hypersensitivity and microglial activation. In contrast, non-NMDA receptors, neurokinin 1 receptor, colony-stimulating factor 1 receptor and P2Y12 receptor were not involved in perineural rHMGB1-induced mechanical hypersensitivity. Furthermore, repeated perineural treatment with an anti-HMGB1 antibody blocked activation of spinal microglia in PSNL mice. Collectively, the current findings demonstrate that increased HMGB1 around injured sciatic nerve might induce nociceptive hypersensitivity through activation of spinal microglia. Thus, HMGB1-dependent mechanisms between the injured sciatic nerve and spinal dorsal horn could be crucial in induction of neuropathic pain.

Inhibition of angiotensin converting enzyme induces mechanical allodynia through increasing substance P expression in mice.

Although emerging evidence shows that angiotensin converting enzyme (ACE) is associated with pain, it is not clear whether inhibition of ACE could affect to nociceptive transmission and which mediators are involved in this process. Here we investigated whether administration of the ACE inhibitors, captopril and enalapril increases the expression of substance P (SP) and whether this increase contributes to the induction of mechanical allodynia in mice. ACE was expressed in the lumbar dorsal root ganglion (DRG) and the superficial dorsal horn (SDH) region of the spinal cord in mice. Either intraperitoneal or intrathecal administration of the ACE inhibitors, captopril and enalapril for 10 days significantly increased the paw withdrawal frequency to innocuous mechanical stimuli and the levels of SP in both the lumbar DRG and the SDH region of the spinal cord dorsal horn. In addition, intraperitoneal administration of the SP receptor (neurokinin-1 receptor) antagonist, L-733,060 suppressed mechanical allodynia that was induced by pretreatment of captopril and enalapril. Intraplantar administration of SP for 3 days induces mechanical allodynia, and this effect was reduced by exogenous ACE administration. These findings demonstrate that inhibition of ACE increases the levels of SP in both the lumbar DRG and spinal cord dorsal horn, ultimately contributing to the induction of mechanical allodynia in mice.

Electroacupuncture may alleviate neuropathic pain via suppressing P2X7R expression.

Neuropathic pain is a severe problem that is difficult to treat clinically. Reducing abnormal remodeling of dendritic spines/synapses and increasing the anti-inflammatory effects in the spinal cord dorsal horn are potential methods to treat this disease. Previous studies have reported that electroacupuncture (EA) could increase the pain threshold after peripheral nerve injury. However, the underlying mechanism is unclear. P2X7 receptors (P2X7R) mediate the activation of microglia and participate in the occurrence and development of neuropathic pain. We hypothesized that the effects of EA on relieving pain may be related to the downregulation of the P2X7R. Spinal nerve ligation (SNL) rats were used as a model in this experiment, and 2'(3')-O-(4-benzoyl)benzoyl ATP (BzATP) was used as a P2X7R agonist. We found that EA treatment decreased dendritic spine density, inhibited synaptic reconstruction and reduced inflammatory response, which is consistent with the decrease in P2X7R expression as well as the improved neurobehavioral performance. In contrast to the beneficial effects of EA, BzATP enhanced abnormal remodeling of dendritic spines/synapses and inflammation. Furthermore, the EA-mediated positive effects were reversed by BzATP, which is consistent with the increased P2X7R expression. These findings indicated that EA improves neuropathic pain by reducing abnormal dendritic spine/synaptic reconstruction and inflammation via suppressing P2X7R expression.