Chronic Constriction Injury Of Sciatic Nerve Research Articles

Role of receptor-interacting protein 1/receptor-interacting protein 3 in inflammation and necrosis following chronic constriction injury of the sciatic nerve.

Nerve damage often leads to nervous system dysfunction and neuropathic pain. The serine-threonine kinases receptor-interacting protein 1 (RIP1) and 3 (RIP3) are associated with inflammation and cell necrosis. This study aimed to explore the role of RIP1 and RIP3 in sciatic nerve chronic constriction injury (CCI) in mice. On a total of thirty mice, sciatic nerve CCI was performed. The paw withdrawal threshold was measured using Von Frey filaments. The mRNA expression and protein levels of inflammatory factors RIP1 and RIP3 in the dorsal root ganglion (DRG), spinal cord (SC) and hippocampus (HIP) were also determined. We found that paw withdrawal threshold was significantly reduced from the second day after the operation, and the levels of tumour necrosis factor-α and interferon-γ in DRG, SC and HIP were significantly increased on the eighth and 14th days in CCI mice. Furthermore, the downstream signalling molecules of RIP1 and RIP3, GTPase dynamin-related protein-1, NLR family pyrin domain containing-3 (NLRP3) and nuclear factor κB-p65 were upregulated. Increased protein levels of programmed cell death protein 1, which indicate cell death of peripheral and central nervous tissue, were induced by CCI of the sciatic nerve. Overall, this study showed that RIP1 and RIP3 were highly expressed in DRG, SC and HIP of the sciatic nerve in CCI mice and may be involved in chronic neuroinflammation and neuronecrosis.

Glial-cytokine-neuronal Adaptations in the Ventral Hippocampus of Rats with Affective Behavioral Changes Following Peripheral Nerve Injury

Nerve damage leads to the development of disabling neuropathic pain in susceptible individuals, where patients present with pain as well as co-morbid behavioral changes, such as anhedonia, decreased motivation and depression. In this study we evaluated whether radial maze behavioral disruptions and glia-cytokine-neuronal adaptations in the hippocampus occurred in individual rats after nerve injury. Exploration behavior and spatial memory were quantified using a radial maze task, while mechanical allodynia was assessed using von Frey testing. Sciatic nerve chronic constriction injury (CCI) reduced withdrawal thresholds in all rats, while pellet-seeking behaviors were altered in some but not all rats. One group, termed ‘No effect’, had no behavioral changes compared to sham rats. Another group, termed ‘Acute effect’, had a temporary alteration to their exploration pattern, displaying more risk-assessment behavior in the early phase post-injury. In a third group, termed ‘Lasting effect’, exploratory behaviors were remarkably different for the entire post-injury period, showing a withdrawal from pellet-seeking. The withdrawal from pellet-seeking was found to be concomitant with distinct glial-cytokine-neuronal adaptations within the contralateral ventral hippocampus, including; increased expression of IL-1β and MCP-1; astrocyte atrophy and decreased area in the dentate gyrus; reactive microglia and increased FosB/ΔFosB expression in the cornu ammonis subfield. Therefore, glial-cytokine-neuronal adaptations in the ventral hippocampus may mediate individual differences in radial maze behavior following CCI. Our data suggest that individual neuroimmune signatures play a significant role in divergent behavioral trajectories following nerve injury, toward functional recovery and coping, or the emergence of ongoing affective state disturbances.

Dexmedetomidine relieves neuropathic pain by inhibiting hyperpolarization-activated cyclic nucleotide-gated currents in dorsal root ganglia neurons.

This study was designed to examine the effect and mechanism of dexmedetomidine (Dex) on neuropathic pain (NP). The NP model was established by performing chronic sciatic nerve constriction injury (CCI). Seven days after CCI surgery, the rats were injected intraperitoneally with Dex, ZD7288 (an HCN channel inhibitor), and saline, respectively. The paw withdrawal threshold to mechanical stimulation and the thermal withdrawal latency tests were performed. After administration, the L4, L5 dorsal root ganglia (DRG) neurons of rats were isolated. In addition, hyperpolarization-activated cyclic nucleotide-gated (HCN) channels subtype plasmids were transfected into human embryonic kidney (HEK)293 cells. Whole-cell clamp recordings were used to examine the properties of HCN currents (Ih) expressed in HEK293 cells and DRG neurons. After surgery, the paw withdrawal threshold to mechanical stimulation and thermal withdrawal latency were reduced, the HCN currents (Ih) amplitude of DRG neurons was increased, and the semiactivated voltage (V1/2) value was decreased in CCI rats (P<0.05). CCI rats treated with Dex or ZD7288 had reduced mechanical and thermal hyperalgesia. The Ih amplitude was lower and the V1/2 value was increased in DRG neurons in CCI rats treated with Dex or ZD7288 (P<0.05). In addition, Dex inhibited HCN1 and HCN2 currents in HEK293 cells; caused a decrease in maximal currents, an increase in the inhibition rate of Ih, and a negative shift in V1/2 (P<0.05). Taken together, our finding suggested that Dex alleviates NP and the effect is likely because of the inhibition of HCN currents.

Procyanidins attenuate neuropathic pain by suppressing matrix metalloproteinase-9/2

BackgroundManagement of neuropathic pain is a real clinical challenge. Despite intense investigation, the mechanisms of neuropathic pain remain substantially unidentified. Matrix metalloproteinase (MMP)-9 and MMP-2 have been reported to contribute to the development and maintenance of neuropathic pain. Therefore, inhibition of MMP-9/2 may provide a novel therapeutic approach for the treatment of neuropathic pain. In this study, we aim to investigate the effect of procyanidins (PC), clinically used health product, on MMP-9/2 in neuropathic pain.MethodsThe nociception was assessed by measuring the incidence of foot withdrawal in response to mechanical indentation in mice. Cell signaling was assayed using gelatin zymography, western blotting, and immunohistochemistry. The BV2 cells were cultured to investigate the effects of PC on microglia.ResultsBoth in vitro and in vivo administration of PC significantly suppresses the activity of MMP-9/2. Oral administration of PC relieves neuropathic pain behaviors induced by chronic constriction sciatic nerve injury (CCI) in mice. Additionally, PC blocks the maturation of interleukin-1β, which is a critical substrate of MMPs, and markedly suppresses CCI-induced MAPK phosphorylation and neuronal and microglia activation, including the reduced phosphorylation of protein kinase C γ and NMDAR1. Furthermore, PC decreases the phosphorylation of p38 mitogen-activated protein kinase and inhibits the translocation of nuclear factor-κB (NF-κB) in microglia.ConclusionsPC is an effective and safe approach to alleviate neuropathic pain via a powerful inhibition on the activation of MMP-9/2.

Chronic constriction injury-induced microRNA-146a-5p alleviates neuropathic pain through suppression of IRAK1/TRAF6 signaling pathway

BackgroundmicroRNA-146a-5p (miRNA-146a-5p) is a key molecule in the negative regulation pathway of TLRs and IL-1 receptor (TIR) signaling. Our recent study demonstrated that MyD88-dependent signaling pathway of TIR in the dorsal root ganglion (DRG) and spinal dorsal horn (SDH) plays a role in peripheral nerve injury-induced neuropathic pain. However, it was not clear whether and how miRNA-146a-5p regulates the TIR pathway of DRG and SDH in the development of neuropathic pain.MethodsThe sciatic nerve chronic constriction injury (CCI) model of rat was used to induce chronic neuropathic pain. The levels and cellular distribution of miRNA-146a-5p were detected with quantitative real-time PCR (qPCR) and fluorescent in situ hybridization (FISH). The RNA level, protein level, and cellular distribution of IRAK1 and TRAF6 that is targeted by miRNA-146a-5p were detected with qPCR, western blot, and immunofluorescent. The pain-related behavioral effect of miRNA-146a-5p was accessed after intrathecal administration. Mechanical stimuli and radiant heat were used to evaluate mechanical allodynia and thermal hyperalgesia.ResultsWe found that the level of miRNA-146a-5p significantly increased in L4-L6 DRGs and SDH after CCI surgery; meanwhile, the protein level of IRAK1 and TRAF6 in DRGs was significantly increased after CCI. Intrathecal injection of miR146a-5p agomir or miRNA-146a-5p antagomir regulates miRNA-146a-5p level of L4-L6 DRGs and SDH. We found that intrathecal injection of miR146a-5p agomir can alleviate mechanical and thermal hyperalgesia in CCI rats and reverse the upregulation of IRAK1 and TRAF6 of L4-L6 DRGs and SDH induced by CCI. We furthermore found that intrathecal injection of miRNA-146a-5p antagomir can exacerbate the mechanical and thermal pain-related behavior of CCI rats and meanwhile increase IRAK1 and TRAF6 of L4-L6 DRGs and SDH expression even further.ConclusionsmiRNA-146a-5p of DRG and SDH can modulate the development of CCI-induced neuropathic pain through inhibition of IRAK1 and TRAF6 in the TIR signaling pathway. Hence, miRNA-146a-5p may serve as a potential therapeutic target for neuropathic pain.

Morphological and Physiological Plasticity of Spinal Lamina II GABA Neurons Is Induced by Sciatic Nerve Chronic Constriction Injury in Mice.

Mice with transgenic insertion of code for enhanced green fluorescent protein (EGFP) at the locus for glutamic acid decarboxylase 67 (GAD67), one of two key enzymes for the synthesis of γ-aminobutyric acid (GABA) were used to test whether the morphological properties of these neurons show plasticity with nerve injury. Physiological properties and the delivery of intracellular label to EGFP-expressing lamina II neurons was done using whole-cell patch-clamp in spinal cord slices from sham and chronic constriction injury (CCI) mice. As well, whole cell recordings were made of non-EGFP labeled cells to ascertain changes in overall inhibitory signaling following CCI. The EGFP labeled neurons in both sham and CCI mice exhibited islet, central and vertical cell morphological profiles but no radial cell profiles were observed. The length of cell dendrites was found to be significantly shorter in CCI mice for all cell profile types. The longest neurites averaged 155.96 ± 18.29 μm in CCI mice compared to 334.93 ± 29.48 μm in sham control mice. No change was observed in either passive or evoked membrane properties of EGFP-expressing neurons in CCI versus sham mice. Meanwhile, the frequency of miniature inhibitory post-synaptic currents of non-EGFP expressing spinal lamina II neurons was significantly reduced. These results suggest that reduced inhibitory output from GABA neurons occurs with nerve injury in part due to altered cell morphology.

N-methyl D-aspartate receptor subtype 2B antagonist, Ro 25-6981, attenuates neuropathic pain by inhibiting postsynaptic density 95 expression

Postsynaptic density-95 (PSD-95) is a synaptic scaffolding protein that plays a crucial role in the development of neuropathic pain. However, the underlying mechanism remains unclear. To address the role of PSD-95 in N-methyl-D-aspartate receptor subtype 2B (NR2B) -mediated chronic pain, we investigated the relationship between PSD-95 activation and NR2B function in the spinal cord, by using a rat model of sciatic nerve chronic constriction injury (CCI). We demonstrate that the expression levels of total PSD-95 and cAMP response element binding protein (CREB), as well as phosphorylated NR2B, PSD-95, and CREB, in the spinal dorsal horn, and the interaction of NR2B with PSD-95 were increased in the CCI animals. Intrathecal injection of the selective NR2B antagonist Ro 25-6981 increased paw withdrawal latency, in a thermal pain assessment test. Moreover, repeated treatment with Ro 25-6981 markedly attenuated the thermal hypersensitivity, and inhibited the CCI-induced upregulation of PSD-95 in the spinal dorsal horn. Furthermore, intrathecal injection of the PSD-95 inhibitor strikingly reversed the thermal and mechanical hyperalgesia. Our results suggest that blocking of NR2B signaling in the spinal cord could be used as a therapeutic candidate for treating neuropathic pain.

MiR-183-5p Alleviates Chronic Constriction Injury-Induced Neuropathic Pain Through Inhibition of TREK-1.

MicroRNAs have been implicated in nerve injury and neuropathic pain. In the previous study we had shown that miR-96 can attenuate neuropathic pain through inhibition of Nav1.3. In this study, we investigated the role of miR-183, a same cluster member of microRNA with miR-96, in neuropathic pain and its potential mechanisms. We found that the expression level of miR-183-5p in dorsal root ganglion was decreased with the development of neuropathic pain induced by chronic constriction sciatic nerve injury (CCI). By contrast, the TREK-1, a K+ channel, was increased. Further investigation identified that intrathecal injection of miR-183-5p mimic efficiently ameliorated neuropathic pain and inhibited the expression of TREK-1, a predicted target gene of miR-183-5p. Luciferase assays confirmed the binding of miR-183-5p and TREK-1. In addition, over-expression of TREK-1 blocked the roles of miR-183-5p in neuropathic pain. Our findings suggested that miR-183-5P participated in the regulation of CCI-induced neuropathic pain through inhibiting the expression of TREK-1.

Gardenoside suppresses the pain in rats model of chronic constriction injury by regulating the P2X3 and P2X7 receptors

Objectives: Here, using rat model, we investigated the roles of gardenoside in the chronic constriction injury (CCI) of the ischiadic nerve.Methods: Bennett and Xie’s unilateral sciatic nerve CCI model was used in this study. A total of 60 rats were divided into control group (CN), sham group (Sham), CCI group, and gardenoside administrated CCI group. An aliquot of 5 mL gardenoside solution was administrated through gavage once per day for 14 d. Mechanical withdrawal threshold (MWT) and the thermal withdrawal latency (TWL) were detected. The levels of inducible nitric oxide synthase (iNOS), interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α) in spinal fluid were detected by ELISA. By using real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blot, we analyzed the expression of P2X purinoceptor 3 and 7 (P2X3 and P2X7 receptors) in different groups. The expression of p-ERK/ERK and p-p38/p38 were also detected by western blot.Results: We found out that gardenoside could significantly improve the sciatica by partially restore the decrease of MWT and TWL in CCI rats. The levels of iNOS, IL-1β, and TNF-α were higher in CCI group (p < .05). The expressions of P2X3 and P2X7 were significantly increased in the CCI rats compared to control rats (p < .05). The levels of p-ERK/ERK and p-p38/p38 were also obviously increased in CCI group (p < .05). After treated with the gardenoside, these increases were decreased.Conclusions: These results indicated that gardenoside may be able to relief CCI-induced neuropathic pain by regulating the P2X3 and the P2X7 expression on the ischiadic nerve.

Gardenoside combined with ozone inhibits the expression of P2X3 and P2X7 purine receptors in rats with sciatic nerve injury.

Neuropathic pain is a severe health problem for which there is a lack of effective therapy. Ozone and Gardenia fruits have been used separately in pain relief for many years; however, their underlying mechanisms remain unclear. To investigate the pain‑relieving effects of combined ozone and Gardenia, a chronic constriction sciatic nerve injury (CCI) rat model was constructed and treated with ozone and gardenoside (Ozo&Gar), which is a compound found in Gardenia fruits. A total of 70 rats were randomly divided into five groups: Control (Ctrl), Ctrl+Ozo&Gar, Sham, CCI, and CCI+Ozo&Gar. The rats in the Ctrl+Ozo&Gar and CCI+Ozo&Gar groups were administered an intravenous injection of 30 µg/ml ozone and 300µmol/l gardenoside. The rats in the Ctrl, Sham and CCI groups were administered the same volume of saline. Pain behavior, mechanical hyperalgesia, thermal hyperalgesia, and the protein expression levels of P2X3 and P2X7 purine receptors in L4‑L5 dorsal root ganglion (DRG) were determined 15 days post‑surgery. The results demonstrated that treatment with a combination of ozone and gardenoside increased mechanical withdrawal threshold and thermal withdrawal latency, thus confirming their pain‑relieving effects. In addition, a significant increase in the mRNA and protein expression levels of P2X3 and P2X7 was detected in the DRG of rats in the CCI group compared with in the control groups; however, following treatment with a combination of ozone and gardenoside, the mRNA and protein expression levels of P2X3 and P2X7 receptors were significantly reduced compared with in the CCI group. These results indicated that the mechanism underlying the pain‑relieving effects of ozone and gardenoside may be mediated by inhibition of P2X3 and P2X7 purine receptors in the DRG. This finding suggested that ozone and gardenoside may be considered potential drug candidates that target P2X3 and P2X7 purine receptors.

Acute- and late-phase matrix metalloproteinase (MMP)-9 activity is comparable in female and male rats after peripheral nerve injury

BackgroundIn the peripheral nerve, pro-inflammatory matrix metalloproteinase (MMP)-9 performs essential functions in the acute response to injury. Whether MMP-9 activity contributes to late-phase injury or whether MMP-9 expression or activity after nerve injury is sexually dimorphic remains unknown.MethodsPatterns of MMP-9 expression, activity and excretion were assessed in a model of painful peripheral neuropathy, sciatic nerve chronic constriction injury (CCI), in female and male rats. Real-time Taqman RT-PCR for MMP-9 and its endogenous inhibitor, tissue inhibitor of metalloproteinase-1 (TIMP-1) of nerve samples over a 2-month time course of CCI was followed by gelatin zymography of crude nerve extracts and purified MMP-9 from the extracts using gelatin Sepharose-beads. MMP excretion was determined using protease activity assay of urine in female and male rats with CCI.ResultsThe initial upsurge in nerve MMP-9 expression at day 1 post-CCI was superseded more than 100-fold at day 28 post-CCI. The high level of MMP-9 expression in late-phase nerve injury was accompanied by the reduction in TIMP-1 level. The absence of MMP-9 in the normal nerve and the presence of multiple MMP-9 species (the proenzyme, mature enzyme, homodimers, and heterodimers) was observed at day 1 and day 28 post-CCI. The MMP-9 proenzyme and mature enzyme species dominated in the early- and late-phase nerve injury, consistent with the high and low level of TIMP-1 expression, respectively. The elevated nerve MMP-9 levels corresponded to the elevated urinary MMP excretion post-CCI. All of these findings were comparable in female and male rodents.ConclusionThe present study offers the first evidence for the excessive, uninhibited proteolytic MMP-9 activity during late-phase painful peripheral neuropathy and suggests that the pattern of MMP-9 expression, activity, and excretion after peripheral nerve injury is universal in both sexes.

A sensitive and selective ELISA methodology quantifies a demyelination marker in experimental and clinical samples

Sciatic nerve chronic constriction injury (CCI) in rodents produces nerve demyelination via proteolysis of myelin basic protein (MBP), the major component of myelin sheath. Proteolysis releases the cryptic MBP epitope, a demyelination marker, which is hidden in the native MBP fold. It has never been established if the proteolytic release of this cryptic MBP autoantigen stimulates the post-injury increase in the respective circulating autoantibodies. To measure these autoantibodies, we developed the ELISA that employed the cryptic 84-104 MBP sequence (MBP84-104) as bait. This allowed us, for the first time, to quantify the circulating anti-MBP84-104 autoantibodies in rat serum post-CCI. The circulating IgM (but not IgG) autoantibodies were detectable as soon as day 7 post-CCI. The IgM autoantibody level continually increased between days 7 and 28 post-injury. Using the rat serum samples, we established that the ELISA intra-assay (precision) and inter-assay (repeatability) variability parameters were 2.87% and 4.58%, respectively. We also demonstrated the ELISA specificity by recording the autoantibodies to the liberated MBP84-104 epitope alone, but not to intact MBP in which the 84-104 region is hidden. Because the 84-104 sequence is conserved among mammals, we tested if the ELISA was applicable to detect demyelination and quantify the respective autoantibodies in humans. Our limited pilot study that involved 16 female multiple sclerosis and fibromyalgia syndrome patients demonstrated that the ELISA was efficient in measuring both the circulating IgG- and IgM-type autoantibodies in patients exhibiting demyelination. We believe that the ELISA measurements of the circulating autoantibodies against the pathogenic MBP84-104 peptide may facilitate the identification of demyelination in both experimental and clinical settings. In clinic, these measurements may assist neurologists to recognize patients with painful neuropathy and demyelinating diseases, and as a result, to personalize their treatment regimens.

The effect of exercise frequency on neuropathic pain and pain-related cellular reactions in the spinal cord and midbrain in a rat sciatic nerve injury model

BackgroundExercise regimens are established methods that can relieve neuropathic pain. However, the relationship between frequency and intensity of exercise and multiple cellular responses of exercise-induced alleviation of neuropathic pain is still unclear. We examined the influence of exercise frequency on neuropathic pain and the intracellular responses in a sciatic nerve chronic constriction injury (CCI) model.Materials and methodsRats were assigned to four groups as follows: CCI and high-frequency exercise (HFE group), CCI and low-frequency exercise (LFE group), CCI and no exercise (No-Ex group), and naive animals (control group). Rats ran on a treadmill, at a speed of 20 m/min, for 30 min, for 5 (HFE) or 3 (LFE) days a week, for a total of 5 weeks. The 50% withdrawal threshold was evaluated for mechanical sensitivity. The activation of glial cells (microglia and astrocytes), expression of brain-derived neurotrophic factor (BDNF) and μ-opioid receptor in the spinal dorsal horn and endogenous opioid in the midbrain were examined using immunohistochemistry. Opioid receptor antagonists (naloxone) were administered using intraperitoneal injection.ResultsThe development of neuropathic pain was related to the activation of glial cells, increased BDNF expression, and downregulation of the μ-opioid receptor in the ipsilateral spinal dorsal horn. In the No-Ex group, neuropathic pain showed the highest level of mechanical hypersensitivity at 2 weeks, which improved slightly until 5 weeks after CCI. In both exercise groups, the alleviation of neuropathic pain was accelerated through the regulation of glial activation, BDNF expression, and the endogenous opioid system. The expression of BDNF and endogenous opioid in relation to exercise-induced alleviation of neuropathic pain differed in the HFE and LFE groups. The effects of exercise-induced alleviation of mechanical hypersensitivity were reversed by the administration of naloxone.ConclusionThe LFE and HFE program reduced neuropathic pain. Our findings indicated that aerobic exercise-induced alleviated neuropathic pain through the regulation of glial cell activation, expression of BDNF in the ipsilateral spinal dorsal horn, and the endogenous opioid system.

Prenatal alcohol exposure is a risk factor for adult neuropathic pain via aberrant neuroimmune function

BackgroundClinical studies show that prenatal alcohol exposure (PAE) results in effects that persist into adulthood. Experimental animal models of moderate PAE demonstrate that young adults with PAE display potentiated sensitivity to light touch, clinically termed allodynia, following sciatic nerve chronic constriction injury (CCI) that coincides with heightened spinal glial, spinal macrophage, and peripheral immune responses. However, basal touch sensitivity and corresponding glial and leukocyte activation are unaltered. Therefore, the current study explored whether the enduring pathological consequences of moderate PAE on sensory processing are unmasked only following secondary neural insult.MethodsIn middle-aged (1 year) Long Evans rats that underwent either prenatal saccharin exposure (control) or moderate PAE, we modified the well-characterized model of sciatic neuropathy, CCI, to study the effects of PAE on neuro-immune responses in adult offspring. Standard CCI manipulation required 4 chromic gut sutures, while a mild version applied a single suture loosely ligated around one sciatic nerve. Spinal glial immunoreactivity was examined using immunohistochemistry. The characterization and functional responses of leukocyte populations were studied using flow cytometry and cell stimulation assays followed by quantification of the proinflammatory cytokines interleukin-1beta (IL-1β) and tumor necrosis factor-alpha (TNF-α). Data were statistically analyzed by ANOVA and unpaired t tests.ResultsThe current report demonstrates that mild CCI generates robust allodynia only in PAE rats, while the pathological effects of PAE following the application of a standard CCI are revealed by enhanced allodynia and elevated spinal glial activation. Additionally, mild CCI increases spinal astrocyte activation but not microglia, suggesting astrocytes play a larger role in PAE-induced susceptibility to aberrant sensory processing. Leukocyte populations from PAE are altered under basal conditions (i.e., prior to secondary insult), as the distribution of leukocyte populations in lymphoid organs and other regions are different from those of controls. Lastly, following in vitro leukocyte stimulation, only PAE augments the immune response to antigen stimulation as assessed by heightened production of TNF-α and IL-1β.ConclusionsThese studies demonstrate PAE may prime spinal astrocytes and peripheral leukocytes that contribute to enduring susceptibility to adult-onset neuropathic pain that is not apparent until a secondary insult later in life.

The histone demethylase JMJD2A regulates the expression of BDNF and mediates neuropathic pain in mice

JMJD2A is a JmjC histone demethylase that catalyzes the demethylation of di- and trimethylated Lys9 and Lys36 in histone H3 (H3K9me2/3 and H3K36me2/3). The role of spinal JMJD2A-dependent histone demethylation in nociception hypersensitivity development remains elusive. Here we reported that the JMJD2A responded to neuropathic pain and participated in the maintenance of neuropathic pain. The mRNA and protein levels of Jmjd2a were significantly increased in the neurons of mouse undergoing neuropathic pain induced by sciatic nerve chronic constrictive injury (CCI) or unilateral spared nerve injury (SNI). Jmjd2a responded to 5-hydroxytryptamine (5-HT) and promoted the expression of the brain-derived neurotrophic factor (Bdnf), which is a protein critically involved in neuropathic pain. JMJD2A bound to the promoter of Bdnf and demethylated H3K9me3 and H3K36me3 at Bdnf promoter to promote the expression of Bdnf. Finally, we showed that JMJD2A promoted the expression of Bdnf during neuropathic pain and neuron-specific knockout of Jmjd2a blocked the hypersensitivity of mice undergoing chronic neuropathic pain induced by CCI and SNI. Taken together, our findings demonstrate that up-regulation of JMJD2A promotes neuropathic pain and it may serve as a promising target for treatment of chronic neuropathic pain.

Adult hippocampal neurogenesis in neuropathic pain and alkyl glycerol ethers treatment

Neuropathic pain manifested by a number of sensory symptoms is often accompanied by disorders of higher nervous activity, such as memory impairment, depression, anxiety, anhedonia, etc. This emphasizes the involvement of supraspinal structures including the hippocampus in neuropathic pain pathogenesis. In the present study, we focused on the impact of chronic neuropathic pain on hippocampal neurogenesis and microglial state. In addition, we test the effect of alkyl glycerol ethers on hippocampal neuronal and microglial plasticity as well as behavioral parameters. Neuropathic pain was induced using the model of sciatic nerve chronic constriction injury. We found an impairment of working memory and locomotor activity in animals with neuropathic pain, which was prevented by alkyl glycerol ethers treatment. Sciatic nerve ligation in mice contributed to the decrease in hippocampal neurogenesis intensity. Alkyl glycerol ethers administration significantly reduced this effect. Neuropathic pain-associated neurogenesis reduction was accompanied by an increased percentage of Iba1-labeled area in the CA1 hippocampal region on the 14th and 28th days after surgery. In addition, we observed a decrease in hippocampal pro-inflammatory microglia marker CD86 immunostaining on day 28 after surgery in alkyl glycerol ethers-treated mice with sciatic nerve ligation. These results are consistent with data on pro- and anti-inflammatory cytokines expression in the hippocampus. Alkyl glycerol ethers administration increased IL-10 and decreased IL-1β hippocampal expression in animals with neuropathic pain. Taken together, these data suggest that neuropathic pain-behavior in rodents is accompanied by changes in microglia polarization, thereby contributing to neurogenesis impairment and cognitive disturbances. Alkyl glycerol ethers prevented M1 microglial activation, contributing to the maintenance of normal neurogenesis levels within the hippocampus and normalizing working memory.

Nortriptyline Enhances Morphine-Conditioned Place Preference in Neuropathic Rats: Role of the Central Noradrenergic System.

Combination drug therapy is commonly used to treat chronic pain conditions such as neuropathic pain, and antidepressant is often used together with opioid analgesics. While rewarding is an intrinsic property of opioid analgesics, it is unknown whether the use of an antidepressant would influence opioid reward, which may contribute to opioid addiction. In this study, we examined whether nortriptyline (a tricyclic antidepressant and a first-line medication for neuropathic pain) would enhance the morphine rewarding property in both naive and chronic constriction sciatic nerve injury (CCI) rats. The rewarding effect of these drugs was assessed using conditioned place preference (CPP). The real-time polymerase chain reaction, western blot, and enzyme-linked immunosorbent assay analysis were used to investigate the function of central noradrenergic system. In naive rats, coadministration of nortriptyline with morphine did not change the acquisition of morphine-induced CPP. However, nortriptyline enhanced the acquisition, delayed the extinction, and augmented the reinstatement of morphine-induced CPP in CCI rats. In CCI rats treated with both nortriptyline and morphine, the expression of α2A-adrenergic receptors, norepinephrine transporter, and tyrosine hydroxylase was markedly decreased in the locus coeruleus, whereas the norepinephrine concentration in the nucleus accumbens was remarkably increased. These results demonstrate that nortriptyline enhanced morphine reward when both drugs were used to treat neuropathic pain in rats and that this behavioral phenotype is likely to be mediated by upregulation of the central noradrenergic system. These findings may have implications in opioid therapy commonly used for chronic pain management.

KLF15 regulates dopamine D2 receptor and participates in mouse models of neuropathic pain

Neuropathic pain is caused by damage to the nervous system, resulting in aberrant pain. The mechanism underlying neuropathic pain remains largely unknown. Krüppel-like factor 15 (KLF15) is a member of the Krüppel-like factor family of transcriptional factors. Here in this study, we aimed to investigate the potential role of the transcriptional factor KLF15 in neuropathic pain. The mRNA and protein levels of Klf15 were significantly increased in the neurons of mouse undergoing neuropathic pain induced by sciatic nerve chronic constrictive injury (CCI) or unilateral spared nerve injury (SNI). In neurons, the upregulation of Klf15 was triggered by the inflammatory factor tumor necrosis factor alpha (TNF-α). As a transcriptional factor, KLF15 promoted the expression of dopamine D2 receptor (Drd2), which is a receptor essentially involved in neuropathic pain. KLF15 bound to the promoter of Drd2 directly and promoted the promoter activity of Drd2. Finally, we showed that knockout of Klf15 repressed the sensitivity in neuropathic pain induced by CCI or SNI. In conclusion, KLF15 is induced in neuropathic pain via a TNF-α-dependent manner and contributes to neuropathic pain partially through promoting the expression of dopamine D2 receptor.

Diosmin reduces chronic constriction injury-induced neuropathic pain in mice

Injury or dysfunction of somatosensory system induces a complex syndrome called neuropathic pain, which still needs adequate pharmacological control. The current pharmacological treatments were in part developed from natural compounds. Flavonoids are natural polyphenolic molecules presenting varied biological activities and low toxicity. The flavonoid diosmin is a safe compound with good tolerability and low toxicity. This study evaluated the antinociceptive effect of diosmin in the sciatic nerve chronic constriction injury (CCI)-induced neuropathic pain model. Male Swiss mice were submitted to CCI and 7 days after, diosmin at 1 or 10 mg/kg was administrated intraperitoneally. Mechanical (electronic analgesimeter) and thermal (hot plate) hyperalgesia were evaluated 1–24 h after treatment. The role of the NO/cGMP/PKG/KATP channel signaling pathway in the analgesic effect of diosmin was evaluated using the pretreatment with L-NAME (an inhibitor of NOS), ODQ (an inhibitor of soluble guanylate cyclase), KT5823 (an inhibitor of PKG), or glibenclamide (an ATP-sensitive K+ channels blocker). Single treatment with diosmin inhibited in a dose-dependent manner CCI-induced mechanical and thermal hyperalgesia by activating the NO/cGMP/PKG/KATP channel signaling pathway and inhibiting spinal cord cytokine (Il-1β and Il-33/St2) and glial cells activation (microglia - Iba-1, oligodendrocytes – Olig2) mRNA expression markers. Daily treatment during 7 days with diosmin inhibited CCI-induced mechanical and thermal hyperalgesia by inhibiting spinal cord cytokine (Il-1β, Tnfα, and Il-33/St2) and glial cells activation (astrocytes – Gfap, Iba-1, and Olig2) markers mRNA expression. In conclusion, diosmin inhibits neuropathic spinal cord nociceptive mechanisms suggesting this flavonoid as a potential therapeutic molecule to reduce nerve lesion-induced neuropathic pain.

Resolving the contributions of anaesthesia, surgery, and nerve injury on brain derived neurotrophic factor expression in the medial prefrontal cortex of male rats in the CCI model of neuropathic pain.

The medial prefrontal cortex (mPFC) is critical for selecting and shaping complex behavioral responses. In rodent models of neuropathic pain there is evidence for both structural and functional changes in the mPFC. Brain derived neurotrophic factor (BDNF) plays a critical role in the normal functioning of the mPFC. It has been suggested that the disruption of complex behaviors and mood seen in some neuropathic pain patients is mediated in part by alterations of BDNF in this cortical region. In Sprague-Dawley rats, mPFC levels of BDNF and TrkB mRNA and protein, were quantified and compared to controls (n = 24) 6 days after either: (a) halothane (1.5%) anaesthesia (n = 12), (b) sham surgery under halothane (n = 12), (c) sciatic nerve chronic constriction injury under halothane (n = 48). The social behaviors of the rats were quantified daily during the experimental period. Halothane anaesthesia increased BDNF and TrkB mRNA bilaterally. These increases were reversed in rats that underwent sham surgical and nerve injury procedures. Further, halothane anaesthesia, surgical procedures, and nerve injury each decreased BDNF protein levels. These results reveal a marked and distinct BDNF expression profile in the mPFC of rats that have undergone each stage of the procedure to produce neuropathic pain by chronic constriction injury of the sciatic nerve. The highly sensitive nature of neurotrophic signalling to general anaesthesia in the mature neuronal circuit of the adult rat brain highlights the importance of careful evaluation and interpretation of data evaluating the effects of experimental procedures on neural substrates.