Dopamine receptor D2, but not D1, mediates the reward circuit from the ventral tegmental area to the central amygdala, which is involved in pain relief.

Pain involves both sensory and affective dimensions. The amygdala is a key player in linking nociceptive stimuli to negative emotional behaviors or affective states. Relief of pain is rewarding and activates brain reward circuits. Whether the reward circuit from the ventral tegmental area (VTA) to the central amygdala (CeA) is involved in pain relief remains unexplored. Using a model of experimental postsurgical pain, we found that pain relief elicited conditioned place preference (CPP), activated CeA-projecting dopaminergic cells in the VTA, and decreased dopaminergic D2 receptor expression in the CeA. Activation of the VTA-CeA neural pathway using optogenetic approaches relieved incisional pain. Administration of a D2 receptor agonist reversed the pain relief elicited by light-induced activation of the VTA-CeA pathway. These findings indicate that the VTA-CeA circuit is involved in pain relief in mice via dopamine receptor D2 in the CeA.

Glucocorticoid Receptor Contributes to Electroacupuncture-Induced Analgesia by Inhibiting Nav1.7 Expression in Rats With Inflammatory Pain Induced by Complete Freund's Adjuvant.

BACKGROUND: While electroacupuncture (EA) has been used traditionally for the treatment of chronic pain, its analgesic mechanisms have not been fully clarified. We observed in an earlier study that EA could reverse inflammatory pain and suppress high Nav1.7 expression. However, the molecular mechanism underlying Nav1.7 expression regulation is unclear. In this study, we studied the relationship between the glucocorticoid receptor (GR) and Nav1.7 and the role of these molecules in EA analgesia. MATERIALS AND METHODS: In this study, we established an inflammatory pain model by intraplantar injection of complete Freund's adjuvant (CFA) in rats. EA stimulation was applied to the ipsilateral "Huantiao" (GB30) and "Zusanli" (ST36) acupoints in the rat model. Western blotting, real-time polymerase chain reaction, immunostaining, intrathecal injection, and chromatin immunoprecipitation (ChIP) assay were performed to determine whether the sodium channel protein Nav1.7 plays a role in CFA-induced pain and whether GR regulates Nav1.7 expression during analgesia following EA stimulation. RESULTS: EA application significantly decreased the paw withdrawal threshold thresholds and thermal paw withdrawal latency and suppressed GR and Nav1.7 expression in the dorsal root ganglion. Moreover, treatment with a GR sense oligonucleotide (OND) markedly reversed these alterations. In contrast, treatment with a GR antisense OND along with EA application exerted a better analgesic effect, which was accompanied by the suppression of Nav1.7 and GR protein expression. The ChIP assay showed that the binding activity of GR to the Nav1.7 promoter was enhanced in CFA injected rats and suppressed in EA-treated rats. CONCLUSIONS: The present study demonstrated that EA exerted anti-hyperalgesic effects by inhibiting GR expression, which led to Nav1.7 expression modulation in the rat model of CFA-induced inflammatory pain.

Epigenetic restoration of voltage-gated potassium channel Kv1.2 alleviates nerve injury-induced neuropathic pain.

Voltage-gated potassium channels (Kv) are important regulators of neuronal excitability for its role of regulating resting membrane potential and repolarization. Recent studies show that Kv channels participate in neuropathic pain, but the detailed underlying mechanisms are far from being clear. In this study, we used siRNA, miR-137 agomir, and antagomir to regulate the expression of Kv1.2 in spinal cord and dorsal root ganglia (DRG) of naïve and chronic constriction injury (CCI) rats. Kv currents and neuron excitability in DRG neurons were examined by patch-clamp whole-cell recording to verify the change in Kv1.2 function. The results showed that Kv1.2 was down-regulated in DRG and spinal dorsal horn (SDH) by CCI. Knockdown of Kv1.2 by intrathecally injecting Kcna2 siRNA induced significant mechanical and thermal hypersensitivity in naïve rats. Concomitant with the down-regulation of Kv1.2 was an increase in the expression of the miR-137. The targeting and regulating of miR-137 on Kcna2 was verified by dual-luciferase reporter system and intrathecal injecting miR-137 agomir. Furthermore, rescuing the expression of Kv1.2 in CCI rats, achieved through inhibiting miR-137, restored the abnormal Kv currents and excitability in DRG neurons, and alleviated mechanical allodynia and thermal hyperalgesia. These results indicate that the miR-137-mediated Kv1.2 impairment is a crucial etiopathogenesis for the nerve injury-induced neuropathic pain and can be a novel potential therapeutic target for neuropathic pain management.

The different mechanisms of peripheral and central TLR4 on chronic postsurgical pain in rats.

Chronic postsurgical pain (CPSP) is a common complication after surgery; however, the underlying mechanisms of CPSP are poorly understood. As one of the most important inflammatory pathways, the Toll-like receptor 4/nuclear factor-kappa B (TLR4/NF-κB) signaling pathway plays an important role in chronic pain. However, the precise role of the TLR4/NF-κB signaling pathway in CPSP remains unclear. In the present study, we established a rat model of CPSP induced by skin/muscle incision and retraction (SMIR) and verified the effects and mechanisms of central and peripheral TLR4 and NF-κB on hyperalgesia in SMIR rats. The results showed that TLR4 expression was increased in both the spinal dorsal horn and dorsal root ganglia (DRGs) of SMIR rats. However, the TLR4 expression pattern in the spinal cord was different from that in DRGs. In the spinal cord, TLR4 was expressed in both neurons and microglia, whereas it was expressed in neurons but not in satellite glial cells in DRGs. Further results demonstrate that the central and peripheral TLR4/NF-κB signaling pathway is involved in the SMIR-induced CPSP by different mechanisms. In the peripheral nervous system, we revealed that the TLR4/NF-κB signaling pathway induced upregulation of voltage-gated sodium channel 1.7 (Nav1.7) in DRGs, triggering peripheral hyperalgesia in SMIR-induced CPSP. In the central nervous system, the TLR4/NF-κB signaling pathway participated in SMIR-induced CPSP by activating microglia in the spinal cord. Ultimately, our findings demonstrated that activation of the peripheral and central TLR4/NF-κB signaling pathway involved in the development of SMIR-induced CPSP.

Protein kinase C-α upregulates sodium channel Nav1.9 in nociceptive dorsal root ganglion neurons in an inflammatory arthritis pain model of rat.

Previous studies have found that increased expression of Nav1.9 and protein kinase C (PKC) contributes to pain hypersensitivity in a couple of inflammatory pain models. Here we want to observe if PKC can regulate the expression of Nav1.9 in dorsal root ganglion (DRG) in rheumatoid arthritis (RA) pain model. A chronic knee joint inflammation model was produced by intra-articular injection of the complete Freund's adjuvant (CFA) in rats. Nociceptive behaviors including mechanical, cold, and heat hyperalgesia were examined. The expression of Nav1.9 and PKCα in DRG was detected by a quantitative polymerase chain reaction, Western blot, and immunofluorescence. The in vitro and in vivo effects of a PKC activator (phorbol 12-myristate 13-acetate [PMA]) and a PKC inhibitor (GF-109203X) on the expression of Nav1.9 were examined. Moreover, the effects of PKC modulators on nociceptive behaviors were studied. Increased mechanical, heat, and cold sensitivity was observed 3 to 14 days after CFA injection. Parallel increases in messenger RNA and protein expression of Nav1.9 and PKCα were found. Immunofluorescence experiments found that Nav1.9 was preferentially colocalized with IB4+DRG neurons in RA rats. In cultured DRG neurons, PMA increased Nav1.9 expression while GF-109203X prevented the effect of PMA. PMA increased Nav1.9 expression in naïve rats while GF-109203X decreased Nav1.9 expression in RA rats. In naïve rats, PMA caused mechanical and cold hyperalgesia. On the other hand, GF-109203X attenuated mechanical and cold hyperalgesia in RA-pain model. Nav1.9 might be upregulated by PKCα in DRG, which contributes to pain hypersensitivity in CFA-induced chronic knee joint inflammation model of RA pain.

MicroRNA-30b regulates expression of the sodium channel Nav1.7 in nerve injury-induced neuropathic pain in the rat.

Voltage-gated sodium channels, which are involved in pain pathways, have emerged as major targets for therapeutic intervention in pain disorders. Nav1.7, the tetrodotoxin-sensitive voltage-gated sodium channel isoform encoded by SCN9A and predominantly expressed in pain-sensing neurons in the dorsal root ganglion, plays a crucial role in nociception. MicroRNAs are highly conserved, small non-coding RNAs. Through binding to the 3' untranslated region of their target mRNAs, microRNAs induce the cleavage and/or inhibition of protein translation. Based on bioinformatics analysis using TargetScan software, we determined that miR-30b directly targets SCN9A To investigate the roles of Nav1.7 and miR-30b in neuropathic pain, we examined changes in the expression of Nav1.7 in the dorsal root ganglion by miR-30b over-expression or knockdown in rats with spared nerve injury. Our results demonstrated that the expression of miR-30b and Nav1.7 was down-regulated and up-regulated, respectively, in the dorsal root ganglion of spared nerve injury rats. MiR-30b over-expression in spared nerve injury rats inhibited SCN9A transcription, resulting in pain relief. In addition, miR-30b knockdown significantly increased hypersensitivity to pain in naive rats. We also observed that miR-30b decreased Nav1.7 expression in PC12 cells. Taken together, our results suggest that miR-30b plays an important role in neuropathic pain by regulating Nav1.7 expression. Therefore, miR-30b may be a promising target for the treatment of chronic neuropathic pain.

Intrathecal injection of fluorocitric acid inhibits the activation of glial cells causing reduced mirror pain in rats.

BACKGROUND: Growing evidence has shown that unilateral nerve injury results in pain hypersensitivity in the ipsilateral and contralateral sides respective to the injury site. This phenomenon is known as mirror image pain (MIP). Glial cells have been indicated in the mechanism of MIP; however, it is not clear how glial cells are involved in MIP. METHODS: To observe phenomenon MIP and the following mechanism, 20 adult male Sprague-Dawley rats (weighing 180-220 g) were separated into two groups: Sham Group (n = 10) and left L5 spinal nerve ligated and sectioned (SNL) group (n = 10). Thermal hyperalgesia and mechanical hypersensitivity were measured for both groups to determine if the SNL model had Mirror image of Pain (MIP). Nav1.7 protein expression in DRG was analyzed using immunohistochemistry and western-blotting. And then to observe the effect of fluorocitrate on MIP, 15 rats were separated into three Groups: Sham Group (n = 5); SNL + FC group: intrathecal injection of Fluorocitric acid(FC) 1 nmol/10 µL (n = 5); SNL + NS group: intrathecal injection of 0.9% Normal Saline (n = 5). Behavior testing, immunocytochemistry, and western-blotting using dorsal root ganglion (DRG) from both sides were then conducted. RESULTS: The results showed pain hypersensitivity in both hind-paws of the SNL animals, Mechanical tests showed the paw withdrawal threshold dropped from 13.30 ± 1.204 g to 2.57 ± 1.963 g at 14 d as will as the ipsilateral paw thermal withdrawal threshold dropped from 16.5 ± 2.236 s to 4.38 ± 2.544 s at 14 d. Mechanical tests showed the contralateral paw withdrawal threshold dropped from 14.01 ± 1.412 to 4.2 ± 1.789 g at 7d will the thermal withdrawal threshold dropped from 16.8 ± 2.176 s to 7.6 ± 1.517 s at 7d. Nav1.7 expression increased and glial cells actived in bilateral side DRG after SNL compared with sham group. After intrathecal injection of fluorocitrate, the glial cell in bilateral DRG were inhibited and the pain behavior were reversed in both hindpaws too. CONCLUSIONS: Fluorocitrate can inhibit the activation of glial cells in spinal cord and DRG, and reduce MIP.

MAPK-ERK-CREB signaling pathway upregulates Nav1.6 in oxaliplatin-induced neuropathic pain in the rat.

The voltage-gated sodium channel subtype Nav1.6 is involved in the electrophysiological changes of primary sensory neurons that occur in oxaliplatin-induced neuropathic pain, but its regulatory mechanism remains unclear. In this study, Western blot, RT-qPCR, immunofluorescence staining, chromatin immunoprecipitation were used to prove the mechanism of MAPK-ERK-CREB signaling pathway participating in oxaliplatin-induced neuropathic pain by regulating Nav1.6. The results showed that p-Raf1 and p-ERK, key molecules in MAPK/ERK pathway, and Nav1.6 were significantly increased in DRGs of oxaliplatin-induced neuropathic pain rats. Inhibition of p-Raf1 and p-ERK respectively not only reduced the expression of Nav1.6 protein in DRGs of OXA rats, but also caused a decrease in Nav1.6 mRNA, which led us to further explore the transcription factor CREB regulated by MAPK/ERK pathway. Results showed that CREB was co-distributed with Nav1.6. Inhibition of CREB resulted in decreased mRNA and protein expression of Nav1.6, and alleviated oxaliplatin-induced neuropathic pain. A chromatin immunoprecipitation experiment proved that OXA caused p-CREB to directly bind to the promoter region of Scn8A, which is the encoding gene for Nav1.6, and promote the transcription of Scn8A. In summary, in this study, we found that oxaliplatin can activate the MAPK/ERK pathway, which promotes the expression and activation of CREB and leads to an increase in Scn8A transcription, and then leads to an increase in Nav1.6 protein expression to enhance neuronal excitability and cause pain. This study provides an experimental basis for the molecular mechanism of sodium channel regulation in oxaliplatin-induced neuropathic pain.

Effects of light illumination and the expression of wee1 on tissue regeneration in adult zebrafish.

The zebrafish exhibits an enhanced capability of regenerating most of its adult tissues. In this study, we examine the roles of light illumination and functional expression of mitosis-specific gene wee1 on adult zebrafish caudal fin regeneration after amputation. During the first 3 days post-amputation (dpa), the caudal fin regenerate rapidly in the day but slowly at night when the fish are kept in a normal light-dark cycle (LD) condition. However, this day-night rhythm of fin regeneration is not seen when the fish are kept in constant dark (DD), constant light (LL), or in fish in which the circadian rhythms are disrupted by random light (RL) exposures. The rate of fin growth reaches the peak levels at 2.5 dpa in LD, but is delayed when the fish are kept in DD, LL or RL conditions. In zebrafish in which the expression of wee1 is blocked by morpholinos, regeneration of the caudal fin is affected. Interestingly, the expression of wee1 also displays robust circadian rhythms. Together, the data suggests that fin regeneration in zebrafish is regulated by both environmental cues and functional gene expressions. Alterations in lighting conditions or inhibition of wee1 expression result in decreases in fin regeneration after injury.

Tissue regeneration after injury in adult zebrafish: the regenerative potential of the caudal fin.

The zebrafish has the potential to regenerate many of its tissues. In this study, we examined caudal fin regeneration in zebrafish that received repeated injuries (fin amputation) at different ages. In zebrafish that received repeated injuries, the potential for caudal fin regeneration, such as tissue growth and the expression of regeneration marker genes (msxb, fgf20a, bmp2b), did not decline in comparison to zebrafish that received only one amputation surgery. The process of initial fin regeneration (e.g., tissue outgrowth and the expression of regeneration marker genes at 7 days post-amputation) did not seem to correlate with age. However, slight differences in fin outgrowth were observed between young and old animals when examined in the late regeneration stages (e.g., 20 and 30 days post-amputation). Together, the data suggest that zebrafish has unlimited regenerative potential in the injured caudal fin.

Perioperative sleep deprivation activates the paraventricular thalamic nucleus resulting in persistent postoperative incisional pain in mice.

Background: The duration of postsurgical pain is closely correlated with perioperative stress. Most patients suffer short-term sleep disorder/deprivation before and/or after surgery, which leads to extended postsurgical pain by an undetermined mechanism. The paraventricular thalamus (PVT) is a critical area that contributes to the regulation of feeding, awakening, and emotional states. However, whether the middle PVT is involved in postoperative pain or the extension of postoperative pain caused by perioperative sleep deprivation has not yet been investigated. Methods: We established a model of postoperative pain by plantar incision with perioperative rapid eye movement sleep deprivation (REMSD) 6 h/day for 3 consecutive days in mice. The excitability of the CaMKIIα+ neurons in the middle PVT (mPVTCaMKIIα) was detected by immunofluorescence and fiber photometry. The activation/inhibition of mPVTCaMKIIα neurons was conducted by chemogenetics. Results: REMSD prolonged the duration of postsurgical pain and increased the excitability of mPVTCaMKIIα neurons. In addition, mPVTCaMKIIα neurons showed increased excitability in response to nociceptive stimuli or painful conditions. However, REMSD did not delay postsurgical pain recovery following the ablation of CaMKIIα neurons in the mPVT. The activation of mPVTCaMKIIα neurons prolonged the duration of postsurgical pain and elicited anxiety-like behaviors. In contrast, inhibition of mPVTCaMKIIα neurons reduced the postsurgical pain after REMSD. Conclusion: Our data revealed that the CaMKIIα neurons in the mPVT are involved in the extension of the postsurgical pain duration induced by REMSD, and represented a novel potential target to treat postoperative pain induced by REMSD.

Role of spinal RIP3 in inflammatory pain and electroacupuncture-mediated analgesic effect in mice.

AIMS: Electroacupuncture (EA) is a potentially useful treatment for inflammatory pain. Receptor-interacting protein 3 (RIP3) triggers the NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome; activation independent of necroptosis has been reported. However, the role of RIP3 in inflammatory pain and its EA-induced analgesic effects remains unclear. MAIN METHODS: Mice were treated with EA (2 Hz, 2 mA) after complete Freund's adjuvant (CFA) pain models were established. Inhibition or activation of spinal RIP3 was achieved by intrathecal administration of GSK-843 (a specific RIP3 inhibitor) or microinjection of lentivirus-RIP3, respectively. Mechanical analgesiometry and thermal analgesiometry were used to assess paw withdrawal threshold and paw withdrawal latency in mice. Quantitative polymerase chain reaction (qRT-PCR) and Western blotting were used to evaluate the expression of RIP3 and NLPR3 in spinal dorsal horn (SDH) of mice. KEY FINDINGS: The expression of spinal RIP3 and NLPR3 increased significantly after CFA injection. Both intrathecal administration of GSK-843 and EA alleviated mechanical and thermal pain behaviors induced by CFA and inhibited the expression of RIP3 and NLRP3 in the SDH of CFA mice. Over-expression of RIP3 induces pain-like symptoms in mice and inhibits the regulatory effects of EA on inflammatory pain. SIGNIFICANCE: Our results indicate that the EA analgesia effect may be related to suppression of RIP3 and NLRP3 expression in the SDH. This study could provide potential insights into the underlying spinal mechanisms involved in the analgesic effect of EA.

Raf1 interacts with OIP5 to participate in oxaliplatin-induced neuropathic pain.

AIMS: Oxaliplatin is an effective anti-cancer platinum-based chemotherapy drug which can cause severe chronic neuropathy, but the molecular mechanism underlying this adverse effect is still unclear. Opa interacting protein 5 (OIP5) is a member of the cancer/testis antigen (CTA) family and is involved in a variety of cancers. Studies have shown that Raf1, which is a serine/threonine-protein kinase, can directly combine with OIP5 to promote its expression. Whether Raf1 and OIP5 can participate in oxaliplatin-induced neuropathic pain has not been reported. MAIN METHODS: In this study, the oxaliplatin-induced neuropathic pain model was prepared by intraperitoneal injection of oxaliplatin. OIP5 and Raf1 were knocked down by intrathecal injection of siRNA against Raf1 and OIP5 (siRaf1, siOIP5). Von Frey fiber and acetone were used to detect pain behavior, and western blot was used to detect the protein expression changes of OIP5 and Raf1 in the dorsal root ganglion (DRG). KEY FINDINGS: The expression levels of p-Raf1 and OIP5 were increased in DRGs of oxaliplatin-induced neuropathic pain rats. Intrathecal administration of siOIP5 to inhibit the expression of OIP5 not only effectively alleviated oxaliplatin-induced mechanical allodynia and cold hyperalgesia, but also decreased the protein expression of Raf1. Intrathecal administration of siRaf1 inhibited the expression of OIP5 and attenuated oxaliplatin-induced neuropathic pain. SIGNIFICANCE: This study confirmed that Raf1 interacts with OIP5 to participate in oxaliplatin-induced neuropathic pain. The restricted expression of OIP5 in normal tissues may make it an ideal drug target for the treatment of oxaliplatin-induced neuropathic pain.

Prognostic Implications of Novel Ten-Gene Signature in Uveal Melanoma.

Background: Uveal melanoma (UM) is the most common primary intraocular cancer in adults. Genomic studies have provided insights into molecular subgroups and oncogenic drivers of UM that may lead to novel therapeutic strategies. Methods: Dataset TCGA-UVM, download from TCGA portal, were taken as the training cohort, and dataset GSE22138, obtained from GEO database, was set as the validation cohort. In training cohort, Kaplan-Meier analysis and univariate Cox regression model were applied to preliminary screen prognostic genes. Besides, the Cox regression model with LASSO was implemented to build a multi-gene signature, which was then validated in the validation cohorts through Kaplan-Meier, Cox, and ROC analyses. In addition, the correlation between copy number aberrations and risk score was evaluated by Spearman test. GSEA and immune infiltrating analyses were conducted for understanding function annotation and the role of the signature in the tumor microenvironment. Results: A ten-gene signature was built, and it was examined by Kaplan-Meier analysis revealing that significantly overall survival, progression-free survival, and metastasis-free survival difference was seen. The ten-gene signature was further proven to be an independent risk factor compared to other clinic-pathological parameters via the Cox regression analysis. Moreover, the receiver operating characteristic curve (ROC) analysis results demonstrated a better predictive power of the UM prognosis that our signature owned. The ten-gene signature was significantly correlated with copy numbers of chromosome 3, 8q, 6q, and 6p. Furthermore, GSEA and immune infiltrating analyses showed that the signature had close interactions with immune-related pathways and the tumor environment. Conclusions: Identifying the ten-gene signature (SIRT3, HMCES, SLC44A3, TCTN1, STPG1, POMGNT2, RNF208, ANXA2P2, ULBP1, and CA12) could accurately identify patients' prognosis and had close interactions with the immunodominant tumor environment, which may provide UM patients with personalized prognosis prediction and new treatment insights.

Voltage-gated sodium channel 1.7 expression decreases in dorsal root ganglia in a spinal nerve ligation neuropathic pain model.

The role of the voltage-gated sodium channel 1.7 (Nav1.7) is unclear in models of neuropathic pain induced by nerve injury. In the present study, we measured expression levels of Nav1.7 in two distinct neuropathic pain models: spinal nerve ligation (SNL) and chronic constriction injury (CCI). In the SNL model, both mRNA and protein levels of Nav1.7 were markedly lower in the L5 dorsal root ganglia (DRG) but were significantly higher in the L4 DRG. Nav1.7 protein levels were notably higher in both L4 and L5 DRGs under CCI conditions. We found that excessive damage of L5 nerves such as SNL reduced expression levels of Nav1.7 in the injured L5 DRG and activated the adjacent uninjured DRG, resulting in Nav1.7 level increases in the adjacent L4 DRG. We confirmed again that Nav1.7 was closely related to neuropathic pain induced by nerve injury. More importantly, our results suggest that tracing the molecular changes exclusively in the L5 DRG in SNL model may not completely explain the pain mechanism; it is necessary to study the adjacent uninjured L4 DRG.

MiR-30b-5p attenuates oxaliplatin-induced peripheral neuropathic pain through the voltage-gated sodium channel Nav1.6 in rats.

Oxaliplatin is a third-generation derivative of platinum that is effective in the treatment of multiple solid tumors. However, it can cause peripheral neuropathic pain, and the molecular mechanisms of this effect remain unknown. We induced a model of peripheral neuropathic pain in rats by intraperitoneally injecting them with oxaliplatin twice a week for 4.5 weeks. We found that both the mRNA and protein expression levels of Nav1.6 (encoded by the gene Scn8a) increased while the miR-30b-5p (shorthand for miR-30b) expression decreased in the dorsal root ganglion (DRG) of treated rats. Using TargetScan and miRanda predictive software, we discovered that Scn8a was a major target of miR-30b. Moreover, we found that miR-30b negatively regulated Scn8a by binding to the Scn8a 3'UTR in PC12 cells. In addition, Nav1.6 and miR-30b were colocalized in the DRG neurons of naive rats. Overexpression of miR-30b using an miR-30b agomir attenuated neuropathic pain induced by oxaliplatin and inhibited both the mRNA and protein expression levels of Nav1.6 both in vitro and in vivo. Conversely, the inhibition of miR-30b with an miR-30b antagomir resulted in neuropathic pain and an increase in the expression of Nav1.6. More importantly, overexpression of miR-30b inhibited the proliferation of LS-174t cells (Colorectal cancer cells). These data suggest that miR-30b contributes to oxaliplatin-induced chronic neuropathic pain through Nav1.6 downregulation and could be a novel therapeutic target for the treatment of oxaliplatin-induced neuropathic pain as a side effect of chemotherapy in cancer patients.

Glucocorticoid receptor inhibit the activity of NF-κB through p38 signaling pathway in spinal cord in the spared nerve injury rats.

AIMS: The glucocorticoid receptors (GRs) are an active regulator in inflammatory responses. The inflammatory reaction plays an important role in neuropathic pain, but the underlying mechanisms that GR regulates the inflammatory responses in neuropathic pain are still unknown. The activation of GRs has been shown to participate in the p38MAPK-mediated suppression of transcription activation. An unanswered question is whether GRs take part in inflammatory responses in neuropathic pain through p38MAPK signaling pathway. MAIN METHODS: The spared nerve injury (SNI) in rats was used as a model of neuropathic pain. Pain sensitivity was tested by von Frey filaments. The expression of GR, p-p38 and NF-κB were detected by Western blot and immunofluorescence. Elisa was used to examine the expression of IL-6 and TNF-α. KEY FINDINGS: Nerve injury led to p38 activation and GR expression decline in spinal cord of SNI rats. Intrathecal injection of the p38MAPK antagonist SB203580 activated GR and decreased NF-κB, resulting in pain relief since 3 days post-operation in SNI rats. Moreover, Intrathecal injection of the GR antagonist RU38486 counteracted the effect of SB203580 on NF-κB expression along with the release of IL-6 and TNF-α. On the contrary, activation of the GR by intrathecal administration of dexamethasone, a GR agonist, inhibited the expression of NF-κB and the release of IL-6 and TNF-α, resulting in pain relief. SIGNIFICANCE: Activation of p38MAPK in spinal cord could downregulate the GR expression and thereby activate NF-κB, thus promoting the release of IL-6 and TNF-α and participating in the development of neuropathic pain.

MicroRNA-182 Alleviates Neuropathic Pain by Regulating Nav1.7 Following Spared Nerve Injury in Rats.

The sodium channel 1.7 (Nav1.7), which is encoded by SCN9A gene, is involved in neuropathic pain. As crucial regulators of gene expression, many miRNAs have already gained importance in neuropathic pain, including miR-182, which is predicted to regulate the SCN9A gene. Nav1.7 expression in L4-L6 dorsal root ganglions (DRGs) can be up regulated by spared nerve injury (SNI), while miR-182 expression was down regulated following SNI model. Exploring the connection between Nav1.7 and miR-182 may facilitate the development of a better-targeted therapy. In the current study, direct pairing of miR-182 with the SCN9A gene was verified using a luciferase assay in vitro. Over-expression of miR-182 via microinjection of miR-182 agomir reversed the abnormal increase of Nav1.7 at both mRNA and protein level in L4-6 DRGs of SNI rats, and significantly attenuated the hypersensitivity to mechanical stimulus in the rats. In contrast, administration of miR-182 antagomir enhanced the Nav1.7 expression at both mRNA and protein level in L4-6 DRGs, companied with the generation of mechanical hypersensitivity in naïve rats. Collectively, we concluded that miR-182 can alleviate SNI- induced neuropathic pain through regulating Nav1.7 in rats.

MiR-30b Attenuates Neuropathic Pain by Regulating Voltage-Gated Sodium Channel Nav1.3 in Rats.

Nav1.3 is a tetrodotoxin-sensitive isoform among voltage-gated sodium channels that are closely associated with neuropathic pain. It can be up-regulated following nerve injury, but its biological function remains uncertain. MicroRNAs (miRNAs) are endogenous non-coding RNAs that can regulate post-transcriptional gene expression by binding with their target mRNAs. Using Target Scan software, we discovered that SCN3A is the major target of miR-30b, and we then determined whether miR-30b regulated the expression of Nav1.3 by transfecting miR-30b agomir through the stimulation of TNF-α or by transfecting miR-30b antagomir in primary dorsal root ganglion (DRG) neurons. The spinal nerve ligation (SNL) model was used to determine the contribution of miR-30b to neuropathic pain, to evaluate changes in Nav1.3 mRNA and protein expression, and to understand the sensitivity of rats to mechanical and thermal stimuli. Our results showed that miR-30b agomir transfection down-regulated Nav1.3 mRNA stimulated with TNF-α in primary DRG neurons. Moreover, miR-30b overexpression significantly attenuated neuropathic pain induced by SNL, with decreases in the expression of Nav1.3 mRNA and protein both in DRG neurons and spinal cord. Activation of Nav1.3 caused by miR-30b antagomir was identified. These data suggest that miR-30b is involved in the development of neuropathic pain, probably by regulating the expression of Nav1.3, and might be a novel therapeutic target for neuropathic pain. Perspective: This study is the first to explore the important role of miR-30b and Nav1.3 in spinal nerve ligation-induced neuropathic pain, and our evidence may provide new insight for improving therapeutic approaches to pain.

[Dexamethason suppresses the activation of rat microglia induced by glutamic acid].

OBJECTIVE: To investigate the effect of dexamethasone (DEX) on the microglia activation induced by glutamic acid (GLU) in rats. METHODS: We isolated and cultured the microglia from the spinal cord of SD infant rats in vitro. The cell purity was tested by immunofluorescence technique. The cells were then randomly divided into 5 groups: Dulbecco' s phosphate buffered saline treatment (DPBS group); GLU treatment (GLU group); DEX pretreatment and then GLU stimulation; simultaneous treatment of DEX and GLU; GLU stimulation followed by DEX treatment. Finally, immunofluorescence technique was used to investigate the expressions of glucocorticoid receptor (GR) and CD11b/c protein. RESULTS: Compared with the DPBS group, GLU group presented the increased expression of CD11b/c protein, the shorten length of cell processes as well as cell shape turning round. Furthermore, compared with the GLU group, the CD11b/c protein expression significantly decreased in the group treated simultaneously with DEX and GLU and the group treated with DEX after GLU stimulation. However, the expression was not different between the GLU group and the group treated with DEX and then stimulated by GLU. CONCLUSION: DEX could inhibit microglia activation induced by GLU, while DEX pretreatment have no such an effect on microglia.