Serotonin receptor 2B induces visceral hyperalgesia in rat model and patients with diarrhea-predominant irritable bowel syndrome.

BACKGROUND: Serotonin receptor 2B (5-HT2B receptor) plays a critical role in many chronic pain conditions. The possible involvement of the 5-HT2B receptor in the altered gut sensation of irritable bowel syndrome with diarrhea (IBS-D) was investigated in the present study. AIM: To investigate the possible involvement of 5-HT2B receptor in the altered gut sensation in rat model and patients with IBS-D. METHODS: Rectosigmoid biopsies were collected from 18 patients with IBS-D and 10 patients with irritable bowel syndrome with constipation who fulfilled the Rome IV criteria and 15 healthy controls. The expression level of the 5-HT2B receptor in colon tissue was measured using an enzyme-linked immunosorbent assay and correlated with abdominal pain scores. The IBS-D rat model was induced by intracolonic instillation of acetic acid and wrap restraint. Alterations in visceral sensitivity and 5-HT2B receptor and transient receptor potential vanilloid type 1 (TRPV1) expression were examined following 5-HT2B receptor antagonist administration. Changes in visceral sensitivity after administration of the TRPV1 antagonist were recorded. RESULTS: Here, we observed greater expression of the 5-HT2B receptor in the colonic mucosa of patients with IBS-D than in that of controls, which was correlated with abdominal pain scores. Intracolonic instillation of acetic acid and wrap restraint induced obvious chronic visceral hypersensitivity and increased fecal weight and fecal water content. Exogenous 5-HT2B receptor agonist administration increased visceral hypersensitivity, which was alleviated by successive administration of a TRPV1 antagonist. IBS-D rats receiving the 5-HT2B receptor antagonist exhibited inhibited visceral hyperalgesia.Moreover, the percentage of 5-HT2B receptor-immunoreactive (IR) cells surrounded by TRPV1-positive cells (5-HT2B receptor I+) and total 5-HT2B receptor IR cells (5-HT2B receptor IT) in IBS-D rats was significantly reduced by the administration of a 5-HT2B receptor antagonist. CONCLUSION: Our finding that increased expression of the 5-HT2B receptor contributes to visceral hyperalgesia by inducing TRPV1 expression in IBS-D patients provides important insights into the potential mechanisms underlying IBS-D-associated visceral hyperalgesia.

Stigmasterol alleviates neuropathic pain by reducing Schwann cell-macrophage cascade in DRG by modulating IL-34/CSF1R.

AIMS: This study aimed to investigate the potential therapeutic applications of stigmasterol for treating neuropathic pain. METHODS: Related mechanisms were investigated by DRG single-cell sequencing analysis and the use of specific inhibitors in cellular experiments. In animal experiments, 32 male Sprague-Dawley rats were randomly divided into the sham operation group, CCI group, ibuprofen group, and stigmasterol group. We performed behavioral tests, ELISA, H&E staining and immunohistochemistry, and western blotting. RESULTS: Cell communication analysis by single-cell sequencing reveals that after peripheral nerve injury, Schwann cells secrete IL-34 to act on CSF1R in macrophages. After peripheral nerve injury, the mRNA expression levels of CSF1R pathway and NLRP3 inflammasome in macrophages were increased in DRG. In vitro studies demonstrated that stigmasterol can reduce the secretion of IL-34 in LPS-induced RSC96 Schwann cells; stigmasterol treatment of LPS-induced Schwann cell-conditioned medium (L-S-CM) does not induce the proliferation and migration of RAW264.7 macrophages; L-S-CM reduces CSF1R signaling pathway (CSF1R, P38MAPK, and NFκB) activation, NLRP3 inflammasome activation, and ROS production. In vivo experiments have verified that stigmasterol can reduce thermal and cold hyperalgesia in rat chronic compressive nerve injury (CCI) model; stigmasterol can reduce IL-1ß, IL-6, TNF-α, CCL2, SP, and PGE2 in serum of CCI rats; immunohistochemistry and western blot confirmed that stigmasterol can reduce the levels of IL-34/CSF1R signaling pathway and NLRP3 inflammasome in DRG of CCI rats. CONCLUSION: Stigmasterol alleviates neuropathic pain by reducing Schwann cell-macrophage cascade in DRG by modulating IL-34/CSF1R axis.

Circular RNA-GRIN2B Suppresses Neuropathic Pain by Targeting the NF-κB/SLICK Pathway.

The role of circular RNAs (circRNAs) in neuropathic pain is linked to the fundamental physiological mechanisms involved. However, the exact function of circRNAs in the context of neuropathic pain is still not fully understood. The functional impact of circGRIN2B on the excitability of dorsal root ganglion (DRG) neurons was investigated using siRNA or overexpression technology in conjunction with fluorescence in situ hybridization and whole-cell patch-clamp technology. The therapeutic efficacy of circGRIN2B in treating neuropathic pain was confirmed by assessing the pain threshold in a chronic constrictive injury (CCI) model. The interaction between circGRIN2B and NF-κB was examined through RNA pulldown, RIP, and mass spectrometry assays. CircGRIN2B knockdown significantly affected the action potential discharge frequency and the sodium-dependent potassium current flux (SLICK) in DRG neurons. Furthermore, knockdown of circGRIN2B dramatically reduced the SLICK channel protein and mRNA expression in vivo and in vitro. Our research confirmed the interaction between circGRIN2B and NF-κB. These findings demonstrated that circGRIN2B promotes the transcription of the SLICK gene by binding to NF-κB. In CCI rat models, the overexpression of circGRIN2B has been shown to hinder the progression of neuropathic pain, particularly by reducing mechanical and thermal hyperalgesia. Additionally, this upregulation significantly diminished the levels of the inflammatory cytokines IL-1ß, IL-6, and TNF-α in the DRG. Upon reviewing these findings, it was determined that circGRIN2B may mitigate the onset of neuropathic pain by modulating the NF-κB/SLICK pathway.

High-fat diets promote peritoneal inflammation and augment endometriosis-associated abdominal hyperalgesia.

Immune dysfunction is one of the central components in the development and progression of endometriosis by establishing a chronic inflammatory environment. Western-style high-fat diets (HFD) have been linked to greater systemic inflammation to cause metabolic and chronic inflammatory diseases, and are also considered an environmental risk factor for gynecologic diseases. Here, we aimed to examine how HFD cause an inflammatory environment in endometriosis and discern their contribution to endometriotic-associated hyperalgesia. Our results showed that HFD-induced obesity enhanced abdominal hyperalgesia that was induced by endometriotic lesions. Peritoneal inflammatory macrophages and cytokine levels increased by lesion induction were elevated by chronic exposure to HFD. Increased expression of pain-related mediators in the dorsal root ganglia was observed after lesion induction under the HFD condition. Although HFD did not affect inflammatory macrophages in the peritoneal cavity without lesion induction, the diversity and composition of the gut microbiota were clearly altered by HFD as a sign of low-grade systemic inflammation. Thus, HFD alone might not establish a local inflammatory environment in the pelvic cavity, but it can contribute to further enhancing chronic inflammation, leading to the exacerbation of endometriosis-associated abdominal hyperalgesia following the establishment and progression of the disease.

Distinct local and global functions of mouse Aß low-threshold mechanoreceptors in mechanical nociception.

The roles of Aß low-threshold mechanoreceptors (LTMRs) in transmitting mechanical hyperalgesia and in alleviating chronic pain have been of great interest but remain contentious. Here we utilized intersectional genetic tools, optogenetics, and high-speed imaging to specifically examine functions of SplitCre labeled mouse Aß-LTMRs in this regard. Genetic ablation of SplitCre-Aß-LTMRs increased mechanical nociception but not thermosensation in both acute and chronic inflammatory pain conditions, indicating a modality-specific role in gating mechanical nociception. Local optogenetic activation of SplitCre-Aß-LTMRs triggered nociception after tissue inflammation, whereas their broad activation at the dorsal column still alleviated mechanical hypersensitivity of chronic inflammation. Taking all data into consideration, we propose a model, in which Aß-LTMRs play distinctive local and global roles in transmitting or alleviating mechanical hyperalgesia of chronic pain, respectively. Our model suggests a strategy of global activation plus local inhibition of Aß-LTMRs for treating mechanical hyperalgesia.

Knockdown siRNA Targeting GPR55 Reveals Significant Differences Between the Anti-inflammatory Actions of KLS-13019 and Cannabidiol.

KLS-13019 was reported previously to reverse paclitaxel-induced mechanical allodynia in a mouse model of chemotherapy-induced peripheral neuropathy (CIPN). Recent studies demonstrated that paclitaxel-induced increases in inflammatory markers (GPR55, NLRP3, and IL-1ß) of dorsal root ganglion (DRG) cultures were shown to be reversed by KLS-13019 treatment. The mechanism of action for KLS-13019-mediated reversal of paclitaxel-induced neuroinflammation now has been explored using GPR55 siRNA. Pre-treatment of DRG cultures with GPR55 siRNA produced a 21% decrease of immunoreactive (IR) area for GPR55 in cell bodies and a 59% decrease in neuritic IR area, as determined by high-content imaging. Using a 24-h reversal treatment paradigm, paclitaxel-induced increases in the inflammatory markers were reversed back to control levels after KLS-3019 treatment. Decreases in these inflammatory markers produced by KLS-13019 were significantly attenuated by GPR55 siRNA co-treatment, with mean IR area responses being attenuated by 56% in neurites and 53% in cell bodies. These data indicate that the percentage decreases in siRNA-mediated attenuation of KLS-13019-related efficacy on the inflammatory markers were similar to the percentage knockdown observed for neuritic GPR55 IR area. Similar studies conducted with cannabidiol (CBD), the parent compound of KLS-13019, produced low efficacy (25%) reversal of all inflammatory markers that were poorly attenuated (29%) by GPR55 siRNA. CBD was shown previously to be ineffective in reversing paclitaxel-induced mechanical allodynia. The present studies indicated significant differences between the anti-inflammatory properties of KLS-13019 and CBD which may play a role in their observed differences in the reversibility of mechanical allodynia in a mouse model of CIPN.

Chronic restraint stress-induced hyperalgesia is modulated by the periaqueductal gray neurons projecting to the rostral ventromedial medulla in mice.

Stress-induced hyperalgesia (SIH) is induced by repeated or chronic exposure to stressful or uncomfortable environments. However, the neural mechanisms involved in the modulatory effects of the periaqueductal gray (PAG) and its associated loops on SIH development hav e not been elucidated. In the present study, we used chronic restraint stress (CRS)-induced hyperalgesia as a SIH model and manipulated neuronal activity via a pharmacogenetic approach to investigate the neural mechanism underlying the effects of descending pain-modulatory pathways on SIH. We found that activation of PAG neurons alleviates CRS-induced hyperalgesia; on the other hand, PAG neurons inhibition facilitates CRS-induced hyperalgesia. Moreover, this modulatory effect is achieved by the neurons which projecting to the rostral ventromedial medulla (RVM). Our data thus reveal the functional role of the PAG-RVM circuit in SIH and provide analgesic targets in the brain for clinical SIH treatment.

Nociceptor spontaneous activity is responsible for fragmenting non-rapid eye movement sleep in mouse models of neuropathic pain.

Spontaneous pain, a major complaint of patients with neuropathic pain, has eluded study because there is no reliable marker in either preclinical models or clinical studies. Here, we performed a comprehensive electroencephalogram/electromyogram analysis of sleep in several mouse models of chronic pain: neuropathic (spared nerve injury and chronic constriction injury), inflammatory (Freund's complete adjuvant and carrageenan, plantar incision) and chemical pain (capsaicin). We find that peripheral axonal injury drives fragmentation of sleep by increasing brief arousals from non-rapid eye movement sleep (NREMS) without changing total sleep amount. In contrast to neuropathic pain, inflammatory or chemical pain did not increase brief arousals. NREMS fragmentation was reduced by the analgesics gabapentin and carbamazepine, and it resolved when pain sensitivity returned to normal in a transient neuropathic pain model (sciatic nerve crush). Genetic silencing of peripheral sensory neurons or ablation of CGRP+ neurons in the parabrachial nucleus prevented sleep fragmentation, whereas pharmacological blockade of skin sensory fibers was ineffective, indicating that the neural activity driving the arousals originates ectopically in primary nociceptor neurons and is relayed through the lateral parabrachial nucleus. These findings identify NREMS fragmentation by brief arousals as an effective proxy to measure spontaneous neuropathic pain in mice.

SIRT3 alleviates painful diabetic neuropathy by mediating the FoxO3a-PINK1-Parkin signaling pathway to activate mitophagy.

INTRODUCTION: Painful diabetic neuropathy (PDN) is a common complication of diabetes. Previous studies have implicated that mitochondrial dysfunction plays a role in the development of PDN, but its pathogenesis and mechanism have not been fully investigated. METHODS: In this study, we used high-fat diet/low-dose streptozotocin-induced rats as a model of type 2 diabetes mellitus. Behavioral testing, whole-cell patch-clamp recordings of dorsal root ganglion (DRG) neurons, and complex sensory nerve conduction velocity studies were used to assess peripheral neuropathy. Mitochondrial membrane potential (MMP), ATP, tissue reactive oxygen species, and transmission electron microscopy were used to evaluate the function and morphology of mitochondria in DRG. Real-time PCR, western blot, and immunofluorescence were performed to investigate the mechanism. RESULTS: We found that damaged mitochondria were accumulated and mitophagy was inhibited in PDN rats. The expression of sirtuin 3 (SIRT3), which is an NAD+-dependent deacetylase in mitochondria, was inhibited. Overexpression of SIRT3 in DRG neurons by intrathecally administered LV-SIRT3 lentivirus ameliorated neurological and mitochondrial dysfunctions. This was evidenced by the reversal of allodynia and nociceptor hyperexcitability, as well as the restoration of MMP and ATP levels. Overexpression of SIRT3 restored the inhibited mitophagy by activating the FoxO3a-PINK1-Parkin signaling pathway. The effects of SIRT3 overexpression, including the reversal of allodynia and nociceptor hyperexcitability, the improvement of impaired mitochondria and mitophagy, and the restoration of PINK1 and Parkin expression, were counteracted when FoxO3a siRNA was intrathecally injected. CONCLUSION: These results showed that SIRT3 overexpression ameliorates PDN via activation of FoxO3a-PINK1-Parkin-mediated mitophagy, suggesting that SIRT3 may become an encouraging therapeutic strategy for PDN.

Repetitive Transcranial Magnetic Stimulation with Sensorimotor Training for the Treatment of Complex Regional Pain Syndrome Type 2 of the Upper Limb Case Report.

This case report describes the use of repetitive transcranial magnetic stimulation (rTMS) combined with sensorimotor training (SMT) to treat an individual with complex regional pain syndrome (CRPS) type 2 with allodynia of the right hand/wrist. After the 9-week intervention, there was a clinically meaningful reduction in pain intensity which continued to 3 months after intervention. Further, clinically meaningful improvements in wrist and hand function and allodynia were observed. Although the use of rTMS for CRPS has been reported, this unique report provides valuable insight into the clinical utility of rTMS plus SMT for the treatment of CRPS and related symptoms.

Electroacupuncture promotes macrophage/microglial M2 polarization and suppresses inflammatory pain through AMPK.

Inflammatory pain, the most prevalent disease globally, remains challenging to manage. Electroacupuncture emerges as an effective therapy, yet its underlying mechanisms are not fully understood. This study investigates whether adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK)-regulated silent information regulator 1 (SIRT1) contributes to electroacupuncture's antinociceptive effects by modulating macrophage/microglial polarization in the spinal dorsal horn of a mouse model of inflammatory pain. In this study, mice, introduced to inflammatory pain through subcutaneous injections of complete freund's adjuvant (CFA) in the plantar area, underwent electroacupuncture therapy every alternate day for 30-min sessions. The assessment of mechanical allodynia and thermal hyperalgesia in these subjects was carried out using paw withdrawal frequency and paw withdrawal latency measurements, respectively. Western blot analysis measured levels of AMPK, phosphorylation-adenosine 5'-monophosphate (AMP)-activated protein kinase, SIRT1, inducible nitric oxide synthase, cluster of differentiation 86, arginase 1, and interleukin 10. In contrast to the group treated solely with CFA, the cohort receiving both CFA and electroacupuncture demonstrated notable decreases in both thermal hyperalgesia and mechanical allodynia. This was accompanied by a marked enhancement in AMPK phosphorylation levels. AMPK knockdown reversed electroacupuncture's analgesic effects and reduced M2 macrophage/microglial polarization enhancement. Additionally, AMPK knockdown significantly weakened electroacupuncture-induced SIRT1 upregulation, and EX-527 injection attenuated electroacupuncture's facilitation of M2 macrophage/microglial polarization without affecting AMPK phosphorylation levels. Furthermore, combining electroacupuncture with SRT1720 enhanced the analgesic effect of SRT1720. Our findings suggest that AMPK regulation of SIRT1 plays a critical role in electroacupuncture's antinociceptive effect through the promotion of M2 macrophage/microglial polarization.

The Serotonin-Mediated Anti-Allodynic Effect of Yokukansan on Paclitaxel-Induced Neuropathic Pain.

Refractory peripheral neuropathy can occur as a side effect in 60-70% of patients receiving Paclitaxel (PTX). Yokukansan (YKS) is a Japanese herbal medicine reported to have analgesic properties for entrapment nerve injuries. Therefore, we investigated the anti-allodynic effect of Yokukansan on Paclitaxel-induced neuropathic pain. All experiments used 6-week-old male Sprague Dawley rats. Mechanical allodynia was evaluated using a dynamic plantar aesthesiometer. A mobile touch-stimulator unit applied progressively increasing force to the mid-plantar region of the hind paw in a vertical direction until the animal withdrew its paw. This was carried out before the Paclitaxel administration and during the first, second, third, and fourth weeks. Using a rat model of PTX-induced neuropathic pain (PTX rat), we injected PTX (intraperitoneally, 2 mg/kg) five times every 2 days. Using the dynamic plantar test, we evaluated the anti-allodynic effect of YKS (orally administered, 1 g/kg). YKS administration on a daily basis significantly enhanced the withdrawal threshold in PTX rats and reduced the expression level of activated microglia immunostaining with Iba1, a specific marker for microglia. The intrathecal administration of WAY-100635 (5-hydroxytryptamine [5-HT]1A receptor antagonist) and Ketanserin (5-HT2A/2C receptor antagonist) inhibited the protective effects of YKS. YKS exhibited an anti-allodynic effect in a rodent model of PTX-induced neuropathic pain by reducing the sensitivity to pain stimuli. These results suggest that Yokukansan may activate 5-HT receptors in the spinal cord, mediating Paclitaxel-induced neuropathic pain.

Assessing the Impact of Influenza Vaccination Timing on Experimental Arthritis: Effects on Disease Progression and Inflammatory Biomarkers.

Numerous studies have indicated a link between vaccines and the exacerbation of autoimmune diseases including rheumatoid arthritis (RA). However, there is no consensus in clinical practice regarding the optimal timing of immunization. Therefore, this study aimed to investigate the impact of the 3Fluart influenza vaccine on the complete Freund's adjuvant (CFA)-induced chronic arthritis rat model and to identify new biomarkers with clinical utility. CFA was injected into the plantar surface of one hind paw and the root of the tail on day 0, and the tail root injection was repeated on day 1. Flu vaccination was performed on day 1 or 7. Paw volume was measured by plethysmometry, mechanonociceptive threshold by dynamic plantar aesthesiometry, neutrophil myeloperoxidase (MPO) activity, and vascular leakage using in vivo optical imaging throughout the 21-day experiment. Inflammatory markers were determined by Western blot and histopathology. CFA-induced swelling, an increase in MPO activity, plasma extravasation in the tibiotarsal joint. Mechanical hyperalgesia of the hind paw was observed 3 days after the injection, which gradually decreased. Co-administration of the flu vaccine on day 7 but not on day 1 resulted in significantly increased heme oxygenase 1 (HO-1) expression. The influenza vaccination appears to have a limited impact on the progression and severity of the inflammatory response and associated pain. Nevertheless, delayed vaccination could alter the disease activity, as indicated by the findings from assessments of edema and inflammatory biomarkers. HO-1 may serve as a potential marker for the severity of inflammation, particularly in the case of delayed vaccination. However, further investigation is needed to fully understand the regulation and role of HO-1, a task that falls outside the scope of the current study.

Puerarin attenuates remifentanil­induced postoperative hyperalgesia via targeting PAX6 to regulate the transcription of TRPV1.

Remifentanil­induced hyperalgesia (RIH) is characterized by the emergence of stimulation­induced pain, including phenomena such as allodynia and thermal hyperalgesia following remifentanil infusion. As a sequence­specific DNA binding transcription factor, PAX6 positively and negatively regulates transcription and is expressed in multiple cell types in the developing and adult central nervous system. It was hypothesized that puerarin could relieve RIH via targeting PAX6 to regulate transcription of transient receptor potential cation channel subfamily V Member 1 (TRPV1). A total of 32 rats were randomly divided into five groups, namely control group, RI group, RI + 10 mg/kg puerarin group (RI + puerarin10), RI + 20 mg/kg puerarin group (RI + puerarin20), and RI + 40 mg/kg puerarin group (RI + puerarin40). Mechanical and thermal hyperalgesia were tested at ­24, 2, 6, 24 and 48 h after remifentanil infusion. Following the sacrifice of rats after the last behavioral test, western blot was used to detect the expression levels of TRPV1 in the tissues; Immunofluorescence staining and western blotting were used to detect the expression of PAX6 in the spinal cord. PharmMapper and JASPAR were used to predict the binding sites of puerarin/PAX6/TRPV1. Chromatin immunoprecipitation­PCR and dual luciferase reporter assay were used to verify the targeting relationship between PAX6 and TRPV1. Immunofluorescence was used to detect the expression levels of TRPV1 and p­NR2B. The results revealed that puerarin (10, 20, 40 mg/kg) dose­dependently reduced thermal and mechanical hyperalgesia from 2 to 48 h after remifentanil infusion. Remifentanil infusion remarkably stimulated the expression of phosphorylated (p­)NR2B. Nevertheless, the increased amount of p­NR2B by RIH was dose­dependently suppressed by puerarin in rats. In conclusion, puerarin was revealed to attenuate postoperative RIH via targeting PAX6 to regulate the transcription of TRPV1.

Blocking Pannexin 1 Channels Alleviates Peripheral Inflammatory Pain but not Paclitaxel-Induced Neuropathy.

BACKGROUND: Pannexin1 (Panx1) is a membrane channel expressed in different cells of the nervous system and is involved in several pathological conditions, including pain and inflammation. At the central nervous system, the role of Panx1 is already well-established. However, in the periphery, there is a lack of information regarding the participation of Panx1 in neuronal sensitization. The dorsal root ganglion (DRG) is a critical structure for pain processing and modulation. For this reason, understanding the molecular mechanism in the DRG associated with neuronal hypersensitivity has become highly relevant to discovering new possibilities for pain treatment. Here, we aimed to investigate the role of Panx1 in acute nociception and peripheral inflammatory and neuropathic pain by using two different approaches. METHODS: Rats were treated with a selective Panx1 blocker peptide (10Panx) into L5-DRG, followed by ipsilateral intraplantar injection of carrageenan, formalin, or capsaicin. DRG neuronal cells were pre-treated with 10Panx and stimulated by capsaicin to evaluate calcium influx. Panx1 knockout mice (Panx1-KO) received carrageenan or capsaicin into the paw and paclitaxel intraperitoneally. The von Frey test was performed to measure the mechanical threshold of rats' and mice's paws before and after each treatment. RESULTS: Pharmacological blockade of Panx1 in the DRG of rats resulted in a dose-dependent decrease of mechanical allodynia triggered by carrageenan, and nociception decreased in the second phase of formalin. Nociceptive behavior response induced by capsaicin was significantly lower in rats treated with Panx1 blockade into DRG. Neuronal cells with Panx1 blockage showed lower intracellular calcium response than untreated cells after capsaicin administration. Accordingly, Panx1-KO mice showed a robust reduction in mechanical allodynia after carrageenan and a lower nociceptive response to capsaicin. A single dose of paclitaxel promoted acute mechanical pain in wildtype (WT) but not in Panx1-KO mice. Four doses of chemotherapy promoted chronic mechanical allodynia in both genotypes, although Panx1-KO mice had significant ablation in the first eight days. CONCLUSION: Our findings suggest that Panx1 is critical for developing peripheral inflammatory pain and acute nociception involving transient receptor potential vanilloid subtype 1 (TRPV1) but is not essential for neuropathic pain chronicity.

Insula→Amygdala and Insula→Thalamus Pathways Are Involved in Comorbid Chronic Pain and Depression-Like Behavior in Mice.

The comorbidity of chronic pain and depression poses tremendous challenges for the treatment of either one because they exacerbate each other with unknown mechanisms. As the posterior insular cortex (PIC) integrates multiple somatosensory and emotional information and is implicated in either chronic pain or depression, we hypothesize that the PIC and its projections may contribute to the pathophysiology of comorbid chronic pain and depression. We show that PIC neurons were readily activated by mechanical, thermal, aversive, and stressful and appetitive stimulation in naive and neuropathic pain male mice subjected to spared nerve injury (SNI). Optogenetic activation of PIC neurons induced hyperalgesia and conditioned place aversion in naive mice, whereas inhibition of these neurons led to analgesia, conditioned place preference (CPP), and antidepressant effect in both naive and SNI mice. Combining neuronal tracing, optogenetics, and electrophysiological techniques, we found that the monosynaptic glutamatergic projections from the PIC to the basolateral amygdala (BLA) and the ventromedial nucleus (VM) of the thalamus mimicked PIC neurons in pain modulation in naive mice; in SNI mice, both projections were enhanced accompanied by hyperactivity of PIC, BLA, and VM neurons and inhibition of these projections led to analgesia, CPP, and antidepressant-like effect. The present study suggests that potentiation of the PIC→BLA and PIC→VM projections may be important pathophysiological bases for hyperalgesia and depression-like behavior in neuropathic pain and reversing the potentiation may be a promising therapeutic strategy for comorbid chronic pain and depression.

Antinociceptive and anti-inflammatory activity of extracts and α-pyrones isolated from Cantinoa stricta.

This study evaluated the composition and the antinociceptive and anti-inflammatory activity of the crude extracts and two isolated compounds, anamarine (ANA) and 10-epi-olguine (eOL), obtained from the leaves of Cantinoa stricta (Lamiaceae). Crude ethanolic extract (EEt) and dichloromethane extract (DCM), selected based on NMR data, were submitted to pharmacological tests in male Swiss mice. The oral administration of EEt and DCM significantly reduced the second phase of formalin-induced nociception (60%), lipopolysaccharide (LPS)-induced mechanical hyperalgesia (90%), and carrageenan (Cg)-induced edema (25%). ANA and eOL, the major compounds in EEt and DCM extracts, administered orally or locally (in the paw), also reduced the LPS-induced mechanical hyperalgesia (Oral ID50 1.9 and 3.9 mg/kg; Local ID50 93.4 and 677.3 ng, respectively) without changing the thermal acute nociception or the motor performance of the animals. Local administration of ANA and eOL also reduced Cg-induced edema (40 and 23%, respectively). These isolated compounds did not change the mechanical hyperalgesia induced by tumor necrosis factor-α, interleukin-1ß, prostaglandin E2, dibutyryl cyclic AMP, or forskolin but reversed the hyperalgesia induced by dopamine, epinephrine, and phorbol 12-myristate 13-acetate. The hyperalgesia induced by epinephrine was reversed in male but not in female mice, in which this response is not dependent on protein kinase C (PKC). These results suggest that C. stricta extracts possess antinociceptive and anti-inflammatory activity which is related to the presence of ANA and eOL. Differently from the known analgesics, these substances seem to exert their action mainly interfering with the sympathetic component of pain, possibly with PKC.

Spinal nerve transection-induced upregulation of SAP97 via promoting membrane trafficking of GluA1-containing AMPA receptors in the dorsal horn contributes to the pathogenesis of neuropathic pain.

Emerging evidence has implicated an important role of synapse-associated protein-97 (SAP97)-regulated GluA1-containing AMPARs membrane trafficking in cocaine restate and in contextual episodic memory of schizophrenia. Herein, we investigated the role of SAP97 in neuropathic pain following lumbar 5 spinal nerve transection (SNT) in rats. Our results showed that SNT led to upregulation of SAP97, enhanced the interaction between SAP97 and GluA1, and increased GluA1-containing AMPARs membrane trafficking in the dorsal horn. Microinjection of AAV-EGFP-SAP97 shRNA in lumbar 5 spinal dorsal horn inhibited SAP97 production, decreased SAP97-GluA1 interaction, reduced the membrane trafficking of GluA1-containing AMPARs, and partially attenuated neuropathic pain following SNT. Intrathecal injections of SAP97 siRNA or NASPM, an antagonist of GluA1-containing AMPARs, also partially reversed neuropathic pain on day 7, but not on day 14, after SNT. Spinal overexpression of SAP97 by AAV-EGFP-SAP97 enhanced SAP97-GluA1 interaction, increased the membrane insertion of GluA1-containing AMPARs, and induced abnormal pain in naïve rats. In addition, treatment with SAP97 siRNA or NASPM i.t. injection alleviated SNT-induced allodynia and hyperalgesia and exhibited a longer effect in female rats. Together, our results indicate that the SNT-induced upregulation of SAP97 via promoting GluA1-containing AMPARs membrane trafficking in the dorsal horn contributes to the pathogenesis of neuropathic pain. Targeting spinal SAP97 might be a promising therapeutic strategy to treatment of chronic pain.

Electroacupuncture attenuates nociceptive behaviors in a mouse model of cancer pain.

Pain is a major symptom in cancer patients, and cancer-induced bone pain (CIBP) is the most common type of moderate and severe cancer-related pain. The current available analgesic treatments for CIBP have adverse effects as well as limited therapeutic effects. Acupuncture is proved effective in pain management as a safe alternative therapy. We evaluated the analgesic effect of acupuncture in treatment of cancer pain and try to explore the underlying analgesic mechanisms. Nude mice were inoculated with cancer cells into the left distal femur to establish cancer pain model. Electroacupuncture (EA) treatment was applied for the xenograft animals. Pain behaviors of mice were evaluated, followed by the detections of neuropeptide-related and inflammation-related indicators in peripheral and central levels. EA treatment alleviated cancer-induced pain behaviors covering mechanical allodynia, thermal hyperalgesia and spontaneous pain, and also down-regulated immunofluorescence expressions of neuropeptide CGRP and p75 in the skin of affected plantar area in xenograft mice, and inhibited expressions of overexpressed neuropeptide-related and inflammation-related protein in the lumbar spinal cord of xenograft mice. Overall, our findings suggest that EA treatment ameliorated cancer-induced pain behaviors in the mouse xenograft model of cancer pain, possibly through inhibiting the expressions of neuropeptide-related and inflammation-related protein in central level following tumor cell xenografts.

Insular cortex stimulation alleviates neuropathic pain through changes in the expression of collapsin response mediator protein 2 involved in synaptic plasticity.

In recent studies, brain stimulation has shown promising potential to alleviate chronic pain. Although studies have shown that stimulation of pain-related brain regions can induce pain-relieving effects, few studies have elucidated the mechanisms of brain stimulation in the insular cortex (IC). The present study was conducted to explore the changes in characteristic molecules involved in pain modulation mechanisms and to identify the changes in synaptic plasticity after IC stimulation (ICS). Following ICS, pain-relieving behaviors and changes in proteomics were explored. Neuronal activity in the IC after ICS was observed by optical imaging. Western blotting was used to validate the proteomics data and identify the changes in the expression of glutamatergic receptors associated with synaptic plasticity. Experimental results showed that ICS effectively relieved mechanical allodynia, and proteomics identified specific changes in collapsin response mediator protein 2 (CRMP2). Neuronal activity in the neuropathic rats was significantly decreased after ICS. Neuropathic rats showed increased expression levels of phosphorylated CRMP2, alpha amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor (AMPAR), and N-methyl-d-aspartate receptor (NMDAR) subunit 2B (NR2B), which were inhibited by ICS. These results indicate that ICS regulates the synaptic plasticity of ICS through pCRMP2, together with AMPAR and NR2B, to induce pain relief.