Objective To explore the mechanism of hyperalgesia priming(HP)and evaluate the intervention effect of electroacupuncture(EA)on pain transition.Methods Fifty-nine male C57 mice were used for 4 separate experiments.In the first experiment,the mice were divided into a control group,a sham sensitization group,and a sensitization group,with 5 mice per group.In the second experiment,the mice were divided into a sham sensitization group,a sensitization group,and a sensitization+EA group,with 5 mice per group.The third experiment consisted of a sensitization group and a sensitization+minocycline group,with 7 mice per group.In the fourth experiment,the mice were divided into a sensitization group,a sensitization+EA+PBS group,and a sensitization+EA+colony-stimulating factor(CSF)group,with 5 mice per group.A HP mouse model was established via consecutive intraplantar injections of carrageenan and prostaglandin E2.EA was applied to bilateral"Zusanli"(ST36)and"Kunlun"(BL60)for 30 min,once daily for 8 d.Mechanical paw withdrawal thresholds(PWTs)were measured at different time points to assess pain transition and the effect of EA.Immunofluorescence was used to detect the activation of microglia marker(Iba1)and astrocyte marker(GFAP)in the spinal cord dorsal horn.Intrathecal injection of minocycline was also administered,and its impacts on Iba1 activation and PWTs were tested.After the experiment,skeleton and Sholl analyses were performed to analyze the morphology of microglia.To further examine the role of CSF-1,the factor was administered intrathecally to assess its influence on the effects of EA.Results In the pain transition mouse model,the PWTs at various time points decreased significantly(P<0.01).After modeling,the microglia was activated(P<0.01),but the astrocyte was not.EA increased the PWTs in pain transition mice(P<0.01)and suppressed microglia activation(P<0.01).Intrathecal minocycline administration inhibited microglia activation,reduced Iba1 expression in the spinal cord dorsal horn(P<0.01),and effectively reversed the PWTs reduction(P<0.01).Subsequent intrathecal CSF-1 injection promoted microglia activation and blocked the analgesic effect of EA in the model mice(P<0.01).Conclusion Microglia activation is critical in pain transition.EA can alleviate hyperalgesia in HP mice by inhibiting microglia activation.

The spinoparabrachial tract is a major ascending pathway that transmits nociceptive signals from the spinal cord to the brain, but whether electroacupuncture (EA) relieves pain-anxiety comorbidity by modulating this pathway remains unclear. Here, we investigated the effects of EA on mechanical allodynia and anxiety-like behaviors in Complete Freund's adjuvant (CFA)-induced inflammatory pain mice using von Frey testing, the open field test, and the elevated plus maze, combined with chemogenetic manipulation of glutamatergic projections from the spinal dorsal horn (SDH) to the lateral parabrachial nucleus (LPBN). EA attenuated CFA-induced mechanical allodynia and anxiety-like behaviors and reduced c-Fos expression in both the SDH and LPBN. Chemogenetic inhibition of LPBN glutamatergic neurons mimicked the analgesic and anxiolytic effects of EA, whereas their activation abolished these effects. Activation of either ipsilateral or contralateral SDH→LPBN glutamatergic projections induced pain- and anxiety-like phenotypes that depended on intact LPBN glutamatergic output. Moreover, selective activation of spinoparabrachial glutamatergic terminals was sufficient to block the therapeutic efficacy of EA. These findings indicate that EA alleviates CFA-induced pain-anxiety comorbidity by inhibiting the SDH-LPBN glutamatergic circuit, identifying a key ascending pathway through which peripheral EA signals modulate central pain and affective processing.

Chemotherapy-induced nausea and vomiting (CINV) remain difficult to control after discharge despite guideline-based antiemetics. This multicenter exploratory single-arm two-stage trial evaluated patient-controlled, real-time transcutaneous electrical acupoint stimulation (TEAS) at Neiguan (PC6) as an adjunct to standard antiemetic therapy in patients with breast cancer. Forty-eight patients who had experienced CINV and were scheduled at enrollment to receive the same chemotherapy and antiemetic regimens during two consecutive cycles were analyzed. Cycle 1 served as the observation period, and cycle 2 added on-demand wrist-worn TEAS initiated by patients at nausea or vomiting onset during 0–120 h after chemotherapy. Compared with the observation period, TEAS was associated with lower nausea frequency and visual analogue scale scores on days 1–5 and higher nausea response rates on days 1–4. Vomiting frequency and severity were lower on days 1–3, but between-period differences were not significant on days 4–5. SF-36 scores increased and SAS/SDS scores decreased on day 5. Two transient local numbness events occurred, with no serious TEAS-related adverse events. These findings suggest that patient-controlled TEAS is feasible and warrants confirmation in randomized sham-controlled trials.

Objective To observe the effect of electro-acupuncture(EA)at different acupoints on chronic inflammatory pain-anxiety comorbidity in mice,and to investigate the effect of naloxone on EA induced activation of neurons of the anterior cingulate cortex(ACC),medial prefrontal cortex(mPFC),paraventricular nucleus of the thalamus(PVT),and mediodorsal thalamus nucleus(MD),so as to explore whether the anti-anxiety effect of EA depends on its analgesic mechanism.Methods In the first part of the experiment,the efficacy of EA at different acupoints was compared.Sixty male C57BL/6 mice were randomly divided into control,model,EA Zusanli(ST36),EA Baihui(GV20),and EA Shengmen(HT7)groups(12 mice/group).In the second part,the impact of naloxone on the analgesic and anxiety effects of EA at ST36 was investigated.Forty male C57BL/6 mice were randomly divided into control,model,EA ST36,and EA ST36+naloxone groups(10 mice/group).Except for those in the control group,all the mice were subjected to a chronic inflammatory pain-anxiety comorbidity by subcutaneous injection of 20 μL emulsified complete Freund's adjuvant(CFA)into the hind paw.Mice in the control group received an equivalent volume of PBS.EA(2 Hz/100 Hz,0.2-0.4 mA)intervention was initiated on the 12th day after modeling,and applied to"Zusanli"(ST36,bilateral)or"Baihui"(GV20),or"Shengmen"(HT7,bilateral).The EA intervention was conducted for 30 min,once daily for 6 d.Naloxone was intraperitoneally injected 30 min before EA intervention.Paw withdrawal thresholds(PWTs)were measured using von Frey filaments.The anxiety-like behaviors were assessed using the open field(OF)test and elevated plus maze(EPM)test,separately.The expression of c-Fos in the ACC,mPFC,PVT,and MD was detected by immunofluorescence staining.Results Compared with the control group,the model group exhibited a significant decrease in the PWTs,number of entries into the open arms and time spent in the open arms in the EPM test,and in the number of entries into the central area,time spent in the central area,and distance traveled in the central area in the OF test(P<0.01),and a significant increase in the number of c-Fos positive neurons in the bilateral ACC,mPFC and MD(P<0.05).The total distance traveled of OF test remained unchanged.Compared with the model group,1)EA at ST36,GV20,and HT7 significantly increased the PWTs(P<0.01,P<0.05),2)EA at ST36 significantly increased the number of entries into the open arms,time spent in the open arms in the EPM test,and increased the number of entries,time spent,and distance traveled in the central area in the OF test(P<0.01 P<0.05),3)EA at GV20 significantly increased the time spent in the open arms in the EPM test(P<0.01),4)EA at HT7 obviously increased the time spent in the open arms of the EPM test and increased the time spent in the central area of the OF test(P<0.01),and 5)EA at ST36 significantly decreased the expression of c-Fos in the bilateral ACC,contralateral mPFC,bilateral PVT,and bilateral MD(P<0.05).Compared with the EA ST36 group,the EA ST36+naloxone group showed a significant decrease in the PWTs(P<0.01),number of entries into the open arms,total distance traveled and time spent in the open arms in the EPM test(P<0.05),and in the number of entries into the central area and time spent in the central area in the OF test(P<0.01).The expression levels of c-Fos in the bilateral ACC and ipsilateral MD were strikingly higher in the EA ST36+naloxone group than those in the EA ST36 group(P<0.05),suggesting a disappearance of the suppressive effect of EA after administration of naloxone.Conclusion EA at ST36 has a better analgesic and emotional relief effects on mice with comorbid pain and emotion compared to EA at GV20 and HT7.The underlying mechanism may involve the inhibition of neuronal excitability in the ACC,mPFC,MD and PVT.Additionally,the emotional relief effect of EA at ST36 is dependent on its analgesic efficacy,which may be related to the inhibition of neuronal excitability in the ACC and MD.

BACKGROUND:The analgesic effects of multiple electroacupuncture (EA) sessions and single EA sessions differ significantly in pain management. Area 24b (A24b) of the anterior cingulate cortex (ACC) is crucial in pain processing. EA relieves pain by targeting and modulating the neuronal activity within this subregion. However, whether the cumulative effect of EA antinociception is connected to A24b mechanisms has remained unclear. METHODS:In our study, we used the Complete Freund's Adjuvant (CFA) model to induce inflammatory pain and the Spared Nerve Injury (SNI) model to induce neuropathic pain, and adult male C57BL/6, FosTRAP, and FosTRAP:Ai9 mice were used as experimental subjects to investigate the cumulative effect of EA antinociception and whether multiple EA sessions and a single EA session regulate different neuronal populations in the A24b. RESULTS:We observed that EA effectively alleviated pain in mice, with three EA sessions yielding superior analgesic effects compared to a single session. Using chemical genetics combined with FosCreER technology to activate EA-TRAPed cells in the A24b, we found that pain relief was more pronounced with three EA sessions. Moreover, chemogenetic inhibition of EA-TRAPed cells in the A24b reversed the analgesic effects of a single EA session but not those of three EA sessions. Fluorescent in situ hybridization results indicated that three EA sessions significantly increased the number of GABAergic neurons in the A24b compared with a single session. Additionally, retrograde tracing revealed that the A24b circuit that monosynaptically innervates EA-TRAPed cells included projections from the central lateral nucleus (CL), lateral mediodorsal thalamic nucleus (MDL), lateral habenula (LHb), dorsal raphe nucleus (DR), caudal linear nucleus of the raphe (CLi), dorsal tuberomamillary nucleus (DTM), periventricular hypothalamic nucleus (Pe) and hippocampal fields CA1, CA2, and CA3. These findings suggest that multiple EA sessions and single EA sessions activated different neuronal populations in the A24b. The enhanced analgesic effect of multiple EA sessions may be attributed to an increase in the proportion of GABAergic neurons within the A24b. CONCLUSIONS:Multiple and single EA sessions recruit distinct neuronal populations in A24b, with the stronger analgesic effect of repeated EA linked to a higher proportion of GABAergic neurons in this region.

Postoperative pain develops after surgical operation. Inadequately managed postoperative pain is oftentimes associated with deterioration in life quality and delayed rehabilitation. Clinical studies confirmed analgesic effects of acupuncture on patients with postoperative pain. We hereby explored mechanisms underlying how acupuncture alleviates postoperative pain. A mouse model of skin incision-induced pain was established to mimic postoperative pain condition. Electroacupuncture (EA) or sham EA was administered at ST36 and BL60 acupoints in model mice. A combination of RNA-Sequencing (RNA-Seq), immunostaining, biochemical assay, in vivo imaging and behavioral test were applied for mechanism investigations. 2/100 Hz EA intervention ameliorated mechanical allodynia and improved cumulative pain of incisional pain model mice. Sham EA exerted no obvious analgesic effect. Incisional pain model mice develop gait impairment, which was improved by EA intervention. RNA-Seq identified EA significantly reduced Il33 gene overexpression in the incised skin tissues of model mice. IL-33 was produced from keratinocytes upon skin incision. EA reduced IL-33 overproduction from keratinocytes, resulting in less macrophage infiltration and less ROS accumulation in the incised skin. Adoptive transfer of macrophages into the incised tissue abrogated the ameliorative effects of EA on macrophage infiltration as well as ROS accumulation in incised skin and further reversed EA-induced analgesia on incisional pain model mice. Additionally, EA intervention did not affect skin wound healing process. These results demonstrate that EA ameliorates incisional pain via suppressing IL-33 overproduction in incised skin tissue. EA-induced suppression of IL-33 overproduction results in less macrophage infiltration and ROS accumulation that contribute to analgesia. Our work helps to support EA as an alternative and green therapy for postoperative pain management.

Objective To observe the effect of electro-acupuncture (EA) at different acupoints on chronic inflammatory pain-anxiety comorbidity in mice, and to investigate the effect of naloxone on EA induced activation of neurons of the anterior cingulate cortex (ACC), medial prefrontal cortex (mPFC), paraventricular nucleus of the thalamus (PVT), and mediodorsal thalamus nucleus (MD), so as to explore whether the anti-anxiety effect of EA depends on its analgesic mechanism. Methods In the first part of the experiment, the efficacy of EA at different acupoints was compared. Sixty male C57BL/6 mice were randomly divided into control, model, EA Zusanli (ST36), EA Baihui (GV20), and EA Shengmen (HT7) groups (12 mice/group). In the second part, the impact of naloxone on the analgesic and anxiety effects of EA at ST36 was investigated. Forty male C57 BL/6 mice were randomly divided into control, model, EA ST36, and EA ST36 + naloxone groups (10 mice/group). Except for those in the control group, all the mice were subjected to a chronic inflammatory pain-anxiety comorbidity by subcutaneous injection of 20 u03BCL emulsified complete Freundu2019s adjuvant (CFA) into the hind paw. Mice in the control group received an equivalent volume of PBS. EA (2 Hz/100 Hz, 0.2u20140.4 mA) intervention was initiated on the 12th day after modeling, and applied to u201CZusanliu201D (ST36, bilateral) or u201CBaihuiu201D (GV20), or u201CShengmenu201D (HT7, bilateral). The EA intervention was conducted for 30 min, once daily for 6 d. Naloxone was intraperitoneally injected 30 min before EA intervention. Paw withdrawal thresholds (PWTs) were measured using von Frey filaments. The anxiety-like behaviors were assessed using the open field (OF) test and elevated plus maze (EPM) test, separately. The expression of c-Fos in the ACC, mPFC, PVT, and MD was detected by immunofluorescence staining. Results Compared with the control group, the model group exhibited a significant decrease in the PWTs, number of entries into the open arms and time spent in the open arms in the EPM test, and in the number of entries into the central area, time spent in the central area, and distance traveled in the central area in the OF test (Pu0026lt;0.01), and a significant increase in the number of c-Fos positive neurons in the bilateral ACC, mPFC and MD (Pu0026lt;0.05). The total distance traveled of OF test remained unchanged. Compared with the model group, 1) EA at ST36, GV20, and HT7 significantly increased the PWTs (Pu0026lt;0.01, Pu0026lt;0.05), 2) EA at ST36 significantly increased the number of entries into the open arms, time spent in the open arms in the EPM test, and increased the number of entries, time spent, and distance traveled in the central area in the OF test (Pu0026lt;0.01 Pu0026lt;0.05), 3) EA at GV20 significantly increased the time spent in the open arms in the EPM test (Pu0026lt;0.01), 4) EA at HT7 obviously increased the time spent in the open arms of the EPM test and increased the time spent in the central area of the OF test (Pu0026lt;0.01), and 5) EA at ST36 significantly decreased the expression of c-Fos in the bilateral ACC, contralateral mPFC, bilateral PVT, and bilateral MD (Pu0026lt;0.05). Compared with the EA ST36 group, the EA ST36 + naloxone group showed a significant decrease in the PWTs (Pu0026lt;0.01), number of entries into the open arms, total distance traveled and time spent in the open arms in the EPM test (Pu0026lt;0.05), and in the number of entries into the central area and time spent in the central area in the OF test (Pu0026lt;0.01). The expression levels of c-Fos in the bilateral ACC and ipsilateral MD were strikingly higher in the EA ST36 + naloxone group than those in the EA ST36 group (Pu0026lt;0.05), suggesting a disappearance of the suppressive effect of EA after administration of naloxone. Conclusion EA at ST36 has a better analgesic and emotional relief effects on mice with comorbid pain and emotion compared to EA at GV20 and HT7. The underlying mechanism may involve the inhibition of neuronal excitability in the ACC, mPFC, MD and PVT. Additionally, the emotional relief effect of EA at ST36 is dependent on its analgesic efficacy, which may be related to the inhibition of neuronal excitability in the ACC and MD.

IntroductionThis bibliometric study systematically analyzes the research landscape of electroacupuncture (EA), focusing on its applications in pain relief and emotional regulation from 2014 to 2024. EA, a contemporary adaptation of traditional acupuncture, has gained significant attention for its potential therapeutic benefits in managing chronic pain and mood disorders.MethodsUsing the Web of Science Core Collection as the primary data source, we identified 537 articles related to EA’s therapeutic effects. The analysis was conducted using bibliometric tools such as VOSviewer and CiteSpace to visualize publication trends, research hotspots, and collaborative networks.ResultsThe study highlights a significant upward trend in research output, with a marked increase in publications from 2019 onwards. China emerged as the leading contributor, accounting for over 60% of the total research output, followed by the United States and South Korea. Key institutions, such as Zhejiang Chinese Medical University and Shanghai University of Traditional Chinese Medicine, have made substantial contributions, emphasizing the importance of traditional Chinese medicine in this research area. Major research themes include the modulation of neurotransmitter systems, the role of endogenous opioids, and the impact of EA on chronic pain and mood disorders. Collaborative networks between countries and institutions are mapped, revealing the centrality of Chinese and American research partnerships.DiscussionThis comprehensive analysis outlines the current state of EA research and identifies gaps and opportunities for future studies, particularly in understanding the mechanistic pathways of EA and its integration into mainstream medical practices. The findings provide a roadmap for enhancing the therapeutic applications of EA and underscore its potential in managing complex conditions involving both physical and emotional components.

Pain is a complex physiological phenomenon driven by dynamic microglial(MG) regulation in the CNS. Aberrant microglial activation mediates neuroinflammation, neuronal hyperexcitability, and synaptic plasticity, exhibiting spatiotemporal and sex-specific heterogeneity. While early pharmacological interventions show time-limited efficacy, electroacupuncture (EA) emerges as a potent non-pharmacological strategy that disrupts microglia-neuron crosstalk via multiple pathways. Critically, EA's efficacy depends on intervention timing, with early application preventing microglial active in neuropathic models. Recent advances reveal CD11c+ microglia as dual regulators of pain resolution and relapse-clearing myelin debris acutely but driving recurrence via MMP-9-mediated synaptic remodeling. This review synthesizes microglial heterogeneity, stage-specific interventions, and EA's translational potential in rebalancing microglial phenotypes for precision pain management.

Evidence indicates that the interplay between pain and anxiety poses clinical challenges for the evaluation and management of chronic pain, yet effective therapies for these comorbidities are limited. This study aimed to elucidate the effects and mechanisms of electroacupuncture (EA) on pain-anxiety comorbidities. Mice injected with Complete Freund's adjuvant (CFA) in the ipsilateral hind paw developed persistent inflammatory pain and anxiety-like behaviors, as assessed by the von Frey, open field, elevated plus maze, and novelty-suppressed feeding tests. EA was administered 12-17d after CFA injection with once daily. rAAV virus and chemogenetics were used to manipulate parvalbumin (PV) interneurons and astrocytes excitation in the anterior cingulate cortex (ACC). Immunofluorescence, morphological analysis, patch clamp and in vivo fiber Ca2+ imaging were used to examine the activation of PV interneurons and astrocytes. The effect of EPCPX (antagonist of A1R) and chemogenetics activated astrocytes on EA analgesia were observed in a subset of mice prior to EA. EA administration alleviated pain and anxiety-like behaviors in CFA mice, activated PV interneurons, and inhibited astrocytes activation in the ACC. Furthermore, both PV interneurons activation and astrocyte inhibition in the ACC elicited effects similar to those elicited by EA on pain and anxiety. Chemogenetic activation of ACC astrocytes reversed the effects of EA. Additionally, astrocyte activation in the ACC suppressed PV interneurons and induced pain-anxiety like behaviors in mice. Adenosine A1 receptors, crucial for mediating the interaction between astrocytes and PV interneurons in the ACC, were also found to be involved in the effects of EA on pain-anxiety comorbidity. These findings reveal that EA alleviates the pain and anxiety comorbidity through a potential mechanism involving the activation of PV interneurons, which are modulated by the inhibition of astrocytes in the ACC, thus providing a promising therapeutic strategy for persistent pain and concurrent anxiety.

Accumulating evidence highlights the anti-inflammatory and analgesic effects of electroacupuncture (EA), yet the underlying mechanisms remain poorly understood. The P2X7 purinergic receptor (P2X7R), located in the peripheral and central nervous systems, has been implicated in the development of chronic inflammatory pain. Inhibition of P2X7R expression has been associated with the analgesic effects of EA. Within the dorsal root ganglia, P2X7R is exclusively expressed in satellite glial cells (SGCs), but its role in the anti-inflammatory effects of EA remains to be elucidated. A chronic inflammatory pain model was established in rats via intraplantar injection of Complete Freund’s Adjuvant (CFA). Western blotting, immunostaining, behavioral assay, pharmacological interventions, AAV-mediated knockdown assays, SGCs culture and real-time cell proliferation analysis were utilized to investigate the cellular mechanisms underlying the effects of EA at the ST36 and BL60 acupuncture points in mitigating inflammatory pain. Rats injected with CFA exhibited long-lasting pain hypersensitivity in the ipsilateral hind paw, accompanied by upregulated expression of P2X7R and TNF-α in the L4-6 DRGs. Pharmacological inhibition or shRNA-mediated knockdown of P2X7R in SGCs significantly attenuated inflammatory pain hypersensitivity, reduced P38 MAPK phosphorylation, and decreased TNF-α levels. EA treatment significantly alleviated CFA-induced pain hypersensitivity and suppressed P2X7R expression alongside its downstream P38 MAPK/TNF-α signaling. The analgesic effects of EA were reversed by the P2X7R agonist BzATP. In vitro findings confirmed that TNF-α secretion by SGCs was markedly elevated in the CFA model but reduced by EA treatment, mimicking the effects of P2X7R antagonism. These findings demonstrate that the inhibition of P2X7R in SGCs and its downstream P38 MAPK/TNF-α signaling pathway contributes to the analgesic effects of EA in chronic inflammatory pain. Targeting peripheral P2X7R in SGCs may provide new insights into the cellular mechanisms of EA’s anti-inflammatory effects.

BackgroundAdolescent insomnia is a prevalent issue with significant implications for mental and physical well-being. Given the increasing incidence of sleep disorders, there is an urgent need for safe and efficient treatment modalities, particularly non-pharmacological interventions. Auricular point therapy, a prominent element of acupuncture in China, is often employed to address insomnia. However, there remains a lack of research on the efficacy of auricular point-vagus nerve stimulation in managing insomnia among teenagers. To evaluate the efficacy of this interventions and explore the regulatory mechanisms of brain function, a randomized trial is planned.Methods/designThis trial is a single-center, single-blind, randomized controlled study. A total of 174 adolescent patients with insomnia will be randomly assigned to either the treatment or control groups. Over a period of 4 weeks, patients in the experimental group will undergo bilateral auricular point-vagus nerve stimulation, while those in the control group will receive bilateral non-auricular acupoint-vagus nerve stimulation. Data collection will occur at baseline, 1 week into the intervention, 4 weeks post-treatment initiation, and 4 weeks post-treatment completion. The primary outcome measures will include the Pittsburgh Sleep Quality Index (PSQI), Adolescent Sleep Hygiene Habits Scale (ASHS), and Adolescent Sleep Assessment Questionnaire (DSM). Secondary outcome measures encompass the Self-Assessment Scale for Anxiety (SAS), Self-Depression Scale (SDS), Quality of Life Scale (SF-36), and Autonomic Composite Score (COMPASS-31). Other indicators will involve the use of functional magnetic resonance imaging (fMRI), heart rate variability (HRV), and polysomnography (PSG).DiscussionThe findings of this study will provide strong support for the use of auricular-vagus nerve stimulation as a safe and effective non-pharmacological intervention for insomnia. This approach will offer advantages over medication by mitigating the risks of addiction and adverse effects associated with prolonged drug therapy. Furthermore, the results will align with prior research that underscores the positive impact of auricular acupuncture point stimulation on enhancing sleep quality.Clinical trial registrationhttps://www.chictr.org.cn, ChiCTR2400087889.

BackgroundPaclitaxel-induced peripheral neuropathy (PIPN) is prevalent among patients receiving paclitaxel chemotherapy, which results in sensory abnormality as well as neuropathic pain. Conventional medications lack effectiveness on PIPN. Clinical trials identified beneficial effects of acupuncture on PIPN among patients receiving chemotherapy. Here we explored the mechanisms underlying how acupuncture might alleviate PIPN.MethodsA mouse model of PIPN was established by repeated paclitaxel application. Electroacupuncture (EA) was applied at ST36 and BL60 acupoints of model mice. Immunostaining, flow cytometry, behavioral assay, in vivo imaging were utilized for effects determination and mechanism exploration.ResultsEA ameliorated mechanical and cold pain hypersensitivities, reduced sensory neuron damage and improved loss in intra-epidermal nerve fibers (IENFs) in model mice. Macrophages infiltration were detected in DRG and sciatic nerve of model mice, which was reduced by EA. EA affected M1-like pro-inflammatory macrophage infiltration in DRG, whereas it did not affect M2-like macrophages. DRG neurons released chemoattractant CCL2 that recruited macrophages via CCR2 to DRG. EA reduced CCL2 overproduction by DRG neurons and reduced macrophage infiltration. Blocking CCR2 mimicked EA's anti-allodynic effect, whereas exogenously applying recombinant CCL2 reversed the ameliorative effect of EA on macrophage infiltration and abolished EA's anti-allodynia on model mice. EA ameliorated other signs of PIPN, including sensory neuron damage, sciatic nerve morphology impairment and IENFs loss. In mice inoculated with breast cancer cells, EA didn't affect paclitaxel-induced antitumor effect.ConclusionsThese findings suggest EA alleviates PIPN by reducing CCL2/CCR2 mediated-pro-inflammatory macrophage infiltration into sensory ganglia as well as the sciatic nerve. Our study supports EA could be used as a potential non-pharmacological therapy for PIPN.

Objective:To explore the peripheral neural mechanism underlying representation along the distribution of stomach meridian induced by intestinal inflammatory reaction using diarrhea predominant-irritable bowel syndrome (IBS-D) mice. Methods:Among 62 healthy male C57BL/6 mice of clean grade, 12 mice were randomly selected and divided into a control group and a model group, 6 mice in each group, additionally, 12 mice were randomly selected and divided into a Tianshu group, a Liangqiu group and a Zusanli group, 4 mice in each group. In the model group, citrobacter was administered orally to establish IBS-D model. In the control group and the model group, the visceral pain threshold was observed using fecal colorectal distension (fCRD) induced electromyography of external oblique muscle, the positive cell number of neutrophil in the colonic muscularis was detected by myeloperoxidase (MPO) staining, the number, location and distribution rule of Evans blue (EB) extravasation points were observed by injection of EB staining solution into the tail vein. In the Tianshu group, the Liangqiu group and the Zusanli group, fluorescent dye Dil was injected at bilateral "Tianshu" (ST25), "Liangqiu" (ST34) and "Zusanli" (ST36) respectively, to observe the dye-positive cell number in different dorsal root ganglion (DRG) segments. In the control group and the model group, the activation of satellite glial cells (SGCs) in different DRG segments was observed by immunofluorescence. Results:Compared with the control group, in the model group, the area under curve of electromyography of external oblique muscle was increased at fCRD of 25, 50 and 75 μL distilled water (P<0.001, P<0.01); the MPO-positive cell number of neutrophil in the colonic muscularis was increased (P<0.01). Few EB extravasation points could be found in the control group, while there were much more EB extravasation points observed in the model group, which was specially distribution in the area of stomach meridian, from "Huaroumen" (ST24) to "Zusanli" (ST36), as well as the surface area dominated by L2-L5 segment of the spinal cord. The Dil-positive cells were mainly exhibited in the DRG of T11, L5 and L4 segments in the Tianshu group, the Liangqiu group and the Zusanli group, respectively. Compared with the control group, the ratio of glial fibrillary acidic protein (GFAP)/glutamine synthetase (GS) co-expression was increased in the DRG of T11, L4 and L5 segments in the model group (P<0.05, P<0.01). Conclusion:The activation of SGCs within DRG of T11, L4 and L5 segments may relate closely to the occurrence of the representation along the stomach meridian distribution in IBS-D mice.

Emotional and cognitive impairments are comorbidities commonly associated with chronic inflammatory pain. To summarize the rules and mechanisms of comorbidities in a complete Freund’s adjuvant (CFA)-induced pain model, we conducted a systematic review of 66 experimental studies identified in a search of three databases (PubMed, Web of Science, and ScienceDirect). Anxiety-like behaviors developed at 1- or 3-days post-CFA induction but also appeared between 2- and 4 weeks post-induction. Pain aversion, pain depression, and cognitive impairments were primarily observed within 2 weeks, 4 weeks, and 2–4 weeks post-CFA injection, respectively. The potential mechanisms underlying the comorbidities between pain and anxiety predominantly involved heightened neuronal excitability, enhanced excitatory synaptic transmission, and neuroinflammation of anterior cingulate cortex (ACC) and amygdala. The primary somatosensory cortex (S1)Glu→caudal dorsolateral striatum (cDLS)GABA, medial septum (MS)CHAT→rACC, rACCGlu→thalamus, parabrachial nucleus (PBN)→central nucleus amygdala (CeA), mediodorsal thalamus (MD)→basolateral amygdala (BLA), insular cortex (IC)→BLA and anteromedial thalamus nucleus (AM)CaMKⅡ→midcingulate cortex (MCC)CaMKⅡ pathways are enhanced in the pain-anxiety comorbidity. The ventral hippocampal CA1 (vCA1)→BLA and BLA→CeA pathways were decreased in the pain-anxiety comorbidity. The BLA→ACC pathway was enhanced in the pain-depression comorbidity. The infralimbic cortex (IL)→locus coeruleus (LC) pathway was enhanced whereas the vCA1→IL pathway was decreased, in the pain-cognition comorbidity. Inflammation/neuroinflammation, oxidative stress, apoptosis, ferroptosis, gut-brain axis dysfunction, and gut microbiota dysbiosis also contribute to these comorbidities.

Diabetic neuropathic pain (DNP) is a diabetic complication that causes severe pain and deeply impacts the quality of the sufferer's daily life. Currently, contemporary clinical treatments for DNP generally exhibit a deficiency in effectiveness. Electroacupuncture (EA) is recognized as a highly effective and safe treatment for DNP with few side effects. Regrettably, the processes via which EA alleviates DNP are still poorly characterized. Transient receptor potential vanilloid 1 (TRPV1) and phosphorylated calcium/calmodulin-dependent protein kinase II (p-CaMKII) are overexpressed on spinal cord dorsal horn (SCDH) in DNP rats, and co-localization is observed between them. Capsazepine, a TRPV1 antagonist, effectively reduced nociceptive hypersensitivity and downregulated the overexpression of phosphorylated CaMKIIα in rats with DNP. Conversely, the CaMKII inhibitor KN-93 did not have any impact on TRPV1. EA alleviated heightened sensitivity to pain caused by nociceptive stimuli and downregulated the level of TRPV1, p-CaMKIIα, and phosphorylated cyclic adenosine monophosphate response element-binding protein (p-CREB) in DNP rats. Intrathecal injection of capsaicin, on the other hand, reversed the above effects of EA. These findings indicated that the CaMKII/CREB pathway on SCDH is located downstream of TRPV1 and is affected by TRPV1. EA alleviates DNP through the TRPV1-mediated CaMKII/CREB pathway.

Comorbid chronic neuropathic pain and anxiety is a common disease that represents a major clinical challenge. The underlying mechanisms of chronic neuropathic pain and anxiety are not entirely understood, which limits the exploration of effective treatment methods. Glutamatergic neurons in the ventrolateral periaqueductal gray (vlPAG) have been implicated in regulating pain, but the potential roles of the vlPAG in neuropathic pain-induced anxiety have not been investigated. Herein, whole-cell recording and immunofluorescence showed that the excitability of CamkIIα neurons in the vlPAG (vlPAG CamkIIα+ neurons) was decreased in mice with spared nerve injury (SNI), while electroacupuncture (EA) activated these neurons. We also showed that chemogenetic inhibition of vlPAG CamkIIα+ neurons resulted in allodynia and anxiety-like behaviors in naive mice. Furthermore, chemogenetic activation of vlPAG CamkIIα+ neurons reduced anxiety-like behaviors and allodynia in mice with SNI, and EA had a similar effect in alleviating these symptoms. Nevertheless, EA combined with chemogenetic activation failed to further relieve allodynia and anxiety-like behaviors. Artificial inhibition of vlPAG CamkIIα+ neurons abolished the analgesic and anxiolytic effects of EA. Overall, our study reveals a novel mechanism of neuropathic pain-induced anxiety and shows that EA may relieve comorbid chronic neuropathic pain and anxiety by activating vlPAG CamkIIα+ neurons. Significance Statement Neuropathic pain is clinically accompanied by anxiety. Both glutamatergic neurons in the ventrolateral periaqueductal gray (vlPAG) and electroacupuncture (EA) have demonstrated analgesic properties. However, the efficacy of these interventions in addressing neuropathic pain and its concomitant anxiety has yet to be fully elucidated. In a mice model of spared nerve injury (SNI), we observed a decreased excitability of vlPAG CamkIIα neurons. Remarkably, EA treatment significantly enhanced the excitability of these neurons. Further, chemogenetic activation of vlPAG CamkIIα+ neurons not only resulted in analgesia but also mitigated anxiety-like behaviors in SNI mice, mirroring the effects observed with EA treatment. Conversely, inhibition of vlPAG CamkIIα+ neurons activity in naive mice reduced pain thresholds and induced anxiety-like behavior, while also negating the beneficial effects of EA. These findings provide novel insights into the mechanistic interplay between chronic neuropathic pain and anxiety, highlighting the therapeutic potential of targeting vlPAG glutamatergic neurons in these conditions.

Excessive deposition of monosodium urate (MSU) crystal in the joint results in gout arthritis, which triggers severe pain and affects life quality. Oxidative stress is a pivotal mechanism that contributes to etiology of gout pain and inflammation. Here we investigated whether activating Nrf2, which plays important roles in regulating endogenous antioxidant response, would attenuate gout arthritis via promoting antioxidant signaling in joint tissues. Gout arthritis model was established by intra-articular injection of MSU (500 μg/ankle) into the right ankle joint of mouse. Pharmacologically activating Nrf2 by activator oltipraz (50, 100 or 150 mg/kg, intraperitoneal) at 1 h before and 5, 23, 47 h after model establishment dose-dependently inhibited joint inflammation, mechanical and heat hypersensitivities in model mice. Oltipraz (100 mg/kg) reversed gait impairments without altering locomotor activity and reduced neutrophil infiltrations in ankle joints. In vitro studies revealed oltipraz (25 μM) inhibited MSU-induced ROS production in mouse macrophages and improved mitochondrial bioenergetics impairments caused by MSU. In vivo ROS imaging combined with biochemical assays confirmed the antioxidant effects of oltipraz on model mice. Nrf2 activation inhibited pro-inflammatory cytokine overproduction in ankle joint and attenuated the overexpression and enhancement in TRPV1 channel in DRG neurons innervating hind limb. Therapeutic effects of oltipraz were abolished by inhibiting Nrf2 or in Nrf2 knockout mice. These results suggest pharmacologically activating Nrf2 alleviates gout pain, gait impairments, inflammation and peripheral sensitization via Nrf2-dependent antioxidant mechanism. Targeting Nrf2 may represent a novel treatment option for gout arthritis.

Objective: Electroacupuncture (EA) is an alternative treatment option for pain. Different frequencies of EA have different pain-relieving effects; however, the central mechanism is still not well understood. Methods: The Fos2A-iCreER (TRAP):Ai9 mice were divided into three groups (sham, 2 Hz, and 100 Hz). The mice were intraperitoneally injected with 4-hydroxytamoxifen (4-OHT) immediately after EA at Zusanli (ST36) for 30 min to record the activated neurons. One week later, the mice were sacrificed, and the number of TRAP-treated neurons activated by EA in the thalamus, amygdala, cortex, and hypothalamus was determined. Results: In the cortex, 2 Hz EA activated more TRAP-treated neurons than 100 Hz EA did in the cingulate cortex area 1 (Cg1) and primary somatosensory cortex (S1), and 2 and 100 Hz EAs did not differ from sham EA. TRAP-treated neurons activated by 2 Hz EA were upregulated in the insular cortex (IC) and secondary somatosensory cortex (S2) compared with those activated by 100 Hz and sham EA. In the thalamus, the number of TRAP-treated neurons activated by 2 Hz EA was elevated in the paraventricular thalamic nucleus (PV) compared with those activated by sham EA. In the ventrolateral thalamic nucleus (VL), the number of TRAP-treated neurons activated by 2 Hz EA was significantly upregulated compared with those activated by 100 Hz EA, and sham EA showed no difference compared with 2 or 100 Hz EA. TRAP-treated neurons were more frequently activated in the ventral posterolateral thalamic nucleus (VPL) by 2 Hz EA than by 100 Hz or sham EA. Conclusions: Low-frequency EA ST36 effectively activates neurons in the Cg1, S1, S2, IC, VPL, PV, and VL. The enhanced excitability of the aforementioned nuclei induced by low-frequency EA may be related to its superior efficacy in the treatment of neuropathological pain.

Objective:Injury can lead to long-term changes that increase the sensitivity of afferent nerve endings to subsequent stimulation and pain can transition from acute to chronic.This phenomenon is known as hyperalgesic priming (HP).This study aimed to understand the mechanisms underlying the effect of electroacupuncture (EA) on HP and optimize acupoint selection for EA to prevent pain transition. Methods:A rat HP model was established using sequential intraplantar injections of carrageenan (Cg) and prostaglandin E2 (PGE2).The pain thresholds were measured using von Frey filaments.EA on bilateral Zusanli (ST36) and Kunlun (BL60) was used to prevent pain transition.The number of mast cells in the ipsilateral hindpaw skin was determined using toluidine blue or fluorescence-labeled avidin staining.The protein expression levels of protein kinase C epsilon (PKCε) in the lumbar dorsal root ganglions (DRGs) were detected by western blotting 24 h after PGE2 injection.Serial pharmacological experiments were conducted to evaluate the relationship between mast cells and pain transition.Finally,EA on the bilateral ST36 and Chongyang (ST42) or a novel combination (ST36 and ST42 on the ipsilateral side,and ST36 and BL60 on the contralateral side) was used to prevent pain transition. Results:Although EA applied to ST36 and BL60 alleviated acute pain induced by Cg injection,it failed to prevent the pain transition caused by PGE2 injection.Mast cell accumulation in the ipsilateral hind paw was observed 7 days after Cg injection.Furthermore,mast cell degranulation may be responsible for PKCε activation in the DRG,a marker of pain transition.EA significantly decreased the number of mast cells in the skin of the ipsilateral hind paw when applied at ST36 and ST42,but not when applied at ST36 and BL60.Furthermore,EA employed to ST36 and ST42 significantly reversed long-term hyperalgesia induced by PGE2 injection,even when administered before injection.However,EA did not alleviate acute pain caused by Cg injection.By using a novel acupoint combination,EA simultaneously alleviated acute pain and prevented pain transition. Conclusions:Our study suggests that mast cells play a critical role in both HP and the transition from acute to chronic pain,whereas EA can prevent pain transition by decreasing the number of mast cells in the local tissue.