Morphine paradoxically prolongs neuropathic pain in rats by amplifying spinal NLRP3 inflammasome activation

Opioid use for pain management has dramatically increased, with little assessment of potential pathophysiological consequences for the primary pain condition. Here, a short course of morphine, starting 10 d after injury in male rats, paradoxically and remarkably doubled the duration of chronic constriction injury (CCI)-allodynia, months after morphine ceased. No such effect of opioids on neuropathic pain has previously been reported. Using pharmacologic and genetic approaches, we discovered that the initiation and maintenance of this multimonth prolongation of neuropathic pain was mediated by a previously unidentified mechanism for spinal cord and pain—namely, morphine-induced spinal NOD-like receptor protein 3 (NLRP3) inflammasomes and associated release of interleukin-1β (IL-1β). As spinal dorsal horn microglia expressed this signaling platform, these cells were selectively inhibited in vivo after transfection with a novel Designer Receptor Exclusively Activated by Designer Drugs (DREADD). Multiday treatment with the DREADD-specific ligand clozapine-N-oxide prevented and enduringly reversed morphine-induced persistent sensitization for weeks to months after cessation of clozapine-N-oxide. These data demonstrate both the critical importance of microglia and that maintenance of chronic pain created by early exposure to opioids can be disrupted, resetting pain to normal. These data also provide strong support for the recent “two-hit hypothesis” of microglial priming, leading to exaggerated reactivity after the second challenge, documented here in the context of nerve injury followed by morphine. This study predicts that prolonged pain is an unrealized and clinically concerning consequence of the abundant use of opioids in chronic pain.

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Abstract P259: Endothelial Acid Sphingomyelinase Gene Determines Inflammasome Activation and Atherosclerotic Lesions in Carotid Arteries During Hypercholesterolemia

Hypertension ◽

10.1161/hyp.66.suppl_1.p259 ◽

2015 ◽

Vol 66 (suppl_1) ◽

Author(s):

Min Xia ◽

Krishna M Bioni ◽

Yang Chen ◽

Xiang Li ◽

Ashley L Pitzer ◽

...

Keyword(s):

Carotid Arteries ◽

Acid Sphingomyelinase ◽

Receptor Protein ◽

Inflammasome Activation ◽

Neointimal Formation ◽

Atherosclerotic Lesions ◽

Junction Protein ◽

Nlrp3 Inflammasomes

Nucleotide oligomerization domain (NOD)-like receptor protein with pyrin domain containing 3 (Nlrp3) inflammasome has been reported to be activated by atherogenic factors, thereby triggering endothelial injury and consequent atherosclerotic lesions in the arterial wall. However, the mechanism activating and regulating Nlrp3 inflammasomes remains poorly understood. The present study tested whether membrane raft (MR) signaling platforms associated with acid sphingomyelinase (ASM) and its product ceramide (Ce) importantly contribute to the activation of Nlrp3 inflammasomes and atherosclerotic lesions during hypercholesterolemia (HC). By confocal microscopy and biochemical analyses, we demonstrated the formation and activation of Nlrp3 inflammasomes in the intima of the carotid arteries of Asm +/+ mice with HC (as shown by a 2-fold increase in caspase-1 activity and a 6-fold enhancement of IL-1β positive stain areas), but not in Asm -/- mice. In endothelium-specific ASM transgenic mice (EC-Asm trg ), this inflammasome formation and activation were enhanced. Correspondingly, HC-induced increases in IL-1β production, ASM expression, Ce level and MR-gp91 phox clustering in the carotid intima were abolished in Asm -/- mice, but enhanced in EC-Asm trg mice. Functionally, endothelium-dependent vasodilation (EDVD) in carotid arteries in vivo (by ultrasound flowmetry) and in vitro (in perfused artery) was impaired by HC in Asm +/+ mice by 33% and 54%, respectively. This endothelial dysfunction was not observed in Asm -/- mice. The endothelial tight junction protein, ZO-1 was reduced by HC in both Asm +/+ and EC-Asm trg mice, but not in Asm -/- mice. It was also found that HC-increased neointimal formation, T-cell infiltration, and fibrosis in 2-week partially ligated carotid arteries (PLCA) occurred in Asm +/+ mice, but not in Asm -/- mice with HC. EC-Asm trg mice even exhibited more severe inflammatory and atherosclerotic lesions. All these results suggest that Asm gene and related MR clustering are essential to endothelial inflammasome activation and dysfunction in carotid arteries, ultimately determining the extent of atherosclerotic lesions.

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Chemogenetic manipulation of microglia inhibits neuroinflammation and neuropathic pain in mice

10.1101/2020.05.21.109538 ◽

2020 ◽

Author(s):

Min-Hee Yi ◽

Yong U. Liu ◽

Kevin Liu ◽

Tingjun Chen ◽

Dale B. Bosco ◽

...

Keyword(s):

Chronic Pain ◽

Neuropathic Pain ◽

Pain Treatment ◽

Interleukin 1 ◽

Spinal Nerve ◽

Synaptic Function ◽

Designer Drugs ◽

Downstream Target ◽

Potential Opportunity

AbstractMicroglia play an important role in the central sensitization and chronic pain. However, a direct connection between microglial function and the development of neuropathic pain in vivo remains incompletely understood. To address this issue, we applied chemogenetic approach by using CX3CR1creER/+:R26LSL-hM4Di/+ transgenic mice to enable expression of inhibitory Designer Receptors Exclusively Activated by Designer Drugs (Gi DREADD) exclusively in microglia. We found that microglial Gi DREADD activation inhibited spinal nerve transection (SNT)-induced microglial reactivity as well as chronic pain initiation and maintenance. Gi DREADD activation downregulated the transcription factor interferon regulatory factor 8 (IRF8) and its downstream target pro-inflammatory cytokine interleukin 1 beta (IL-1β). Using in vivo spinal cord recording, we found that activation of microglial Gi DREADD attenuated synaptic transmission following SNT. Our results demonstrate that microglial Gi DREADD reduces neuroinflammation, synaptic function and neuropathic pain after peripheral nerve injury. Thus, chemogenetic approaches provide a potential opportunity for interrogating microglial function and neuropathic pain treatment.

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Optogenetic activation of spinal microglia triggers chronic pain in mice

PLoS Biology ◽

10.1371/journal.pbio.3001154 ◽

2021 ◽

Vol 19 (3) ◽

pp. e3001154

Author(s):

Min-Hee Yi ◽

Yong U. Liu ◽

Anthony D. Umpierre ◽

Tingjun Chen ◽

Yanlu Ying ◽

...

Keyword(s):

Chronic Pain ◽

Microglial Activation ◽

Red Light ◽

Inflammasome Activation ◽

Pain Hypersensitivity ◽

Microglial Phenotype ◽

C Fiber ◽

Inward Currents ◽

Calcium Elevation

Spinal microglia are highly responsive to peripheral nerve injury and are known to be a key player in pain. However, there has not been any direct evidence showing that selective microglial activation in vivo is sufficient to induce chronic pain. Here, we used optogenetic approaches in microglia to address this question employing CX3CR1creER/+: R26LSL-ReaChR/+ transgenic mice, in which red-activated channelrhodopsin (ReaChR) is inducibly and specifically expressed in microglia. We found that activation of ReaChR by red light in spinal microglia evoked reliable inward currents and membrane depolarization. In vivo optogenetic activation of microglial ReaChR in the spinal cord triggered chronic pain hypersensitivity in both male and female mice. In addition, activation of microglial ReaChR up-regulated neuronal c-Fos expression and enhanced C-fiber responses. Mechanistically, ReaChR activation led to a reactive microglial phenotype with increased interleukin (IL)-1β production, which is likely mediated by inflammasome activation and calcium elevation. IL-1 receptor antagonist (IL-1ra) was able to reverse the pain hypersensitivity and neuronal hyperactivity induced by microglial ReaChR activation. Therefore, our work demonstrates that optogenetic activation of spinal microglia is sufficient to trigger chronic pain phenotypes by increasing neuronal activity via IL-1 signaling.

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Usefulness of Olanzapine as an Adjunct to Opioid Treatment and for the Treatment of Neuropathic Pain

Anesthesiology ◽

10.1097/aln.0b013e31823c7e56 ◽

2012 ◽

Vol 116 (1) ◽

pp. 159-169 ◽

Cited By ~ 27

Author(s):

Kazuhiro Torigoe ◽

Kae Nakahara ◽

Mahardian Rahmadi ◽

Kazumi Yoshizawa ◽

Hiroshi Horiuchi ◽

...

Keyword(s):

Chronic Pain ◽

Neuropathic Pain ◽

Sleep Disturbance ◽

Brain Tissue ◽

Gastrointestinal Transit ◽

Thermal Hyperalgesia ◽

Nausea And Vomiting ◽

Dependent Manner ◽

Antiemetic Effect

Background The use of opioids for pain management is often associated with nausea and vomiting. Although conventional antipsychotics are often used to counter emesis, they can be associated with extrapyramidal symptoms. However, chronic pain can induce sleep disturbance. The authors investigated the effects of the atypical antipsychotic olanzapine on morphine-induced emesis and the sleep dysregulation associated with chronic pain. Methods A receptor binding assay was performed using mouse whole brain tissue. The emetic response in ferrets was evaluated by counting retching and vomiting behaviors. Catalepsy in mice was evaluated by placing both of their forepaws over a horizontal bar. Released dopamine was measured by an in vivo microdialysis study. Sleep disturbance in mice in a neuropathic pain-like state was assayed by electroencephalogram and electromyogram recordings. Results Olanzapine showed high affinity for muscarinic M1 receptor in brain tissue. Olanzapine decreased morphine-induced nausea and vomiting in a dose-dependent manner. However, olanzapine at a dose that had an antiemetic effect (0.03 mg/kg) did not induce catalepsy or hyperglycemia. In addition, olanzapine at this dose had no effect on the morphine-induced release of dopamine or inhibition of gastrointestinal transit. Finally, olanzapine inhibited thermal hyperalgesia and completely alleviated the sleep disturbance induced by sciatic nerve ligation. Conclusion These findings suggest that olanzapine may be useful for the treatment of morphine-induced emesis and as an adjunct for the treatment of neuropathic pain associated with sleep disturbance.

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Dual Role of Triptolide in Interrupting the NLRP3 Inflammasome Pathway to Attenuate Cardiac Fibrosis

International Journal of Molecular Sciences ◽

10.3390/ijms20020360 ◽

2019 ◽

Vol 20 (2) ◽

pp. 360 ◽

Cited By ~ 15

Author(s):

Xi-Chun Pan ◽

Ya Liu ◽

Yan-Yan Cen ◽

Ya-Lan Xiong ◽

Jing-Mei Li ◽

...

Keyword(s):

Nlrp3 Inflammasome ◽

Cardiac Fibrosis ◽

Cardiac Fibroblasts ◽

Dual Role ◽

Receptor Protein ◽

Inflammasome Activation ◽

Collagen Production ◽

Antifibrotic Effect ◽

Myeloid Differentiation Factor

In a previous paper, we reported that triptolide (TP), a commonly used immunomodulator, could attenuate cardiac hypertrophy. This present study aimed to further explore the inhibition of cardiac fibrosis by TP and the possible mechanism from the perspective of the NOD-like receptor protein 3 (NLRP3) inflammasome. Hematoxylin-eosin and Masson’s staining, immunohistochemistry, and immunofluorescence were performed to observe cardiac fibrotic changes in mice and mouse cardiac fibroblasts (CFs). The Western blot, colocalization, and immunoprecipitation were applied to detect protein expression and interactions. Results suggested that TP dose-dependently inhibited cardiac fibrosis induced by isoproterenol and collagen production of CFs induced by angiotensin II. TP exhibited an antifibrotic effect via inhibiting activation of the NLRP3 inflammasome, which sequentially decreased IL-1β maturation, myeloid differentiation factor 88 (MyD88)-related phosphorylation of c-Jun N-terminal kinase (JNK), extracellular regulated protein kinase 1/2 (ERK1/2), and TGF-β1/Smad signaling, and ultimately resulted in less collagen production. Moreover, TP showed no antifibrotic effect in Nlrp3-knockout CFs. Notably, TP inhibited the expression of NLRP3 and apoptosis-associated speck-like proteins containing a caspase recruitment domain (ASC) as well as inflammasome assembly, by interrupting the NLRP3-ASC interaction to inhibit inflammasome activation. Finally, TP indeed inhibited the NLRP3-TGFβ1-Smad pathway in vivo. Conclusively, TP was found to play a dual role in interrupting the activation of the NLRP3 inflammasome to attenuate cardiac fibrosis.

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GPER/β-Alanine Positive Interaction in The Dorsal Root Ganglion Uncovers Potential Mechanisms: Mediating Continuous Neuronal Sensitisation and Neuroinflammation Responses in Neuropathic Pain

10.21203/rs.3.rs-1090097/v1 ◽

2021 ◽

Author(s):

Zhenzhen Xu ◽

Wanli Xie ◽

Yiqi Feng ◽

Yanting Wang ◽

Yuyao He ◽

...

Keyword(s):

Chronic Pain ◽

Estrogen Receptor ◽

Neuropathic Pain ◽

Dorsal Root Ganglion ◽

Dorsal Root ◽

Neuronal Response ◽

Male Rats ◽

Patch Clamp Technique ◽

Positive Interaction ◽

The Impact

Abstract Background: The pathogenesis of neuropathic pain and the reasons for the prolonged unhealing are still unknown. Increasing evidence suggests that estrogen sex differences play a role in pain sensitivity, but few studies focused on the role of estrogen receptor which maybe an important molecular component contributing to peripheral pain transduction. We aimed to investigate the impact of oestrogen receptors in nociceptive neuronal response in the dorsal root ganglion (DRG) and spinal dorsal horn using a spared nerve injury (SNI) rat model of chronic pain. Methods: We used a class of estrogen receptors antagonists and agonists intrathecal (i.t.) administrated to male rats with SNI or normal rats to identify the main receptor. Moreover, we applied genes identified through genomic metabolic analysis to determine the key metabolism point and elucidate potential mechanisms mediating continuous neuronal sensitisation and neuroinflammation responses in neuropathic pain. The excitability of DRG neurons was detected using the patch clamp technique. Immunohistochemistry, Western blotting, qPCR and behavioral testing were used to assess the expressions, cellular distributions, and actions of main receptor and its related signaling molecules.Results: Increasing the expression and function of G protein-coupled estrogen receptor (GPER), but not estrogen receptor-α (ERα) and estrogen receptor-β (ERβ), in the DRG, but not the dorsal spinal cord, contributed to SNI-induced neuronal sensitisation. Inhibiting GPER expression in the DRG alleviated SNI-induced pain behaviours and neuroinflammation by downregulating IL-1β and IL-6 expression as well as restoring GABAα2 expression simultaneously. Additionally, the positive interaction between GPER and β-alanine, β-alanine accumulation enhances pain sensation and promotes chronic pain development. Conclusion: GPER activation in the DRG causes a positive interaction of β-alanine with IL-1β and IL-6 expression and represses GABAα2 involved in post-SNI neuropathic pain development. Blocking GPER and eliminating β-alanine in the DRG may prevent neuropathic pain development.

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Koumine Suppresses IL-1β Secretion and Attenuates Inflammation Associated With Blocking ROS/NF-κB/NLRP3 Axis in Macrophages

Frontiers in Pharmacology ◽

10.3389/fphar.2020.622074 ◽

2021 ◽

Vol 11 ◽

Author(s):

Yufei Luo ◽

Bojun Xiong ◽

Haiping Liu ◽

Zehong Chen ◽

Huihui Huang ◽

...

Keyword(s):

Nlrp3 Inflammasome ◽

Inflammatory Diseases ◽

Inhibitory Effect ◽

Receptor Protein ◽

Mechanistic Study ◽

Inflammasome Activation ◽

Nlrp3 Inflammasome Activation ◽

Underlying Mechanisms

Koumine (KM), one of the primary constituents of Gelsemium elegans, has been used for the treatment of inflammatory diseases such as rheumatoid arthritis, but whether KM impacts the activation of the NOD-like receptor protein 3 (NLRP3) inflammasome remains unknown. This study aimed to explore the inhibitory effect of KM on NLRP3 inflammasome activation and the underlying mechanisms both in vitro using macrophages stimulated with LPS plus ATP, nigericin or monosodium urate (MSU) crystals and in vivo using an MSU-induced peritonitis model. We found that KM dose-dependently inhibited IL-1β secretion in macrophages after NLRP3 inflammasome activators stimulation. Furthermore, KM treatment efficiently attenuated the infiltration of neutrophils and suppressed IL-1β production in mice with MSU-induced peritonitis. These results indicated that KM inhibited NLRP3 inflammasome activation, and consistent with this finding, KM effectively inhibited caspase-1 activation, mature IL-1β secretion, NLRP3 formation and pro-IL-1β expression in LPS-primed macrophages treated with ATP, nigericin or MSU. The mechanistic study showed that, KM exerted a potent inhibitory effect on the NLRP3 priming step, which decreased the phosphorylation of IκBα and p65, the nuclear localization of p65, and the secretion of TNF-α and IL-6. Moreover, the assembly of NLRP3 was also interrupted by KM. KM blocked apoptosis-associated speck-like protein containing a CARD (ASC) speck formation and its oligomerization and hampered the NLRP3-ASC interaction. This suppression was attributed to the ability of KM to inhibit the production of reactive oxygen species (ROS). In support of this finding, the inhibitory effect of KM on ROS production was completely counteracted by H2O2, an ROS promoter. Our results provide the first indication that KM exerts an inhibitory effect on NLRP3 inflammasome activation associated with blocking the ROS/NF-κB/NLRP3 signal axis. KM might have potential clinical application in the treatment of NLRP3 inflammasome-related diseases.

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CLEC5A knockdown protects against the cardiac dysfunction after Myocardial infarction by suppressing macrophage polarization, NLRP3 inflammasome activation and pyroptosis

Biochemistry and Cell Biology ◽

10.1139/bcb-2020-0672 ◽

2021 ◽

Author(s):

Xin Wang ◽

Yu Hu ◽

Yaguang Wang ◽

Dapeng Shen ◽

Guizhou Tao

Keyword(s):

Myocardial Infarction ◽

Nlrp3 Inflammasome ◽

Cardiac Dysfunction ◽

Macrophage Polarization ◽

Left Ventricular ◽

Receptor Protein ◽

Inflammasome Activation ◽

M1 Polarization

Increasing evidence has shown that NOD-like receptor protein 3 (NLRP3) inflammasome and pyroptotic cell death play vital roles in the pathophysiology of myocardial infarction (MI), a common cardiovascular disease characterized with cardiac dysfunction. C-type lectin member 5A (CLEC5A) is reported to strongly associate with activation of NLRP3 inflammasome and pyroptosis. In this study, in vivo MI model was established by the ligation of left anterior descending coronary artery on male C57BL/6 mice, and CLEC5A knockdown was further achieved by intra-myocardial injection of adenovirus delivering shRNA-CLEC5A. CLEC5A was found to be highly expressed in left ventricular of MI mice, while CLEC5A knockdown conversely alleviated the cardiac dysfunction in MI mice. Besides, MI-induced classical activation of macrophages was significantly inhibited after CLEC5A silencing. Additionally, CLEC5A knockdown dramatically inhibited MI-triggered activation of NLRP3 inflammasome, pyroptosis and NF-κB signaling in left ventricular of mice. In vitro experiment further validated that CLEC5A knockdown suppressed M1 polarization in LPS/IFNγ-stimulated RAW264.7 cells, and inhibited the polarized RAW264.7-induced activation of NLRP3 inflammasome/pyroptosis signaling in co-cultured cardiomyocytes. In conclusion, CLEC5A knockdown protects against the MI-induced cardiac dysfunction by regulating macrophage polarization, NLRP3 inflammasome and cell pyroptosis.

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Mechanism of Ginkgolide B Amelioration of Neuropathic Pain Induced by Chronic Constriction Injury

Current Topics in Nutraceutical Research ◽

10.37290/ctnr2641-452x.19:359-365 ◽

2021 ◽

Vol 19 (3) ◽

pp. 359-365

Author(s):

Guizhen Yan ◽

Aobo Ma ◽

Man Huang ◽

Yuan Zhang

Keyword(s):

Neuropathic Pain ◽

Protein Expression ◽

Nlrp3 Inflammasome ◽

Chronic Constriction Injury ◽

Nerve Fibers ◽

Fibrous Structure ◽

Receptor Protein ◽

Inflammasome Activation ◽

Ginkgolide B ◽

Nlrp3 Inflammasome Activation

Activation of NOD-like receptor protein 3 (NLRP3) inflammasome is implicated in the pathogenesis of neuropathic pain. Ginkgolide B contributes to the suppression of NLRP3 inflammasome activation to prevent hypoxic-ischemic brain injury. However, the role of ginkgolide B on neuropathic pain has not been reported yet. We have shown that administration of ginkgolide B lowered pain threshold measured by paw-withdrawal threshold and paw-withdrawal latency in rats subjected to chronic constriction injury. Nerve fibers in rats postchronic constriction injury were swollen, and the fibrous structure was disordered. Treatment with ginkgolide B attenuated the nerve fiber swelling and reduced the disordered fibrous structure. Ginkgolide B dosage dependently attenuated chronic constriction injury-induced increase of proinflammatory cytokines. Protein expression of NLRP3 and its downstream targets (caspase 1 and IL-1β) were increased by chronic constriction injury and reduced by ginkgolide B. Lastly, ginkgolide B counteracted with the promotive effects of chronic constriction injury on protein expression of TLR4 and p-NF-κB. In conclusion, ginkgolide B demonstrated anti-inflammatory and antinociceptive effects in rats' model with neuropathic pain by suppression of TLR4-NF-κB-mediated NLRP3 inflammasome activation.

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Pain Syndromes Other Than Headache

10.2310/neuro.6177 ◽

2016 ◽

Author(s):

Edgar L. Ross

Keyword(s):

Chronic Pain ◽

Neuropathic Pain ◽

Sensory Processing ◽

Pain Syndrome ◽

The Body ◽

Spinal Cord Dorsal Horn ◽

Opioid Use ◽

Nerve Injuries ◽

Pain Syndromes ◽

Chronic Pain Syndromes

Pain is experienced within a complex biologic, emotional, psychological, and social context that may defy physical examination, diagnostic procedures, and laboratory tests. This chapter aims to empower internists to improve their medical practices in pain management. It provides a scientific background that covers nociception and how sensory processing occurs at multiple levels in the body. Clinical assessment is detailed, as well as diagnostic categories that include mixed or uncertain chronic pain syndromes (back pain, fibromyalgia, postamputation pain, pain from cancer and bone) and neuropathic pain syndromes (polyneuropathy, mononeuropathy multiplex, ganglionopathy, genetic disorders, focal and regional syndromes). Treatment of chronic pain can be surgical or interventional. Pharmacologic treatment for acute and chronic nociceptive pain includes special considerations for geriatric and terminal patients. For treatment of neuropathic pain, medications are the major component. One tables lists iatrogenic nerve injuries that can cause posttraumatic neuralgia and complex regional pain syndrome. Other tables detail stepwise pharmacologic management of neuropathic pain and cite recommendations on opioid use from the Centers for Disease Control and Prevention. One figure illustrates how pain transducers monitor and influence tissue conditions. Other figures show sensory processing in the spinal cord dorsal horn, physical findings in the feet of patients with bilateral foot pain from small-fiber polyneuropathy, illustrate how examination can identify specific nerve injuries causing chronic pain, and provide classification of chronic pain syndromes. This chapter contains 82 references.

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