Electroacupuncture Regulates Pain Transition Through Inhibiting PKCε and TRPV1 Expression in Dorsal Root Ganglion.
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ABSTRACT:
SUBMITTER: Fang J
PROVIDER: S-EPMC8329384 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Background:Acute pain can transition to chronic pain, presenting a major clinical challenge. Electroacupuncture (EA) can partly prevent the transition from acute to chronic pain. However, little is known about the mechanisms underlying the effect of EA. This study investigated the effect of EA on pain transition and the activation of metabotropic glutamate receptor 5 (mGluR5)-protein kinase C epsilon (PKC?) signaling pathway in the dorsal root ganglia (DRG). Methods:The hyperalgesic priming model was established by the sequential intraplantar injection of carrageenan (1%, 100 ?L) and prostaglandin E2 (PGE2) into the left hind paw of rats. EA treatment (2/100 Hz, 30 min, once/day) was applied at bilateral Zusanli (ST36) and Kunlun (BL60) acupoints in rats. Von Frey filaments were used to investigate the mechanical withdrawal threshold (MWT) at different time points. The protein expression levels of mGluR5 and PKC? in the ipsilateral L4-L6 DRGs of rats were detected by Western blot. Some pharmacological experiments were performed to evaluate the relationship between mGluR5, PKC? and the MWT. It was also used to test the effects of EA on the expression levels of mGluR5 and PKC? and changes in the MWT. Results:Sequential injection of carrageenan and PGE2 significantly decreased the MWT of rats and up-regulated the expression level of mGluR5 and PKC? in the ipsilateral L4-L6 DRGs. EA can reverse the hyperalgesic priming induced by sequential injection of carrageenan/PGE and down-regulate the protein expression of mGluR5 and PKC?. Glutamate injection instead of PGE2 can mimic the hyperalgesic priming model. Pharmacological blocking of mGluR5 with specific antagonist MTEP can prevent the hyperalgesic priming and inhibit the activation of PKC? in DRGs. Furthermore, EA also produced analgesic effect on the hyperalgesic priming rats induced by carrageenan/mGluR5 injection and inhibited the high expression of PKC?. Sham EA produced none analgesic and regulatory effect. Conclusion:EA can regulate pain transition and it may relate with its inhibitory effect on the activation of mGluR5-PKC? signaling pathway in the DRGs.
| S-EPMC7311359 | biostudies-literature
Project description:BackgroundNicotinamide adenine dinucleotide phosphate oxidase 2 (NOX2)-induced oxidative stress, including the production of reactive oxygen species (ROS) and hydrogen peroxide, plays a pivotal role in neuropathic pain. Although the activation and plasma membrane translocation of protein kinase C (PKC) isoforms in dorsal root ganglion (DRG) neurons have been implicated in multiple pain models, the interactions between NOX2-induced oxidative stress and PKC remain unknown.MethodsA spared nerve injury (SNI) model was established in adult male rats. Pharmacologic intervention and AAV-shRNA were applied locally to DRGs. Pain behavior was evaluated by Von Frey tests. Western blotting and immunohistochemistry were performed to examine the underlying mechanisms. The excitability of DRG neurons was recorded by whole-cell patch clamping.ResultsSNI induced persistent NOX2 upregulation in DRGs for up to 2 weeks and increased the excitability of DRG neurons, and these effects were suppressed by local application of gp91-tat (a NOX2-blocking peptide) or NOX2-shRNA to DRGs. Of note, the SNI-induced upregulated expression of PKCε but not PKC was decreased by gp91-tat in DRGs. Mechanical allodynia and DRG excitability were increased by ψεRACK (a PKCε activator) and reduced by εV1-2 (a PKCε-specific inhibitor). Importantly, εV1-2 failed to inhibit SNI-induced NOX2 upregulation. Moreover, the SNI-induced increase in PKCε protein expression in both the plasma membrane and cytosol in DRGs was attenuated by gp91-tat pretreatment, and the enhanced translocation of PKCε was recapitulated by H2O2 administration. SNI-induced upregulation of PKCε was blunted by phenyl-N-tert-butylnitrone (PBN, an ROS scavenger) and the hydrogen peroxide catalyst catalase. Furthermore, εV1-2 attenuated the mechanical allodynia induced by H2O2 CONCLUSIONS: NOX2-induced oxidative stress promotes the sensitization of DRGs and persistent pain by increasing the plasma membrane translocation of PKCε.
| S-EPMC8101139 | biostudies-literature
Project description:Transient receptor potential vanilloid type 1 (TRPV1) is a plasma membrane Ca(2+) channel involved in transduction of painful stimuli. Dorsal root ganglion (DRG) neurons express ectopic but functional TRPV1 channels in the endoplasmic reticulum (ER) (TRPV1(ER)). We have studied the properties of TRPV1(ER) in DRG neurons and HEK293T cells expressing TRPV1. Activation of TRPV1(ER) with capsaicin or other vanilloids produced an increase of cytosolic Ca(2+) due to Ca(2+) release from the ER. The decrease of [Ca(2+)](ER) was directly revealed by an ER-targeted aequorin Ca(2+) probe, expressed in DRG neurons using a herpes amplicon virus. The sensitivity of TRPV1(ER) to capsaicin was smaller than the sensitivity of the plasma membrane TRPV1 channels. The low affinity of TRPV1(ER) was not related to protein kinase A- or C-mediated phosphorylations, but it was due to inactivation by cytosolic Ca(2+) because the sensitivity to capsaicin was increased by loading the cells with the Ca(2+) chelator BAPTA. Decreasing [Ca(2+)](ER) did not affect the sensitivity of TRPV1(ER) to capsaicin. Disruption of the TRPV1 calmodulin-binding domains at either the C terminus (Delta35AA) or the N terminus (K155A) increased 10-fold the affinity of TRPV1(ER) for capsaicin, suggesting that calmodulin is involved in the inactivation. The lack of TRPV1 sensitizers, such as phosphatidylinositol 4,5-bisphosphate, in the ER could contribute to decrease the affinity for capsaicin. The low sensitivity of TRPV1(ER) to agonists may be critical for neuron health, because otherwise Ca(2+) depletion of ER could lead to ER stress, unfolding protein response, and cell death.
| S-EPMC2781673 | biostudies-literature
Project description:BackgroundAnnexin A1 (ANXA1) exerts anti-nociceptive effect through ANXA1 receptor formyl peptide receptor 2 (FPR2/ALX (receptor for lipoxin A4), FPR2) at the dorsal root ganglia (DRG) level. However, the mechanisms remain elucidated. By using radiant heat, hot/cold plate, tail flick, von Frey, and Randall-Selitto tests to detect nociception in intact and chemical (capsaicin, menthol, mustard oil, formalin or CFA) injected AnxA1 conditional knockout (AnxA1-/-) mice, applying calcium imaging and patch clamp recordings in cultured DRG neurons to measure neuronal excitability, conducting immunofluorescence and western blotting to detect the protein levels of TRPV1, FPR2 and its downstream molecules, and performing double immunofluorescence and co-immunoprecipitation to investigate the interaction between Calmodulin (CaM) and TRPV1; we aim to uncover the molecular and cellular mechanisms of ANXA1's role in antinociception.ResultsAnxA1-/- mice exhibited significant sensitivity to noxious heat (mean ± SD, 6.2 ± 1.0 s vs. 9.9 ± 1.6 s in Hargreaves test; 13.6 ± 1.5 s vs. 19.0 ± 1.9 s in hot plate test; n = 8; P < 0.001), capsaicin (101.0 ± 15.3 vs. 76.2 ± 10.9; n = 8; P < 0.01), formalin (early phase: 169.5 ± 32.8 s vs. 76.0 ± 21.9 s; n = 8; P < 0.05; late phase: 444.6 ± 40.1 s vs. 320.4 ± 33.6 s; n = 8; P < 0.01) and CFA (3.5 ± 0.8 s vs. 5.9 ± 1.4 s; n = 8; P < 0.01). In addition, we found significantly increased capsaicin induced Ca2+ response, TRPV1 currents and neuronal firing in AnxA1 deficient DRG neurons. Furthermore, ANXA1 mimic peptide Ac2-26 robustly increased intracellular Ca2+, inhibited TRPV1 current, activated PLCβ and promoted CaM-TRPV1 interaction. And these effects of Ac2-26 could be attenuated by FPR2 antagonist Boc2.ConclusionsSelective deletion of AnxA1 in DRG neurons enhances TRPV1 sensitivity and deteriorates noxious heat or capsaicin induced nociception, while ANXA1 mimic peptide Ac2-26 desensitizes TRPV1 via FPR2 and the downstream PLCβ-Ca2+-CaM signal. This study may provide possible target for developing new analgesic drugs in inflammatory pain.
| S-EPMC8393810 | biostudies-literature
Project description:The purinergic 2X7 (P2X7) receptor expressed in satellite glial cells (SGCs) is involved in the inflammatory response, and transient receptor potential vanilloid 1 (TRPV1) participates in the process of neurogenic inflammation, such as that in diabetic neuropathic pain (DNP) and peripheral neuralgia. The main purpose of this study was to explore the role of the P2X7 receptor in DNP hypersensitivity mediated by TRPV1 in the rat and its possible mechanism. A rat model of type 2 diabetes mellitus-related neuropathic pain (NPP) named the DNP rat model was established in this study. The mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) of DNP rats were increased after intrathecal injection of the P2X7 receptor antagonist A438079, and the mRNA and protein levels of TRPV1 in the dorsal root ganglion (DRG) were decreased in DNP rats treated with A438079 compared to untreated DNP rats; in addition, A438079 also decreased the phosphorylation of p38 and extracellular signal-regulated kinase 1/2 (ERK1/2) in the DNP group. Based on these results, the P2X7 receptor might be involved in DNP mediated by TRPV1.
| S-EPMC8215290 | biostudies-literature
Project description:Collateral branches from axons are key components of functional neural circuits that allow neurons to connect with multiple synaptic targets. Like axon growth and guidance, formation of collateral branches depends on the regulation of microtubules, but how such regulation is coordinated to ensure proper circuit development is not known. Based on microarray analysis, we have identified a role for microtubule-associated protein 7 (MAP7) during collateral branch development of dorsal root ganglion (DRG) sensory neurons. We show that MAP7 is expressed at the onset of collateral branch formation. Perturbation of its expression by overexpression or shRNA knockdown alters axon branching in cultured DRG neurons. Localization and time-lapse imaging analysis reveals that MAP7 is enriched at branch points and colocalizes with stable microtubules, but enters the new branch with a delay, suggesting a role in branch maturation. We have also investigated a spontaneous mutant mouse that expresses a truncated MAP7 and found a gain-of-function phenotype both in vitro and in vivo Further domain analysis suggests that the amino half of MAP7 is responsible for branch formation, suggesting a mechanism that is independent of its known interaction with kinesin. Moreover, this mouse exhibits increased pain sensitivity, a phenotype that is consistent with increased collateral branch formation. Therefore, our study not only uncovers the first neuronal function of MAP7, but also demonstrates the importance of proper microtubule regulation in neural circuit development. Furthermore, our data provide new insights into microtubule regulation during axonal morphogenesis and may shed light on MAP7 function in neurological disorders.SIGNIFICANCE STATEMENT Neurons communicate with multiple targets by forming axonal branches. In search of intrinsic factors that control collateral branch development, we identified a role for microtubule-associated protein 7 (MAP7) in dorsal root ganglion sensory neurons. We show that MAP7 expression is developmentally regulated and perturbation of this expression alters branch formation. Cell biological analysis indicates that MAP7 promotes branch maturation. Analysis of a spontaneous mouse mutant suggests a molecular mechanism for branch regulation and the potential influence of collateral branches on pain sensitivity. Our studies thus establish the first neuronal function of MAP7 and demonstrate its role in branch morphogenesis and neural circuit function. These findings may help in our understanding of the contribution of MAP7 to neurological disorders and nerve regeneration.
| S-EPMC5299575 | biostudies-literature
Project description:Neuropathic pain encompasses a diverse array of clinical entities affecting 7-10% of the population, which is challenging to adequately treat. Several promising therapeutics derived from molecular discoveries in animal models of neuropathic pain have failed to translate following unsuccessful clinical trials suggesting the possibility of important cellular-level and molecular differences between animals and humans. Establishing the extent of potential differences between laboratory animals and humans, through direct study of human tissues and/or cells, is likely important in facilitating translation of preclinical discoveries to meaningful treatments. Patch-clamp electrophysiology and RNA-sequencing was performed on dorsal root ganglia taken from patients with variable presence of radicular/neuropathic pain. Findings establish that spontaneous action potential generation in dorsal root ganglion neurons is associated with radicular/neuropathic pain and radiographic nerve root compression. Transcriptome analysis suggests presence of sex-specific differences and reveals gene modules and signalling pathways in immune response and neuronal plasticity related to radicular/neuropathic pain that may suggest therapeutic avenues and that has the potential to predict neuropathic pain in future cohorts.
| S-EPMC6487328 | biostudies-literature
Project description:BackgroundPrimary and metastatic cancers that affect bone are frequently associated with severe and intractable pain. The mechanisms underlying the development of bone cancer pain are largely unknown. The aim of this study was to determine whether enhanced excitability of primary sensory neurons contributed to peripheral sensitization and tumor-induced hyperalgesia during cancer condition. In this study, using techniques of whole-cell patch-clamp recording associated with immunofluorescent staining, single-cell reverse-transcriptase PCR and behavioral test, we investigated whether the intrinsic membrane properties and the excitability of small-sized dorsal root ganglion (DRG) neurons altered in a rat model of bone cancer pain, and whether suppression of DRG neurons activity inhibited the bone cancer-induced pain.ResultsOur present study showed that implantation of MRMT-1 tumor cells into the tibial canal in rats produced significant mechanical and thermal hyperalgesia in the ipsilateral hind paw. Moreover, implantation of tumor cells provoked spontaneous discharges and tonic excitatory discharges evoked by a depolarizing current pulse in small-sized DRG neurons. In line with these findings, alterations in intrinsic membrane properties that reflect the enhanced neuronal excitability were observed in small DRG neurons in bone cancer rats, of which including: 1) depolarized resting membrane potential (RMP); 2) decreased input resistance (Rin); 3) a marked reduction in current threshold (CT) and voltage threshold (TP) of action potential (AP); 4) a dramatic decrease in amplitude, overshot, and duration of evoked action potentials as well as in amplitude and duration of afterhyperpolarization (AHP); and 5) a significant increase in the firing frequency of evoked action potentials. Here, the decreased AP threshold and increased firing frequency of evoked action potentials implicate the occurrence of hyperexcitability in small-sized DRG neurons in bone cancer rats. In addiotion, immunofluorescent staining and single-cell reverse-transcriptase PCR revealed that in isolated small DRG neurons, most neurons were IB4-positive, or expressed TRPV1 or CGRP, indicating that most recorded small DRG neurons were nociceptive neurons. Finally, using in vivo behavioral test, we found that blockade of DRG neurons activity by TTX inhibited the tumor-evoked mechanical allodynia and thermal hyperalgesia in bone cancer rats, implicating that the enhanced excitability of primary sensory neurons underlied the development of bone cancer pain.ConclusionsOur present results suggest that implantation of tumor cells into the tibial canal in rats induces an enhanced excitability of small-sized DRG neurons that is probably as results of alterations in intrinsic electrogenic properties of these neurons. Therefore, alterations in intrinsic membrane properties associated with the hyperexcitability of primary sensory neurons likely contribute to the peripheral sensitization and tumor-induced hyperalgesia under cancer condition.
| S-EPMC3379961 | biostudies-literature
Project description:Itch, a sensation eliciting a desire to scratch, is distinct from but not completely independent of pain. Inspiring achievements have been made in the characterization of itch-related receptors and neurotransmitters, but the molecular mechanisms controlling the development of pruriceptors remain poorly understood. Here, our RNAseq and in situ hybridization data show that the transcription factor Tlx3 is required for the expression of a majority of itch-related molecules in the dorsal root ganglion (DRG). As a result, Tlx3F/F;Nav1.8-cre mice exhibit significantly attenuated acute and dry skin-induced chronic itch. Furthermore, our study indicates that TRPV1 plays a pivotal role in the chronic itch evoked by dry skin and allergic contact dermatitis (ACD). The mutants also display impaired response to cold and inflammatory pain and elevated response to capsaicin, whereas the responses to acute mechanical, thermal stimuli and neuropathic pain remain normal. In Tlx3F/F;Nav1.8-cre mice, TRPV1 is derepressed and expands predominantly into IB4+ non-peptidergic (NP) neurons. Collectively, our data reveal a molecular mechanism in regulating the development of pruriceptors and controlling itch and pain sensations.
| S-EPMC5487456 | biostudies-literature
Project description:Paralleling the activation of dorsal horn microglia after peripheral nerve injury is a significant expansion and proliferation of macrophages around injured sensory neurons in dorsal root ganglia (DRG). Here we demonstrate a critical contribution of DRG macrophages, but not those at the nerve injury site, to both the initiation and maintenance of the mechanical hypersensitivity that characterizes the neuropathic pain phenotype. In contrast to the reported sexual dimorphism in the microglial contribution to neuropathic pain, depletion of DRG macrophages reduces nerve injury-induced mechanical hypersensitivity and expansion of DRG macrophages in both male and female mice. However, fewer macrophages are induced in the female mice and deletion of colony-stimulating factor 1 from sensory neurons, which prevents nerve injury-induced microglial activation and proliferation, only reduces macrophage expansion in male mice. Finally, we demonstrate molecular cross-talk between axotomized sensory neurons and macrophages, revealing potential peripheral DRG targets for neuropathic pain management.
| S-EPMC6959328 | biostudies-literature