Phospholipase C-mediated hydrolysis of phosphatidylinositol 4,5-bisphosphate generates diacylglycerol, inositol 1,4,5-trisphosphate and protons, all of which can regulate TRPV1 activity via different ...mechanisms. Here we explored the possibility that the diacylglycerol metabolites 2-arachidonoylglycerol and 1-arachidonoylglycerol, and not metabolites of these monoacylglycerols, activate TRPV1 and contribute to this signaling cascade. 2-Arachidonoylglycerol and 1-arachidonoylglycerol activated native TRPV1 on vascular sensory nerve fibers and heterologously expressed TRPV1 in whole cells and inside-out membrane patches. The monoacylglycerol lipase inhibitors methylarachidonoyl-fluorophosphonate and JZL184 prevented the metabolism of deuterium-labeled 2-arachidonoylglycerol and deuterium-labeled 1-arachidonoylglycerol in arterial homogenates, and enhanced TRPV1-mediated vasodilator responses to both monoacylglycerols. In mesenteric arteries from TRPV1 knock-out mice, vasodilator responses to 2-arachidonoylglycerol were minor. Bradykinin and adenosine triphosphate, ligands of phospholipase C-coupled membrane receptors, increased the content of 2-arachidonoylglycerol in dorsal root ganglia. In HEK293 cells expressing the phospholipase C-coupled histamine H1 receptor, exposure to histamine stimulated the formation of 2-AG, and this effect was augmented in the presence of JZL184. These effects were prevented by the diacylglycerol lipase inhibitor tetrahydrolipstatin. Histamine induced large whole cell currents in HEK293 cells co-expressing TRPV1 and the histamine H1 receptor, and the TRPV1 antagonist capsazepine abolished these currents. JZL184 increased the histamine-induced currents and tetrahydrolipstatin prevented this effect. The calcineurin inhibitor ciclosporin and the endogenous "entourage" compound palmitoylethanolamide potentiated the vasodilator response to 2-arachidonoylglycerol, disclosing TRPV1 activation of this monoacylglycerol at nanomolar concentrations. Furthermore, intracerebroventricular injection of JZL184 produced TRPV1-dependent antinociception in the mouse formalin test. Our results show that intact 2-arachidonoylglycerol and 1-arachidonoylglycerol are endogenous TRPV1 activators, contributing to phospholipase C-dependent TRPV1 channel activation and TRPV1-mediated antinociceptive signaling in the brain.
Patients with cholestatic disease exhibit pruritus and analgesia, but the mechanisms underlying these symptoms are unknown. We report that bile acids, which are elevated in the circulation and ...tissues during cholestasis, cause itch and analgesia by activating the GPCR TGR5. TGR5 was detected in peptidergic neurons of mouse dorsal root ganglia and spinal cord that transmit itch and pain, and in dermal macrophages that contain opioids. Bile acids and a TGR5-selective agonist induced hyperexcitability of dorsal root ganglia neurons and stimulated the release of the itch and analgesia transmitters gastrin-releasing peptide and leucine-enkephalin. Intradermal injection of bile acids and a TGR5-selective agonist stimulated scratching behavior by gastrin-releasing peptide- and opioid-dependent mechanisms in mice. Scratching was attenuated in Tgr5-KO mice but exacerbated in Tgr5-Tg mice (overexpressing mouse TGR5), which exhibited spontaneous pruritus. Intraplantar and intrathecal injection of bile acids caused analgesia to mechanical stimulation of the paw by an opioid-dependent mechanism. Both peripheral and central mechanisms of analgesia were absent from Tgr5-KO mice. Thus, bile acids activate TGR5 on sensory nerves, stimulating the release of neuropeptides in the spinal cord that transmit itch and analgesia. These mechanisms could contribute to pruritus and painless jaundice that occur during cholestatic liver diseases.
Pannexin-1 (Panx1) is a large-pore membrane channel involved in the release of ATP and other signaling mediators. Little is known about the expression and functional role of Panx1 in the dorsal root ...ganglion (DRG) in the development of chronic neuropathic pain. In this study, we determined the epigenetic mechanism involved in increased Panx1 expression in the DRG after nerve injury. Spinal nerve ligation in rats significantly increased the mRNA and protein levels of Panx1 in the DRG but not in the spinal cord. Immunocytochemical labeling showed that Panx1 was primarily expressed in a subset of medium and large DRG neurons in control rats and that nerve injury markedly increased the number of Panx1-immunoreactive DRG neurons. Nerve injury significantly increased the enrichment of two activating histone marks (H3K4me2 and H3K9ac) and decreased the occupancy of two repressive histone marks (H3K9me2 and H3K27me3) around the promoter region of Panx1 in the DRG. However, nerve injury had no effect on the DNA methylation level around the Panx1 promoter in the DRG. Furthermore, intrathecal injection of the Panx1 blockers or Panx1-specific siRNA significantly reduced pain hypersensitivity induced by nerve injury. In addition, siRNA knockdown of Panx1 expression in a DRG cell line significantly reduced caspase-1 release induced by neuronal depolarization. Our findings suggest that nerve injury increases Panx1 expression levels in the DRG through altered histone modifications. Panx1 up-regulation contributes to the development of neuropathic pain and stimulation of inflammasome signaling.
Background: Pannexin-1 can release many signaling molecules, and blocking pannexin-1 at the spinal cord level reduces chronic pain.
Results: Nerve injury increases pannexin-1 expression in primary sensory neurons via histone modifications. Pannexin-1 knockdown reduces pain hypersensitivity.
Conclusion: Pannexin-1 up-regulation in primary sensory neurons contributes to neuropathic pain.
Significance: Understanding the molecular mechanism of neuronal plasticity will improve treatments for neuropathic pain.
Following peripheral axon injury, dysregulation of non-coding microRNAs (miRs) occurs in dorsal root ganglia (DRG) sensory neurons. Here we show that DRG neuron cell bodies release extracellular ...vesicles, including exosomes containing miRs, upon activity. We demonstrate that miR-21-5p is released in the exosomal fraction of cultured DRG following capsaicin activation of TRPV1 receptors. Pure sensory neuron-derived exosomes released by capsaicin are readily phagocytosed by macrophages in which an increase in miR-21-5p expression promotes a pro-inflammatory phenotype. After nerve injury in mice, miR-21-5p is upregulated in DRG neurons and both intrathecal delivery of a miR-21-5p antagomir and conditional deletion of miR-21 in sensory neurons reduce neuropathic hypersensitivity as well as the extent of inflammatory macrophage recruitment in the DRG. We suggest that upregulation and release of miR-21 contribute to sensory neuron-macrophage communication after damage to the peripheral nerve.
The integration of somatosensory information is generally assumed to be a function of the central nervous system (CNS). Here we describe fully functional GABAergic communication within rodent ...peripheral sensory ganglia and show that it can modulate transmission of pain-related signals from the peripheral sensory nerves to the CNS. We found that sensory neurons express major proteins necessary for GABA synthesis and release and that sensory neurons released GABA in response to depolarization. In vivo focal infusion of GABA or GABA reuptake inhibitor to sensory ganglia dramatically reduced acute peripherally induced nociception and alleviated neuropathic and inflammatory pain. In addition, focal application of GABA receptor antagonists to sensory ganglia triggered or exacerbated peripherally induced nociception. We also demonstrated that chemogenetic or optogenetic depolarization of GABAergic dorsal root ganglion neurons in vivo reduced acute and chronic peripherally induced nociception. Mechanistically, GABA depolarized the majority of sensory neuron somata, yet produced a net inhibitory effect on the nociceptive transmission due to the filtering effect at nociceptive fiber T-junctions. Our findings indicate that peripheral somatosensory ganglia represent a hitherto underappreciated site of somatosensory signal integration and offer a potential target for therapeutic intervention.
•Platins, taxanes, vincalkaloids, proteasome inhibitors are effective anticancer drugs.•Clinical use causes peripheral neurotoxicity impairing patients’ quality of life.•Several mechanisms are ...involved in the development of peripheral neurotoxicity.•Mechanisms knowledge is useful for the development of neuroprotective strategies.
Cisplatin, oxaliplatin, paclitaxel, vincristine and bortezomib are some of the most effective drugs successfully employed (alone or in combinations) as first-line treatment for common cancers. However they often caused severe peripheral neurotoxicity and neuropathic pain. Structural deficits in Dorsal Root Ganglia and sensory nerves caused symptoms as sensory loss, paresthesia, dysaesthesia and numbness that result in patient’ suffering and also limit the life-saving therapy. Several scientists have explored the various mechanisms involved in the onset of chemotherapy-related peripheral neurotoxicity identifying molecular targets useful for the development of selected neuroprotective strategies. Dorsal Root Ganglia sensory neurons, satellite cells, Schwann cells, as well as neuronal and glial cells in the spinal cord, are the preferential sites in which chemotherapy neurotoxicity occurs. DNA damage, alterations in cellular system repairs, mitochondria changes, increased intracellular reactive oxygen species, alterations in ion channels, glutamate signalling, MAP-kinases and nociceptors ectopic activation are among the events that trigger the onset of peripheral neurotoxicity and neuropathic pain. In the present work we review the role of the main players in determining the pathogenesis of anticancer drugs-induced peripheral neuropathy.
The objective of study was to investigate the inhibitory effect of sinomenine on neuropathic pain on dorsal root ganglia (DRG). The DRG cell line and spinal nerve ligation (SNL) model were used in ...this study. The effect of sinomenine on the cell viability was examined by MTT assay. The expression of p38 MAPK, NF-κB, c-fos, SP and TNF-α was detected by using immunofluorescence and immunohistochemistry assay. We also assessed the level of p-CaMKII, COX-2, p-CREB, IL-17A, TLR4 and IL-1β via western blotting and RT-qPCR. Compared to the controls, sinomenine showed a protective effect on TNF-α-induced apoptosis on DRG cells in a dose-dependent manner, with an increase of cell viability and a decrease of reactive oxygen species level as well as LDH release. Parallelly, sinomenine treatment significantly reduced the expression of various factors related to stress and inflammation, including p38 MAPK, NF-κB, c-fos, p-CAMKII, COX-2, p-CREB, TLR4 and IL-17A in DRG cells in vitro. Furthermore, we found that administration of sinomenine significantly reduced mechanical withdrawal threshold and thermal withdrawal latency and inhibited the inflammation and activation of p38 signaling in SNL rats. It is noting that combined therapy of sinomenine and pulsed radiofrequency exhibited higher efficacy of dorsal root ganglia inflammation than single treatment as well as the combination of oxycodone and pulsed radiofrequency. Sinomenine inhibited the apoptosis of DRG cell by regulating p38 MAPK/CREB signalling pathway, which provides evidence to alleviate neuropathic pain in clinic.
Reactive oxygen species (ROS) contribute to tissue damage and remodelling mediated by the inflammatory response after injury. Here we show that ROS, which promote axonal dieback and degeneration ...after injury, are also required for axonal regeneration and functional recovery after spinal injury. We find that ROS production in the injured sciatic nerve and dorsal root ganglia requires CX3CR1-dependent recruitment of inflammatory cells. Next, exosomes containing functional NADPH oxidase 2 complexes are released from macrophages and incorporated into injured axons via endocytosis. Once in axonal endosomes, active NOX2 is retrogradely transported to the cell body through an importin-β1-dynein-dependent mechanism. Endosomal NOX2 oxidizes PTEN, which leads to its inactivation, thus stimulating PI3K-phosporylated (p-)Akt signalling and regenerative outgrowth. Challenging the view that ROS are exclusively involved in nerve degeneration, we propose a previously unrecognized role of ROS in mammalian axonal regeneration through a NOX2-PI3K-p-Akt signalling pathway.
NF-κB has an essential role in the initiation and progression of pancreatic cancer and specifically mediates the induction of epithelial–mesenchymal transition and invasiveness. In this study, we ...demonstrate the importance of activated NF-κB signaling in EMT induction, lymphovascular metastasis, and neural invasion. Modulation of NF-κB activity was accomplished through the specific NF-κB inhibitor (BAY 11-7085), triptolide, and Minnelide treatment, as well as overexpression of IKBα repressor and IKK activator plasmids. In the classical lymphovascular metastatic cascade, inhibition of NF-κB decreased the expression of several EMT transcription factors (SNAI1, SNAI2, and ZEB1) and mesenchymal markers (VIM and CDH2) and decreased in vitro invasion, which was rescued by IKK activation. This was further demonstrated in vivo via BAY 11-7085 treatment in a orthotopic model of pancreatic cancer. In vivo NF-κB inhibition decreased tumor volume; decreased tumor EMT gene expression, while restoring cell–cell junctions; and decreasing overall metastasis. Furthermore, we demonstrate the importance of active NF-κB signaling in neural invasion. Triptolide treatment inhibits Nerve Growth Factor (NGF) mediated, neural-tumor co-culture in vitro invasion, and dorsal root ganglia (DRG) neural outgrowth through a disruption in tumor-neural cross talk. In vivo, Minnelide treatment decreased neurotrophin expression, nerve density, and sciatic nerve invasion. Taken together, this study demonstrates the importance of NF-κB signaling in the progression of pancreatic cancer through the modulation of EMT induction, lymphovascular invasion, and neural invasion.
Mechanical allodynia, induced by normally innocuous low-threshold mechanical stimulation, represents a cardinal feature of neuropathic pain. Blockade or ablation of high-threshold, small-diameter ...unmyelinated group C nerve fibers (C-fibers) has limited effects on mechanical allodynia. Although large, myelinated group A fibers, in particular Aβ-fibers, have previously been implicated in mechanical allodynia, an A-fiber-selective pharmacological blocker is still lacking. Here we report a new method for targeted silencing of A-fibers in neuropathic pain. We found that Toll-like receptor 5 (TLR5) is co-expressed with neurofilament-200 in large-diameter A-fiber neurons in the dorsal root ganglion (DRG). Activation of TLR5 with its ligand flagellin results in neuronal entry of the membrane-impermeable lidocaine derivative QX-314, leading to TLR5-dependent blockade of sodium currents, predominantly in A-fiber neurons of mouse DRGs. Intraplantar co-application of flagellin and QX-314 (flagellin/QX-314) dose-dependently suppresses mechanical allodynia after chemotherapy, nerve injury, and diabetic neuropathy, but this blockade is abrogated in Tlr5-deficient mice. In vivo electrophysiology demonstrated that co-application of flagellin/QX-314 selectively suppressed Aβ-fiber conduction in naive and chemotherapy-treated mice. TLR5-mediated Aβ-fiber blockade, but not capsaicin-mediated C-fiber blockade, also reduced chemotherapy-induced ongoing pain without impairing motor function. Finally, flagellin/QX-314 co-application suppressed sodium currents in large-diameter human DRG neurons. Thus, our findings provide a new tool for targeted silencing of Aβ-fibers and neuropathic pain treatment.
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