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Spinal disinhibition has been hypothesized to underlie pain hypersensitivity in neuropathic pain. Apparently contradictory mechanisms have been reported, raising questions on the best target to produce analgesia. Here, we show that nerve injury is associated with a reduction in the number of inhibitory synapses in the spinal dorsal horn. Paradoxically, this is accompanied by a BDNF-TrkB-mediated upregulation of synaptic GABARs and by an α1-to-α2GABAR subunit switch, providing a mechanistic rationale for the analgesic action of the α2,3GABAR benzodiazepine-site ligand L838,417 after nerve injury. Yet, we demonstrate that impaired Cl extrusion underlies the failure of L838,417 to induce analgesia at high doses due to a resulting collapse in Cl gradient, dramatically limiting the benzodiazepine therapeutic window. In turn, enhancing KCC2 activity not only potentiated L838,417-induced analgesia, it rescued its analgesic potential at high doses, revealing a novel strategy for analgesia in pathological pain, by combined targeting of the appropriate GABAR-subtypes and restoring Cl homeostasis.
Learn More >The mechanical head-withdrawal threshold (MHWT) was significantly reduced following inferior alveolar nerve transection (IANX) in rats. Nitrate and nitrite synthesis was dramatically increased in the trigeminal ganglion (TG) at 6 h after the IANX. The relative number of neuronal nitric oxide synthase (nNOS)-immunoreactive (IR) cells was significantly higher in IANX rats compared to sham-operated and N-propyl-L-arginine (NPLA)-treated IANX rats. On day 3 after NPLA administration, the MHWT recovered considerably in IANX rats. Following L-arginine injection into the TG, the MHWT was significantly reduced within 15 min, and the mean number of TG cells encircled by glial fibrillary acidic protein (GFAP)-IR cells was substantially higher. The relative number of nNOS-IR cells encircled by GFAP-IR cells was significantly increased in IANX rats. In contrast, after NPLA injection into the TG, the relative number of GFAP-IR cells was considerably reduced in IANX rats. Fluorocitrate administration into the TG significantly reduced the number of GFAP-IR cells and prevented the MHWT reduction in IANX rats. The present findings suggest that following IANX, satellite glial cells are activated via nitric oxide (NO) signaling from TG neurons. The spreading satellite glial cell activation within the TG results in mechanical hypersensitivity of face regions not directly associated with the trigeminal nerve injury.
Learn More >Metastatic breast cancer is prevalent worldwide, and one of the most common sites of metastasis are long bones. Of patients with disease, the major symptom is pain, yet current medications fail to adequately result in analgesic efficacy and present major undesirable adverse effects. In our study we investigate the potential of a novel monoacylglycerol lipase (MAGL) inhibitor, MJN110, in a murine model of cancer induced bone pain (CIBP). Literature has previously demonstrated that MAGL inhibitors function to increase the endogenous concentrations of 2-arachydonylglycerol, which then activate CB1 and CB2 receptors inhibiting inflammation and pain. We demonstrate that administration of MJN110 significantly and dose-dependently alleviates spontaneous pain behavior during acute administration compared to vehicle control. In addition, the MJN110 maintains its efficacy in a chronic dosing paradigm over the course of 7 days without signs of receptor sensitization. In vitro analysis of MJN110 demonstrated a dose dependent and significant decrease in cell viability of 66.1 breast adenocarcinoma cells and to a greater extent than KML29, an alternate MAGL inhibitor, or the CB2 agonist JWH015. Chronic administration of the compound did not appear to affect tumor burden evidenced by radiograph or histological analysis. Together, these data support the application for MJN110 as a novel therapeutic for cancer induced bone pain. SIGNIFICANCE STATEMENT: Current standard of care for metastatic breast cancer pain is opioid-based therapies with adjunctive chemotherapy, which have highly addictive and other deleterious side effects. The need for effective, non-opioid based therapies is essential and harnessing the endogenous cannabinoid system is proving to be a new target to treat various types of pain conditions. We present a novel drug targeting the endogenous cannabinoid system that is effective at reducing pain in a mouse model of metastatic breast cancer to bone.
Learn More >Central poststroke pain (CPSP) is one of the neuropathic pain syndromes that can occur following stroke involving the somatosensory system. However, the underlying mechanism of CPSP remains largely unknown. Here, we established a CPSP mouse model by inducing a focal hemorrhage in the thalamic ventrobasal complex and confirmed the development of mechanical allodynia. In this model, microglial activation was observed in the somatosensory cortex, as well as in the injured thalamus. By using a CSF1 receptor inhibitor, we showed that microglial depletion effectively prevented allodynia development in our CPSP model. In the critical phase of allodynia development, c-fos-positive neurons increased in the somatosensory cortex, accompanied by ectopic axonal sprouting of the thalamocortical projection. Furthermore, microglial ablation attenuated both neuronal hyperactivity in the somatosensory cortex and circuit reorganization. These findings suggest that microglia play a crucial role in the development of CPSP pathophysiology by promoting sensory circuit reorganization.
Learn More >In the pain matrix, the insular cortex (IC) is mainly involved in discriminative sensory and motivative emotion. Abnormal signal transmission from injury site causes neuropathic pain, which generates enhanced synaptic plasticity. The mammalian target of rapamycin (mTOR) complex is the key regulator of protein synthesis; it is involved in the modulation of synaptic plasticity. To date, there has been no report on the changes in optical signals in the IC under neuropathic condition after treatment with mTOR inhibitors, such as Torin1 and XL388. Therefore, we aimed to determine the pain-relieving effect of mTOR inhibitors (Torin1 and XL388) and observe the changes in optical signals in the IC after treatment. Mechanical threshold was measured in adult male Sprague-Dawley rats after neuropathic surgery, and therapeutic effect of inhibitors was assessed on post-operative day 7 following the microinjection of Torin1 or XL388 into the IC. Optical signals were acquired to observe the neuronal activity of the IC in response to peripheral stimulation before and after treatment with mTOR inhibitors. Consequently, the inhibitors showed the most effective alleviation 4 h after microinjection into the IC. In optical imaging, peak amplitudes of optical signals and areas of activated regions were reduced after treatment with Torin1 and XL388. However, there were no significant optical signal changes in the IC before and after vehicle application. These findings suggested that Torin1 and XL388 are associated with the alleviation of neuronal activity that is excessively manifested in the IC, and is assumed to diminish synaptic plasticity.
Learn More >Pain and emotional distress have a reciprocal relation. The amygdala has been implicated in emotional processing. The central nucleus of amygdala (CeA) receives nociceptive information from the dorsal horn of spinal cord, and responsible for the central plasticity in chronic pain. Neuropathic pain is a type of severe chronic pain and can be strongly influenced by emotional components. Plastic changes in CeA may play a key role in the development and/or maintenance of neuropathic pain. We studied the expression levels of proteins in CeA of spinal nerve transection (SNT) model rats. Total tissue lysate proteins were separated by two dimensional-gel electrophoresis (2D-PAGE). Gels from different time points were compared using Progenesis SameSpot software, and the spots with Fold Change greater than 2 were excised for the protein identification by mass spectrometry. We identified more than 50 cytosolic proteins as significantly altered in their expressions in CeA of SNT rats, and most of these changes have been validated at mRNA levels by qRT-PCR. We also identified more than 40 membrane proteins as notably up- or down-regulated in CeA of SNT model rats relative to a control using stable isotope dimethyl labeling nano-LC-MS/MS based proteomics and found that one of such protein, doublecortin, is specifically localized in the membrane fraction without changes in total amount of the protein. Immunohistochemistry showed that doublecortin is expressed in processes in CeA of rats 7 and 21 days post SNT surgery, suggesting doublecorin is one of the proteins which may contribute to the plastic changes in CeA in the neuropathic pain model. These dysregulated proteins may play roles in reciprocal relationships between pain and psychological distress in the amygdala and contribute to central sensitization.
Learn More >There exists a limited number of studies investigating the correlation between spinal adenosine A1 receptors and Vincristine-induced peripheral neuropathy (VIPN). This study explored the role of intrathecal N6-(2-phenylisopropyl)-adenosine R-(-)isomer (R-PIA) in neuropathic pain induced in rats by administering vincristine (100 μg/kg i.p.) for 10 days (two 5-day cycles with a 2-day pause).
Learn More >G protein-coupled receptors can signal through both G proteins and ß-arrestin2. For the μ-opioid receptor, early experimental evidence from a single study suggested that G protein signalling mediates analgesia whereas ß-arrestin2 signalling mediates respiratory depression and constipation. Consequently, for more than a decade much research effort has been focused on developing biased μ-opioid agonists that preferentially target G protein signalling over β-arrestin signalling as it was believed that such drugs would be analgesics devoid of respiratory depressant activity. However, the prototypical compounds that have been developed based on this concept have so far failed in clinical and preclinical development.
Learn More >Cancer-induced pain (CIP) is a kind of chronic pain that occurs during cancer progression over time. However, the mechanisms are largely unknown, and clinical treatment remains challenging. LncRNAs have been reported to play critical roles in various biological processes, including chronic pain. The aim of our study was to investigate whether lncRNAs participate in the development of CIP by regulating the expression levels of some molecules related to pain modulation. The CIP model was established by injecting Walker 256 mammary gland tumor cells into the tibial canal of rats. In this study, we found that lncRNA-NONRATT021203.2 was increased in the CIP rats and that lncRNA-NONRATT021203.2-siRNA could relieve hyperalgesia in these rats. For elucidation of the underlying mechanism, we showed that lncRNA-NONRATT021203.2 could target C-X-C motif chemokine ligand 9 (CXCL9), which was increased in the CIP rats, and that CXCL9-siRNA could relieve hyperalgesia. At the same time, silencing lncRNA-NONRATT021203.2 expression decreased the mRNA and protein levels of CXCL9. Immunofluorescence analysis showed that CXCL9 was mainly expressed in the CGRP-positive and IB4-positive DRG neurons. Further research showed that lncRNA-NONRATT021203.2 and CXCL9 were colocalized in the DRG neurons. Our data suggested that lncRNA-NONRATT021203.2 participated in the CIP in rats and likely mediates the upregulation of CXCL9. The present study provided us with a new potential target for the clinical treatment of cancer-induced pain.
Learn More >Dipyrone is an analgesic pro-drug used clinically to control moderate pain with a high analgesic efficacy and low toxicity. Dipyrone is hydrolyzed to 4-methylaminoantipyrine (4-MAA), which is metabolized to 4-aminoantipyrine (4-AA). Here, were investigate the involvement of peripheral cannabinoid CB and opioid receptor activation in the local antihyperalgesic effect of dipyrone and 4-MAA. The inflammatory agent, carrageenan was administered to the hindpaw of male Wistar rats, and the mechanical nociceptive threshold was quantified by electronic von Frey test. Dipyrone or 4-MAA were locally administered 2.5 h after carrageenan. Following dipyrone injection, hindpaw tissue was harvested and its hydrolysis to 4-MAA was analyzed by mass spectrometry (MS). The selective CB receptor antagonist (AM630), naloxone (a non-selective opioid receptor antagonist), nor-BNI (a selective kappa-opioid receptor), CTOP (a selective mu-opioid receptor), or naltrindole (a selective delta-opioid receptor) was administered 30 min prior to 4-MAA. The results demonstrate that carrageenan-induced mechanical hyperalgesia was inhibited by dipyrone or 4-MAA in a dose-dependent manner. Dipyrone administered to the hindpaw was completely hydrolyzed to 4-MAA. The antihyperalgesic effect of 4-MAA was completely reversed by AM630, naloxone and nor-BNI, but not by CTOP or naltrindole. These data suggest that the local analgesic effect of dipyrone is mediated by its hydrolyzed bioactive form, 4-MAA and, at least in part, depends on CB receptor and kappa-opioid receptor activation. In conclusion, the analgesic effect of dipyrone may involve a possible interaction between the cannabinoid and opioid system in peripheral tissue.
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