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Despite being routinely used for pain management, opioid use is limited due to adverse effects such as development of tolerance and paradoxical pain, including thermal hyperalgesia and mechanical allodynia. Evidence indicates that continued morphine administration causes increased expression of proinflammatory mediators such as tumor necrosis factor-alpha (TNF-α). The objectives of the present study were to determine the effects of B1 (-[(1-benzimidazol-2-yl)methyl]-4-methoxyaniline) and B8 (-{4-[(1-benzimidazol-2-yl)methoxy]phenyl}acetamide), benzimidazole derivatives, on thermal nociception and mechanical allodynia during repeated morphine (intraperitoneal; 5 mg/kg twice daily for 6 days)-induced paradoxical pain and TNF-α expression in the spinal cord in mice. Our data indicate that administration of benzimidazole derivatives attenuated morphine-induced thermal hyperalgesia and mechanical allodynia. Benzimidazole derivatives also reduced TNF-α expression in mice. Taken together, these results suggest that benzimidazole derivatives might be useful for the treatment of neuroinflammatory consequences of continued morphine administration and could be potential drug candidates for the management of opioid-induced paradoxical pain.
Learn More >Acupuncture therapy is effective for relieving postoperative pain. Our previous study showed that electroacupuncture (EA) at Futu (LI18) and Hegu (LI4)-Neiguan (PC6) could alleviate incisional neck pain, which was related with its effect in upregulating γ-aminobutyric acid (GABA) expression in cervical (C3-6) dorsal root ganglions (DRGs); but whether its receptor subsets GABAα2R and GABAR1 in C3-6 DRGs are involved in EA analgesia or not, it remains unknown.
Learn More >Sigma-1 (σ) receptor antagonists are promising tools for neuropathic pain treatment, but it is unknown whether σ receptor inhibition ameliorates the neuropathic signs induced by nerve transection, in which the pathophysiological mechanisms and response to drug treatment differ from other neuropathic pain models. In addition, σ antagonism ameliorates inflammatory pain through modulation of the endogenous opioid system, but it is unknown whether this occurs during neuropathic pain. We investigated the effect of σ inhibition on the painful hypersensitivity associated with the spared nerve injury (SNI) model in mice. Wild-type (WT) mice developed prominent cold (acetone test), mechanical (von Frey test), and heat hypersensitivity (Hargreaves test) after SNI. σ receptor knockout (ခσ-KO) mice did not develop cold allodynia and showed significantly less mechanical allodynia, although they developed heat hyperalgesia after SNI. The systemic acute administration of the selective σ receptor antagonist S1RA attenuated all three types of SNI-induced hypersensitivity in WT mice. These ameliorative effects of S1RA were reversed by the administration of the σ agonist PRE-084, and were absent in σ-KO mice, indicating the selectivity of S1RA-induced effects. The opioid antagonist naloxone and its peripherally restricted analog naloxone methiodide prevented S1RA-induced effects in mechanical and heat hypersensitivity, but not in cold allodynia, indicating that opioid-dependent and -independent mechanisms are involved in the effects of this σ antagonist. The repeated administration of S1RA twice a day during 10 days reduced SNI-induced cold, mechanical, and heat hypersensitivity without inducing analgesic tolerance during treatment. These effects were observed up to 12 h after the last administration, when S1RA was undetectable in plasma or brain, indicating long-lasting pharmacodynamic effects. These data suggest that σ antagonism may have therapeutic value for the treatment of neuropathic pain induced by the transection of peripheral nerves.
Learn More >Peripheral nerve injury can induce neuroplastic changes in the central nervous system and result in neuropathic pain. This study investigated functional involvement in dorsal paratrigeminal nucleus (dPa5) and nucleus tractus solitarii (NTS) neurons projecting to the parabrachial nucleus (PBN) after trigeminal nerve injury. Anatomical quantification was performed based on phosphorylated extracellular signal-regulated kinase (pERK) expression underlying orofacial neuropathic pain associated with infraorbital nerve chronic constriction injury (ION-CCI) in rats. ION-CCI rats exhibited heat and mechanical hypersensitivity in the ipsilateral upper lip. After injection of retrograde tracer fluorogold (FG) into the contralateral PBN, ION-CCI rats received capsaicin or noxious mechanical stimulation to the upper lip. The total number of FG-labeled neurons in dPa5 and NTS did not change after ION-CCI, and pERK expression in dPa5 did not differ between sham and ION-CCI rats. In the NTS contralateral to ION-CCI, the number of pERK-immunoreactive neurons and percentage of pERK-immunoreactive FG-labeled PBN projection neurons were increased after capsaicin stimulation in ION-CCI rats. The present findings suggest that enhanced noxious inputs from the NTS to the PBN after trigeminal nerve injury modulates PBN neuron activity, which accompanies the affective components of orofacial neuropathic pain.
Learn More >P2X receptors constitute a gene family of cation channels gated by extracellular ATP. They mediate fast ionotropic purinergic signaling in neurons and non-excitable cell types in vertebrates. The highly calcium-permeable P2X4 subtype has been shown to play a significant role in cardiovascular physiology, inflammatory responses and neuro-immune communication. We previously reported the discovery of a P2X4-selective antagonist, the small organic compound BX430, with submicromolar potency for human P2X4 receptors and marked species-dependence (Ase et al., 2015). The present study investigates the molecular basis of P2X4 inhibition by the non-competitive blocker BX430 using a structural and functional approach relying on mutagenesis and electrophysiology. We provide evidence for the critical contribution of a single hydrophobic residue located in the ectodomain of P2X4 channel subunits, Ile312 in human P2X4, which determines blockade by BX430. We also show that the nature of this extracellular residue in various vertebrate P2X4 orthologs underlies their specific sensitivity or resistance to the inhibitory effects of BX430. Taking advantage of high-resolution crystallographic data available on zebrafish P2X4, we used molecular dynamics simulation to model the docking of BX430 on an allosteric binding site around Ile315 (zebrafish numbering) in the ectodomain of P2X4. We also observed that the only substitution I312D (human numbering) that renders P2X4 silent by itself has also a profound silencing effect on all other P2X subtypes tested when introduced at homologous positions. The generic impact of this aspartate mutation on P2X function indicates that the pre-TM2 subregion involved is conserved functionally and defines a novel allosteric inhibitory site present in all P2X receptor channels. This conserved structure-channel activity relationship might be exploited for the rational design of potent P2X subtype-selective antagonists of therapeutic value.
Learn More >Up to two-thirds of patients affected by spinal cord injury (SCI) develop central neuropathic pain (CNP), which has a high impact on their quality of life. Most of the patients are largely refractory to current treatments, and new pharmacological strategies are needed. Recently, it has been shown that the acute administration of the σ1R antagonist MR309 (previously developed as E-52862) at 28 days after spinal cord contusion results in a dose-dependent suppression of both mechanical allodynia and thermal hyperalgesia in wild-type CD-1 Swiss female mice. The present work was addressed to determine whether MR309 might exert preventive effects on CNP development by repeated administration during the first week after SCI in mice. To this end, the MR309 (16 or 32 mg/kg i.p.) modulation on both thermal hyperalgesia and mechanical allodynia development were evaluated weekly up to 28 days post-injury. In addition, changes in pro-inflammatory cytokine (TNF-α, IL-1β) expression and both the expression and activation (phosphorylation) of the N-methyl-D-aspartate receptor subunit 2B (NR2B-NMDA) and extracellular signal-regulated kinases (ERK1/2) were analyzed. The repeated treatment of SCI-mice with MR309 resulted in significant pain behavior attenuation beyond the end of the administration period, accompanied by reduced expression of central sensitization-related mechanistic correlates, including extracellular mediators (TNF-α and IL-1β), membrane receptors/channels (NR2B-NMDA) and intracellular signaling cascades (ERK/pERK). These findings suggest that repeated MR309 treatment after SCI may be a suitable pharmacologic strategy to modulate SCI-induced CNP development.
Learn More >Pain (nociceptive) input caudal to a spinal contusion injury can undermine long-term recovery and increase tissue loss (secondary injury). Prior work suggests that nociceptive stimulation has this effect because it fosters the breakdown of the blood-spinal cord barrier (BSCB) at the site of injury, allowing blood to infiltrate the tissue. The present study examined whether these effects impact tissue rostral and caudal to the site of injury. In addition, the study evaluated whether cutting communication with the brain, by means of a rostral transection, affects the development of hemorrhage. Eighteen hours after rats received a lower thoracic (T11-12) contusion injury, half underwent a spinal transection at T2. Noxious electrical stimulation (shock) was applied 6 h later. Cellular assays showed that, in non-transected rats, nociceptive stimulation increased hemoglobin content, activated pro-inflammatory cytokines and engaged signals related to cell death at the site of injury. These effects were not observed in transected animals. In the next experiment, the spinal transection was performed at the time of contusion injury. Nociceptive stimulation was applied 24 h later and tissue was sectioned for microscopy. In non-transected rats, nociceptive stimulation increased the area of hemorrhage and this effect was blocked by spinal transection. These findings imply that the adverse effect of noxious stimulation depends upon spared ascending fibers and the activation of rostral (brain) systems. If true, stimulation should induce less hemorrhage after a severe contusion injury that blocks transmission to the brain. To test this, rats were given a mild, moderate, or severe, injury and electrical stimulation was applied 24 h later. Histological analyses of longitudinal sections showed that nociceptive stimulation triggered less hemorrhage after a severe contusion injury. The results suggest that brain-dependent processes drive pain-induced hemorrhage after spinal cord injury (SCI).
Learn More >Oral cancer patients report severe function-induced pain; severity is greater in females. We hypothesize that a neutrophil-mediated endogenous analgesic mechanism is responsible for sex differences in nociception secondary to oral squamous cell carcinoma (SCC). Neutrophils isolated from the cancer-induced inflammatory microenvironment contain β-endorphin protein and are identified by the Ly6G immune marker. We previously demonstrated that male mice with carcinogen-induced oral SCC exhibit less nociceptive behavior and a higher concentration of neutrophils in the cancer microenvironment compared to female mice with oral SCC. Oral cancer cells secrete granulocyte colony stimulating factor (G-CSF), a growth factor that recruits neutrophils from bone marrow to the cancer microenvironment. We found that recombinant G-CSF (rG-CSF, 5 μg/mouse, intraperitoneal) significantly increased circulating Ly6G neutrophils in the blood of male and female mice within 24 h of administration. In an oral cancer supernatant mouse model, rG-CSF treatment increased cancer-recruited Ly6G neutrophil infiltration and abolished orofacial nociceptive behavior evoked in response to oral cancer supernatant in both male and female mice. Local naloxone treatment restored the cancer mediator-induced nociceptive behavior. We infer that rG-CSF-induced Ly6G neutrophils drive an endogenous analgesic mechanism. We then evaluated the efficacy of chronic rG-CSF administration to attenuate oral cancer-induced nociception using a tongue xenograft cancer model with the HSC-3 human oral cancer cell line. Saline-treated male mice with HSC-3 tumors exhibited less oral cancer-induced nociceptive behavior and had more β-endorphin protein in the cancer microenvironment than saline-treated female mice with HSC-3 tumors. Chronic rG-CSF treatment (2.5 μg/mouse, every 72 h) increased the HSC-3 recruited Ly6G neutrophils, increased β-endorphin protein content in the tongue and attenuated nociceptive behavior in female mice with HSC-3 tumors. From these data, we conclude that neutrophil-mediated endogenous opioids warrant further investigation as a potential strategy for oral cancer pain treatment.
Learn More >Recent findings from rodent studies suggest that high-fat diet (HFD) increases hyperalgesia independent of obesity status. Furthermore, weight loss interventions such as voluntary physical activity (PA) for adults with obesity or overweight was reported to promote pain reduction in humans with chronic pain. However, regardless of obesity status, it is not known whether HFD intake and sedentary (SED) behavior is underlies chronic pain susceptibility. Moreover, differential gene expression in the nucleus accumbens (NAc) plays a crucial role in chronic pain susceptibility. Thus, the present study used an adapted model of the inflammatory prostaglandin E2 (PGE2)-induced persistent hyperalgesia short-term (PH-ST) protocol for mice, an HFD, and a voluntary PA paradigm to test these hypotheses. Therefore, we performed an analysis of differential gene expression using a transcriptome approach of the NAc. We also applied a gene ontology enrichment tools to identify biological processes associated with chronic pain susceptibility and to investigate the interaction between the factors studied: diet (standard diet vs. HFD), physical activity behavior (SED vs. PA) and PH-ST (PGE vs. saline). Our results demonstrated that HFD intake and sedentary behavior promoted chronic pain susceptibility, which in turn was prevented by voluntary physical activity, even when the animals were fed an HFD. The transcriptome of the NAc found 2,204 differential expression genes and gene ontology enrichment analysis revealed 41 biologic processes implicated in chronic pain susceptibility. Taking these biological processes together, our results suggest that genes related to metabolic and mitochondria stress were up-regulated in the chronic pain susceptibility group (SED-HFD-PGE), whereas genes related to neuroplasticity were up-regulated in the non-chronic pain susceptibility group (PA-HFD-PGE). These findings provide pieces of evidence that HFD intake and sedentary behavior provoked gene expression changes in the NAc related to promotion of chronic pain susceptibility, whereas voluntary physical activity provoked gene expression changes in the NAc related to prevention of chronic pain susceptibility. Finally, our findings confirmed previous literature supporting the crucial role of voluntary physical activity to prevent chronic pain and suggest that low levels of voluntary physical activity would be helpful and highly recommended as a complementary treatment for those with chronic pain.
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