Target For Treatment Of Pain Research Articles

Role of mu opioid receptor (MOR) agonist efficacy as a determinant of opioid antinociception in a novel assay of pain-depressed behavior in female and male mice.

Intermediate efficacy mu opioid receptor (MOR) agonists have potential to retain analgesic effectiveness while improving safety, but the optimal MOR efficacy for effective and safe opioid analgesia is unknown. Preclinical assays of pain-depressed behavior can assess effects of opioids and other candidate analgesics on pain-related behavioral depression, which is a common manifestation of clinically relevant pain and target of pain treatment. Accordingly, the present study goal was to validate a novel assay of pain-depressed locomotor behavior in mice and evaluate the role of MOR efficacy as a determinant of opioid analgesic effects and related safety measures. Male and female ICR mice were tested in a locomotor chamber consisting of 2 compartments connected by a doorway that contained a 1-inch-tall barrier. Dependent measures during 15-min behavioral sessions included crosses between compartments (which required vertical activity to surmount the barrier) and total movement counts (which required horizontal activity to break photobeams in each compartment). Intraperitoneal injection of lactic acid (IP acid) produced a concentration- and time-dependent depression of both endpoints. Optimal blockade of IP acid-induced behavioral depression with minimal motor impairment was achieved with intermediate-efficacy MOR treatments that also produced less gastrointestinal-transit inhibition and respiratory depression than the high-efficacy MOR agonist fentanyl. Sex differences in treatment effects were rare. Overall, these findings validate a novel procedure for evaluating opioids and other candidate analgesic effects on pain-related behavioral depression in mice and support continued research with intermediate-efficacy MOR agonists as a strategy to retain opioid analgesic effectiveness with improved safety.

Design, development and evaluation of a prodrug-type TrkA-selective inhibitor with antinociceptive effects in vivo

Tropomyosin receptor kinase A (TrkA) is considered a therapeutic target in pain treatment, and several TrkA inhibitors have been designed and developed as potential analgesics. The subselectivity toward TrkA is closely associated with safety and efficacy and was achieved by developing an allosteric inhibitor to occupy the less conserved non-ATP binding site. Druggable modifications of potent TrkA inhibitors could be challenging due to the unique requirement of functional groups that supply potency but are accompanied by poor druggability. We developed carboxyl acid 5 as a potent (IC50 = 22.4 nM) and subselective TrkA inhibitor (>8000-fold toward TrkB and TrkC). We also converted 5 to corresponding prodrug 39 as a useful tool compound in future analgesic development with promising antinociceptive effects in both hot plate testing and formalin-induced pain models with a suitable ED50 value (7.8 mg/kg).

Interactions between the Nociceptin and Toll-like Receptor Systems.

Nociceptin and the nociceptin receptor (NOP) have been described as targets for treatment of pain and inflammation, whereas toll-like receptors (TLRs) play key roles in inflammation and impact opioid receptors and endogenous opioids expression. In this study, interactions between the nociceptin and TLR systems were investigated. Human THP-1 cells were cultured with or without phorbol myristate acetate (PMA 5 ng/mL), agonists specific for TLR2 (lipoteichoic acid, LTA 10 µg/mL), TLR4 (lipopolysaccharide, LPS 100 ng/mL), TLR7 (imiquimod, IMQ 10 µg/mL), TLR9 (oligonucleotide (ODN) 2216 1 µM), PMA+TLR agonists, or nociceptin (0.01–100 nM). Prepronociceptin (ppNOC), NOP, and TLR mRNAs were quantified by RT-qPCR. Proteins were measured using flow cytometry. PMA upregulated ppNOC mRNA, intracellular nociceptin, and cell membrane NOP proteins (all p < 0.05). LTA and LPS prevented PMA’s upregulating effects on ppNOC mRNA and nociceptin protein (both p < 0.05). IMQ and ODN 2216 attenuated PMA’s effects on ppNOC mRNA. PMA, LPS, IMQ, and ODN 2216 increased NOP protein levels (all p < 0.05). PMA+TLR agonists had no effects on NOP compared to PMA controls. Nociceptin dose-dependently suppressed TLR2, TLR4, TLR7, and TLR9 proteins (all p < 0.01). Antagonistic effects observed between the nociceptin and TLR systems suggest that the nociceptin system plays an anti-inflammatory role in monocytes under inflammatory conditions.

Senso-Immunologic Prospects for Complex Regional Pain Syndrome Treatment.

Complex regional pain syndrome (CRPS) is a chronic pain syndrome that occurs in tissue injuries as the result of surgery, trauma, or ischemia. The clinical features of this severely painful condition include redness and swelling of the affected skin. Intriguingly, it was recently suggested that transient receptor potential ankyrin 1 (TRPA1) is involved in chronic post-ischemia pain, a CRPS model. TRPA1 is a non-selective cation channel expressed in calcitonin gene-related peptide (CGRP)-positive primary nociceptors that becomes highly activated in ischemic conditions, leading to the generation of pain. In this review, we summarize the history of TRPA1 and its involvement in pain sensation, inflammation, and CRPS. Furthermore, bone atrophy is also thought to be a characteristic clinical sign of CRPS. The altered bone microstructure of CRPS patients is thought to be caused by aggravated bone resorption via enhanced osteoclast differentiation and activation. Although TRPA1 could be a target for pain treatment in CRPS patients, we also discuss the paradoxical situation in this review. Nociceptor activation decreases the risk of bone destruction via CGRP secretion from free nerve endings. Thus, TRPA1 inhibition could cause severe bone atrophy. However, the suitable therapeutic strategy is controversial because the pathologic mechanisms of bone atrophy in CRPS are unclear. Therefore, we propose focusing on the remission of abnormal bone turnover observed in CRPS using a recently developed concept: senso-immunology.

Enterochromaffin Cells: Sentinels to Gut Microbiota in Hyperalgesia?

In recent years, increasing studies have been conducted on the mechanism of gut microbiota in neuropsychiatric diseases and non-neuropsychiatric diseases. The academic community has also recognized the existence of the microbiota-gut-brain axis. Chronic pain has always been an urgent difficulty for human beings, which often causes anxiety, depression, and other mental symptoms, seriously affecting people’s quality of life. Hyperalgesia is one of the main adverse reactions of chronic pain. The mechanism of gut microbiota in hyperalgesia has been extensively studied, providing a new target for pain treatment. Enterochromaffin cells, as the chief sentinel for sensing gut microbiota and its metabolites, can play an important role in the interaction between the gut microbiota and hyperalgesia through paracrine or neural pathways. Therefore, this systematic review describes the role of gut microbiota in the pathological mechanism of hyperalgesia, learns about the role of enterochromaffin cell receptors and secretions in hyperalgesia, and provides a new strategy for pain treatment by targeting enterochromaffin cells through restoring disturbed gut microbiota or supplementing probiotics.

Plasma-Based Inflammatory Signatures in Patients with Sickle Cell Disease during Baseline Health and Acute Pain

INTRODUCTION: Pain, the most common complication of sickle cell disease (SCD), presents as both sudden acute pain and chronic daily pain. However, there is wide variability in frequency and presentation of pain despite inheritance of the same monogenic gene defect. SCD has long been recognized as a chronic inflammatory condition. The ongoing effect of repeated vaso-occlusion, ischemia-reperfusion injury and hemolysis contribute to further SCD inflammation and likely pain. Regulation of the immune response can potentially modulate the inflammatory impact on pain. The collective balance of these inflammatory mediators in union in SCD patients and how this balance may change during baseline health and acute pain is unknown. The objective of this work was to determine the balance between patients' inflammatory and immune regulatory response and examine whether this balance changes during acute pain in patients with SCD. METHODS: We conducted a cross sectional analysis involving 3 cohorts: patients with SCD who were in their baseline health state, patients with SCD who had an acute pain episode and healthy African American controls. We used a novel bioassay originally developed for use in type 1 diabetes and applied to cystic fibrosis, inflammatory bowel disease and influenza to determine the inflammatory/immune regulatory response. This response was calculated as a composite Inflammatory Index (I.I.com) from these 3 patient cohorts. Patient plasma was co-cultured with cryopreserved PBMCs from a healthy donor to induce transcription (Figure 1). We identified informative transcripts that differentiate SCD patients from healthy controls thereby defining the disease-specific plasma-induced signature and retained ones differentially expressed between patients with SCD and controls that exhibit a fold change &amp;gt;1.4, ANOVA p-value of &amp;lt;0.05 and an FDR &amp;lt;10%. The data were subjected to ontological analyses for quantitative interpretation with Database for Annotation, Visualization and Integrated Discovery (DAVID) and Ingenuity Pathway Analysis (IPA). Our scoring strategy used the degree of induction of genes in inflammatory and regulatory ontological classes. The composite I.I.com was calculated using the average ratio between the mean log intensity of the genes classified as being "inflammatory" versus "regulatory". Independent samples Student's t-test was used to compare the mean I.I.com between 1) SCD baseline health cohort and controls and 2) SCD baseline health cohort and SCD acute pain cohort. RESULTS: Plasma from 16 patients with SCD in baseline health, 27 patients with SCD with an acute pain episode, and 45 African American controls were collected and analyzed. The average age of the study population was 12.6 (SD=3.6) years old and 52.3% were female. Quantitative scoring of plasma-induced signatures showed SCD patients had significantly higher mean I.I.com during baseline health compared to controls (0.713 vs. -1.235-12, p=5.4625-11). In addition, patients with SCD during acute pain episodes had significantly higher I.I.com than patients in baseline health (1.282 vs. 0.713, p=5.2051-8) (Figure 2). Heat map in Figure 3 shows differential gene expression between the cohorts; green and red colors in heat maps represent lower or higher relative expression respectively. CONCLUSION: Our findings show distinct immune signatures in SCD patients compared to controls and distinct signatures in SCD patients during acute pain episodes as compared to baseline health. The novel assay used to assess the inflammatory and immune regulatory gene expression in the three cohorts studied allowed for the determination of the balance between the two immune states. The imbalance between inflammation and immune regulation shown in our results in SCD patients of SCD pain. Further investigation into the specific inflammatory pathways that contribute to altered immune response could lead to novel targets for pain treatment. Disclosures Mucalo: NIH/NINDS: Research Funding; NIH/NHLBI: Research Funding. Jia:NIH/NHBLI: Research Funding; NIH/NINDS: Research Funding. Panepinto:NINDS: Research Funding; HRSA: Research Funding; NINDS: Research Funding; NHLBI: Research Funding. Roethle:NIH/NHLBI: Research Funding; NIH/NINDS: Research Funding. Hessner:NIH/NHLBI: Research Funding; NIH/NINDS: Research Funding. Brandow:NIH / NHLBI: Research Funding; Greater Milwaukee Foundation: Research Funding.

Acoustic Waves in Axonal Membrane and Caveolins are the New Targets for Pain Treatment with High Frequency Ultrasound.

Reciprocal interaction between electrical and mechanical waves observed in axonal membrane during its excitation leads to a paradigm shift in pain research making the uncoupling of electro-mechanical signals an interesting target in pain treatment. This uncoupling can be realized either through direct disturbance of the mechanical surface waves in axonal membrane or through shifting of the thermodynamic state of this membrane far from its phase transition point. Both effects can be effectively realized through application of the very high frequency ultrasound waves. Additional target for application of ultrasound in pain treatment is the caveolin-1, which is abundantly present in Schwann cells as well as in the non-axonal tissues. Both targets demonstrate frequency-dependent reactions, thus making a very high frequency ultrasound a promising treatment modality in pain treatment.

Status of peripheral sodium channel blockers for non-addictive pain treatment.

The effective and safe treatment of pain is an unmet health-care need. Current medications used for pain management are often only partially effective, carry dose-limiting adverse effects and are potentially addictive, highlighting the need for improved therapeutic agents. Most common pain conditions originate in the periphery, where dorsal root ganglion and trigeminal ganglion neurons feed pain information into the CNS. Voltage-gated sodium (NaV) channels drive neuronal excitability and three subtypes - NaV1.7, NaV1.8 and NaV1.9 - are preferentially expressed in the peripheral nervous system, suggesting that their inhibition might treat pain while avoiding central and cardiac adverse effects. Genetic and functional studies of human pain disorders have identified NaV1.7, NaV1.8 and NaV1.9 as mediators of pain and validated them as targets for pain treatment. Consequently, multiple NaV1.7-specific and NaV1.8-specific blockers have undergone clinical trials, with others in preclinical development, and the targeting of NaV1.9, although hampered by technical constraints, might also be moving ahead. In this Review, we summarize the clinical and preclinical literature describing compounds that target peripheral NaV channels and discuss the challenges and future prospects for the field. Although the potential of peripheral NaV channel inhibition for the treatment of pain has yet to be realized, this remains a promising strategy to achieve non-addictive analgesia for multiple pain conditions.

Epoxy Fatty Acids Are Promising Targets for Treatment of Pain, Cardiovascular Disease and Other Indications Characterized by Mitochondrial Dysfunction, Endoplasmic Stress and Inflammation.

Bioactive lipid mediators resulting from the metabolism of polyunsaturated fatty acids (PUFA) are controlled by many pathways that regulate the levels of these mediators and maintain homeostasis to prevent disease. PUFA metabolism is driven primarily through three pathways. Two pathways, the cyclooxygenase (COX) and lipoxygenase (LO) enzymatic pathways, form metabolites that are mostly inflammatory, while the third route of metabolism results from the oxidation by the cytochrome P450 enzymes to form hydroxylated PUFA and epoxide metabolites. These epoxygenated fatty acids (EpFA) demonstrate largely anti-inflammatory and beneficial properties, in contrast to the other metabolites formed from the degradation of PUFA. Dysregulation of these systems often leads to chronic disease. Pharmaceutical targets of disease focus on preventing the formation of inflammatory metabolites from the COX and LO pathways, while maintaining the EpFA and increasing their concentration in the body is seen as beneficial to treating and preventing disease. The soluble epoxide hydrolase (sEH) is the major route of metabolism of EpFA. Inhibiting its activity increases concentrations of beneficial EpFA, and often disease states correlate to mutations in the sEH enzyme that increase its activity and decrease the concentrations of EpFA in the body. Recent approaches to increasing EpFA include synthetic mimics that replicate biological activity of EpFA while preventing their metabolism, while other approaches focus on developing small molecule inhibitors to the sEH. Increasing EpFA concentrations in the body has demonstrated multiple beneficial effects in treating many diseases, including inflammatory and painful conditions, cardiovascular disease, neurological and disease of the central nervous system. Demonstration of efficacy in so many disease states can be explained by the fundamental mechanism that EpFA have of maintaining healthy microvasculature and preventing mitochondrial and endoplasmic reticulum stress. While there are no FDA approved methods that target the sEH or other enzymes responsible for metabolizing EpFA, current clinical efforts to test for efficacy by increasing EpFA that include inhibiting the sEH or administration of EpFA mimics that block metabolism are in progress.

Extracellular signal-regulated kinases (ERK) 1 and 2 as a key molecule in pain research.

Pain can be divided into nociceptive, inflammatory, and neuropathic pain. It is important to understanding the molecular mechanism of pain signaling in the development of pain relief therapies. Twenty years ago, extracellular signal-regulated kinases (ERK) 1 and 2, which are members of the mitogen-activated protein kinase superfamily, were identified as molecules activated in neurons by the exposure of peripheral tissues to noxious stimuli. Further studies have revealed that peripheral nerve injury induces ERK activation in glial cells, sensory neurons, and second-order neurons, albeit at different time points. Moreover, inhibition of ERK suppresses pathological pain in animals with peripheral nerve injury. Therefore, ERK is currently recognized as an important molecule in pain signaling and a potential novel target for pain treatment. This review introduces recent advances in revealing the regulation of ERK in pain research.

Pharmacological Blockade of Spinal CXCL3/CXCR2 Signaling by NVP CXCR2 20, a Selective CXCR2 Antagonist, Reduces Neuropathic Pain Following Peripheral Nerve Injury.

Recently, the role of CXCR2 in nociception has been noted. Our studies provide new evidence that the intrathecal administration of its CINC ligands (Cytokine-Induced Neutrophil Chemoattractant; CXCL1-3) induces pain-like behavior in naïve mice, and the effect occurring shortly after administration is associated with the neural location of CXCR2, as confirmed by immunofluorescence. RT-qPCR analysis showed, for the first time, raised levels of spinal CXCR2 after chronic constriction injury (CCI) of the sciatic nerve in rats. Originally, on day 2, we detected escalated levels of the spinal mRNA of all CINCs associated with enhancement of the protein level of CXCL3 lasting until day 7. Intrathecal administration of CXCL3 neutralizing antibody diminished neuropathic pain on day 7 after CCI. Interestingly, CXCL3 is produced in lipopolysaccharide-stimulated microglial, but not astroglial, primary cell cultures. We present the first evidence that chronic intrathecal administrations of the selective CXCR2 antagonist, NVP CXCR2 20, attenuate neuropathic pain symptoms and CXCL3 expression after CCI. Moreover, in naïve mice, this antagonist prevented CXCL3-induced hypersensitivity. However, NVP CXCR2 20 did not diminish glial activation, thus not enhancing morphine/buprenorphine analgesia. These results provide novel insight into the crucial role of CXCR2 in neuropathy based on CXCL3 modulation, which may become a potential therapeutic target in pain treatment.

The impact of changes in serum levels of metalloproteinase-2 and metalloproteinase-9 on pain perception in patients with disc herniation before and after surgery.

Purpose: The aim of our research was to investigate the link between serum levels of metalloproteinase-2 (MMP-2) and MMP-9, and the degree of pain experienced before and 1 and 3 months after microdiscectomy in 70 patients with disc herniation (DH).Patients and methods: The control group (group C) consisted of 70 healthy subjects and the DH group consisted of 70 patients with sciatica pain caused by lumbar DH. Before (DH0) and 1 and 3 months after surgery, the patients were assessed in terms of the following biochemical parameters: MMP-2, tissue inhibitors of metalloproteinases-2 (TIMP-2), MMP-2/TIMP-2, MMP-9, TIMP-1, and MMP-9/TIMP1, and the following clinical parameters: Numeric Rating Scale for the back (NRS-B) and the leg (NRS-L) and the Pain Rating Index (PRI) and Present Pain Intensity (PPI) of the McGill Pain Questionnaire.Results: No statistically significant correlations were observed following the biochemical and clinical assessments performed in group C and the DH group before surgery. After surgery (1 month), higher levels of TIMP-1 correlated with higher levels of NRS-B (rs =0.27; p<0.05). At 3 months after surgery higher levels of TIMP-2 and lower levels of MMP-2/TIMP-2 were correlated with higher levels of NRS-L (rs =0.27, p<0.05 and rs =−0.31, p<0.05, respectively) and higher levels of TIMP-2 were correlated with higher PRI scores (rs =0.27; p<0.005) and PPI scores (rs =0.35; p<0.01).Conclusion: The results showed that MMPs are involved in DH and play a significant role in the perception of pain after DH surgery. However, the value of MMPs as a potential therapeutic target in pain treatment should be considered cautiously.

Biophysical and Pharmacological Profiling of Multiple Voltage-Gated Sodium Channel Subtypes on QPatch II

Voltage - gated sodium channels (VGSC) are responsible for the initiation and propagation of action potentials in excitable cells. VGSC have been identified as excellent drug targets for treatment of pain, epilepsy and to other neurological disorders. Early compounds, however, were developed using empirical approaches. The identification of the molecular identity of VGSC in combination with technological advances, such as the automated patch clamp technique, provide the basis for a rational design of subtype – selective compounds. To date, 9 functional mammalian isoforms (NaV1.1–1.9) have been described in the literature. The various subtypes differ in their expression pattern and exhibit distinct biophysical and pharmacological profiles. All have in common that they produce a transient inward current in response to membrane depolarization. During this process the VGSC transitions from a closed to and open into an inactivated state. Interestingly, inhibitor compounds often exhibit different pharmacological profiles dependent upon the ion channel conformational state. In the present study, the second generation QPatch (QPatch II; Sophion Bioscience) was used in combination with adaptive voltage protocols to investigate state-dependent inhibition of tetrodotoxin (TTX) and tetracaine on 8 different VGSC subtypes (NaV1.1-8). A first step was to determine the half inactivation potential V½(inactivation) for each individual cell. This value was then used during the next steps as preconditioning pulse. Such an adaptive protocol allowed to determine IC50 values for both the closed and the inactivated state and reduce heterogeneity of the cells. Both IC50 values and biophysical parameters of the different subtypes align well with literature values.

Impaired Junctions and Invaded Macrophages in Oral Epithelia With Oral Pain.

Recurrent or chronic oral pain is a great burden for patients. Recently, the links between epithelial barrier loss and disease were extended to include initiation and propagation. To explore the effects of pathohistological changes in oral epithelia on pain, we utilized labial mucosa samples in diagnostic labial gland biopsies from patients with suspected Sjögren's syndrome (SS), because they frequently experience pain and discomfort. In most labial mucosa samples from patients diagnosed with SS, disseminated epithelial cellular edema was prevalent as ballooning degeneration. The disrupted epithelia contained larger numbers of infiltrating macrophages in patients with oral pain than in patients without pain. Immunohistochemistry revealed that edematous areas were distinct from normal areas, with disarranged cell-cell adhesion molecules (filamentous actin, E-cadherin, β-catenin). Furthermore, edematous areas were devoid of immunostaining for transient receptor potential channel vanilloid 4 (TRPV4), a key molecule in adherens junctions. In an investigation on whether impaired TRPV4 affect cell-cell adhesion, calcium stimulation induced intimate cell-cell contacts among oral epithelial cells from wild-type mice, while intercellular spaces were apparent in cells from TRPV4-knockout mice. The present findings highlight the relationship between macrophages and epithelia in oral pain processing, and identify TRPV4-mediated cell-cell contacts as a possible target for pain treatment.

A Functional Kinase Short Interfering Ribonucleic Acid Screen Using Protease-Activated Receptor 2-Dependent Opening of Transient Receptor Potential Vanilloid-4.

Protease-activated receptor 2 (PAR2) is a proinflammatory G-protein coupled receptor (GPCR) that is activated by inflammatory proteases, and its activation initiates signaling pathways that modulate the nonselective cation channel transient receptor potential vanilloid-4 (TRPV4). PAR2-dependent opening of TRPV4 has been attributed to kinase activation, but the identity of the responsible enzymes is unknown. Deciphering the signaling pathways involved in the PAR2-dependent opening of TRPV4 may yield new targets for pain treatment. This study has identified specific kinases that are involved in opening TRPV4, using a selective screen of short interfering ribonucleic acid (siRNA) SMARTpools, which individually targeted all human kinases, in human embryonic kidney 293 (HEK293) cells that stably express inducible TRPV4. This screen is unique because it uses a real-time assay measuring intracellular calcium with Fura-2AM dye. From the primary screen, subsequent confirmation screen, and on-target messenger ribonucleic acid expression analysis, we identified two kinases as crucial to the PAR2-dependent opening of TRPV4 in HEK293 cells, mitogen-activated protein kinase 13 and with no lysine kinase 4. In conclusion, this study describes a powerful new application of siRNA knockdown to identity signaling molecules that are responsible for the PAR2-dependent opening of TRPV4, which will help elucidate this signaling process.

Ion channel TRPA1 is a promising therapeutic target for treatment of pain

TRPA1 is an ankyrin receptor of TRP family. It is a non-selective, calcium-permeable cation channel that is highly expressed by a subset of small diameter sensory neurons with cell bodies in the dorsal root, trigeminal, nodose and jugular ganglia. In the last time information about the role of TRPA1 in pain and cold sensitivity, as well as in the formation and maintenance of inflammation is increasing in scientific literature. Given this information, the interest for search and study of pharmacological agents, which selectively blocked of TRPA1 and reduced the severity of pain and inflammation is increasing.

A novel disorder reveals clathrin heavy chain-22 is essential for human pain and touch development.

Congenital inability to feel pain is very rare but the identification of causative genes has yielded significant insights into pain pathways and also novel targets for pain treatment. We report a novel recessive disorder characterized by congenital insensitivity to pain, inability to feel touch, and cognitive delay. Affected individuals harboured a homozygous missense mutation in CLTCL1 encoding the CHC22 clathrin heavy chain, p.E330K, which we demonstrate to have a functional effect on the protein. We found that CLTCL1 is significantly upregulated in the developing human brain, displaying an expression pattern suggestive of an early neurodevelopmental role. Guided by the disease phenotype, we investigated the role of CHC22 in two human neural crest differentiation systems; human induced pluripotent stem cell-derived nociceptors and TRKB-dependant SH-SY5Y cells. In both there was a significant downregulation of CHC22 upon the onset of neural differentiation. Furthermore, knockdown of CHC22 induced neurite outgrowth in neural precursor cells, which was rescued by stable overexpression of small interfering RNA-resistant CHC22, but not by mutant CHC22. Similarly, overexpression of wild-type, but not mutant, CHC22 blocked neurite outgrowth in cells treated with retinoic acid. These results reveal an essential and non-redundant role for CHC22 in neural crest development and in the genesis of pain and touch sensing neurons.

Ion Channel Modulation in Spinal/Trigeminal Synaptic Plasticity

Ion Channel Modulation in Spinal/Trigeminal Synaptic Plasticity

MrgC agonism at central terminals of primary sensory neurons inhibits neuropathic pain

Chronic neuropathic pain is often refractory to current pharmacotherapies. The rodent Mas-related G-protein-coupled receptor subtype C (MrgC) shares substantial homogeneity with its human homologue, MrgX1, and is located specifically in small-diameter dorsal root ganglion neurons. However, evidence regarding the role of MrgC in chronic pain conditions has been disparate and inconsistent. Accordingly, the therapeutic value of MrgX1 as a target for pain treatment in humans remains uncertain. Here, we found that intrathecal injection of BAM8-22 (a 15-amino acid peptide MrgC agonist) and JHU58 (a novel dipeptide MrgC agonist) inhibited both mechanical and heat hypersensitivity in rats after an L5 spinal nerve ligation (SNL). Intrathecal JHU58-induced pain inhibition was dose dependent in SNL rats. Importantly, drug efficacy was lost in Mrg-cluster gene knockout (Mrg KO) mice and was blocked by gene silencing with intrathecal MrgC siRNA and by a selective MrgC receptor antagonist in SNL rats, suggesting that the drug action is MrgC dependent. Further, in a mouse model of trigeminal neuropathic pain, microinjection of JHU58 into ipsilateral subnucleus caudalis inhibited mechanical hypersensitivity in wild-type but not Mrg KO mice. Finally, JHU58 attenuated the miniature excitatory postsynaptic currents frequency both in medullary dorsal horn neurons of mice after trigeminal nerve injury and in lumbar spinal dorsal horn neurons of mice after SNL. We provide multiple lines of evidence that MrgC agonism at spinal but not peripheral sites may constitute a novel pain inhibitory mechanism that involves inhibition of peripheral excitatory inputs onto postsynaptic dorsal horn neurons in different rodent models of neuropathic pain.

Mu Opioid Receptors on Primary Afferent Nav1.8 Neurons Contribute to Opiate-Induced Analgesia: Insight from Conditional Knockout Mice

Opiates are powerful drugs to treat severe pain, and act via mu opioid receptors distributed throughout the nervous system. Their clinical use is hampered by centrally-mediated adverse effects, including nausea or respiratory depression. Here we used a genetic approach to investigate the potential of peripheral mu opioid receptors as targets for pain treatment. We generated conditional knockout (cKO) mice in which mu opioid receptors are deleted specifically in primary afferent Nav1.8-positive neurons. Mutant animals were compared to controls for acute nociception, inflammatory pain, opiate-induced analgesia and constipation. There was a 76% decrease of mu receptor-positive neurons and a 60% reduction of mu-receptor mRNA in dorsal root ganglia of cKO mice. Mutant mice showed normal responses to heat, mechanical, visceral and chemical stimuli, as well as unchanged morphine antinociception and tolerance to antinociception in models of acute pain. Inflammatory pain developed similarly in cKO and controls mice after Complete Freund’s Adjuvant. In the inflammation model, however, opiate-induced (morphine, fentanyl and loperamide) analgesia was reduced in mutant mice as compared to controls, and abolished at low doses. Morphine-induced constipation remained intact in cKO mice. We therefore genetically demonstrate for the first time that mu opioid receptors partly mediate opiate analgesia at the level of Nav1.8-positive sensory neurons. In our study, this mechanism operates under conditions of inflammatory pain, but not nociception. Previous pharmacology suggests that peripheral opiates may be clinically useful, and our data further demonstrate that Nav1.8 neuron-associated mu opioid receptors are feasible targets to alleviate some forms of persistent pain.