Pannexin-1 channel inhibition alleviates opioid withdrawal in rodents by modulating locus coeruleus to spinal cord circuitry.

Opioid withdrawal is a liability of chronic opioid use and misuse, impacting people who use prescription or illicit opioids. Hyperactive autonomic output underlies many of the aversive withdrawal symptoms that make it difficult to discontinue chronic opioid use. The locus coeruleus (LC) is an important autonomic centre within the brain with a poorly defined role in opioid withdrawal. We show here that pannexin-1 (Panx1) channels expressed on microglia critically modulate LC activity during opioid withdrawal. Within the LC, we found that spinally projecting tyrosine hydroxylase (TH)-positive neurons (LCspinal) are hyperexcitable during morphine withdrawal, elevating cerebrospinal fluid (CSF) levels of norepinephrine. Pharmacological and chemogenetic silencing of LCspinal neurons or genetic ablation of Panx1 in microglia blunted CSF NE release, reduced LC neuron hyperexcitability, and concomitantly decreased opioid withdrawal behaviours in mice. Using probenecid as an initial lead compound, we designed a compound (EG-2184) with greater potency in blocking Panx1. Treatment with EG-2184 significantly reduced both the physical signs and conditioned place aversion caused by opioid withdrawal in mice, as well as suppressed cue-induced reinstatement of opioid seeking in rats. Together, these findings demonstrate that microglial Panx1 channels modulate LC noradrenergic circuitry during opioid withdrawal and reinstatement. Blocking Panx1 to dampen LC hyperexcitability may therefore provide a therapeutic strategy for alleviating the physical and aversive components of opioid withdrawal.

Expression of GAD2 in excitatory neurons projecting from the ventrolateral periaqueductal gray to the locus coeruleus.

The ventrolateral periaqueductal gray (vlPAG) functionally projects to diverse brain regions, including the locus coeruleus (LC). Excitatory projections from the vlPAG to the LC are well described, while few studies have indicated the possibility of inhibitory projections. Here, we quantified the relative proportion of excitatory and inhibitory vlPAG-LC projections in male and female mice, and found an unexpected overlapping population of neurons expressing both GAD2 and VGLUT2. Combined in vitro optogenetic stimulation and electrophysiology of LC neurons revealed that vlPAG neurons expressing channelrhodopsin-2 under the GAD2 promoter release both GABA and glutamate. Subsequent experiments identified a population of GAD2+/VGLUT2+ vlPAG neurons exclusively releasing glutamate onto LC neurons. Altogether, we demonstrate that ∼25% of vlPAG-LC projections are inhibitory, and that there is a significant GAD2 expressing population of glutamatergic projections. Our findings have broad implications for the utility of GAD2-Cre lines within midbrain and brainstem regions, and especially within the PAG.

Targeting Pannexin-1 Channels: Addressing the 'Gap' in Chronic Pain.

Chronic pain complicates many diseases and is notoriously difficult to treat. In search of new therapeutic targets, pannexin-1 (Panx1) channels have sparked intense interest as a key mechanism involved in a variety of chronic pain conditions. Panx1 channels are transmembrane proteins that release ions and small molecules, such as adenosine triphosphate (ATP). They are expressed along important nodes of the pain pathway, modulating activity of diverse cell types implicated in the development and progression of chronic pain caused by injury or pathology. This review highlights advances that have unlocked the core structure and machinery controlling Panx1 function with a focus on understanding and treating chronic pain.

Sex Differences in Plasma, Adipose Tissue, and Central Accumulation of Cannabinoids, and Behavioral Effects of Oral Cannabis Consumption in Male and Female C57BL/6 Mice.

BACKGROUND: Cannabis edibles are an increasingly popular form of cannabis consumption. Oral consumption of cannabis has distinct physiological and behavioral effects compared with injection or inhalation. An animal model is needed to understand the pharmacokinetics and physiological effects of oral cannabis consumption in rodents as a model for human cannabis edible use. METHODS: Adult male and female C57BL/6 mice received a single dose of commercially available cannabis oil (5 mg/kg Δ9-tetrahydrocannabinol [THC]) by oral gavage. At 0.5, 1, 2, 3, and 6 hours post exposure, plasma, hippocampus, and adipose tissue were collected for THC, 11-OH-THC, and THC-COOH measures. RESULTS: We report delayed time to peak THC and 11-OH-THC concentrations in plasma, brain, and adipose tissue, which is consistent with human pharmacokinetics studies. We also found sex differences in the cannabis tetrad: (1) female mice had a delayed hypothermic effect 6 hours post consumption, which was not present in males; (2) females had stronger catalepsy than males; (3) males were less mobile following cannabis exposure, whereas female mice showed no difference in locomotion but an anxiogenic effect at 3 hours post exposure; and (4) male mice displayed a longer-lasting antinociceptive effect of oral cannabis. CONCLUSIONS: Oral cannabis consumption is a translationally relevant form of administration that produces similar physiological effects as injection or vaping administration and thus should be considered as a viable approach for examining the physiological effects of cannabis moving forward. Furthermore, given the strong sex differences in metabolism of oral cannabis, these factors should be carefully considered when designing animal studies on the effects of cannabis.

Differential regulation of Cav 3.2 and Cav 2.2 calcium channels by CB1 receptors and cannabidiol.

BACKGROUND AND PURPOSE: Cannabinoids are a promising therapeutic avenue for chronic pain. However, clinical trials often fail to report analgesic efficacy of cannabinoids. Inhibition of voltage gate calcium (Cav ) channels is one mechanism through which cannabinoids may produce analgesia. We hypothesized that cannabinoids and cannabinoid receptor agonists target different types of Cav channels through distinct mechanisms. EXPERIMENTAL APPROACH: Electrophysiological recordings from tsA-201 cells expressing either Cav 3.2 or Cav 2.2 were used to assess inhibition by HU-210 or cannabidiol (CBD) in the absence and presence of the CB1 receptor. Homology modelling assessed potential interaction sites for CBD in both Cav 2.2 and Cav 3.2. Analgesic effects of CBD were assessed in mouse models of inflammatory and neuropathic pain. KEY RESULTS: HU-210 (1 µM) inhibited Cav 2.2 function in the presence of CB1 receptor but had no effect on Cav 3.2 regardless of co-expression of CB1 receptor. By contrast, CBD (3 µM) produced no inhibition of Cav 2.2 and instead inhibited Cav 3.2 independently of CB1 receptors. Homology modelling supported these findings, indicating that CBD binds to and occludes the pore of Cav 3.2, but not Cav 2.2. Intrathecal CBD alleviated thermal and mechanical hypersensitivity in both male and female mice, and this effect was absent in Cav 3.2 null mice. CONCLUSION AND IMPLICATIONS: Our findings reveal differential modulation of Cav 2.2 and Cav 3.2 channels by CB1 receptors and CBD. This advances our understanding of how different cannabinoids produce analgesia through action at different voltage-gated calcium channels and could influence the development of novel cannabinoid-based therapeutics for treatment of chronic pain.

Analgesic, Anesthetic, and Addiction Clinical Trial Translations, Innovations, Opportunities, and Networks-American Pain Society-American Academy of Pain Medicine Pain Taxonomy Diagnostic Criteria for Acute Needle Pain.

Needle procedures are among the most common causes of pain and distress for individuals seeking health care. While needle pain is especially problematic for children needle pain and associated fear also has significant impact on adults and can lead to avoidance of appropriate medical care. Currently there is not a standard definition of needle pain. A taxonomy, or classification system, for acute needle pain would aid research efforts and enhance clinical care. To meet this need, the Analgesic, Anesthetic, and Addiction Clinical Trial Translations, Innovations, Opportunities, and Networks public-private partnership with the U.S. Food and Drug Administration, the American Pain Society, and the American Academy of Pain Medicine formed the Analgesic, Anesthetic, and Addiction Clinical Trial Translations, Innovations, Opportunities, and Networks-American Pain Society-American Academy of Pain Medicine Pain Taxonomy initiative. One of the goals of this initiative was to develop taxonomies for acute pain disorders, including needle pain. To accomplish this, a working group of experts in needle pain was convened. Based on available literature and expert opinion, the working group used a 5-dimenional structure (diagnostic criteria, common features, modulating factors, impact and/or functional consequences, and putative mechanisms) to develop an acute pain taxonomy that is specific needle pain. As part of this, a set of 4 diagnostic criteria, with 2 modifiers to account for the influence of needle associated fear, are proposed to define the types of acute needle pain. PERSPECTIVE: This article presents a taxonomy for acute needle pain. This taxonomy could help to standardize definitions of acute pain in clinical studies of patients undergoing needle procedures.

Spinal A3 adenosine receptor activation acutely restores morphine antinociception in opioid tolerant male rats.

Opioids are potent analgesics, but their pain-relieving effects diminish with repeated use. The reduction in analgesic potency is a hallmark of opioid analgesic tolerance, which hampers opioid pain therapy. In the central nervous system, opioid analgesia is critically modulated by adenosine, a purine nucleoside implicated in the beneficial and detrimental actions of opioid medications. Here, we focus on the A3 adenosine receptor (A3 AR) in opioid analgesic tolerance. Intrathecal administration of the A3 AR agonist MRS5698 with daily systemic morphine in male rats attenuated the reduction in morphine antinociception over 7 days. In rats with established morphine tolerance, intrathecal MRS5698 partially restored the antinociceptive effects of morphine. However, when MRS5698 was discontinued, these animals displayed a reduced antinociceptive response to morphine. Our results suggest that MRS5698 acutely and transiently potentiates morphine antinociception in tolerant rats. By contrast, in morphine-naïve rats MRS5698 treatment did not impact thermal nociceptive threshold or affect antinociceptive response to a single injection of morphine. Furthermore, we found that morphine-induced adenosine release in cerebrospinal fluid was blunted in tolerant animals, but total spinal A3 AR expression was not affected. Collectively, our findings indicate that spinal A3 AR activation acutely potentiates morphine antinociception in the opioid tolerant state.

Maternal and iatrogenic neonatal opioid withdrawal syndrome: Differences and similarities in recognition, management, and consequences.

Opioids are potent analgesics used to manage pain in both young and old, but the increased use in the pregnant population has significant individual and societal implications. Infants dependent on opioids, either through maternal or iatrogenic exposure, undergo neonatal opioid withdrawal syndrome (NOWS), where they may experience withdrawal symptoms ranging from mild to severe. We present a detailed and original review of NOWS caused by maternal opioid exposure (mNOWS) and iatrogenic opioid intake (iNOWS). While these two entities have been assessed entirely separately, recognition and treatment of the clinical manifestations of NOWS overlap. Neonatal risk factors such as age, genetic predisposition, drug type, and clinical factors like type of opioid, cumulative dose of opioid exposure, and disease status affect the incidence of both mNOWS and iNOWS, as well as their severity. Recognition of withdrawal is dependent on clinical assessment of symptoms, and the use of clinical assessment tools designed to determine the need for pharmacotherapy. Treatment of NOWS relies on a combination of non-pharmacological therapies and pharmacological options. Long-term consequences of opioids and NOWS continue to generate controversy, with some evidence of anatomic brain changes, but conflicting animal and human clinical evidence of significant cognitive or behavioral impacts on school-age children. We highlight the current knowledge on clinically relevant recognition, treatment, and consequences of NOWS, and identify new advances in clinical management of the neonate. This review brings a unique clinical perspective and critically analyzes gaps between the clinical problem and our preclinical understanding of NOWS.

Role of Primary Afferents in Arthritis Induced Spinal Microglial Reactivity.

Increased afferent input resulting from painful injury augments the activity of central nociceptive circuits via both neuron-neuron and neuron-glia interactions. Microglia, resident immune cells of the central nervous system (CNS), play a crucial role in the pathogenesis of chronic pain. This study provides a framework for understanding how peripheral joint injury signals the CNS to engage spinal microglial responses. During the first week of monosodium iodoacetate (MIA)-induced knee joint injury in male rats, inflammatory and neuropathic pain were characterized by increased firing of peripheral joint afferents. This increased peripheral afferent activity was accompanied by increased Iba1 immunoreactivity within the spinal dorsal horn indicating microglial activation. Pharmacological silencing of C and A afferents with co-injections of QX-314 and bupivacaine, capsaicin, or flagellin prevented the development of mechanical allodynia and spinal microglial activity after MIA injection. Elevated levels of ATP in the cerebrospinal fluid (CSF) and increased expression of the ATP transporter vesicular nucleotide transporter (VNUT) in the ipsilateral spinal dorsal horn were also observed after MIA injections. Selective silencing of primary joint afferents subsequently inhibited ATP release into the CSF. Furthermore, increased spinal microglial reactivity, and alleviation of MIA-induced arthralgia with co-administration of QX-314 with bupivacaine were recapitulated in female rats. Our results demonstrate that early peripheral joint injury activates joint nociceptors, which triggers a central spinal microglial response. Elevation of ATP in the CSF, and spinal expression of VNUT suggest ATP signaling may modulate communication between sensory neurons and spinal microglia at 2 weeks of joint degeneration.

Identifying the Neurodevelopmental Differences of Opioid Withdrawal.

Stopping opioid medications can result in a debilitating withdrawal syndrome in chronic users. Opioid withdrawal can occur at all ages, but mechanistic understanding of this condition is predominantly derived from adult studies. Here, we examined whether there are age-dependent differences in the behavioural phenotype and cellular indices of opioid withdrawal. We tested this by assessing the behavioural and cFos response (a surrogate marker for neuronal activation) to morphine withdrawal in C57BL/6J mice across key developmental stages-neonatal, adolescent, and adulthood. Mice in all age groups received escalating doses of morphine (10-50 mg/kg) over 5 days and withdrawal was precipitated by a single injection of the opioid receptor antagonist naloxone (2 mg/kg) two hours after the last morphine dose. In adult and adolescent mice, withdrawal behaviours were robust, with age-related differences in autonomic and somatic signs. In both groups, cFos expression was increased in spinally projecting neurons within the Periaqueductal Grey (PAG), Rostro-ventromedial Medulla (RVM), and Locus Coeruleus. Neonatal animals displayed both a distinct behavioural withdrawal and cFos expression profile. Notably, in young animals cFos expression was increased within the PAG and LC, but decreased in the RVM. In summary, naloxone challenge precipitated robust opioid withdrawal behaviours across all developmental stages with neonatal animals displaying differences in withdrawal behaviours and unique neuronal activation patterns within key brainstem regions.

Cannabinoids in Chronic Pain: Therapeutic Potential Through Microglia Modulation.

Chronic pain is a complex sensory, cognitive, and emotional experience that imposes a great personal, psychological, and socioeconomic burden on patients. An estimated 1.5 billion people worldwide are afflicted with chronic pain, which is often difficult to treat and may be resistant to the potent pain-relieving effects of opioid analgesics. Attention has therefore focused on advancing new pain therapies directed at the cannabinoid system because of its key role in pain modulation. Endocannabinoids and exogenous cannabinoids exert their actions primarily through Gi/o-protein coupled cannabinoid CB1 and CB2 receptors expressed throughout the nervous system. CB1 receptors are found at key nodes along the pain pathway and their activity gates both the sensory and affective components of pain. CB2 receptors are typically expressed at low levels on microglia, astrocytes, and peripheral immune cells. In chronic pain states, there is a marked increase in CB2 expression which modulates the activity of these central and peripheral immune cells with important consequences for the surrounding pain circuitry. Growing evidence indicate that interventions targeting CB1 or CB2 receptors improve pain outcomes in a variety of preclinical pain models. In this mini-review, we will highlight recent advances in understanding how cannabinoids modulate microglia function and its implications for cannabinoid-mediated analgesia, focusing on microglia-neuron interactions within the spinal nociceptive circuitry.

A dynamic role for dopamine receptors in the control of mammalian spinal networks.

Dopamine is well known to regulate movement through the differential control of direct and indirect pathways in the striatum that express D1 and D2 receptors respectively. The spinal cord also expresses all dopamine receptors; however, how the specific receptors regulate spinal network output in mammals is poorly understood. We explore the receptor-specific mechanisms that underlie dopaminergic control of spinal network output of neonatal mice during changes in spinal network excitability. During spontaneous activity, which is a characteristic of developing spinal networks operating in a low excitability state, we found that dopamine is primarily inhibitory. We uncover an excitatory D1-mediated effect of dopamine on motoneurons and network output that also involves co-activation with D2 receptors. Critically, these excitatory actions require higher concentrations of dopamine; however, analysis of dopamine concentrations of neonates indicates that endogenous levels of spinal dopamine are low. Because endogenous levels of spinal dopamine are low, this excitatory dopaminergic pathway is likely physiologically-silent at this stage in development. In contrast, the inhibitory effect of dopamine, at low physiological concentrations is mediated by parallel activation of D2, D3, D4 and α2 receptors which is reproduced when endogenous dopamine levels are increased by blocking dopamine reuptake and metabolism. We provide evidence in support of dedicated spinal network components that are controlled by excitatory D1 and inhibitory D2 receptors that is reminiscent of the classic dopaminergic indirect and direct pathway within the striatum. These results indicate that network state is an important factor that dictates receptor-specific and therefore dose-dependent control of neuromodulators on spinal network output and advances our understanding of how neuromodulators regulate neural networks under dynamically changing excitability.

Spinal interleukin-6 contributes to central sensitisation and persistent pain hypersensitivity in a model of juvenile idiopathic arthritis.

Pain is the most debilitating symptom in juvenile idiopathic arthritis. As pain correlates poorly to the extent of joint pathology, therapies that control joint inflammation are often inadequate as analgesics. We test the hypothesis that juvenile joint inflammation leads to sensitisation of nociceptive circuits in the central nervous system, which is maintained by cytokine expression in the spinal cord. Here, transient joint inflammation was induced in postnatal day (P)21 and P40 male Sprague-Dawley rats with a single intra-articular ankle injection of complete Freund's adjuvant. Hindpaw mechanical pain sensitivity was assessed using von Frey hair and weight bearing tests. Spinal neuron activity was measured using in vivo extracellular recording and immunohistochemistry. Joint and spinal dorsal horn TNFα, IL1ß and IL6 protein expression was quantified using western blotting. We observed greater mechanical hyperalgesia following joint inflammation in P21 compared to P40 rats, despite comparable duration of swelling and joint inflammatory cytokine levels. This is mirrored by spinal neuron hypersensitivity, which also outlasted the duration of active joint inflammation. The cytokine profile in the spinal cord differed at the two ages: prolonged upregulation of spinal IL6 was observed in P21, but not P40 rats. Finally, spinal application of anti-IL-6 antibody (30 ng) reduced the mechanical hyperalgesia and neuronal activation. Our results indicate that persistent upregulation of pro-inflammatory cytokines in the spinal dorsal horn is associated with neuronal sensitisation and mechanical hyperalgesia in juvenile rats, beyond the progress of joint pathology. In addition, we provide proof of concept that spinal IL6 is a key target for treating persistent pain in JIA.

Pannexin 1 Channels as a Therapeutic Target: Structure, Inhibition, and Outlook.

Pannexin 1 (Panx1) channels are transmembrane proteins that release adenosine triphosphate and play an important role in intercellular communication. They are widely expressed in somatic and nervous system tissues, and their activity has been associated with many pathologies such as stroke, epilepsy, inflammation, and chronic pain. While there are a variety of small molecules known to inhibit Panx1, currently little is known about the mechanism of channel inhibition, and there is a dearth of sufficiently potent and selective drugs targeting Panx1. Herein we provide a review of the current literature on Panx1 structural biology and known pharmacological agents that will help provide a basis for rational development of Panx1 chemical modulators.

Chronic Morphine-Induced Changes in Signaling at the A3 Adenosine Receptor Contribute to Morphine-Induced Hyperalgesia, Tolerance, and Withdrawal.

Treating chronic pain by using opioids, such as morphine, is hampered by the development of opioid-induced hyperalgesia (OIH; increased pain sensitivity), antinociceptive tolerance, and withdrawal, which can contribute to dependence and abuse. In the central nervous system, the purine nucleoside adenosine has been implicated in beneficial and detrimental actions of morphine, but the extent of their interaction remains poorly understood. Here, we demonstrate that morphine-induced OIH and antinociceptive tolerance in rats is associated with a twofold increase in adenosine kinase (ADK) expression in the dorsal horn of the spinal cord. Blocking ADK activity in the spinal cord provided greater than 90% attenuation of OIH and antinociceptive tolerance through A3 adenosine receptor (A3AR) signaling. Supplementing adenosine signaling with selective A3AR agonists blocked OIH and antinociceptive tolerance in rodents of both sexes. Engagement of A3AR in the spinal cord with an ADK inhibitor or A3AR agonist was associated with reduced dorsal horn of the spinal cord expression of the NOD-like receptor pyrin domain-containing 3 (60%-75%), cleaved caspase 1 (40%-60%), interleukin (IL)-1ß (76%-80%), and tumor necrosis factor (50%-60%). In contrast, the neuroinhibitory and anti-inflammatory cytokine IL-10 increased twofold. In mice, A3AR agonists prevented the development of tolerance in a model of neuropathic pain and reduced naloxone-dependent withdrawal behaviors by greater than 50%. These findings suggest A3AR-dependent adenosine signaling is compromised during sustained morphine to allow the development of morphine-induced adverse effects. These findings raise the intriguing possibility that A3AR agonists may be useful adjunct to opioids to manage their unwanted effects. SIGNIFICANCE STATEMENT: The development of hyperalgesia and antinociceptive tolerance during prolonged opioid use are noteworthy opioid-induced adverse effects that reduce opioid efficacy for treating chronic pain and increase the risk of dependence and abuse. We report that in rodents, these adverse effects are due to reduced adenosine signaling at the A3AR, resulting in NOD-like receptor pyrin domain-containing 3-interleukin-1ß neuroinflammation in spinal cord. These effects are attenuated by A3AR agonists, suggesting that A3AR may be a target for therapeutic intervention with selective A3AR agonist as opioid adjuncts.

Chronic morphine regulates TRPM8 channels via MOR-PKCß signaling.

Postoperative shivering and cold hypersensitivity are major side effects of acute and chronic opioid treatments respectively. TRPM8 is a cold and menthol-sensitive channel found in a subset of dorsal root ganglion (DRG) nociceptors. Deletion or inhibition of the TRPM8 channel was found to prevent the cold hyperalgesia induced by chronic administration of morphine. Here, we examined the mechanisms by which morphine was able to promote cold hypersensitivity in DRG neurons and transfected HEK cells. Mice daily injected with morphine for 5 days developed cold hyperalgesia. Treatment with morphine did not alter the expressions of cold sensitive TREK-1, TRAAK and TRPM8 in DRGs. However, TRPM8-expressing DRG neurons isolated from morphine-treated mice exhibited hyperexcitability. Sustained morphine treatment in vitro sensitized TRPM8 responsiveness to cold or menthol and reduced activation-evoked desensitization of the channel. Blocking phospholipase C (PLC) as well as protein kinase C beta (PKCß), but not protein kinase A (PKA) or Rho-associated protein kinase (ROCK), restored channel desensitization. Identification of two PKC phosphorylation consensus sites, S1040 and S1041, in the TRPM8 and their site-directed mutation were able to prevent the MOR-induced reduction in TRPM8 desensitization. Our results show that activation of MOR by morphine 1) promotes hyperexcitability of TRPM8-expressing neurons and 2) induces a PKCß-mediated reduction of TRPM8 desensitization. This MOR-PKCß dependent modulation of TRPM8 may underlie the onset of cold hyperalgesia caused by repeated administration of morphine. Our findings point to TRPM8 channel and PKCß as important targets for opioid-induced cold hypersensitivity.

TRPV1 promotes opioid analgesia during inflammation.

Pain and inflammation are inherently linked responses to injury, infection, or chronic diseases. Given that acute inflammation in humans or mice enhances the analgesic properties of opioids, there is much interest in determining the inflammatory transducers that prime opioid receptor signaling in primary afferent nociceptors. Here, we found that activation of the transient receptor potential vanilloid type 1 (TRPV1) channel stimulated a mitogen-activated protein kinase (MAPK) signaling pathway that was accompanied by the shuttling of the scaffold protein ß-arrestin2 to the nucleus. The nuclear translocation of ß-arrestin2 in turn prevented its recruitment to the µ-opioid receptor (MOR), the subsequent internalization of agonist-bound MOR, and the suppression of MOR activity that occurs upon receptor desensitization. Using the complete Freund's adjuvant (CFA) inflammatory pain model to examine the role of TRPV1 in regulating endogenous opioid analgesia in mice, we found that naloxone methiodide (Nal-M), a peripherally restricted, nonselective, and competitive opioid receptor antagonist, slowed the recovery from CFA-induced hypersensitivity in wild-type, but not TRPV1-deficient, mice. Furthermore, we showed that inflammation prolonged morphine-induced antinociception in a mouse model of opioid receptor desensitization, a process that depended on TRPV1. Together, our data reveal a TRPV1-mediated signaling pathway that serves as an endogenous pain-resolution mechanism by promoting the nuclear translocation of ß-arrestin2 to minimize MOR desensitization. This previously uncharacterized mechanism may underlie the peripheral opioid control of inflammatory pain. Dysregulation of the TRPV1-ß-arrestin2 axis may thus contribute to the transition from acute to chronic pain.

Kappa Opioid Receptors Drive a Tonic Aversive Component of Chronic Pain.

Pain is a multidimensional experience and negative affect, or how much the pain is "bothersome", significantly impacts the sufferers' quality of life. It is well established that the κ opioid system contributes to depressive and dysphoric states, but whether this system contributes to the negative affect precipitated by the occurrence of chronic pain remains tenuous. Using a model of persistent pain, we show by quantitative real-time-PCR, florescence in situ hybridization, Western blotting and GTPgS autoradiography an upregulation of expression and the function of κ opioid receptors (KORs) and its endogenous ligand dynorphin in the mesolimbic circuitry in animals with chronic pain compared with surgical controls. Using in vivo microdialysis and microinjection of drugs into the mesolimbic dopamine system, we demonstrate that inhibiting KORs reinstates evoked dopamine release and reward-related behaviors in chronic pain animals. Chronic pain enhanced KOR agonist-induced place aversion in a sex-dependent manner. Using various place preference paradigms, we show that activation of KORs drives pain aversive states in male but not female mice. However, KOR antagonist treatment was effective in alleviating anxiogenic and depressive affective-like behaviors in both sexes. Finally, ablation of KORs from dopamine neurons using AAV-TH-cre in KORloxP mice prevented pain-induced aversive states as measured by place aversion assays. Our results strongly support the use of KOR antagonists as therapeutic adjuvants to alleviate the emotional, tonic-aversive component of chronic pain, which is argued to be the most significant component of the pain experience that impacts patients' quality of life.SIGNIFICANCE STATEMENT We show that KORs are sufficient to drive the tonic-aversive component of chronic pain; the emotional component of pain that is argued to significantly impact a patient's quality of life. The impact of our study is broadly relevant to affective disorders associated with disruption of reward circuitry and thus likely contributes to many of the devastating sequelae of chronic pain, including the poor response to treatment of many patients, debilitating affective disorders (other disorders including anxiety and depression that demonstrate high comorbidity with chronic pain) and substance abuse. Indeed, coexisting psychopathology increases pain intensity, pain-related disability and effectiveness of treatments (Jamison and Edwards, 2013).