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HSK21542 injection is a new peripheral kappa opioid receptor (KOR) agonist. To evaluate its safety, tolerability, pharmacokinetics and pharmacodynamics, this study was conducted in healthy volunteers, consisting of two parts: a single ascending dose (0.2–3.375 μg/kg, 15‐min infusion) and different infusion durations (0.2 and 1 μg/kg, 2‐ or 15‐min infusion). The area under the plasma concentration‐time curve (AUC) and peak concentration (Cmax) of HSK21542 were dose‐linear among 0.2–3.375 μg/kg. After intravenous injection, HSK21542 was rapidly eliminated with a half‐life (t1/2) of 1.5 h, and the majority (48.02%) of the dose was excreted unchanged in urine. Pharmacodynamic results showed that HSK21542 increased prolactin release and reached a peak at 1–2 h after administration but had no significant effect on vasopressin levels. There was a brief increase in urine volume within the initial 2 h after administration. HSK21542 was well tolerated; most of the adverse effects (AEs) in the trial group were grade 1, and only 2 cases (4.0%) were grade 2. The main AE was paresthesia, which appeared in 42% (21/50) in the trial group. No serious adverse event (SAE) was observed. No subject withdrew early due to AEs. These results suggest that HSK21542 may be a potential treatment for pain and pruritic conditions. [ABSTRACT FROM AUTHOR]

Many drugs of abuse, including alcohol, disrupt long-term synaptic depression (LTD) at dorsal striatal glutamate synapses. This disruption is common to many forms of LTD that are mediated by G protein coupled receptors (GPCRs) that signal through the inhibitory G i/o class of G proteins. A loss of LTD is thought to mediate behavioral changes associated with the development of substance use disorders. We have previously shown in multiple studies that LTD mediated by the G i/o -coupled mu opioid receptor is disrupted by in vivo opioid and alcohol exposure in adolescent and adult mice. One of our previous studies suggested that LTD mediated by delta and kappa opioid receptors was resistant to the LTD-disrupting properties of in vivo opioid exposure. We hypothesized that delta and kappa opioid receptor-mediated LTD would be exceptions to the generalizable observation that forms of dorsal striatal G i/o -coupled receptor LTD are disrupted by drugs of abuse. Specifically, we predicted that these forms of LTD would be resistant to the deleterious effects of alcohol consumption, just as they were resistant to opioid exposure. Indeed, in adult male mice that drank alcohol for 3 weeks, delta and kappa opioid receptor-mediated LTD at glutamatergic inputs to direct pathway and indirect pathway medium spiny neurons in the dorsolateral striatum was unaffected by alcohol. These data demonstrate that alcohol effects on GPCR-mediated LTD are not generalizable across all types of G i/o -coupled GPCRs. • Kappa and delta opioid receptor long-term depression occur at glutamate inputs to both subtypes of medium spiny neurons in dorsolateral striatum. • Neither kappa or delta opioid receptor long-term synaptic depression is sensitive to previous alcohol drinking. • Alcohol effects on opioid long-term depression are not generalizable across all subtypes of opioid receptors. [ABSTRACT FROM AUTHOR]

Abstract Objective Kappa opioid receptor (KOR) signaling is involved in joint development and inflammation in Osteoarthritis (OA), while the biochemical mechanism remains unclarified. This study aims to investigate downstream molecular events of KOR activation, to provide novel perspectives in OA pathology. Methods U50,488H, a selective KOR agonist, was intra-articularly injected in mice upon destabilization of the medial meniscus (DMM) as OA models, with PBS injection as control. The behavioral and histological evaluation was assessed by hot plate test and red solid green staining, respectively. Alterations in mRNA and protein expression were assessed by RNA-seq, RT-qPCR, immunohistochemistry and western blotting (WB) in chondrocytes treated with TNF-α or TNF-α + U50,488H. Proteins interacted with KOR were explored using proximity labeling followed by mass spectrometry and then testified by co-immunoprecipitation (Co-IP) assay and immunofluorescence (IF). Results OA-induced pain was reduced and cartilage degeneration was alleviated upon KOR activation in DMM mice. In chondrocytes, activation of KOR reversed the upregulation of MMPs, IL-6, IL-1β and phosphorylated(p-) STAT3, stimulated by TNF-α, while the expression of NF-κB, MAPKs and AKT signaling weren't reversed. RNA-seq and IF results presented that KOR activation evidently reduced STAT3 nuclear translocation in chondrocytes upon TNF-α stimuli. The reduction may be resulted from the binding of KOR and STAT3 in the plasma membrane, revealed by proximity labeling and Co-IP results. Conclusions KOR activation protects cartilage from OA, and this protective effect is mainly exerted via sequestering STAT3 on the plasma membrane, resulting in inactivation of STAT3-dependent immune responses which otherwise contributes to OA.

We recently developed a series of nalfurafine analogs (TK10, TK33, and TK35) that may serve as non-addictive candidate analgesics. These compounds are mixed-action agonists at the kappa and delta opioid receptors (KOR and DOR, respectively) and produce antinociception in a mouse warm-water tail-immersion test while failing to produce typical mu opioid receptor (MOR)-mediated side effects. The warm-water tail-immersion test is an assay of pain-stimulated behavior vulnerable to false-positive analgesic-like effects by drugs that produce motor impairment. Accordingly, this study evaluated TK10, TK33, and TK35 in a recently validated assay of pain-related behavioral depression in mice that are less vulnerable to false-positive effects. For comparison, we also evaluated the effects of the MOR agonist/analgesic hydrocodone (positive control), the neurokinin 1 receptor (NK1R) antagonist aprepitant (negative control), nalfurafine as a selective KOR agonist, SNC80 as a selective DOR agonist, and a nalfurafine/SNC80 mixture. Intraperitoneal injection of dilute lactic acid (IP lactic acid) served as a noxious stimulus to depress vertical and horizontal locomotor activity in male and female ICR mice. IP lactic acid-induced locomotor depression was alleviated by hydrocodone but not by aprepitant, nalfurafine, SNC80, the nalfurafine/SNC80 mixture, or the KOR/DOR agonists. These results suggest that caution is warranted in advancing mixed-action KOR/DOR agonists as candidate analgesics. [ABSTRACT FROM AUTHOR]

Objective: Kappa opioid receptor (KOR) signaling is involved in joint development and inflammation in Osteoarthritis (OA), while the biochemical mechanism remains unclarified. This study aims to investigate downstream molecular events of KOR activation, to provide novel perspectives in OA pathology. Methods: U50,488H, a selective KOR agonist, was intra-articularly injected in mice upon destabilization of the medial meniscus (DMM) as OA models, with PBS injection as control. The behavioral and histological evaluation was assessed by hot plate test and red solid green staining, respectively. Alterations in mRNA and protein expression were assessed by RNA-seq, RT-qPCR, immunohistochemistry and western blotting (WB) in chondrocytes treated with TNF-α or TNF-α + U50,488H. Proteins interacted with KOR were explored using proximity labeling followed by mass spectrometry and then testified by co-immunoprecipitation (Co-IP) assay and immunofluorescence (IF). Results: OA-induced pain was reduced and cartilage degeneration was alleviated upon KOR activation in DMM mice. In chondrocytes, activation of KOR reversed the upregulation of MMPs, IL-6, IL-1β and phosphorylated(p-) STAT3, stimulated by TNF-α, while the expression of NF-κB, MAPKs and AKT signaling weren't reversed. RNA-seq and IF results presented that KOR activation evidently reduced STAT3 nuclear translocation in chondrocytes upon TNF-α stimuli. The reduction may be resulted from the binding of KOR and STAT3 in the plasma membrane, revealed by proximity labeling and Co-IP results. Conclusions: KOR activation protects cartilage from OA, and this protective effect is mainly exerted via sequestering STAT3 on the plasma membrane, resulting in inactivation of STAT3-dependent immune responses which otherwise contributes to OA. [ABSTRACT FROM AUTHOR]

MeShell Green,1 Charles A Veltri,1 Oliver Grundmann1,2 1College of Pharmacy, Department of Pharmaceutical Sciences, Midwestern University, Glendale, AZ, USA; 2College of Pharmacy, Department of Medicinal Chemistry, University of Florida, Gainesville, FL, USACorrespondence: Oliver Grundmann, Department of Medicinal Chemistry, College of Pharmacy, University of Florida, 1345 Center Drive, Room P3-20, Gainesville, FL, 32611, USA, Tel +1-352-246-4994, Fax +1-352-392-9455, Email grundman@ufl.eduAbstract: Nalmefene hydrochloride was first discovered as an opioid antagonist derivative of naltrexone in 1975. It is among the most potent opioid antagonists currently on the market and is differentiated from naloxone and naltrexone by its partial agonist activity at the kappa-opioid receptor which may benefit in the treatment of alcohol use disorder. Oral nalmefene has been approved in the European Union for treatment of alcohol use disorder since 2013. As of 2023, nalmefene is available in the United States as an intranasal spray for reversal of opioid overdose but is not approved for alcohol or opioid use disorder as a maintenance treatment. The substantially longer half-life of nalmefene and 5-fold higher binding affinity to opioid receptors makes it a superior agent over naloxone in the reversal of high potency synthetic opioids like fentanyl and the emerging nitazenes. Nalmefene presents with a comparable side effect profile to other opioid antagonists and should be considered for further development as a maintenance treatment for opioid and other substance use disorders.Keywords: opioid antagonist, withdrawal, substance use disorder, kappa opioid receptor

Selective adult social attachments, or 'pair bonds', represent central relationships for individuals in a number of social species, including humans. Loss of a pair mate has emotional consequences that may or may not diminish over time, and that often translate into impaired psychological and physical health. In this paper, we review the literature on the neuroendocrine mechanisms for the emotional consequences of partner loss, with a special focus on hypothesized interactions between oxytocin, corticotropin-releasing hormone and the κ opioid system. This article is part of the theme issue 'Interplays between oxytocin and other neuromodulators in shaping complex social behaviours'.

Multiple sclerosis (MS) afflicts millions of people worldwide. While multiple therapeutics have recently been developed and approved as treatment agents, they are not 100% effective. Recent developments investigating the endogenous opioid system involvement in MS has revealed that agonists of the kappa opioid receptor (KOR) have beneficial effects in both animal models of MS (and demyelinating disorders more generally) as well as in vitro models of remyelination. Several groups have contributed to this development. We summarize here the findings of these published studies, with comparisons of the effects and discussion of similarities and differences. The effects of KOR agonists involve both neuroimmunomodulation as well as remyelination, in different populations of cells. The compelling findings in MS model systems using KOR agonists strongly indicate that further investigations at both mechanistic and translational therapeutic levels are warranted.

Multiple sclerosis (MS) afflicts millions of people worldwide. While multiple therapeutics have recently been developed and approved as treatment agents, they are not 100% effective. Recent developments investigating the endogenous opioid system involvement in MS has revealed that agonists of the kappa opioid receptor (KOR) have beneficial effects in both animal models of MS (and demyelinating disorders more generally) as well as in vitro models of remyelination. Several groups have contributed to this development. We summarize here the findings of these published studies, with comparisons of the effects and discussion of similarities and differences. The effects of KOR agonists involve both neuroimmunomodulation as well as remyelination, in different populations of cells. The compelling findings in MS model systems using KOR agonists strongly indicate that further investigations at both mechanistic and translational therapeutic levels are warranted. [ABSTRACT FROM AUTHOR]

We recently developed a series of nalfurafine analogs (TK10, TK33, and TK35) that may serve as non-addictive candidate analgesics. These compounds are mixed-action agonists at the kappa and delta opioid receptors (KOR and DOR, respectively) and produce antinociception in a mouse warm-water tail-immersion test while failing to produce typical mu opioid receptor (MOR)-mediated side effects. The warm-water tail-immersion test is an assay of pain-stimulated behavior vulnerable to false-positive analgesic-like effects by drugs that produce motor impairment. Accordingly, this study evaluated TK10, TK33, and TK35 in a recently validated assay of pain-related behavioral depression in mice that are less vulnerable to false-positive effects. For comparison, we also evaluated the effects of the MOR agonist/analgesic hydrocodone (positive control), the neurokinin 1 receptor (NK1R) antagonist aprepitant (negative control), nalfurafine as a selective KOR agonist, SNC80 as a selective DOR agonist, and a nalfurafine/SNC80 mixture. Intraperitoneal injection of dilute lactic acid (IP lactic acid) served as a noxious stimulus to depress vertical and horizontal locomotor activity in male and female ICR mice. IP lactic acid-induced locomotor depression was alleviated by hydrocodone but not by aprepitant, nalfurafine, SNC80, the nalfurafine/SNC80 mixture, or the KOR/DOR agonists. These results suggest that caution is warranted in advancing mixed-action KOR/DOR agonists as candidate analgesics.

Prior studies examining the effects of cocaine on the dynorphin/kappa opioid receptor (Dyn/KOR) system primarily focus on non-contingent cocaine exposure, but the effects of self-administration, which more closely reflects human drug-taking behaviors, are not well studied. In this study we characterized the effects of escalated intravenous cocaine self-administration on the functional state of the Dyn/KOR system and its interaction with mesolimbic dopamine signaling. Rats self-administered cocaine in an extended access, limited intake cocaine procedure, in which animals obtained 40 infusions per day (1.5 mg/kg/inf) for 5 consecutive days to ensure comparable consumption levels. Following single day tests of cue reactivity and progressive ratio responding, quantitative real-time polymerase chain reaction was used to measure levels of Oprk and Pdyn transcripts in the ventral tegmental area and nucleus accumbens. Additionally, after self-administration, ex vivo fast-scan cyclic voltammetry in the NAc was used to examine the ability of the KOR agonist U50,488 to inhibit dopamine release. We found that KOR-induced inhibition of dopamine release was enhanced in animals that self-administered cocaine compared to controls, suggesting upregulated Dyn/KOR activity after cocaine self-administration. Furthermore, expression levels of Pdyn in the nucleus accumbens and ventral tegmental area, and Oprk in the nucleus accumbens, were elevated in cocaine animals compared to controls. Additionally, Pdyn expression in the nucleus accumbens was negatively correlated with progressive ratio breakpoints, a measure of motivation to self-administer cocaine. Overall, these data suggest that cocaine self-administration elevates KOR/Dyn system activity in the mesolimbic dopamine pathway.

Kappa opioid receptors (KOPr) are involved in the response to stress. KOPr are also targets for the treatment of stress-related psychiatric disorders including depression, anxiety, and addiction although effects of KOPr are often sex-dependent. Here we investigated c-Fos expression in a range of brain regions in male and female mice following an acute stressor, and a single injection of KOPr agonist. Using adult C57BL/6 c-Fos-GFP transgenic mice and quantitative fluorescence microscopy, we identified brain regions activated in response to a challenge with the KOPr agonist U50,488 (20 mg/kg) or an acute stress (15 min forced swim stress, FSS). In male mice, U50,488 increased expression of c-Fos in the prelimbic area of the prefrontal cortex (PFCx), nucleus accumbens (NAcc), and basolateral nuclei of the amygdala (BLA). In contrast, in female mice U50,488 only activated the BLA but not the PFCx or the NAcc. FSS increased activation of PFCx, NAcc, and BLA in males while there was no activation of the PFCx in female mice. In both sexes, the KOPr antagonist norBNI significantly blocked U50,488-induced, but not stress-induced activation of brain regions. In separate experiments, activated cells were confirmed as non-GABAergic neurons in the PFCx and NAcc. Together these data demonstrate sex differences in activation of brain regions that are key components of the 'reward' circuitry. These differential responses may contribute to sex differences in stress-related psychiatric disorders and in the treatment of depression, anxiety, and addiction. [ABSTRACT FROM AUTHOR]

The macrocyclic tetrapeptide CJ-15,208 (cyclo[Phe-D-Pro-Phe-Trp]) and its D-Trp isomer exhibit kappa opioid receptor (KOR) antagonism which prevents stress-induced reinstatement of extinguished cocaine-conditioned place preference. Here, we evaluated the effects of substitution of Trp and D-Trp on the peptides' opioid activity, antinociceptive tolerance, and the ability to prevent relapse to extinguished drug-CPP. Six analogs were synthesized using a combination of solid-phase peptide synthesis and cyclization in solution. The analogs were evaluated in vitro for opioid receptor affinity in radioligand competition binding assays, efficacy in the [35S]GTPγS assay, metabolic stability in mouse liver microsomes, and for opioid activity and selectivity in vivo in the mouse 55 °C warm-water tail-withdrawal assay. Potential liabilities of locomotor impairment, respiratory depression, acute tolerance, and conditioned place preference (CPP) were also assessed in vivo, and the ameliorating effect of analogs on the reinstatement of extinguished cocaine-place preference was assessed. Substitutions of other D-amino acids for D-Trp did not affect (or in one case increased) KOR affinity, while two of the three substitutions of an L-amino acid for Trp decreased KOR affinity. In contrast, all but one substitution increased mu opioid receptor (MOR) affinity in vitro. The metabolic stabilities of the analogs were similar to those of their respective parent peptides, with analogs containing a D-amino acid being much more rapidly metabolized than those containing an L-amino acid in this position. In vivo, CJ-15,208 analogs demonstrated antinociception, although potencies varied over an 80-fold range and the mediating opioid receptors differed by substitution. KOR antagonism was lost for all but the D-benzothienylalanine analog, and the 2′-naphthylalanine analog instead demonstrated significant delta opioid receptor (DOR) antagonism. Introduction of DOR antagonism coincided with reduced acute opioid antinociceptive tolerance and prevented stress-induced reinstatement of extinguished cocaine-CPP. [ABSTRACT FROM AUTHOR]

Pain is complex and is a unique experience for individuals in that no two people will have exactly the same physiological and emotional response to the same noxious stimulus or injury. Pain is composed of two essential processes: a sensory component that allows for discrimination of the intensity and location of a painful stimulus and an emotional component that underlies the affective, motivational, unpleasant, and aversive response to a painful stimulus. Kappa opioid receptor (KOR) activation in the periphery and throughout the neuroaxis modulates both of these components of the pain experience. In this chapter we focus on recent findings that KORs contribute to the emotional, aversive nature of chronic pain, including how expression in the limbic circuitry contributes to anhedonic states and components of opioid misuse disorder. While the primary focus is on preclinical pain models, we also highlight clinical or human research where there is strong evidence for KOR involvement in negative affective states associated with chronic pain and opioid misuse.

Kappa opioid receptor (KOR) agonists have preclinical antipsychostimulant effects; however, adverse side effects have limited their therapeutic development. In this preclinical study, conducted in Sprague Dawley rats, B6-SJL mice, and non-human primates (NHPs), we evaluated the G-protein-biased analogue of salvinorin A (SalA), 16-bromo salvinorin A (16-BrSalA), for its anticocaine effects, side effects, and activation of cellular signaling pathways. 16-BrSalA dose-dependently decreased the cocaine-primed reinstatement of drug-seeking behavior in a KOR-dependent manner. It also decreased cocaine-induced hyperactivity, but had no effect on responding for cocaine on a progressive ratio schedule. Compared to SalA, 16-BrSalA had an improved side effect profile, with no significant effects in the elevated plus maze, light–dark test, forced swim test, sucrose self-administration, or novel object recognition; however, it did exhibit conditioned aversive effects. 16-BrSalA increased dopamine transporter (DAT) activity in HEK-293 cells coexpressing DAT and KOR, as well as in rat nucleus accumbens and dorsal striatal tissue. 16-BrSalA also increased the early phase activation of extracellular-signal-regulated kinases 1 and 2, as well as p38 in a KOR-dependent manner. In NHPs, 16-BrSalA caused dose-dependent increases in the neuroendocrine biomarker prolactin, similar to other KOR agonists, at doses without robust sedative effects. These findings highlight that G-protein-biased structural analogues of SalA can have improved pharmacokinetic profiles and fewer side effects while maintaining their anticocaine effects. [ABSTRACT FROM AUTHOR]

Striatal kappa opioid receptor (KOR) availability in 48 subjects with Alcohol Use Disorder (AUD) was previously found to be associated with degree of drinking following a week of naltrexone treatment (de Laat et al. Biological Psychiatry, 86(11), 864-871, 2019). The purpose of the current study was to determine if spectral clustering applied to previously acquired KOR images (with [11C]LY2795050 PET) could identify meaningful groupings of different responses to naltrexone and to assess the robustness of the finding. Spectral clustering was applied to 6 features (regional volume of distribution values, VT) per AUD subject to produce 3 classes of subjects with different mean responses to naltrexone. Response to naltrexone was quantified as the difference in drinks consumed in an established lab-based alcohol drinking paradigm (Krishnan-Sarin et al. Biological Psychiatry, 62(6), 694-697, 2007) prior to, and after a week of naltrexone treatment. Clustering was applied exclusively to features of the image data with no a priori knowledge of the subjects' responses. Separation of classes was tested using a 1-way analysis of variance (ANOVA) with drink reduction as the outcome of interest. To assess robustness of the result, the size of the training set was varied by using successively reduced subsets of the data. Clustering resulted in significantly different groupings of drink reduction. The finding was robust to initialization of the spectral clustering procedure and was replicable for different random subsets of training subjects. Finding: Spectral clustering of kappa PET images separates AUD subjects into behaviorally distinct groups expressing distinct responses to naltrexone. [ABSTRACT FROM AUTHOR]

Chronic kidney disease-associated pruritus (CKD-aP) is often experienced by patients with CKD receiving dialysis. Approximately 40% of hemodialysis patients are 'moderately' to 'extremely bothered' by itching, associated with reduced quality of life, poor sleep quality, and depression as well as worse clinical outcomes, including increased medication use, infections, hospitalizations, and mortality. This review covers the pathophysiology and treatment landscape of CKD-aP, and the development, clinical efficacy, and safety profile of difelikefalin. We summarize the existing evidence, and discuss both the position of difelikefalin in the treatment pathway and potential future developments. Difelikefalin is a kappa opioid receptor agonist, with a primary mode of action that is outside of the central nervous system and provides an improved safety profile compared with other opioid agonists, with limited potential for abuse and dependency. Difelikefalin has demonstrated efficacy, tolerability, and safety profile in several large-scale clinical trials in more than 1,400 hemodialysis patients with CKD-aP treated for up to 64 weeks. Difelikefalin is the only approved treatment for CKD-aP in the U.S.A and Europe; other treatments are used off-label, have limited proof of efficacy in large-scale clinical trials in this patient population, and may present an increased risk of toxicity in patients with CKD. [ABSTRACT FROM AUTHOR]

Nalfurafine, a moderately selective kappa opioid receptor (KOR) agonist, is used in Japan for treatment of itch without causing dysphoria or psychotomimesis. Here we characterized the pharmacology of compound 42B, a 3-dehydroxy analogue of nalfurafine and compared with that of nalfurafine. Nalfurafine and 42B acted as full KOR agonists and partial μ opioid receptor (MOR) agonists, but 42B showed much lower potency for both receptors and lower KOR/MOR selectivity, different from previous reports. Molecular modeling revealed that water-mediated hydrogen-bond formation between 3-OH of nalfurafine and KOR accounted for its higher KOR potency than 42B. The higher potency of both at KOR over MOR may be due to hydrogen-bond formation between nonconserved Y7.35 of KOR and their carbonyl groups. Both showed modest G protein signaling biases. In mice, like nalfurafine, 42B produced antinociceptive and antiscratch effects and did not cause conditioned place aversion (CPA) in the effective dose ranges. Unlike nalfurafine, 42B caused motor incoordination and hypolocomotion. As both agonists showed G protein biases, yet produced different effects on locomotor activity and motor incoordination, the findings and those in the literature suggest caution in correlating in vitro biochemical data with in vivo behavior effects. The factors contributing to the disconnect, including pharmacodynamic and pharmacokinetic issues, are discussed. In addition, our results suggest that among the KOR-induced adverse behaviors, CPA can be separated from motor incoordination and hypolocomotion.

Abstract Background Adverse pathophysiological and behavioral outcomes related to mild traumatic brain injury (mTBI), posttraumatic stress disorder (PTSD), and chronic pain are common following blast exposure and contribute to decreased quality of life, but underlying mechanisms and prophylactic/treatment options remain limited. The dynorphin/kappa opioid receptor (KOR) system helps regulate behavioral and inflammatory responses to stress and injury; however, it has yet to be investigated as a potential mechanism in either humans or animals exposed to blast. We hypothesized that blast-induced KOR activation mediates adverse outcomes related to inflammation and affective behavioral response. Methods C57Bl/6 adult male mice were singly or repeatedly exposed to either sham (anesthesia only) or blast delivered by a pneumatic shock tube. The selective KOR antagonist norBNI or vehicle (saline) was administered 72 h prior to repetitive blast or sham exposure. Serum and brain were collected 10 min or 4 h post-exposure for dynorphin A-like immunoreactivity and cytokine measurements, respectively. At 1-month post-exposure, mice were tested in a series of behavioral assays related to adverse outcomes reported by humans with blast trauma. Results Repetitive but not single blast exposure resulted in increased brain dynorphin A-like immunoreactivity. norBNI pretreatment blocked or significantly reduced blast-induced increase in serum and brain cytokines, including IL-6, at 4 h post exposure and aversive/anxiety-like behavioral dysfunction at 1-month post-exposure. Conclusions Our findings demonstrate a previously unreported role for the dynorphin/KOR system as a mediator of biochemical and behavioral dysfunction following repetitive blast exposure and highlight this system as a potential prophylactic/therapeutic treatment target.

Increased vigilance in settings of potential threats or in states of vulnerability related to pain is important for survival. Pain disrupts sleep and conversely, sleep disruption enhances pain, but the underlying mechanisms remain unknown. Chronic pain engages brain stress circuits and increases secretion of dynorphin, an endogenous ligand of the kappa opioid receptor (KOR). We therefore hypothesized that hypothalamic dynorphin/KOR signalling may be a previously unknown mechanism that is recruited in pathological conditions requiring increased vigilance. We investigated the role of KOR in wakefulness, non-rapid eye movement (NREM) sleep and rapid eye movement (REM) sleep in freely moving naïve mice and in mice with neuropathic pain induced by partial sciatic nerve ligation using EEG/EMG recordings. Systemic continuous administration of U69,593, a KOR agonist, over 5 days through an osmotic minipump decreased the amount of NREM and REM sleep and increased sleep fragmentation in naïve mice throughout the light-dark sleep cycle. We used KORcre mice to selectively express a Gi-coupled designer receptor activated by designer drugs (Gi-DREADD) in KORcre neurons of the hypothalamic paraventricular nucleus, a key node of the hypothalamic-pituitary-adrenal stress response. Sustained activation of Gi-DREADD with clozapine-N-oxide delivered in drinking water over 4 days, disrupted sleep in these mice in a similar way as systemic U69,593. Mice with chronic neuropathic pain also showed disrupted NREM and total sleep that was normalized by systemic administration of two structurally different KOR antagonists, norbinaltorphimine and NMRA-140, currently in phase II clinical development, or by CRISPR/Cas9 editing of paraventricular nucleus KOR, consistent with endogenous KOR activation disrupting sleep in chronic pain. Unexpectedly, REM sleep was diminished by either systemic KOR antagonist or by CRISPR/Cas9 editing of paraventricular nucleus KOR in sham-operated mice. Our findings reveal previously unknown physiological and pathophysiological roles of dynorphin/KOR in eliciting arousal. Physiologically, dynorphin/KOR signalling affects transitions between sleep stages that promote REM sleep. Furthermore, while KOR antagonists do not promote somnolence in the absence of pain, they normalized disrupted sleep in chronic pain, revealing a pathophysiological role of KOR signalling that is selectively recruited to promote vigilance, increasing chances of survival. Notably, while this mechanism is likely beneficial in the short-term, disruption of the homeostatic need for sleep over longer periods may become maladaptive resulting in sustained pain chronicity. A novel approach for treatment of chronic pain may thus result from normalization of chronic pain-related sleep disruption by KOR antagonism. [ABSTRACT FROM AUTHOR]

The serotonin and kappa opioid receptor (KOR) systems are strongly implicated in disorders of negative affect, such as anxiety and depression. KORs expressed on axon terminals inhibit the release of neurotransmitters, including serotonin. The substantia nigra pars reticulata (SNr) is involved in regulating affective behaviors. It receives the densest serotonergic innervation in the brain and has high KOR expression; however, the influence of KORs on serotonin transmission in this region is yet to be explored. Here, we used ex vivo fast-scan cyclic voltammetry (FSCV) to investigate the effects of a KOR agonist, U50, 488 (U50), and a selective serotonin reuptake inhibitor, escitalopram, on serotonin release and reuptake in the SNr. U50 alone reduced serotonin release and uptake, and escitalopram alone augmented serotonin release and slowed reuptake, while pretreatment with U50 blunted both the release and uptake effects of escitalopram. Here, we show that the KOR influences serotonin signaling in the SNr in multiple ways and short-term activation of the KOR alters serotonin responses to escitalopram. These interactions between KORs and serotonin may contribute to the complexity in the responses to treatments for disorders of negative affect. Ultimately, the KOR system may prove to be a promising pharmacological target, alongside traditional antidepressant treatments. [ABSTRACT FROM AUTHOR]

Marine cyanobacteria are a rich source of bio-active metabolites that have been utilized as leads for drug discovery and pharmacological tools for basic science research. Here, we describe the re-isolation of a well-known metabolite, barbamide, from Curaçao on three different occasions and the characterization of barbamide's biological interactions with targets of the mammalian nervous system. Barbamide was originally discovered as a molluscicidal agent from a filamentous marine cyanobacterium. In our hands, we found little evidence of toxicity against mammalian cell cultures. However, barbamide showed several affinities when screened for binding affinity for a panel of 45 receptors and transporters known to be involved in nociception and sensory neuron activity. We found high levels of binding affinity for the dopamine transporter, the kappa opioid receptor, and the sigma receptors (sigma-1 and sigma-2 also known as transmembrane protein 97; TMEM97). We tested barbamide in vitro in isolated sensory neurons from female mice to explore its functional impact on calcium flux in these cells. Barbamide by itself had no observable impact on calcium flux. However, barbamide enhanced the effect of the TRPV1 agonist capsaicin and enhanced store-operated calcium entry (SOCE) responses after depletion of intracellular calcium. Overall, these results demonstrate the biological potential of barbamide at sensory neurons with implications for future drug development projects surrounding this molecule. [ABSTRACT FROM AUTHOR]

The natural product Salvinorin A (SalA) was the first nitrogen-lacking agonist discovered for the opioid receptors and exhibits high selectivity for the kappa opioid receptor (KOR) turning SalA into a promising analgesic to overcome the current opioid crisis. Since SalA's suffers from poor pharmacokinetic properties, particularly the absence of gastrointestinal bioavailability, fast metabolic inactivation, and subsequent short duration of action, the rational design of new tailored analogs with improved clinical usability is highly desired. Despite being known for decades, the binding mode of SalA within the KOR remains elusive as several conflicting binding modes of SalA were proposed hindering the rational design of new analgesics. In this study, we rationally determined the binding mode of SalA to the active state KOR by in silico experiments (docking, molecular dynamics simulations, dynophores) in the context of all available mutagenesis studies and structure-activity relationship (SAR) data. To the best of our knowledge, this is the first comprehensive evaluation of SalA's binding mode since the determination of the active state KOR crystal structure. SalA binds above the morphinan binding site with its furan pointing toward the intracellular core while the C2-acetoxy group is oriented toward the extracellular loop 2 (ECL2). SalA is solely stabilized within the binding pocket by hydrogen bonds (C210ECL2, Y3127.35, Y3137.36) and hydrophobic contacts (V1182.63, I1393.33, I2946.55, I3167.39). With the disruption of this interaction pattern or the establishment of additional interactions within the binding site, we were able to rationalize the experimental data for selected analogs. We surmise the C2-substituent interactions as important for SalA and its analogs to be experimentally active, albeit with moderate frequency within MD simulations of SalA. We further identified the non-conserved residues 2.63, 7.35, and 7.36 responsible for the KOR subtype selectivity of SalA. We are confident that the elucidation of the SalA binding mode will promote the understanding of KOR activation and facilitate the development of novel analgesics that are urgently needed. [ABSTRACT FROM AUTHOR]

Background: Adverse pathophysiological and behavioral outcomes related to mild traumatic brain injury (mTBI), posttraumatic stress disorder (PTSD), and chronic pain are common following blast exposure and contribute to decreased quality of life, but underlying mechanisms and prophylactic/treatment options remain limited. The dynorphin/kappa opioid receptor (KOR) system helps regulate behavioral and inflammatory responses to stress and injury; however, it has yet to be investigated as a potential mechanism in either humans or animals exposed to blast. We hypothesized that blast-induced KOR activation mediates adverse outcomes related to inflammation and affective behavioral response.Methods: C57Bl/6 adult male mice were singly or repeatedly exposed to either sham (anesthesia only) or blast delivered by a pneumatic shock tube. The selective KOR antagonist norBNI or vehicle (saline) was administered 72 h prior to repetitive blast or sham exposure. Serum and brain were collected 10 min or 4 h post-exposure for dynorphin A-like immunoreactivity and cytokine measurements, respectively. At 1-month post-exposure, mice were tested in a series of behavioral assays related to adverse outcomes reported by humans with blast trauma.Results: Repetitive but not single blast exposure resulted in increased brain dynorphin A-like immunoreactivity. norBNI pretreatment blocked or significantly reduced blast-induced increase in serum and brain cytokines, including IL-6, at 4 h post exposure and aversive/anxiety-like behavioral dysfunction at 1-month post-exposure.Conclusions: Our findings demonstrate a previously unreported role for the dynorphin/KOR system as a mediator of biochemical and behavioral dysfunction following repetitive blast exposure and highlight this system as a potential prophylactic/therapeutic treatment target. [ABSTRACT FROM AUTHOR]

Immediately following spinal cord injury (SCI) patients experience pain associated with injury to the spinal cord and nerves as well as with accompanying peripheral injuries. This pain is usually treated with opioids, and most commonly with morphine. However, in a rodent model we have shown that, irrespective of the route of administration, morphine administered in the acute phase of SCI undermines long-term locomotor recovery. Our previous data suggest that activation of kappa opioid receptors (KORs) mediates these negative effects. Blocking KORs with norbinaltorphimine (norBNI), prior to a single dose of epidural morphine, prevented the morphine-induced attenuation of locomotor recovery. Because numerous cellular changes occur with chronic opioid administration compared with a single dose, the current study tested whether norBNI was also effective in a more clinically relevant paradigm of repeated, intravenous morphine administration after SCI. We hypothesized that blocking KOR activation during repeated, intravenous morphine administration would also protect recovery. Supporting this hypothesis, we found that blocking KOR activation in young, male rats prevented the negative effects of morphine on locomotor recovery, although neither norBNI nor morphine had an effect on long-term pain at the doses used. We also found that norBNI treatment blocked the adverse effects of morphine on lesion size. These data suggest that a KOR antagonist given in conjunction with morphine may provide a clinical strategy for effective analgesia without compromising locomotor recovery after SCI. [ABSTRACT FROM AUTHOR]

Aim: Endogenous dynorphin signaling via kappa opioid receptors (KORs) plays a key role in producing the depressive and aversive consequences of stress. We investigated the behavioral effects of the dynorphin/KOR system in the ventral pallidum (VP) and studied the underlying mechanisms. Methods: To investigate the effects of dynorphin on the VP, we conducted behavioral experiments after microinjection of drugs or shRNA and brain‐slice electrophysiological recordings. Histological tracing and molecular biological experiments were used to identify the distribution of KORs and the possible sources of dynorphin projections to the VP. Results: An elevated dynorphin concentration and increased KOR activity were observed in the VP after acute stress. Infusion of dynorphin‐A into the VP produced depressive‐like phenotypes including anhedonia and despair and anxiety behaviors, but did not alter locomotor behavior. Mechanistically, dynorphin had an inhibitory effect on VP neurons—reducing their firing rate and inhibiting excitatory transmission—through direct activation of KORs and modulation of downstream G‐protein‐gated inwardly rectifying potassium (GIRK) channels and high‐voltage gated calcium channels (VGCCs). Tracing revealed direct innervation of VP neurons by dynorphin‐positive projections; potential sources of these dynorphinergic projections include the nucleus accumbens, amygdala, and hypothalamus. Blockade of dynorphin/KOR signaling in the VP by drugs or viral knock‐down of KORs significantly reduced despair behavior in rats. Conclusions: Endogenous dynorphinergic modulation of the VP plays a critical role in mediating depressive reactions to stress. [ABSTRACT FROM AUTHOR]

Kappa opioid receptor (KOR) agonists are potentially useful as analgesic and anti-pruritic agents, for prevention and treatment of substance use disorders, and for treatment of demyelinating diseases. However, side effects of KOR agonists, including psychotomimesis, dysphoria, and sedation, have caused early termination of clinical trials. Understanding the signaling mechanisms underlying the beneficial therapeutic effects and the adverse side effects may help in the development of KOR agonist compounds. In this review, we summarize the current knowledge in this regard in five sections. First, studies conducted on mutant mouse lines (GRK3-/-, p38alpha MAPK-/-, b-arrestin2-/-, phosphorylation-deficient KOR) are summarized. In addition, the abilities of four distinct KOR agonists, which have analgesic and anti-pruritic effects with different side effect profiles, to cause KOR phosphorylation are discussed. Second, investigations on the KOR agonist nalfurafine, both in vitro and in vivo are reviewed. Nalfurafine was the first KOR full agonist approved for clinical use and in the therapeutic dose range it did not produce significant side effects associated with typical KOR agonists. Third, large-scale high-throughput phosphoproteomic studies without a priori hypotheses are described. These studies have revealed that KOR-mediated side effects are associated with many signaling pathways. Fourth, several novel G proteinbiased KOR agonists that have been characterized for in vitro biochemical properties and agonist biases and in vivo behavior effects are described. Lastly, possible mechanisms underlying KOR-mediated CPA, hypolocomotion and motor incoordination are discussed. Overall, it is agreed upon that the analgesic and anti-pruritic effects of KOR agonists are mediated via G protein signaling. However, there is no consensus on the mechanisms underlying their side effects. GRK3, p38 MAPK, b-arrestin2, mTOR pathway, CB1 cannabinoid receptor and protein kinase C have been implicated in one side effect or another. For drug discovery, after initial in vitro characterization, in vivo pharmacological characterizations in various behavior tests are still the most crucial steps and dose separation between beneficial therapeutic effects and adverse side effects are the critical determinant for the compounds to be moved forward for clinical development. [ABSTRACT FROM AUTHOR]

Abstract Background As a class of the opioid receptors, the kappa opioid receptor (KOR) has been verified to be a potential biomarker and therapeutic target for human malignant tumors. However, a thorough understanding of whether KOR affects progression of esophageal squamous cell carcinoma (ESCC) is still lacking. This study focused on exploring the effect of knocking down KOR in ESCC and its underlying mechanism. Methods Bioinformatics analysis was used to compare the different expression level of OPRK1 (KOR gene) in tumor and adjacent normal tissues, and predict the relationship between KOR expression and overall survival. RNA-sequence analysis was performed to detect the altered functions and mechanisms after down regulating KOR. The in vitro and in vivo assays were used to detect the effects of down-regulated KOR on cell proliferation, migration and invasion. Substrate gel zymography and 3D cell culture assays were used to find the effect of KOR knockdown on the degradation of extracellular matrix (ECM), and immunefluorescence was performed to detect the altered cytoskeleton. Western blotting and immunohistochemistry were used to explore the underlying mechanism pathway. Results Bioinformatics analysis revealed that the expression of OPRK1 was lower in tumor tissue than that in adjacent normal tissues, and lowered expression of KOR was associated with poorer overall survival. The in vitro assays demonstrated that down-regulation of KOR enhanced ESCC proliferation, metastasis and invasion. Western blotting revealed that down-regulation of KOR could activate PDK1-AKT signaling pathway, which actively regulated the cancer progression. Down-regulation of KOR enhanced the formation of invadopodia, secretion of matrix metalloproteinase-2 (MMP2) and rearrangement of cytoskeleton, which were positively related with the invasion of ESCC. KOR knockdown enhanced the tumor invasion and elevated the AKT phosphorylation in nude mice. The AKT kinase inhibition could reverse the effect of down-regulation of KOR. Conclusion KOR might act as a tumor suppressor in ESCC and down-regulation of KOR could enhance the ESCC tumor phenotype. Graphical abstract Video Abstract