Challenges for opioid receptor nomenclature: IUPHAR Review 9.

UNLABELLED: Recent developments in the study of the structure and function of opioid receptors raise significant challenges for the definition of individual receptor types and the development of a nomenclature that precisely describes isoforms that may subserve different functions in vivo. Presentations at the 2013 meeting of the International Narcotics Research Conference in Cairns, Australia, considered some of the new discoveries that are now unravelling the complexities of opioid receptor signalling. Variable processing of opioid receptor messenger RNAs may lead to the presence of several isoforms of the µ receptor. Each opioid receptor type can function either as a monomer or as part of a homo- or heterodimer or higher multimer. Additionally, recent evidence points to the existence of agonist bias in the signal transduction pathways activated through µ receptors, and to the presence of regulatory allosteric sites on the receptors. This brief review summarizes the recent discoveries that raise challenges for receptor definition and the characterization of signal transduction pathways activated by specific receptor forms. LINKED ARTICLES: This article is part of a themed section on Opioids: New Pathways to Functional Selectivity. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-2.

Interleukin-1 beta-induced up-regulation of opioid receptors in the untreated and morphine-desensitized U87 MG human astrocytoma cells.

BACKGROUND: Interleukin-1beta (IL-1ß) is a pro-inflammatory cytokine that can be produced in the central nervous system during inflammatory conditions. We have previously shown that IL-1ß expression is altered in the rat brain during a morphine tolerant state, indicating that this cytokine may serve as a convergent point between the immune challenge and opiate mediated biological pathways. We hypothesized that IL-1ß up-regulates opioid receptors in human astrocytes in both untreated and morphine-desensitized states. METHODS: To test this hypothesis, we compared the basal expression of the mu (MOR), delta (DOR), and kappa (KOR) opioid receptors in the human U87 MG astrocytic cell line to SH-SY5Y neuronal and HL-60 immune cells using absolute quantitative real time RT-PCR (AQ-rt-RT-PCR). To demonstrate that IL-1ß induced up-regulation of the MOR, DOR and KOR, U87 MG cells (2 x 105 cells/well) were treated with IL-1ß (20 ng/mL or 40 ng/mL), followed by co-treatment with interleukin-1 receptor antagonist protein (IL-1RAP) (400 ng/mL or 400 ng/mL). The above experiment was repeated in the cells desensitized with morphine, where U87 MG cells were pre-treated with 100 nM morphine. The functionality of the MOR in U87 MG cells was then demonstrated using morphine inhibition of forksolin-induced intracellular cAMP, as determined by radioimmunoassay. RESULTS: U87 MG cells treated with IL-1ß for 12 h showed a significant up-regulation of MOR and KOR. DOR expression was also elevated, although not significantly. Treatment with IL-1ß also showed a significant up-regulation of the MOR in U87 MG cells desensitized with morphine. Co-treatment with IL-1ß and interleukin-1 receptor antagonist protein (IL-1RAP) resulted in a significant decrease in IL-1ß-mediated MOR up-regulation. CONCLUSION: Our results indicate that the pro-inflammatory cytokine, IL-1ß, affects opiate-dependent pathways by up-regulating the expression of the MOR in both untreated and morphine-desensitized U87 MG.

Study on the activation of the opioid receptors by a set of morphine derivatives in a well-defined assay system.

As a first step in our search for new opiates, we have established cellular assays to monitor opioid receptor activation and study the activities of a set of morphine derivatives. Intracellular calcium changes were monitored in human embryonic kidney-293 T cells expressing individual opioid receptors upon cotransfection with a chimeric G protein. This assay was validated by comparing the potencies of the endogenous peptides to reported values. All of the opiates were found to interact with the three opioid receptor subtypes but with a range of differences in efficacies and potencies. Most of the opiates preferentially acted at the µ receptor. None of the opiates showed a preference for the δ receptor. Only oripavine and its precursor thebaine showed a preference for the κ over the µ receptor. The results indicate that the opiates with a C-3 hydroxyl group or C-6 ketone group but in the presence of a 7, 8-single bond exhibit higher activity. It is noteworthy that the 6-O-methyl group seems to improve the selectivity for κ receptor. This is the first comparative and comprehensive report on the activation of the three different opioid receptors by a set of morphine derivatives in a well-defined assay system. These data can serve as a basis for the characterization of novel opiates.

Opioid receptor subtypes: fact or artifact?

There is a vast amount of pharmacological evidence favouring the existence of multiple subtypes of opioid receptors. In addition to the primary classification of µ (mu: MOP), δ (delta: DOP), κ (kappa: KOP) receptors, and the nociceptin/orphanin FQ peptide receptor (NOP), various groups have further classified the pharmacological µ into µ(1-3), the δ into δ(1-2)/δ(complexed/non-complexed), and the κ into κ(1-3). From an anaesthetic perspective, the suggestions that µ(1) produced analgesia and µ(2) produced respiratory depression are particularly important. However, subsequent to the formal identification of the primary opioid receptors (MOP/DOP/KOP/NOP) by cloning and the use of this information to produce knockout animals, evidence for these additional subtypes is lacking. Indeed, knockout of a single gene (and hence receptor) results in a loss of all function associated with that receptor. In the case of MOP knockout, analgesia and respiratory depression is lost. This suggests that further sub-classification of the primary types is unwise. So how can the wealth of pharmacological data be reconciled with new molecular information? In addition to some simple misclassification (κ(3) is probably NOP), there are several possibilities which include: (i) alternate splicing of a common gene product, (ii) receptor dimerization, (iii) interaction of a common gene product with other receptors/signalling molecules, or (iv) a combination of (i)-(iii). Assigning variations in ligand activity (pharmacological subtypes) to one or more of these molecular suggestions represents an interesting challenge for future opioid research.

Opioid peptides in peripheral pain control.

Opioids have a long history of therapeutic use as a remedy for various pain states ranging from mild acute nociceptive pain to unbearable chronic advanced or end-stage disease pain. Analgesia produced by classical opioids is mediated extensively by binding to opioid receptors located in the brain or the spinal cord. Nevertheless, opioid receptors are also expressed outside the CNS in the periphery and may become valuable assets in eliciting analgesia devoid of shortcomings typical for the activation of their central counterparts. The discovery of endogenous opioid peptides that participate in the formation, transmission, modulation and perception of pain signals offers numerous opportunities for the development of new analgesics. Novel peptidic opioid receptor analogs, which show limited access through the blood brain barrier may support pain therapy requiring prolonged use of opioid drugs.

Involvement of opioid receptors in the CGRP-induced antinociception in the nucleus accumbens of rats.

It has been demonstrated that calcitonin gene-related peptide (CGRP) plays important roles in the modulation of nociception in the nucleus accumbens (NAc) of rats. The present study is performed to explore the possible involvement of opioid receptors in the CGRP-induced antinociception in the NAc of rats. Intra-NAc administration of CGRP induces significant increases in the hindpaw withdrawal latency (HWL) to noxious thermal and mechanical stimulation in rats. Interestingly, the CGRP-induced antinociceptive effects are inhibited by following intra-NAc injection of the opioid receptor antagonist naloxone, suggesting that the opioid receptors are involved in the CGRP-induced antinociception in the NAc of rats. Furthermore, the CGRP-induced antinociception is attenuated by intra-NAc injection of mu-opioid receptor (MOR) antagonist beta-funaltrexamine (beta-FNA) and kappa-opioid receptor (KOR) antagonist nor-binaltorphimine (nor-BNI), but not by delta-receptor (DOR) antagonist naltrindole. In the present study, we also demonstrated that there was no significant difference between the CGRP-induced antinociception and the morphine-induced antinociception in the NAc in rats. The results of the present study demonstrate that both mu- and kappa-opioid receptors are involved in the CGRP-induced antinociception in the NAc of rats.

Function of the opioid system during inflammation in carp.

The opioid system is involved in modulation of both innate and acquired immune responses, thus altering resistance to a variety of infectious agents. We sequenced and characterized carp opioid receptors (MOR, DOR, and KOR) and found their regulated expression in piscine leukocytes. Moreover, both in vivo and in vitro opioids affect activity of leukocytes and expression of inflammatory molecules, especially chemokines and chemokine receptors. Our data indicate an evolutionary conserved role for the opioid system in immune regulation.

Multiple ligands in opioid research.

The observation in 1979 that opioid receptors interact, led to the design of bivalent ligands in an attempt to improve selectivity and affinity towards the different subtypes( i.e. mu, delta, and kappa). Dimers of monovalent 'parent' opioid structures have been evaluated and include: (a) endogenous (e.g enkephalins) or exogenous (e.g dermorphin) peptide dimer analogues (b) mixed peptidic -non-peptidic bivalent ligands and (c) dual non-peptidic dimers. Chimeric structures, using an opioid pharmacophore in combination with a a non-opioid pharmacophore, have also been prepared. The common aim in all these studies is to improve the pharmacological profile of potential analgesics to minimize common opioid-induced side effects, such as physical dependence and tolerance. Here we present a brief overview efforts to develop bivalent opioid ligands for use in pain-related research.

Roles of opioid receptor subtypes on the antinociceptive effect of intrathecal sildenafil in the formalin test of rats.

Recently, it has been known that the antinociception of sildenafil, a phosphodiesterase 5 inhibitor, is mediated through the opioid receptors. There are common three types of opioid receptors mu, delta, and kappa. We characterized the role of subtypes of opioid receptor for the antinociception of sildenafil at the spinal level. Intrathecal catheters were placed for drug delivery and formalin solution (5%, 50 microl) was injected for induction of nociception within male SD rats. The effect of mu opioid receptor antagonist (CTOP), delta opioid receptor antagonist (naltrindole), and kappa opioid receptor antagonist (GNTI) on the activity of sildenafil was examined. Intrathecal sildenafil decreased the flinching responses during phases 1 and 2 in the formalin test. Intrathecal CTOP and naltrindole reversed the antinociception of sildenafil during both phases in the formalin test. Intrathecal GNTI reversed the effect of sildenafil during phase 2, but not phase 1. These results suggest that sildenafil is effective to acute pain and the facilitated pain state at the spinal level. Both mu and delta opioid receptors are involved. However, it seems that kappa opioid receptors play in the effect of sildenafil.

Decreased spinal cord opioid receptor mRNA expression and antinociception in a Theiler's murine encephalomyelitis virus model of multiple sclerosis.

Multiple sclerosis patients typically experience increased pain that is relatively insensitive to opiate treatment. The mechanistic basis for this increased nociception is currently poorly understood. In the present study, we utilized the Theiler's murine encephalomyelitis virus (TMEV) model of MS to examine possible changes in spinal cord opioid receptor mRNA over the course of disease progression. TMEV infection led to significantly decreased mu, delta and kappa opioid receptor mRNA expression as analyzed by quantitative real-time PCR in both male and female mice at days 90, 150 and 180 post-infection (PI). Since opioid receptor mRNA expression decreased in TMEV mice, we examined whether opiate analgesia is also altered. TMEV infected female mice had significantly decreased opiate analgesia in thermal nociceptive tests beginning at day 90 PI, while TMEV-infected male mice did not display significantly decreased opiate analgesia until day 120 PI. The novel finding that opioid receptor expression is significantly decreased in the spinal cord of TMEV mice could explain the increased nociception and loss of opiate analgesia observed in both TMEV mice and multiple sclerosis patients.

Opioid systems in the dentate gyrus.

Opiate drugs alter cognitive performance and influence hippocampal excitability, including long-term potentiation (LTP) and seizure activity. The dentate gyrus (DG) contains two major opioid peptides, enkephalins and dynorphins, which have opposing effects on excitability. Enkephalins preferentially bind to delta- and mu-opioid receptors (DORs and MORs) while dynorphins preferentially bind to kappa-opioid receptors (KORs). Opioid receptors can also be activated by exogenous opiate drugs such as the MOR agonist morphine. Enkephalins are contained in the mossy fiber pathway, in the lateral perforant path (PP) and in scattered GABAergic interneurons. MORs and DORs are predominantly in distinct subpopulations of GABAergic interneurons known to inhibit granule cells, and are present at low levels within granule cells. MOR and DOR agonists increase excitability and facilitate LTP in the molecular layer. Anatomical and physiological evidence is consistent with somatodendritic and axon terminal targeting of both MORs and DORs. Dynorphins are in the granule cells, most abundantly in mossy fibers but also in dendrites. KORs have been localized to granule cell mossy fibers, supramammillary afferents to granule cells, and PP terminals. KOR agonists, including endogenous dynorphins, diminish the induction of LTP. Recent evidence indicates that opiates and opioids also modulate other processes in the hippocampal formation, including adult neurogenesis, the actions of gonadal hormones, and development of neonatal transmitter systems.

Relative contribution of peripheral versus central opioid receptors to antinociception.

Opioid effects are mediated by central and peripheral opioid receptors. Here we examine the relative contribution of each receptor population to antinociception elicited by systemically administered centrally penetrating opioids, and by loperamide (a peripherally restricted opioid). Nociception (abdominal writhes) was induced by intraperitoneally (i.p.) injected 0.6% acetic acid in mice. We analyzed opioid receptor expression in peritoneum by immunohistochemistry, antinociception after i.p. injected agonists at mu (morphine, loperamide)-, delta (SNC80)- and kappa (U50488)-receptors, and its reversibility by subcutaneously (s.c.) administered centrally penetrating antagonists beta-funaltrexamine (mu), naltrindole (delta) and nor-binaltorphimine (kappa), and by the peripherally restricted antagonist naloxone methiodide (NLXM). NLXM was also injected intracerebroventricularly (i.c.v.) before i.p. loperamide. Mu-, kappa- and, to a lesser degree, delta-receptors were expressed on peripheral nerve terminals in the peritoneum. The anatomical distribution of the opioid receptor staining was very similar to the staining for calcitonin gene-related peptide, a marker of sensory neurons. Morphine, U50488 and, to a lesser degree, SNC80 blocked acetic and acid induced writhes. These effects were reversed by beta-funaltrexamine, nor-binaltorphimine and naltrindole, respectively. NLXM (s.c.) reversed antinociceptive effects of morphine, SNC80 and U50488 by 57%, 80% and 47%, respectively. Loperamide (0.05 mg/kg)-induced antinociception was reversed by s.c. beta-funaltrexamine and NLXM. Loperamide (0.1 mg/kg)-induced antinociception was completely blocked by s.c. beta-funaltrexamine but was only attenuated (by 50%) by s.c. or i.c.v. NLXM. In conclusion, systemically administered centrally penetrating mu-, delta- and kappa-agonists produced a substantial part of antinociception through peripheral opioid receptors. Higher dose loperamide-induced antinociception involved also central opioid receptors.

Adverse effects of opioids on the central nervous systems of palliative care patients.

Opioids, defined as drugs that stimulate opioid receptors, are primarily used in the treatment of moderate to severe pain. They induce central nervous system (CNS) adverse effects which can be divided into three groups. The first group includes effects that lower the level of consciousness-sedation, drowsiness and sleep disturbance. The second group affects the thinking process and the ability to react-cognitive impairment, psychomotor impairment, delirium, hallucinations, dreams and nightmares. The third group is of the direct toxic effects of opioids on neurons and includes myoclonus (perhaps), hyperalgesia and tolerance. This review addresses the incidence, possible mechanisms, and treatment of each of these groups of opioid-induced adverse effects.

Relationship of opioid receptors with GABAergic neurons in the rat inferior colliculus.

The inferior colliculus is a critical structure for processing auditory information and receives ascending and descending synaptic auditory projections. In addition to GABAergic and glutamatergic innervations, other neurotransmitter systems are also reported in the inferior colliculus, including opioid peptides. In the present study, the relative distribution of each type of opioid receptor, mu (MOR), delta (DOR) and kappa (KOR) within GABAergic neurons in the inferior colliculus was examined. GABA immunoreactivity was expressed by small, medium and large neurons and distributed in the central nucleus and the pericentral nucleus of the inferior colliculus. Immunostaining for MOR, DOR and KOR receptors was found in both disc-shaped cells and stellate cells. Punctiform beta-endorphin immunolabelling was observed in the proximity of GABA-positive neurons. Co-localization of GABA and MOR receptors was observed in neurons and nerve terminals in the central nucleus, dorsal cortex and external cortex of the inferior colliculus. Quantification of the co-localization patterns determined that a higher proportion of GABA neurons was associated with MOR receptors compared with KOR or DOR receptors.

Atypical anxiolytic-like response to naloxone in benzodiazepine-resistant 129S2/SvHsd mice: role of opioid receptor subtypes.

RATIONALE: Mice of many 129 substrains respond to environmental novelty with behavioural suppression and high levels of anxiety-like behaviour. Although resistant to conventional anxiolytics, this behavioural phenotype may involve stress-induced release of endogenous opioids. OBJECTIVES: To assess the effects of opioid receptor blockade on behavioural reactions to novelty stress in a chlordiazepoxide-resistant 129 substrain. MATERIALS AND METHODS: Experiment 1 contrasted the effects of the broad-spectrum opioid receptor antagonist naloxone (1.0-10.0 mg/kg) in C57BL/6JOlaHsd and 129S2/SvHsd mice exposed to the elevated plus-maze. Experiments 2-4 examined the responses of 129S2/SvHsd mice to the mu-selective opioid receptor antagonist beta-funaltrexamine (2.5-10.0 mg/kg), the delta-selective antagonist naltrindole (2.5-10.0 mg/kg) and the kappa-selective antagonist nor-binaltorphimine (2.5-5.0 mg/kg). RESULTS: 129 mice displayed higher levels of anxiety-like behaviour and lower levels of general exploration relative to their C57 counterparts. Although naloxone failed to alter the behaviour of C57 mice, both doses of this antagonist produced behaviourally selective reductions in open-arm avoidance in 129 mice. Surprisingly, none of the more selective opioid receptor antagonists replicated this effect of naloxone: beta-funaltrexamine was devoid of behavioural activity, naltrindole suppressed rearing (all doses) and increased immobility (10 mg/kg), while nor-binaltorphimine (5 mg/kg) nonspecifically increased percent open arm entries. CONCLUSIONS: Recent evidence suggests differential involvement of opioid receptor subtypes in the anxiolytic efficacy of diverse compounds including conventional benzodiazepines. The insensitivity of 129 mice to the anxiolytic action of chlordiazepoxide, coupled with their atypical anxiolytic response to naloxone (but not more selective opioid receptor antagonists), suggests an abnormality in anxiety-related neurocircuitry involving opioid-GABA interactions.

Multiple opioid receptors on immune cells modulate intracellular signaling.

In 1979, Joseph Wybran reported his insights into the existence of different opioid receptor subtypes on T-cells. He observed that morphine and methionine enkephalin had different effects on human T-cell rosetting to sheep red blood cells. Since that time, a wide array of laboratories have shown that opiate alkyloids and opioid peptides exert pleiotropic effects on immune cell function. These compounds are immunomodulators, modifying immune responses to extracellular stimuli such as mitogens, antigens, and antibodies that cross-link the T-cell receptor. It has been demonstrated that cells involved in host defense and immunity express mRNA transcripts encoding the various opioid receptors originally described in neuronal tissues. Molecular imaging approaches have demonstrated the regulated expression of both delta and kappa opioid receptors, predominantly on T-cells. Moreover, atypical opiate and opioid binding sites are present on these cells. This review will consider the evidence for both classical and atypical opioid receptors and their effects on signaling within immune cells; our emphasis is the T-cell and its delta opioid receptor.

[Opioid receptors and their selective ligands]. / Receptory opioidowe i ich selektywne ligandy peptydowe.

Opioid receptors (micro, delta, and kappa) belong to a large family of G protein-coupled receptors and play an important physiological role. Stimulation of these receptors triggers analgesic effects and affects the function of gastrointestinal tract. The discovery of opioid peptides, which are endogenous ligands of opioid receptors, including delta-selective enkephalins, kappa-selective dynorphins, and micro-selective endomorphins, initiated their structure-activity relationship studies. For the last 30 years, hundreds of analogs of opioid peptides have been synthesized in an effort to obtain the compounds more active, selective, and resistant to biodegradation than the endogenous ligands. Different unnatural amino acids, as well as cyclisation procedures, leading to conformationaly restricted analogs, were employed. All these modifications resulted in obtaining very selective agonists and antagonists with high affinity at micro-, dlta-, and kappa-opioid receptors, which are extremely useful tools in further studies on the pharmacology of opioid receptors in a mammalian organism.

The endogenous opioid system and clinical pain management.

The endogenous opioid system is one of the most studied innate pain-relieving systems. This system consists of widely scattered neurons that produce three opioids: beta-endorphin, the met- and leu-enkephalins, and the dynorphins. These opioids act as neurotransmitters and neuromodulators at three major classes of receptors, termed mu, delta, and kappa, and produce analgesia. Like their endogenous counterparts, the opioid drugs, or opiates, act at these same receptors to produce both analgesia and undesirable side effects. This article examines some of the recent findings about the opioid system, including interactions with other neurotransmitters, the location and existence of receptor subtypes, and how this information drives the search for better analgesics. We also consider how an understanding of the opioid system affects clinical responses to opiate administration and what the future may hold for improved pain relief. The goal of this article is to assist clinicians to develop pharmacological interventions that better meet their patient's analgesic needs.