Action Of Endogenous Opioid Peptides Research Articles

Positive Allosteric Modulation of the Mu Opioid Receptor

Opioid therapeutics, such as morphine, that act at the mu‐opioid receptor (MOR) are the clinical standard for patients struggling to manage symptoms associated with pain. It is widely understood that although opioids are effective at treating pain, their use leads to the development of severe adverse effects, such as constipation, addiction, and respiratory depression. Thus, there is a clear need for a safer alternative to manage pain. One such alternative is to enhance the effects of the body’s endogenous opioid system by positive allosteric modulation (PAM) of MOR. A known PAM, BMS‐986122, enhances MOR agonist potency in cellular models and MOR agonist mediated antinociception in vivo. In addition, this PAM is active alone in a variety of mouse pain assays by promoting the activity of endogenous opioid peptides. Moreover, at an effective antinociceptive dose, BMS‐986122 alone produces less severe adverse effects than morphine as determined by measures of constipation, respiratory depression, and conditioned place preference. However, we do not yet know how the overall pharmacology of opioids is affected by PAMs or if all opioid drugs are equally sensitive to PAM modulation. Here we compare the ability of BMS‐986122 to enhance the action of three structurally diverse opioid drugs, morphine, methadone, and fentanyl, in an acute pain assay and in an assay examining respiratory depression using CD‐1 male and female mice. We show that BMS‐986122 increases the antinociceptive effects of the opioid therapeutics in the warm water tail withdrawal assay without promoting the ability of the drugs to lower blood oxygen levels or heart rate. Future work will assess the effects of BMS‐986122 in additional acute and chronic pain models and with a more diverse group of opioids. If additional experiments support the concept that BMS‐986122 enhances MOR‐mediated antinociception but not MOR‐mediated respiratory depression, this will validate the development of MOR‐PAMs as standalone pain medications and support the use of PAMs as opioid‐sparing drugs for the effective management of pain.

Positive allosteric modulation of the mu-opioid receptor produces analgesia with reduced side effects

Positive allosteric modulators (PAMs) of the mu-opioid receptor (MOR) have been hypothesized as potentially safer analgesics than traditional opioid drugs. This is based on the idea that PAMs will promote the action of endogenous opioid peptides while preserving their temporal and spatial release patterns and so have an improved therapeutic index. However, this hypothesis has never been tested. Here, we show that a mu-PAM, BMS-986122, enhances the ability of the endogenous opioid Methionine-enkephalin (Met-Enk) to stimulate G protein activity in mouse brain homogenates without activity on its own and to enhance G protein activation to a greater extent than β-arrestin recruitment in Chinese hamster ovary (CHO) cells expressing human mu-opioid receptors. Moreover, BMS-986122 increases the potency of Met-Enk to inhibit GABA release in the periaqueductal gray, an important site for antinociception. We describe in vivo experiments demonstrating that the mu-PAM produces antinociception in mouse models of acute noxious heat pain as well as inflammatory pain. These effects are blocked by MOR antagonists and are consistent with the hypothesis that in vivo mu-PAMs enhance the activity of endogenous opioid peptides. Because BMS-986122 does not bind to the orthosteric site and has no inherent agonist action at endogenously expressed levels of MOR, it produces a reduced level of morphine-like side effects of constipation, reward as measured by conditioned place preference, and respiratory depression. These data provide a rationale for the further exploration of the action and safety of mu-PAMs as an innovative approach to pain management.

Opioidergic regulation of contractile activity of lymphatic vessels

The aim is to study the effect of endogenous opioid peptides on the contractile activity of lymphatic vessels in the white rat. Materials and methods . Under isometric conditions, the dynamics of spontaneous contractile activity of isolated annular segments of rat lymphatic vessels under the action of endogenous opioid peptides (beta-endorphin and endomorphin-1) was record; possible mechanisms of opiates action were studied. Results . The range of β-endorphin level 1×10 –11 –1×10 –8 M resulted in a decrease of the minute productivity of the lymphatic vessels, by 22%, mainly due to a decreasing of single contractions frequency. In the presence of blockers of voltage-gated and ATP-sensitive potassium channels and blockers of eNOS, the inhibitory effect was not manifest. Endomorphin-1, used in the concentration range of 1×10 –10 –1×10 –8 M, stimulated the contractile activity of the lymphatic vessels by an average of 15 %. The stimulatory effect was not manifest when using endomorphin-1 in the presence of a selective NK1 receptor antagonist. Conclusions . The inhibitory effect of β-endorphin is associated with the activation of voltage-dependent and ATP-sensitive potassium channels, endothelium-(NO)-dependent mechanisms. The action of endomorphin-1, stimulating lymphangionic motility, can be carry out through interaction with NK receptors.

Regulator of G-Protein Signaling (RGS) Protein Modulation of Opioid Receptor Signaling as a Potential Target for Pain Management.

Opioid drugs are the gold standard for the management of pain, but their use is severely limited by dangerous and unpleasant side effects. All clinically available opioid analgesics bind to and activate the mu-opioid receptor (MOR), a heterotrimeric G-protein-coupled receptor, to produce analgesia. The activity of these receptors is modulated by a family of intracellular RGS proteins or regulators of G-protein signaling proteins, characterized by the presence of a conserved RGS Homology (RH) domain. These proteins act as negative regulators of G-protein signaling by serving as GTPase accelerating proteins or GAPS to switch off signaling by both the Gα and βγ subunits of heterotrimeric G-proteins. Consequently, knockdown or knockout of RGS protein activity enhances signaling downstream of MOR. In this review we discuss current knowledge of how this activity, across the different families of RGS proteins, modulates MOR activity, as well as activity of other members of the opioid receptor family, and so pain and analgesia in animal models, with particular emphasis on RGS4 and RGS9 families. We discuss inhibition of RGS proteins with small molecule inhibitors that bind to sensitive cysteine moieties in the RH domain and the potential for targeting this family of intracellular proteins as adjuncts to provide an opioid sparing effect or as standalone analgesics by promoting the activity of endogenous opioid peptides. Overall, we conclude that RGS proteins may be a novel drug target to provide analgesia with reduced opioid-like side effects, but that much basic work is needed to define the roles for specific RGS proteins, particularly in chronic pain, as well as a need to develop newer inhibitors.

Characterization of a Multiple-Scan-Rate Voltammetric Waveform for Real-Time Detection of Met-Enkephalin.

Opioid peptides are critically involved in a variety of physiological functions necessary for adaptation and survival, and as such, understanding the precise actions of endogenous opioid peptides will aid in identification of potential therapeutic strategies to treat a variety of disorders. However, few analytical tools are currently available that offer both the sensitivity and spatial resolution required to monitor peptidergic concentration fluctuations in situ on a time scale commensurate with that of neuronal communication. Our group has developed a multiple-scan-rate waveform to enable real-time voltammetric detection of tyrosine containing neuropeptides. Herein, we have evaluated the waveform parameters to increase sensitivity to methionine-enkephalin (M-ENK), an endogenous opioid neuropeptide implicated in pain, stress, and reward circuits. M-ENK dynamics were monitored in adrenal gland tissue, as well as in the dorsal striatum of anesthetized and freely behaving animals. The data reveal cofluctuations of catecholamine and M-ENK in both locations and provide measurements of M-ENK dynamics in the brain with subsecond temporal resolution. Importantly, this work also demonstrates how voltammetric waveforms can be customized to enhance detection of specific target analytes, broadly speaking.

Enkephalinase inhibitors, potential therapeutics for the future treatment of diarrhea predominant functional gastrointestinal disorders.

The endogenous opioid system (EOS) is considered being a crucial element involved in the pathophysiology of irritable bowel syndrome (IBS) as it regulates gastrointestinal (GI) homeostasis through modulation of motility and water and ion secretion/absorption. Along with opioid receptors (ORs), the following components of EOS can be distinguished: 1. endogenous opioid peptides (EOPs), namely enkephalins, endorphins, endomorphins and dynorphins, and 2. peptidases, which regulate the metabolism (synthesis and degradation) of EOPs. Enkephalins, which are δ-opioid receptors agonists, induce significant effects in the GI tract as they act as potent pro-absorptive neurotransmitters. The action of enkephalins and other EOPs is limited, since EOPs are easily and rapidly inactivated by a natural metalloendopeptidase (enkephalinase/neprilysin) and aminopeptidase N. Studies show that the activity of EOPs can be enhanced by inhibition of these enzymes. In this review, we discuss the antidiarrheal and antinociceptive potential of enkephalinase inhibitors. Furthermore, our review is to answer the question whether enkephalinase inhibitors may be helpful in the future treatment of diarrhea predominant functional GI disorders.

Blockade of analgesic effects following systemic administration of N-methyl-kyotorphin, NMYR and arginine in mice deficient of preproenkephalin or proopiomelanocortin gene

Kyotorphin is a unique biologically active neuropeptide (l-tyrosine-l-arginine), which is reported to have opioid-like analgesic actions through a release of Met-enkephalin from the brain slices. N-methyl-l-tyrosine-l-arginine (NMYR), an enzymatically stable mimetic of kyotorphin, successfully caused potent analgesic effects in thermal and mechanical nociception tests in mice when it was given through systemic routes. NMYR analgesia was abolished in μ-opioid receptor-deficient (MOP-KO) mice, and by intracerebroventricular (i.c.v.) injection of naloxone and of N-methyl l-leucine-l-arginine (NMLR), a kyotorphin receptor antagonist. In the Ca2+-mobilization assay using CHO cells expressing Gαqi5 and hMOPr or hDOPr, however, the addition of kyotorphin neither activated MOPr-mechanisms, nor affected the concentration-dependent activation of DAMGO- or Met-Enkephalin-induced MOPr activation, and Met-enkephalin-induced DOPr activation. NMYR-analgesia was significantly attenuated in preproenkephalin (PENK)- or proopioimelanocortin (POMC)-KO mice. The systemic administration of arginine, which is reported to elevate the level of endogenous kyotorphin selectively in midbrain and medulla oblongata, pain-related brain regions, caused significant analgesia, and the analgesia was reversed by i.c.v. injection of NMLR or naloxone. In addition, PENK- and POMC-KO mice also attenuated the arginine-induced analgesia. All these findings suggest that NMYR and arginine activate brain kyotorphin receptor in direct and indirect manner, respectively and both compounds indirectly cause the opioid-like analgesia through the action of endogenous opioid peptides.

Targeting immune-driven opioid analgesia by sigma-1 receptors: Opening the door to novel perspectives for the analgesic use of sigma-1 antagonists

Immune cells have a known role in pronociception, since they release a myriad of inflammatory algogens which interact with neurons to facilitate pain signaling. However, these cells also produce endogenous opioid peptides with analgesic potential. The sigma-1 receptor is a ligand-operated chaperone that modulates neurotransmission by interacting with multiple protein partners, including the μ-opioid receptor. We recently found that sigma-1 antagonists are able to induce opioid analgesia by enhancing the action of endogenous opioid peptides of immune origin during inflammation. This opioid analgesia is seen only at the inflamed site, where immune cells naturally accumulate. In this article we review the difficulties of targeting the opioid system for selective pain relief, and discuss the dual role of immune cells in pain and analgesia. Our discussion creates perspectives for possible novel therapeutic uses of sigma-1 antagonists as agents able to maximize the analgesic potential of the immune system.

Allostery at opioid receptors: modulation with small molecule ligands.

This article is part of a themed section on Emerging Areas of Opioid Pharmacology. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.14/issuetoc.

Opioid receptor expression in human brain and peripheral tissues using absolute quantitative real-time RT-PCR

BackgroundThe actions of endogenous opioid peptides are mediated by 3 main classes of opioid receptors; mu (MOR), kappa (KOR), and delta (DOR). MethodsWe developed an absolute quantitative real-time reverse transcriptase PCR (AQ-rt-RT-PCR) assay to quantify MOR, DOR, and KOR mRNA in 22 human tissues. ResultsMOR mRNA was greatly enriched (12–20×106copies/μg) in the cerebellum, nucleus accumbens, and caudate nucleus; moderate (6×106copies/μg) in the dorsal root ganglion, spinal cord, and adrenal gland; low (2×104copies/μg) in the pancreas and small intestine; and absent in the lung, spleen, kidney, heart, skeletal muscle, liver, and thymus. High levels (>8.8×106copies/μg) of DOR mRNA were expressed in the brain and dorsal root ganglion; moderate (1.5×106copies/μg) in the adrenal gland and pancreas; low (2×104–6.5×105copies/μg in the cerebellum, spinal cord, small intestine, skeletal muscle, thymus, lung, and kidney); and very low (3.8×103copies/μg) in the heart. DOR mRNA was not detected in the spleen or liver. KOR mRNA was moderate (1×106copies/μg) in brain regions and dorsal root ganglion, but low (1.6–7×105copies/μg) in the cerebellum, temporal lobe and all other peripheral tissues. ConclusionsOur data demonstrate that the AQ-rt-RT-PCR is a highly reproducible and precise method to study the expression of opioid receptors in various tissues and under different disease conditions.

Expression and immunolocalization of the mu-opioid receptor in human sperm cells

The expression of the mu-opioid receptor in human sperm cells was investigated by immunocytochemistry and immunoblot analyses. Results demonstrated that the receptor is localized on the acrosomal region and the neck portion of the sperm head. These results suggest a possible role for the mu-opioid receptor in mediating the action of endogenous opioid peptides on sperm cells during the complex and poorly characterized cellular events that enable spermatozoa to fertilize oocytes.

Opioids, the Enteric Nervous System, and Postoperative Ileus

Postoperative ileus, which is characterized by quiescence of the intestinal musculature, reflects the operation of a neural program in the enteric nervous system (ENS) in which the synapses that form the propulsive motor circuits are “locked” into an inactive state. In the inactive state, a component of the inhibitory motor innervation of the musculature remains active, and as a consequence, responsiveness of the muscle to electrical signals from pacemaker cells called interstitial cells of Cajal is suppressed. Physiological ileus is the normal absence of motility that occurs for varying lengths of time in different digestive states. Physiological motor quiescence becomes pathological when the synapses in the neural microcircuits for the motor programs in the ENS program library are rendered inoperative for abnormally long periods. In this state of paralytic ileus, the basic circuits are “locked” in an inactive state, while unremitting activity of inhibitory motor neurons continuously suppresses autogenic muscular activity. Release and actions of endogenous opioid peptides within the ENS inactivate the synaptic microcircuits and might account for the accompanying paralytic ileus. This hypothesis is based in part on the known actions of opioid peptides and opiates on the cellular neurophysiology of enteric neurons. The prototype opiate is morphine, which acts to delay gastric emptying and intestinal transit, to suppress intestinal secretion of water and electrolytes, and to suppress transport of bile into the duodenum—all of which are related to suppression of neuronal excitability and synaptic transmission in the ENS.

Mu Opioid Receptor (OPRM-1) Polymorphisms among Patients with Sickle Cell Disease.

Opioid analgesics form an important component of the management of acute and chronic pain in patients with sickle cell disease (SCD). Chronic opioid use, sometimes associated with dependence and addiction in a subset of patients, may pose difficult management problems. This adds to the sometime antagonistic relationship that can develop between providers and patients with SCD. Pain management can become a sociological and psychological issue in the management of SCD. The Mu opioid receptor (OPRM-1) is the primary site of action of endogenous opioid peptides (enkephalins and endorphins) and of opioid analgesics, such as morphine, methadone, fentanyl, heroin, and related compounds. Rapid activation of the mu receptor results in a euphoric effect, conferring the reinforcing or rewarding effects of opioids and thus contributing to the development of addiction. It has been known that there is variation between individuals in sensitivity to opioids suggesting potential variation in the receptor protein and the gene. Some recent data have shown that polymorphisms in the OPRM-1 gene affect pain threshold and tolerance as well as opioid requirements for optimal pain control. In an effort to unravel the complex issues surrounding pain frequency, pain tolerance, opioid usage, and opioid addiction in this patient population, we conducted a study of our adult sickle cell patients for the frequency of the two common cSNPs and another SNP in the IVSII (G691C) of the OPRM-1 gene. DNA samples from randomly selected patients were used in this study. The OPRM-1 gene was PCR amplified and subjected to cycle sequencing on an ABI Prism automated sequencer. The results showed that of the 97 adult SCD patients screened for the C17T polymorphism (GenBank accession #AY292291 and 292290), 67 (69.7%) had the wild type (CC), 28 (29.1%) were heterozygous (CT), and 11 (11.4%) were homozygous (TT) for the SNP. This represents an allele frequency for the T of 0.26. Of the 39 sequenced samples for the A1189G, all (100%) showed the wild type (AA). These results closely resemble those reported by Bond et al (PNAS , 95;9608, 1998) for the frequency of the two cSNPs in the African-American population (allele frequency of 0.21 for the C17T and 0.016 for the A118G). Of the 16 samples screened for IVS II G691C polymorphism (GenBank accession #AY299483), 6 (37.5%) had the wild type (GG), 9 (56.3%) were heterozygous (GC), and 1 (6.2%) was homozygous (CC). Our results among SCD patients show a high frequency of C17T polymorphism in the OPRM-1 gene. Detailed studies in higher numbers of SCD patients and clinical correlations with pain frequency and threshold, opioid usage, opioid abuse and drug seeking behavior will be undertaken. It is expected that these studies will clarify the role of OPRM-1 polymorphisms as a genetic modifier associated with pain frequency, with tolerance as well as opioid use and abuse.

Effects of morphine administration and its withdrawal on rat brain aminopeptidase activities

The endogenous opioid neuropeptide system seems to be involved in the neural processes which underlie drug addiction. Several studies have reported that the administration of morphine induces changes in the levels and/or activity of endogenous opioid peptides (enkephalin, dynorphin) and their precursors in specific brain regions of the adult CNS. The aim of this work was to study the effects of chronic morphine exposure and its withdrawal on certain aminopeptidases capable of degrading opioid peptides in brain areas including the amygdala, hypothalamus, hippocampus, striatum and brain cortices. In animals treated with morphine, aminopeptidase N presented higher enzyme activity levels in the striatum, the hypothalamus and the amygdala compared to control animals, although statistically significant differences were observed only in the case of the striatum. In addition, the activity of soluble puromycin-sensitive aminopeptidase (PSA) was found to be higher in the frontal cortex of these rats. In contrast, rats experiencing withdrawal symptoms presented decreased levels of aminopeptidase activity in certain brain areas. Thus, the activity of aminopeptidase N in the hippocampus and soluble puromycin-sensitive aminopeptidase in the frontal cortex were found to be lower in rats experiencing naloxone precipitated withdrawal symptoms, compared to the corresponding controls. Finally, the activity of the three studied aminopeptidases in vitro was unaltered by incubation with morphine, suggesting that the observed effects are not due to a direct action of this opioid upon the aminopeptidases. The results of the present report indicate that aminopeptidases may play an important role in the processes of tolerance and withdrawal associated with morphine administration.

Blockade of mu-opioid receptors reveals the hyperalgesic effect of orphanin FQ/nociceptin in the rat hot plate test.

Orphanin FQ (OFQ, also known as nociceptin) has been proposed to oppose the antinociceptive effect of endogenous opioid peptides in the brain. We sought to determine whether, conversely, the endogenous opioid peptides counteract a pronociceptive action of OFQ. In testing this hypothesis, naloxone, a non-selective opioid receptor antagonist, was used to block the action of endogenous opioid peptides. We then examined whether OFQ would produce hyperalgesia in the absence of such an endogenous opioidergic tone. Neither naloxone (1 mg kg(-1); s.c.) nor OFQ (up to 30 nmol; i.c.v.) alone induced any significant change in mean hot plate latency. However, OFQ dose-dependently produced hyperalgesia in rats pretreated with naloxone, implying that OFQ can indeed produce hyperalgesia once an endogenous opioidergic tone is inhibited. In subsequent studies, we used subtype selective opioid receptor antagonists to determine which class of opioid receptor is involved in this response. The effect of naloxone was reproduced using the selective mu-opioid receptor antagonist CTOP (D-Phe-Cyc-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2), but not by administration of the delta-opioid receptor antagonist, naltrindole (NTI) or the kappa-opioid receptor antagonist nor-binaltorphimine (nor-BNI). These results suggest that endogenous opioid peptides acting at the mu-, but not kappa- or delta-opioid receptor may be counteracting the hyperalgesic effect of OFQ in rats.

Modulation of humoral immune responses in the rat by centrally applied Met–Enk and opioid receptor antagonists: functional interactions of brain OP1, OP2 and OP3 receptors

We have previously demonstrated that central application of leucine–enkephalin (Leu–Enk) elicits potentiation and suppression of humoral immune responses through OP1 (δ) and OP2 (κ) receptors, respectively. Interestingly, both effects were found to be additionally dependent on OP3 (μ) receptor function. In the present study, we have further investigated whether opioid receptor interactions underlie the immunomodulatory effects of endogenous opioids as well as exogenously applied methionine–enkephalin (Met–Enk). For that purpose, the plaque-forming cell (PFC) response was determined in rats injected intracerebroventricularly (i.c.v.) with opioid receptor-selective antagonists and Met–Enk. Application of the OP1 antagonist ICI 174864, but not naltrindole, resulted in suppression of the PFC response. In contrast, i.c.v. injection of the OP2 selective antagonist nor-binaltorphimine (nor-BNI) significantly potentiated the PFC response. Both effects, presumably mediated by endogenous opioid peptides, were antagonized by the OP3 receptor antagonist β-funaltrexamine (β-FNA) at a dose that was devoid of immunomodulatory activity. The immunopotentiation of the PFC response induced by Met–Enk was reversed by OP1 receptor antagonists, naltrindole and ICI 174864, but not by β-FNA or nor-BNI.On the basis of these and previous findings, it may be concluded that central OP3 receptors are permissive for the central immunomodulatory action of endogenous opioid peptides and Leu–Enk. In contrast, the central immunoenhancing effect of Met–Enk appears to be mediated through OP3-independent OP1 receptors.

Gender differences in opioid-mediated analgesia: animal and human studies.

Gender differences in opioid-mediated analgesia: animal and human studies.

Effects of naloxone infusion on nocturnal prolactin secretion and Fos/FRA expression in pregnant rats.

Dopamine (DA) secreted by tuberoinfundibular dopaminergic (TIDA) neurons in the hypothalamus is the major inhibitory factor controlling prolactin (PRL) secretion from the anterior pituitary. Endogenous opioid peptides (EOPs), mainly the neuropeptide beta-endorphin, have a stimulatory influence on PRL secretion. During the first half of pregnancy in rats, PRL secretion is characterized by two daily surges, the nocturnal surge and the diurnal surge. We tested the hypothesis that EOPs are critical stimulatory factors in regulating the nocturnal PRL surge, possibly via inhibition of TIDA neuronal activity. Naloxone (NAL), an opioid antagonist, was continuously infused (0.2 mg/10 microL/min i.v.) during the expected time of the nocturnal PRL surge in day 8 pregnant rats. Radioimmunoassay (RIA) was used to measure plasma PRL levels, and the immunocytochemical (ICC) staining of Fos/FRA was performed to detect changes in transcriptional activity of neurons in the hypothalamus. ICC of tyrosine hydroxylase (TH), the rate-limiting enzyme for DA synthesis, was performed to visualize TIDA neurons in the arcuate nucleus. The results showed that the nocturnal surge of PRL was markedly delayed and dampened in NAL-treated rats (p < 0.01). The numbers of both Fos/FRA and (Fos/FRA)/TH dual-staining neurons increased in the arcuate nucleus following NAL treatment (p < 0.05 for both comparisons). These data indicate that EOPs are critical stimulatory factors for the nocturnal PRL surge and that the actions of EOPs are partially mediated via decreasing TIDA neuronal activity.

Regulation of Mu Opioid Receptor mRNA Levels by Activation of Protein Kinase C in Human SH-SY5Y Neuroblastoma Cells

The mu opioid receptor (MuOR) is a member of the superfamily of G protein-coupled receptors that mediates the analgesic actions of endogenous opioid peptides and the narcotic alkaloid derivatives of morphine. Activation and translocation of protein kinase C (PKC) by N-methyl-D-aspartate receptor stimulation correlates with resistance to opioid drugs in experimental states of neuropathic pain, but the cellular mechanisms of resistance have not been identified. One possibility is that PKC activation regulates MuOR mRNA expression and thus the ability to generate functional receptors. Using a human neuroblastoma cell line, the authors tested the hypothesis that phorbol ester activation of PKC regulates MuOR mRNA levels. SH-SY5Y cells were maintained in a continuous monolayer culture and treated with phorbol esters or other agents before extraction of total cellular RNA. Slot-blot hybridization was used to measure the level of MuOR mRNA using 32P-labeled MuOR cDNA probes under high-stringency conditions. Autoradiograms were analyzed by scanning and densitometry. MuOR mRNA levels decreased in a dose- and time-dependent manner after tetradecanoyl phorbol acetate (TPA) was administered to activate PKC. The nadir, a level of approximately 50% of control, was at 2-8 h, followed by gradual recovery. The actions of TPA were blocked by pretreatment with the selective PKC inhibitor bisindolylmaleimide, but not by inhibition of protein synthesis with cycloheximide or anisomycin. The combination of TPA treatment and transcription inhibition with actinomycin D was associated with a transient increase in MuOR mRNA. Mu opioid receptor mRNA levels are regulated by activation of PKC in a neuronal model. Protein kinase C effects which decrease MuOR mRNA levels appear largely independent of new protein synthesis, and cytotoxicity does not account for the findings. Plasticity of MuOR gene expression may contribute to variations in clinical responses to opioid analgesics in clinical states such as neuropathic pain.

Mechanisms of actions of opioids and non-steroidal anti-inflammatory drugs.

Opioids and non-steroidal anti-inflammatory drugs (NSAIDs) are the commonest drugs used to treat pain. Opioids mimic the actions of endogenous opioid peptides by interacting with mu, delta or kappa opioid receptors. The opioid receptors are coupled to G1 proteins and the actions of the opioids are mainly inhibitory. They close N-type voltage-operated calcium channels and open calcium-dependent inwardly-rectifying potassium channels. This results in hyperpolarization and a reduction in neuronal excitability. They also decrease intracellular cAMP which modulates the release of nociceptive neurotransmitters (e.g. substance P). Inhibition of prostaglandin synthesis by cyclooxygenase is the principal mode of the analgesic and anti-inflammatory actions of NSAIDs. Cyclo-oxygenase is inhibited irreversibly by aspirin and reversibly by other NSAIDs. The widespread inhibition of cyclo-oxygenase is responsible for many of the adverse effects of these drugs. NSAIDs also reduce prostaglandin production within the CNS. This is the main action of paracetamol.