dermorphin and Pain

Strong opioid analgesics, including morphine, are the mainstays for treating moderate to severe acute pain and alleviating chronic cancer pain. However, opioid-related adverse effects, including nausea or vomiting, sedation, respiratory depression, constipation, pruritus (itch), analgesic tolerance, and addiction and abuse liability, are problematic. In addition, the use of opioids to relieve chronic noncancer pain is controversial due to the "opioid crisis" characterized by opioid misuse or abuse and escalating unintentional death rates due to respiratory depression. Hence, considerable research internationally has been aimed at the "Holy Grail" of the opioid analgesic field, namely the discovery of novel and safer opioid analgesics with improved opioid-related adverse effects. In this Perspective, medicinal chemistry strategies are addressed, where structurally diverse nonmorphinan-based opioid ligands derived from natural sources were deployed as lead molecules. The current state of play, clinical or experimental status, and novel opioid ligand discovery approaches are elaborated in the context of retaining analgesia with improved safety and reduced adverse effects, especially addiction liability.

Topics: Analgesics, Opioid; Animals; Biological Products; Cell Line, Tumor; Chemistry, Pharmaceutical; Drug Discovery; Humans; Ligands; Pain; Peptides; Receptors, Opioid, delta; Receptors, Opioid, kappa; Receptors, Opioid, mu

Opioids, such as morphine, are the most effective treatment for pain but their efficacy is diminished with the development of tolerance following repeated administration. Recently, we found that morphine activated ERK in opioid-tolerant but not in naïve rats, suggesting that morphine activation of μ-opioid receptors is altered following repeated morphine administration. Here, we have tested the hypothesis that μ-opioid receptor activation of ERK in the ventrolateral periaqueductal gray (vlPAG) is dependent on dynamin, a protein implicated in receptor endocytosis.. Rats were made tolerant to repeated microinjections of morphine into the vlPAG. The effects of dynamin on ERK activation and antinociception were assessed by microinjecting myristoylated dominant-negative dynamin peptide (Dyn-DN) or a scrambled control peptide into the vlPAG. Microinjection of a fluorescent dermorphin analogue (DERM-A594) into the vlPAG was used to monitor μ-opioid receptor internalization.. Morphine did not activate ERK and Dyn-DN administration had no effect on morphine-induced antinociception in saline-pretreated rats. In contrast, morphine-induced ERK activation in morphine-pretreated rats that was blocked by Dyn-DN administration. Dyn-DN also inhibited morphine antinociception. Finally, morphine reduced DERM-A594 internalization only in morphine-tolerant rats indicating that μ-opioid receptors were internalized and unavailable to bind DERM-A594.. Repeated morphine administration increased μ-opioid receptor activation of ERK signalling via a dynamin-dependent mechanism. These results demonstrate that the balance of agonist signalling to G-protein and dynamin-dependent pathways is altered, effectively changing the functional selectivity of the agonist-receptor complex.. 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.

Topics: Analgesics, Opioid; Animals; Drug Tolerance; Dynamins; Extracellular Signal-Regulated MAP Kinases; Hot Temperature; Male; Morphine; Opioid Peptides; Pain; Periaqueductal Gray; Rats, Sprague-Dawley; Receptors, Opioid, mu

Microinjection of opioids into the ventrolateral periaqueductal gray (vlPAG) produces antinociception in part by binding to mu-opioid receptors (MOPrs). Although both high and low efficacy agonists produce antinociception, low efficacy agonists such as morphine produce limited MOPr internalization suggesting that MOPr internalization and signaling leading to antinociception are independent. This hypothesis was tested in awake, behaving rats using DERM-A594, a fluorescently labeled dermorphin analog, and internalization blockers. Microinjection of DERM-A594 into the vlPAG produced both antinociception and internalization of DERM-A594. Administration of the irreversible opioid receptor antagonist beta-chlornaltrexamine (beta-CNA) prior to DERM-A594 microinjection reduced both the antinociceptive effect and the number of DERM-A594 labeled cells demonstrating that both effects are opioid receptor-mediated. Pretreatment with the internalization blockers dynamin dominant-negative inhibitory peptide (dynamin-DN) and concanavalinA (ConA) attenuated both DERM-A594 internalization and antinociception. Microinjection of dynamin-DN and ConA also decreased the antinociceptive potency of the unlabeled opioid agonist dermorphin when microinjected into the vlPAG as demonstrated by rightward shifts in the dose-response curves. In contrast, administration of dynamin-DN had no effect on the antinociceptive effect of microinjecting the GABA(A) receptor antagonist bicuculline into the vlPAG. The finding that dermorphin-induced antinociception is attenuated by blocking receptor internalization indicates that key parts of opioid receptor-mediated signaling depend on internalization.

Topics: Analgesics, Opioid; Animals; Bicuculline; Concanavalin A; Dynamins; Fluorescent Dyes; GABA-A Receptor Antagonists; Male; Microinjections; Naltrexone; Neurons; Opioid Peptides; Pain; Pain Measurement; Peptides; Periaqueductal Gray; Rats; Rats, Sprague-Dawley; Receptors, Opioid, mu

Syntheses are described of new dermorphin and [D-Ala2]deltorphin I analogues in which the phenylalanine, the tyrosine or the valine residues have been substituted by the corresponding N-alkylglycine residues. Structural investigations by CD measurements in different solvents and preliminary pharmacological experiments were carried out on the resulting peptide-peptoid hybrids. The contribution from aromatic side chain residues is prominent in the CD spectra of dermorphin analogues and the assignment of a prevailing secondary structure could be questionable. In the CD spectra of deltorphin analogues the aromatic contribution is lower and the dichroic curves indicate the predominance of random conformer populations. The disappearance of the aromatic contribution in the [Ntyr1,D-Ala2]-deltorphin spectrum could be explained in terms of high conformational freedom of the N-terminal residue. The kinetics of degradation of the synthetic peptoids digestion by rat and human plasma enzymes were compared with that of [Leu5]-enkephalin. The binding to opioid receptors was tested on crude membrane preparations from CHO cells stably transfected with the mu- and delta-opioid receptors. The biological potency of peptoids was compared with that of dermorphin in GPI preparations and with that of deltorphin I in MVD preparations. All the substitutions produced a dramatic decrease in the affinity of the peptide-peptoid hybrids for both the mu- and delta-opioid receptors. Nval5 and/or Nval6 containing hybrids behaved as mu-opioid receptor agonists and elicit a dose-dependent analgesia (tail-flick test) when injected i.c.v. in rats.

Topics: Analgesics, Opioid; Animals; CHO Cells; Cricetinae; Cricetulus; Guinea Pigs; Male; Mice; Muscle Contraction; Muscle, Smooth; N-substituted Glycines; Oligopeptides; Opioid Peptides; Pain; Protein Conformation; Rats; Receptors, Opioid, delta; Receptors, Opioid, mu

Although injury-induced afferent discharge declines significantly over time, experimental neuropathic pain persists unchanged for long periods. These observations suggest that processes that initiate experimental neuropathic pain may differ from those that maintain such pain. Here, the role of descending facilitation arising from developing plasticity in the rostral ventromedial medulla (RVM) in the initiation and maintenance of experimental neuropathic pain was explored. Tactile and thermal hypersensitivity were induced in rats by spinal nerve ligation (SNL). RVM lidocaine blocked SNL-induced tactile and thermal hypersensitivity on post-SNL days 6-12 but not on post-SNL day 3. Lesion of RVM cells expressing mu-opioid receptors with dermorphin-saporin did not prevent the onset of SNL-induced tactile and thermal hypersensitivity, but these signs reversed to baseline levels beginning on post-SNL day 4. Similarly, lesions of the dorsolateral funiculus (DLF) did not prevent the onset of SNL-induced tactile and thermal hypersensitivity, but these signs reversed to baseline levels beginning on post-SNL day 4. Lesions of the DLF also blocked the SNL-induced increase in spinal dynorphin content, which has been suggested to promote neuropathic pain. These data distinguish mechanisms that initiate the neuropathic state as independent of descending supraspinal influences and additional mechanism(s) that require supraspinal facilitation to maintain such pain. In addition, the data indicate that these time-dependent descending influences can underlie some of the SNL-induced plasticity at the spinal level. Such time-dependent descending influences driving associated spinal changes, such as the upregulation of dynorphin, are key elements in the maintenance, but not initiation, of neuropathic states.

Topics: Animals; Behavior, Animal; Denervation; Dynorphins; Immunotoxins; Kinetics; Lidocaine; Ligation; Male; Medulla Oblongata; Microinjections; N-Glycosyl Hydrolases; Oligopeptides; Opioid Peptides; Pain; Plant Proteins; Rats; Rats, Sprague-Dawley; Ribosome Inactivating Proteins, Type 1; Saporins; Spinal Cord; Spinal Nerves

The solid phase procedure, based on the Fmoc chemistry, was used to prepare some opioid deltorphin (H-Tyr-D-Ala-Phe-Asp-Val-Val-Gly-NH2, DEL C) and dermorphin (H-Tyr-D-Ala-Phe-Gly-Tyr-Pro-Ser-NH2, DER) analogues in which a D-glucopyranosyl moiety is beta-O-glycosidically linked to a Thr4 or Thr7 side chain. Their activities were determined in binding studies based on displacement of mu- and delta-receptor selective radiolabels from rat brain membrane synaptosomes, in guinea pig ileum and rabbit jejenum bioassays, and, in vivo, by a mouse tail-flick test after intracerebroventricular (icv) and subcutaneous (sc) administrations. The glyco analogues modified at position 4 displayed low opioid properties, while Thr7-glycosylated peptides retained high delta- or mu-selectivity and remarkable activity in vivo. In particular, as systemic antinociceptive agents, the latter glucoside-bearing compounds were more potent than the parent unglycosylated peptide counterparts, showing a high blood to brain rate of influx which may be due to the glucose transporter GLUT-1.

Topics: Amino Acid Sequence; Analgesics, Opioid; Animals; Binding, Competitive; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalins; Glycopeptides; Guinea Pigs; Ileum; In Vitro Techniques; Indicators and Reagents; Jejunum; Kinetics; Male; Mice; Muscle Contraction; Muscle, Smooth; Oligopeptides; Opioid Peptides; Pain; Rabbits; Rats; Rats, Sprague-Dawley; Receptors, Opioid; Structure-Activity Relationship; Synaptosomes

Topics: Administration, Intranasal; Amino Acid Sequence; Analgesics, Opioid; Animals; Electric Stimulation; Molecular Sequence Data; Oligopeptides; Oncorhynchus mykiss; Opioid Peptides; Pain; Pain Threshold

Three naturally occurring dermorphin-like peptides from the skin of the frog Phyllomedusa bicolor, the related carboxyl-terminal amides, and some substituted analogs were synthesized, their binding profiles to opioid receptors were determined, and their biological activities were studied in isolated organ preparations and intact animals. The opioid binding profile revealed a very high selectivity of these peptides for mu sites and suggested the existence of two receptor subtypes, of high and low affinity. The peptides tested acted as potent mu opioid agonists on isolated organ preparations. They were several times more active in inhibiting electrically evoked contractions in guinea pig ileum than in mouse vas deferens. When injected into the lateral brain ventricle or peritoneum of rats, the high-affinity-site-preferring ligand, [Lys7-NH2]dermorphin, behaved as a potent analgesic agent. By contrast, the low-affinity-site-preferring ligand, [Trp4,Asn7-NH2]dermorphin, produced a weak antinociception but an intense catalepsy.

Topics: Amino Acid Sequence; Analgesics, Opioid; Animals; Anura; Binding, Competitive; Brain; Catalepsy; Cerebral Ventricles; Electric Stimulation; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalins; Guinea Pigs; Injections, Intraventricular; Kinetics; Molecular Sequence Data; Myenteric Plexus; Naloxone; Nociceptors; Oligopeptides; Opioid Peptides; Pain; Rats; Receptors, Opioid; Receptors, Opioid, delta; Receptors, Opioid, kappa; Receptors, Opioid, mu; Skin; Skin Physiological Phenomena; Structure-Activity Relationship

Dermorphin, Tyr-D-Ala-Phe-Gly-Tyr-Pro-Ser-NH2 is an extraordinarily potent and highly mu-selective opioid heptapeptide isolated from amphibian skin. It is unique among peptides synthesized by animal cells in having an amino acid residue in the D-configuration. At least two different preprodermorphin cDNAs were cloned from skin of Phylomedusa sauvagei; their predicted amino acid sequences contained four to five homologous repeats of 35 amino acids, each repeat including one copy of the dermorphin progenitor sequence. Tyr-Ala-Phe-Gly-Tyr-Pro-Ser-Gly, flanked by Lys-Arg at the amino end and by Glu-Ala-Lys-Lys at the carboxyl end [Science (Wash. D. C.) 238:200-202 (1987)]. The D-Ala in position 2 in dermorphin is encoded by a usual Ala codon in the precursor sequence. Of the two prodermorphin molecules, one has a dermorphin copy replaced with a distinct heptapeptide same processing signals. Assuming the same pathway as for the release of dermorphin, processing of this precursor may yield, beside dermorphin, a copy of a new peptide, Tyr-D-Met-Phe-His-Leu-Met-Asp-NH2. We have synthetized this peptide together with its (L-Met2)-counterpart and evaluated their respective opioid receptor selectivity in the mouse vas deferens and guinea pig ileum assays and in rat brain membrane binding assays. Overall, the data collected demonstrate that the putative prodermorphin product Tyr-D-Met-Phe-His-Leu-Met-Asp-NH2 named dermenkephalin, behaves as a potent delta opioid agonist exhibiting high affinity and high selectivity for the delta opioid receptor. Prodermorphin, thus, offers a surprising example of an opioid biosynthetic precursor that might simultaneously generate highly potent and fully selective agonists for the mu- (morphine) and the delta (enkephalin) opioid receptors, respectively. In addition, because dermenkephalin has no structural features in common with the sequence of all the hitherto known opioid peptides, it should be a useful tool for identifying conformational determinants for high affinity and selective binding of opioids to the delta receptor.

Topics: Amino Acid Sequence; Analgesia; Animals; Binding, Competitive; Brain; Cell Membrane; Cerebral Ventricles; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalin, D-Penicillamine (2,5)-; Enkephalins; Injections, Intraventricular; Kinetics; Male; Mice; Oligopeptides; Opioid Peptides; Pain; Rats; Receptors, Opioid; Receptors, Opioid, delta

The intrathecal administration of dermorphin, an endogenous heptopeptide first discovered in amphibia, produces dose-dependent selective inhibitions of C fibre-evoked responses in rat dorsal horn nociceptive neurones (ED50 0.11 micrograms). Naloxone (10 micrograms) but not ICI 174,864 (125 micrograms) antagonised the effects of the peptide. A beta-fibre-evoked activity was relatively unaffected. Thus dermorphin can profoundly inhibit nociceptive afferent input in the spinal cord, and in this preparation is more potent (approximately 40X) than morphine.

Topics: Analgesics, Opioid; Animals; Evoked Potentials; Kinetics; Nerve Fibers; Neurons; Oligopeptides; Opioid Peptides; Pain; Physical Stimulation; Rats; Rats, Inbred Strains; Receptors, Opioid; Receptors, Opioid, mu; Reference Values; Spinal Cord

Studies on the characteristics of spinally administered dermorphin, a novel heptapeptide isolated from the skin of South American Phyllomedusa frogs, indicated that this agent is 3-5000X more active spinal morphine on the hot plate, tail flick and writhing tests. This agent displays naloxone reversibility and cross tolerance to spinal morphine, and possesses all of the characteristics of a mu opiate receptor agonist.

Topics: Analgesics, Opioid; Animals; Drug Tolerance; Injections, Spinal; Male; Morphine; Naloxone; Oligopeptides; Opioid Peptides; Pain; Rats; Rats, Inbred Strains; Receptors, Opioid; Spinal Cord

Bilateral electrolytic lesions of caudate nucleus were observed to potentiate the cataleptic response, and to prevent the muscular rigidity produced by intracerebroventricular injection or infusion of dermorphin. These results suggest that caudate nucleus plays an inhibitory role in controlling the cataleptic response and its integrity is essential for full development of dermorphin-induced rigidity.

Topics: Animals; Catalepsy; Caudate Nucleus; Corpus Striatum; Injections, Intraventricular; Male; Muscle Rigidity; Oligopeptides; Opioid Peptides; Pain; Rats

Dermorphin is the representative of a new class of potent opioid peptides occurring in amphibian skin and possesses the unique feature of having a D-Ala residue incorporated in the peptide molecule. The effect of dermorphin on the spontaneous and evoked neuronal activity by a nociceptive stimulus was studied in the nucleus lateralis anterior and ventrobasal complex of the rat thalamus. The high firing frequency induced by nociceptive stimuli was blocked when dermorphin was injected intraperitoneally at the dose of 1.5 mg/kg. The action starts about 10 min after injection and lasts on average for 120 min. Naloxone, a specific opioid antagonist, injected i.p. at a dose of 1 mg/kg antagonized the effect of dermorphin. The dermorphin time-course is about twice that of morphine (1.5 mg/kg i.p.) under the same experimental conditions.

Topics: Action Potentials; Analgesics; Animals; Male; Naloxone; Oligopeptides; Opioid Peptides; Pain; Rats; Rats, Inbred Strains; Thalamic Nuclei

D-Arg containing dipeptides, H-Tyr-D-Arg-OMe and H-Tyr (Et)-D-Arg-OMe, and D-Arg2 substituted N-terminal tetrapeptides of dermorphin, H-Tyr-D-Arg-Phe-Gly-OEt and H-Tyr (Et)-D-Arg-Phe-Gly-OEt administered intracerebroventricularly exhibited dose-dependent antinociceptive activities in mice as measured by the tail pressure and phenylbenzoquinone writhing tests. The effects of these peptides used were significantly antagonized by the pretreatment with naloxone, indicating that these effects must be produced through opioid receptors. Furthermore, it is of conspicuous interest that the effects of tetrapeptides revealed in infinitestimal order (ED50 = 12.5 and 355.0 pmole in the tail pressure test and 3.1 and 53.0 pmole in the phenylbenzoquinone writhing test, respectively) and was much more potent and prolonged than those of morphine, not to mention dipeptides used. However, judging from the difference of peak times and the degree of the antagonism by naloxone, it was suggested that dipeptides and tetrapeptides used might act on different sites of action in the central nervous system.

Topics: Analgesia; Animals; Benzoquinones; Dipeptides; Dose-Response Relationship, Drug; Male; Mice; Morphine; Naloxone; Narcotics; Oligopeptides; Opioid Peptides; Pain; Quinones; Time Factors

Dermorphin (I) and [D-Ala2, Phe3, Gly4, Tyr5, Pro6]-beta c-EP (II) have been synthesized by the solid-phase method (beta c-EP, camel beta-endorphin). Positions 1 through 7 of II correspond to the sequence of I. Relative potencies of synthetic peptides in the mouse tail-flick test for analgesia by the intracerebroventricular route were: human beta-endorphin, 100; camel beta-endorphin, 164; I, 450; II, 440. The dermorphin was about 670 times more potent than morphine in the assay. Peptide II represents a rare instance where the enkephalin moiety of beta-endorphin has been altered to produce a more potent analgesic.

Topics: Amino Acid Sequence; Animals; Biological Assay; Indicators and Reagents; Mice; Narcotics; Oligopeptides; Opioid Peptides; Pain; Structure-Activity Relationship