enkephalin--ala(2)-mephe(4)-gly(5)- and Sciatica

Opiate analgesia in the spinal cord is impaired during neuropathic pain. We hypothesized that this is caused by a decrease in μ-opioid receptor inhibition of neurotransmitter release from primary afferents. To investigate this possibility, we measured substance P release in the spinal dorsal horn as neurokinin 1 receptor (NK1R) internalization in rats with chronic constriction injury (CCI) of the sciatic nerve. Noxious stimulation of the paw with CCI produced inconsistent NK1R internalization, suggesting that transmission of nociceptive signals by the injured nerve was variably impaired after CCI. This idea was supported by the fact that CCI produced only small changes in the ability of exogenous substance P to induce NK1R internalization or in the release of substance P evoked centrally from site of nerve injury. In subsequent experiments, NK1R internalization was induced in spinal cord slices by stimulating the dorsal root ipsilateral to CCI. We observed a complete loss of the inhibition of substance P release by the μ-opioid receptor agonist [D-Ala(2), NMe-Phe(4), Gly-ol(5)]-enkephalin (DAMGO) in CCI rats but not in sham-operated rats. In contrast, DAMGO still inhibited substance P release after inflammation of the hind paw with complete Freund's adjuvant and in naïve rats. This loss of inhibition was not due to μ-opioid receptor downregulation in primary afferents, because their colocalization with substance P was unchanged, both in dorsal root ganglion neurons and primary afferent fibers in the dorsal horn. In conclusion, nerve injury eliminates the inhibition of substance P release by μ-opioid receptors, probably by hindering their signaling mechanisms.

Topics: Analgesics, Opioid; Animals; Disease Models, Animal; Dose-Response Relationship, Drug; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Ganglia, Spinal; Hyperalgesia; Inflammation; Male; Neurons; Pain; Rats; Rats, Sprague-Dawley; Receptors, Neurokinin-1; Receptors, Opioid, mu; Sciatic Nerve; Sciatica; Spinal Cord; Substance P

Neuropathic pain is a debilitating condition that is often difficult to treat using conventional pharmacological interventions and the exact mechanisms involved in the establishment and maintenance of this type of chronic pain have yet to be fully elucidated. The present studies examined the effect of chronic nerve injury on μ-opioid receptors and receptor-mediated G-protein activity within the supraspinal brain regions involved in pain processing of mice. Chronic constriction injury (CCI) reduced paw withdrawal latency, which was maximal at 10 days post-injury. [d-Ala2,(N-Me)Phe4,Gly5-OH] enkephalin (DAMGO)-stimulated [(35)S]GTPγS binding was then conducted at this time point in membranes prepared from the rostral ACC (rACC), thalamus and periaqueductal grey (PAG) of CCI and sham-operated mice. Results showed reduced DAMGO-stimulated [(35)S]GTPγS binding in the thalamus and PAG of CCI mice, with no change in the rACC. In thalamus, this reduction was due to decreased maximal stimulation by DAMGO, with no difference in EC(50) values. In PAG, however, DAMGO E(max) values did not significantly differ between groups, possibly due to the small magnitude of the main effect. [(3)H]Naloxone binding in membranes of the thalamus showed no significant differences in B(max) values between CCI and sham-operated mice, indicating that the difference in G-protein activation did not result from differences in μ-opioid receptor levels. These results suggest that CCI induced a region-specific adaptation of μ-opioid receptor-mediated G-protein activity, with apparent desensitization of the μ-opioid receptor in the thalamus and PAG and could have implications for treatment of neuropathic pain.

Topics: Analgesics, Opioid; Animals; Constriction; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Interactions; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; GTP-Binding Proteins; Guanosine 5'-O-(3-Thiotriphosphate); Hyperalgesia; Male; Mice; Naloxone; Narcotic Antagonists; Pain Threshold; Protein Binding; Receptors, Opioid, mu; Sciatica; Sulfur Isotopes; Thalamus; Time Factors; Tritium

Recent clinical studies have demonstrated that when opioids are used to control pain, psychological dependence is not a major problem. In this study, we further investigated the mechanisms that underlie the suppression of opioid reward under neuropathic pain in rodents. Sciatic nerve ligation suppressed a place preference induced by the selective mu-opioid receptor agonist [d-Ala(2), N-MePhe(4), Gly-ol(5)] enkephalin (DAMGO) and reduced both the increase in the level of extracellular dopamine by s.c. morphine in the nucleus accumbens and guanosine-5'-o-(3-[(35)S]thio) triphosphate ([(35)S]GTPgammaS) binding to membranes of the ventral tegmental area (VTA) induced by DAMGO. These effects were eliminated in mice that lacked the beta-endorphin gene. Furthermore, intra-VTA injection of a specific antibody to the endogenous mu-opioid peptide beta-endorphin reversed the suppression of the DAMGO-induced rewarding effect by sciatic nerve ligation in rats. These results provide molecular evidence that nerve injury results in the continuous release of endogenous beta-endorphin to cause the dysfunction of mu-opioid receptors in the VTA. This phenomenon could explain the mechanism that underlies the suppression of opioid reward under a neuropathic pain-like state.

Topics: Analysis of Variance; Animals; Behavior, Animal; beta-Endorphin; Conditioning, Operant; Disease Models, Animal; Dose-Response Relationship, Drug; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Female; Guanosine 5'-O-(3-Thiotriphosphate); Male; Mice; Mice, Knockout; Morphine; Narcotics; Pain Measurement; Protein Binding; Reaction Time; Reward; Sciatica; Time Factors; Tyrosine 3-Monooxygenase

Phosphorylation of specific sites in the second intracellular loop and in the C-terminal domain have previously been suggested to cause desensitization and internalization of the mu-opioid receptor (MOP-R). To assess sites of MOP-R phosphorylation in vivo, affinity-purified, phosphoselective antibodies were raised against either phosphothreonine-180 in the second intracellular loop (MOR-P1) or the C-terminal domain of MOP-R containing phosphothreonine-370 and phosphoserine-375 (MOR-P2). We found that MOR-P2-immunoreactivity (IR) was significantly increased within the striatum of wild-type C57BL/6 mice after injection of the agonist fentanyl. Pretreatment with the antagonist naloxone blocked the fentanyl-induced increase. Furthermore, mutant mice lacking MOP-R showed only non-specific nuclear MOR-P2-IR before or after fentanyl treatment, confirming the specificity of the MOR-P2 antibodies. To assess whether MOP-R phosphorylation occurs following endogenous opioid release, we induced chronic neuropathic pain by partial sciatic nerve ligation (pSNL), which caused a significant increase in MOR-P2-IR in the striatum. pSNL also induced signs of mu opioid receptor tolerance demonstrated by a rightward shift in the morphine dose response in the tail withdrawal assay and by a reduction in morphine conditioned place preference (CPP). Mutant mice selectively lacking all forms of the beta-endorphin peptides derived from the proopiomelanocortin (Pomc) gene did not show increased MOR-P2-IR, decreased morphine antinociception, or reduced morphine CPP following pSNL. In contrast gene deletion of either proenkephalin or prodynorphin opioids did not block the effects of pSNL. These results suggest that neuropathic pain caused by pSNL in wild-type mice activates the release of the endogenous opioid beta-endorphin, which subsequently induces MOP-R phosphorylation and opiate tolerance.

Topics: Analgesics, Opioid; Analysis of Variance; Animals; Behavior, Animal; beta-Endorphin; Cell Line, Transformed; Conditioning, Operant; Corpus Striatum; Drug Interactions; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Green Fluorescent Proteins; Humans; Hyperalgesia; Mice; Mice, Knockout; Mutagenesis; Naloxone; Narcotic Antagonists; Phosphorylation; Phosphothreonine; Receptors, Opioid, mu; Sciatica; Transfection

Clinically, it has been reported that chronic pain induces depression, anxiety, and reduced quality of life. The endogenous opioid system has been implicated in nociception, anxiety, and stress. The present study was undertaken to investigate whether chronic pain could induce anxiogenic effects and changes in the opioidergic function in the amygdala in mice. We found that either injection of complete Freund's adjuvant (CFA) or neuropathic pain induced by sciatic nerve ligation produced a significant anxiogenic effect at 4 weeks after the injection or surgery. Under these conditions, the selective mu-opioid receptor agonist [D-Ala2,N-MePhe4,Gly5-ol]-enkephalin (DAMGO)- and the selective delta-opioid receptor agonist (+)-4-[(alphaR)-alpha-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide (SNC80)-stimulated [35S]GTPgammaS binding in membranes of the amygdala was significantly suppressed by CFA injection or nerve ligation. CFA injection was associated with a significant increase in the kappa-opioid receptor agonist 2-(3,4-dichlorophenyl)-N-methyl-N-[(1S)-1-phenyl-2-(1-pyrrolidinyl)ethyl]acetamide hydrochloride (ICI199,441)-stimulated [35S]GTPgammaS binding in membranes of the amygdala. The intracerebroventricular administration and microinjection of a selective mu-opioid receptor antagonist, a selective delta-opioid receptor antagonist, and the endogenous kappa-opioid receptor ligand dynorphin A caused a significant anxiogenic effect in mice. We also found that thermal hyperalgesia induced by sciatic nerve ligation was reversed at 8 weeks after surgery. In the light-dark test, the time spent in the lit compartment was not changed at 8 weeks after surgery. Collectively, the present data constitute the first evidence that chronic pain has an anxiogenic effect in mice. This phenomenon may be associated with changes in opioidergic function in the amygdala.

Topics: Amygdala; Analgesics, Opioid; Analysis of Variance; Animals; Anxiety; Behavior, Animal; Benzamides; Chronic Disease; Diazepam; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Interactions; Dynorphins; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Freund's Adjuvant; Guanosine 5'-O-(3-Thiotriphosphate); Injections, Intraventricular; Male; Maze Learning; Mice; Mice, Inbred C57BL; Naltrexone; Narcotic Antagonists; Narcotics; Pain; Pain Measurement; Piperazines; Protein Binding; Pyrrolidines; Rats; Rats, Sprague-Dawley; Reaction Time; Sciatica; Somatostatin; Sulfur Isotopes; Time Factors; Tranquilizing Agents

The role of the adenosine A1 receptor in nociception was assessed using mice lacking the A1 receptor (A1R-/-) and in rats. Under normal conditions, the A1R-/- mice exhibited moderate heat hyperalgesia in comparison to the wild-type mice (A1R+/+). The mechanical and cold sensitivity were unchanged. The antinociceptive effect of morphine given intrathecally (i.t.), but not systemically, was reduced in A1R-/- mice and this reduction in the spinal effect of morphine was not associated with a decrease in binding of the mu-opioid ligand DAMGO in the spinal cord. A1R-/- mice also exhibited hypersensitivity to heat, but not mechanical stimuli, after localized inflammation induced by carrageenan. In mice with photochemically induced partial sciatic nerve injury, the neuropathic pain-like behavioral response to heat or cold stimulation were significantly increased in the A1R-/-mice. Peripheral nerve injury did not change the level of adenosine A1 receptor in the dorsal spinal cord in rats and i.t. administration of R-PIA effectively alleviated pain-like behaviors after partial nerve injury in rats and in C57/BL/6 mice. Taken together, these data suggest that the adenosine A1 receptor plays a physiological role in inhibiting nociceptive input at the spinal level in mice. The C-fiber input mediating noxious heat is inhibited more than other inputs. A1 receptors also contribute to the antinociceptive effect of spinal morphine. Selective A1 receptor agonists may be tested clinically as analgesics, particularly under conditions of neuropathic pain.

Topics: Adenosine A1 Receptor Agonists; Adenosine A1 Receptor Antagonists; Analgesics, Opioid; Analysis of Variance; Animals; Behavior, Animal; Dose-Response Relationship, Drug; Drug Administration Routes; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Female; Functional Laterality; Hyperalgesia; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Morphine; Nociceptors; Pain Measurement; Photochemistry; Protein Binding; Radioligand Assay; Rats; Reaction Time; Receptor, Adenosine A1; Sciatica; Statistics, Nonparametric; Time Factors; Xanthines

Co-localization of opioid and melanocortin receptor expression, especially at the spinal cord level in the dorsal horn and in the gray matter surrounding the central canal led to the suggestion that melanocortins might play a role in nociceptive processes. In the present studies, we aimed to determine the effects of melanocortins, administered intrathecally, on allodynia, and to ascertain whether there is an interaction between opioid and melanocortin systems at the spinal cord level. Neuropathic pain was induced by chronic constriction injury (CCI) of the right sciatic nerve in rats. Tactile allodynia was assessed using von Frey filaments, while thermal hyperlagesia was evaluated in cold water allodynia test. In the present experiments, melanocortin receptor antagonist, SHU9119 was much more potent than mu-opioid receptor agonist, morphine after their intrathecal (i.th.) administration in neuropathic rats. SHU9119 alleviated allodynia in a comparable manner to DAMGO, a selective and potent mu-opioid receptor agonist. Administration of melanocortin receptor agonist, melanotan-II (MTII) increased the sensitivity to tactile and cold stimulation. Moreover, we demonstrated that the selective blockade of mu-opioid receptor by cyprodime (CP) enhanced antiallodynic effect of SHU9119 as well as pronociceptive action of MTII, whereas the combined administration of mu receptor agonist (DAMGO) and SHU9119 significantly reduced the analgesic effect of those ligands. DAMGO also reversed the proallodynic effect of melanocortin receptor agonist, MTII. In conclusion, it seems that the endogenous opioidergic system acts as a functional antagonist of melanocortinergic system, and mu-opioid receptor activity appears to be involved in the modulation of melanocortin system function.

Topics: alpha-MSH; Analgesics, Opioid; Animals; Chronic Disease; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Injections, Spinal; Male; Melanocyte-Stimulating Hormones; Morphinans; Morphine; Nociceptors; Oligopeptides; Rats; Rats, Wistar; Receptor, Melanocortin, Type 4; Receptors, Corticotropin; Receptors, Opioid, mu; Sciatica; Spinal Cord