Naloxone-reversible Antinociception Research Articles

Binding preference at the μ-opioid receptor underlies distinct pharmacology of cyclopropyl versus valeryl analogs of fentanyl

Illicitly manufactured fentanyl is driving the current opioid crisis, and various fentanyl analogs are appearing in recreational drug markets worldwide. To assess the potential health risks posed by fentanyl analogs, it is necessary to understand structure-activity relationships for these compounds. Here we compared the pharmacology of two structurally related fentanyl analogs implicated in opioid overdose: cyclopropylfentanyl and valerylfentanyl. Cyclopropylfentanyl has a three-carbon ring attached to the carbonyl group on the fentanyl scaffold, whereas valerylfentanyl has a four-carbon chain at the same position. In vitro assays examining μ-opioid receptor (MOR) coupling to G proteins in CHO cells showed that cyclopropylfentanyl is a full agonist (EC50 = 8.6 nM, %Emax = 113%), with potency and efficacy similar to fentanyl (EC50 = 10.3 nM, %Emax = 113%). By contrast, valerylfentanyl is a partial agonist at MOR (EC50 = 179.8 nM, %Emax = 60%). Similar results were found in assays assessing MOR-mediated β-arrestin recruitment in HEK cells. In vivo studies in male CD-1 mice demonstrated that both fentanyl analogs induce naloxone-reversible antinociception and respiratory suppression, but cyclopropylfentanyl is 100-times more potent as an antinociceptive agent (ED50 = 0.04 mg/kg, s. c.) than valerylfentanyl (ED50 = 4.0 mg/kg, s. c.). Molecular simulation results revealed that the alkyl chain of valerylfentanyl cannot be well accommodated by the active state of MOR and may transition the receptor toward an inactive state, converting the fentanyl scaffold to a partial agonist. Taken together, our results suggest that cyclopropylfentanyl presents much greater risk of adverse effects when compared to valerylfentanyl. Moreover, the summed findings may provide clues to the design of therapeutic opioids with reduced adverse side effects.

Demethylating Drugs as Novel Analgesics for Cancer Pain

In this study, we evaluated the analgesic potential of demethylating drugs on oral cancer pain. Although demethylating drugs could affect expression of many genes, we focused on the mu-opioid receptor (OPRM1) gene pathway, because of its role in pain processing. We determined the antinociceptive effect of OPRM1 re-expression in a mouse oral cancer model. Using a mouse oral cancer model, we determined whether demethylating drugs produced antinociception through re-expression of OPRM1. We then re-expressed OPRM1 with adenoviral transduction and determined if, and by what mechanism, OPRM1 re-expression produced antinociception. To determine the clinical significance of OPRM1 on cancer pain, we quantified OPRM1 methylation in painful cancer tissues and nonpainful contralateral normal tissues of patients with oral cancer, and nonpainful dysplastic tissues of patients with oral dysplasia. We demonstrated that OPRM1 was methylated in cancer tissue, but not normal tissue, of patients with oral cancer, and not in dysplastic tissues from patients with oral dysplasia. Treatment with demethylating drugs resulted in mechanical and thermal antinociception in the mouse cancer model. This behavioral change correlated with OPRM1 re-expression in the cancer and associated neurons. Similarly, adenoviral-mediated OPRM1 re-expression on cancer cells resulted in naloxone-reversible antinociception. OPRM1 re-expression on oral cancer cells in vitro increased β-endorphin secretion from the cancer, and decreased activation of neurons that were treated with cancer supernatant. Our study establishes the regulatory role of methylation in cancer pain. OPRM1 re-expression in cancer cells produces antinociception through cancer-mediated endogenous opioid secretion. Demethylating drugs have an analgesic effect that involves OPRM1.

Acetaminophen (Paracetamol)

Therapeutic Reviews aim to provide essential independent information for health professionals about drugs used in palliative and hospice care. Additional content is available on www.palliativedrugs.com. Country-specific books (Hospice and Palliative Care Formulary USA, and Palliative Care Formulary, British and Canadian editions) are also available and can be ordered from www.palliativedrugs.com. The series editors welcome feedback on the articles (hq@palliativedrugs.com).

Effects of a combination of ketanserin and propranolol on inflammatory hyperalgesia in rats

Pain management is still challenging in clinic as current analgesics either are not very effective or produce serious adverse effects. This study aimed to examine if old drugs could display the new use and to develop a novel therapy for inflammatory pain. Injection of carrageenan in hindpaw evoked hyperalgesia detected by noxious heat stimulation. Intraplantar (i.pl.) injection of the 5-HT1A receptor antagonist WAY-100635 increased paw withdrawal latency (PWL) above normal level (hypoalgesia) during the late phase of carrageenan-evoked inflammation. The hypoalgesia was completely abolished by systemic injection of naloxone chloride and naloxone methiodide. Moreover, i.pl. injection of a combination of WAY-100635 and ketanserin, a 5-HT2A receptor antagonist, at their minimal doses attenuated hyperalgesia in the late phase of carrageenan-evoked inflammation. Subcutaneous (s.c.) injection of both ketanserin and propranolol dose-dependently inhibited carrageenan-evoked hyperalgesia. The treatment with a combination of ketanserin and propranolol by s.c. injection abolished carrageenan-evoked hyperalgesia at the doses, at which the drugs failed to alter the hypersensitivity when they were given alone. Furthermore, the combination of ketanserin and propranolol was also effective in relieving arthritic hyperalgesia and muscle pain at a minimal dose. The present study suggests that the activation of 5-HT1A receptors suppressed naloxone-reversible antinociception contributing to the maintenance of inflammatory pain, and that the concomitant blockade of 5-HT1A and 5-HT2A receptors in the periphery produced synergistic effects on inflammatory hyperalgesia. It is proposed that the combination of ketanserin and propranolol injected s.c. could be a promising therapy for relieving inflammatory pain with minimal side effects.

Synthesis, Conformation, and Biological Characterization of a Sugar Derivative of Morphine that is a Potent, Long-Lasting, and Nontolerant Antinociceptive

A synthetic mannoside derivative, namely, 6-morphinyl-alpha-D-mannopyranoside, shows a naloxone-reversible antinociception that is 100-fold more potent and twice as long lasting compared to morphine when administered intraperitoneally to rats in paw pressure and tail flick tests. The compound does not produce tolerance and binds to rat mu opioid receptors with twice the affinity of morphine. NMR studies suggest that differences of activity between the derivative and its parent compound M6G might be related to their differing molecular dynamic behavior.

YFa, a chimeric opioid peptide, induces kappa‐specific antinociception with no tolerance development during 6 days of chronic treatment

Our previous study showed that YGGFMKKKFMRFamide (YFa), a chimeric peptide of Met-enkephalin, and Phe-Met-Arg-Phe-NH2 induced naloxone-reversible antinociception and attenuated the development of tolerance to morphine analgesia. In continuation, the present study investigated which specific opioid receptors-mu, delta or kappa-mediate the observed YFa antinociception pharmacologically using specific antagonists and whether chronic administration of YFa at 26.01 micromol/kg per day induces tolerance and its effect on the expression of mu and kappa opioid receptors from day 4 to day 6, with endomorphine-1 (EM-1) and saline taken as positive and negative controls, respectively. Quantitative differential expression analysis was carried out by real-time reverse-transcriptase polymerase chain reaction, and the corresponding changes in protein levels were assessed by Western blot. A pharmacological investigation revealed that nor-binaltorphimine, a specific kappa opioid receptor-1 (KOR1) antagonist, completely antagonized the antinociception induced by 39.01 micromol/kg of YFa. Importantly, its chronic intraperitoneal administration did not result in significant tolerance over 6 days, whereas EM-1 induced significant tolerance after day 4. Differential expression analysis revealed that EM-1 caused up-regulation of mu opioid receptor-1 on day 4, followed by down-regulation on later days. Interestingly, YFa treatment caused a decrease on day 4, followed by an increase in the expression of KOR1 from day 5 onward. In conclusion, YFa induces kappa-specific antinociception, with no development of tolerance during 6 days of chronic treatment, which further articulates new directions for improved designing of peptide-based analgesics that may be devoid of adverse effects like tolerance.

Antinociception by Spinal and Systemic Oxycodone: Why Does the Route Make a Difference?

The pharmacology of oxycodone is poorly understood despite its growing clinical use. The discrepancy between its good clinical effectiveness after systemic administration and the loss of potency after spinal administration led the authors to study the pharmacodynamic effects of oxycodone and its metabolites using in vivo and in vitro models in rats. Male Sprague-Dawley rats were used in hot-plate, tail-flick, and paw-pressure tests to study the antinociceptive properties of morphine, oxycodone, and its metabolites oxymorphone and noroxycodone. Mu-opioid receptor agonist-stimulated GTPgamma[S] autoradiography was used to study G-protein activation induced by morphine, oxycodone, and oxymorphone in the rat brain and spinal cord. Spontaneous locomotor activity was measured to assess possible sedation or motor dysfunction. Naloxone and the selective kappa-opioid receptor antagonist nor-binaltorphimine were used to study the opioid receptor selectivity of the drugs. Oxycodone showed lower efficacy and potency to stimulate GTPgamma[S] binding in the spinal cord and periaqueductal gray compared with morphine and oxymorphone. This could relate to the fact that oxycodone produced only weak naloxone-reversible antinociception after intrathecal administration. It also suggests that the metabolites may have a role in oxycodone-induced analgesia in rats. Intrathecal oxymorphone produced strong long-lasting antinociception, whereas noroxycodone produced antinociception with very high doses only. Subcutaneous administration of oxycodone and oxymorphone produced thermal and mechanical antinociception that was reversed by naloxone but not by nor-binaltorphimine. Oxymorphone was more potent than oxycodone, particularly in the hot-plate and paw-pressure tests. The low intrathecal potency of oxycodone in rats seems be related to its low efficacy and potency to stimulate mu-opioid receptor activation in the spinal cord.

Endogenous opioid-mediated antinociception in cholestatic mice is peripherally, not centrally, mediated

Cholestasis is associated with naloxone reversible antinociception and opiate receptor antagonists are used clinically to treat pruritus. Pain and pruritus are closely interrelated and opioids modulate both sensations. Therefore, we undertook a series of experiments to characterize opioid-mediated antinociception in cholestasis and determine if it occurs inside or outside the CNS. Antinociception scores to both thermal and mechanical stimuli were determined in mice with cholestasis due to bile duct resection vs sham controls. Cholestatic mice demonstrated significant antinociception to both stimuli compared to controls, which was reversible by the opiate receptor antagonist naloxone. The experiments were repeated with a naloxone derivative, which does not cross the blood-brain-barrier (i.e. naloxone methiodide) with similar results, indicating an opioid antinociceptive effect mediated outside of the CNS. Experiments with intraplantar injections of low dose naloxone methiodide confirmed that cholestasis-associated antinociception occurs at the level of cutaneous nerve endings. These findings were supported by findings of increased dermal met-enkephalin expression in cholestatic mice. Cholestasis in mice is associated with antinociception due to local effects of endogenous opioids (i.e. met-enkephalin) at the level of sensory nerve endings. These findings may have direct implications in the management of cholestasis associated pruritus.

Control of inflammatory pain by chemokine-mediated recruitment of opioid-containing polymorphonuclear cells

Opioid-containing leukocytes can counteract inflammatory hyperalgesia. Under stress or after local injection of corticotropin releasing factor (CRF), opioid peptides are released from leukocytes, bind to opioid receptors on peripheral sensory neurons and mediate antinociception. Since polymorphonuclear cells (PMN) are the predominant opioid-containing leukocyte subpopulation in early inflammation, we hypothesized that PMN and their recruitment by chemokines are important for peripheral opioid-mediated antinociception at this stage. Rats were intraplantarly injected with complete Freund's adjuvant (CFA). Using flow cytometry, immunohistochemistry, and ELISA, leukocyte subpopulations, chemokine receptor (CXCR2) expression on opioid-containing leukocytes and the CXCR2 ligands keratinocyte-derived chemokine (KC), macrophage inflammatory protein-2 (MIP-2) and cytokine-induced neutrophil chemoattractant-2 (CINC-2) were quantified. Paw pressure threshold (PPT) was determined before and after intraplantar and subcutaneous injection of CRF with or without naloxone. PMN depletion was achieved by intravenous injection of an antiserum. Chemokines were blocked by intraplantar injection of anti-MIP-2 and/or anti-KC antiserum. We found that at 2 h post CFA (i) intraplantar but not subcutaneous injection of CRF produced dose-dependent and naloxone-reversible antinociception ( P<0.05, ANOVA). (ii) Opioid-containing leukocytes in the paw and CRF-induced antinociception were reduced after PMN depletion ( P<0.05, t-test). (iii) Opioid-containing leukocytes mostly expressed CXCR2. MIP-2 and KC, but not CINC-2 were detectable in inflamed but not in noninflamed tissue ( P<0.05, ANOVA). (iv) Combined but not single blockade of MIP-2 and KC reduced the number of opioid-containing leukocytes and peripheral opioid-mediated antinociception ( P<0.05, t-test; P>0.05, ANOVA). In summary, in early inflammation peripheral opioid-mediated antinociception is critically dependent on PMN and their recruitment by CXCR2 chemokines.

Pharmacological and functional biochemical properties of d-Ala 2- d-Nle 5-enkephalin-Arg-Phe

Tyr- d-Ala-Gly-Phe- d-Nle-Arg-Phe (DADN) a synthetic analogue of the endogenous Met-enkephalin-Arg-Phe (Tyr-Gly-Gly-Phe-Met-Arg-Phe; MERF), was investigated in radioligand binding assays, [ 35S]GTPγS stimulation experiments as well as in in vivo algesiometric tests. Binding properties of [ 3H]DADN were measured in crude membrane fractions of rat spinal cord tissues and in homogenates of Chinese hamster ovary (CHO) cells selectively expressing δ-, κ-or μ-opioid receptors. The highest affinity for [ 3H]DADN binding was observed in membranes from CHO cells transfected with μ-opioid receptors confirming the μ-selectivity of the peptide. Unlabeled DADN was also investigated in functional biochemical experiments by measuring opioid receptor-mediated G-protein activation in rat brain membrane fractions. The peptide stimulated the activity of the regulatory G-proteins in a concentration dependent manner, and the stimulation was efficiently inhibited in the presence of μ-receptor specific antagonist ligands further supporting the selectivity profile of DADN. Intrathecally administered DADN produced a dose-related, naloxone-reversible antinociception in rat hot water tail-flick tests. Among the selective opioid antagonists tested, the δ-selective naltrindole (NTI) and the κ-specific norbinaltorphimine (norBNI) showed only slight blocking effects compared with naloxone. The results obtained in the in vitro agonist-stimulated [ 35S]GTPγS binding assays are in good agreement with the opioid agonist effect seen in the in vivo pain test.

Effect of D-arginine on antinociception induced by kyotorphin, Tyr-cav, L-cav and Tyr(Cl2)-cav in rats.

Kyotorphin (Kyo) is synthesized in specific brain regions where it may modulate synaptic transmission. Kyo directly excites cortical neurons, and indirectly exerts opioid actions to produce analgesia via the release of [Met]-enkephalin. Kyo is formed by a specific enzyme from L-tyrosine, L-arginine (L-arg) and ATP in the presence of Mg2+. Kyo and its analogues Tyr-cav and Tyr(Cl2)-cav exert naloxone-reversible antinociception in the paw-pressure test. The aim of the present study was to investigate the effect of D-arginine (D-arg) on the analgesic effects of Kyo, Tyr-cav and Tyr(Cl2)-cav during acute pain. The changes in the nociceptive effects were examined in male Wistar rats using the tail flick (TF) and hot plate (HP) tests. Kyo, Tyr-cav, Tyr(Cl2)-cav, L-canavanine (L-cav) and D-arg were applied in rats intracerebroventricularly (i.c.v.) at a dose of 20 microg/20 microl. Kyo, Tyr-cav, Tyr(Cl2)-cav and L-cav exerted antinociceptive activity in both tests used. Applied alone, D-arg had no antinociceptive activity in TF and HP tests. D-arg decreased the TF and HP latency of the Kyo, Tyr-cav, Tyr(Cl2)-cav and L-cav. Taken together, these results reveal D-arg as a potential inhibitor of the investigated peptides during acute pain.

μ Receptor and G i2α antisense attenuate [ d-Met 2]-FMRFamide antinociception in mice

FMRFamide (Phe-Met-Arg-Phe-NH 2) and several analogs produce centrally-mediated, naloxone-reversible antinociception, but have minimal affinity for opioid receptor (sub)types. In the present study, the antinociception in mice (55°C tail-flick test) produced by supraspinal (intracerebroventricular; ICV) administration of [ d-Met 2]-FMRFamide (a stable analog of FMRFamide) was attenuated by pretreatment with ICV oligodeoxyribonucleotide antisense to the opioid μ receptor or by antisense to the G i2α G-protein subunit. These data suggest that [ d-Met 2]-FMRFamide produces its antinociception via an opioid interneuron.

Is fatigue associated with cholestasis mediated by altered central neurotransmission?

Is fatigue associated with cholestasis mediated by altered central neurotransmission?

A pharmacological profile of the novel, peripherally‐selective k‐opioid receptor agonist, EMD 61753

1. The pharmacological properties of the novel diarylacetamide kappa-opioid receptor agonist, EMD 61753, have been compared with those of ICI 197067 (a centrally-acting kappa agonist) and ICI 204448 (a peripherally-selective kappa agonist). 2. EMD 61753 binds with high affinity (IC50 5.6 nM) and selectivity (kappa:mu:delta:sigma binding ratio 1:536:125: > 1,786) to kappa-opioid receptors and is a full and potent (IC50 54.5 nM) agonist in an in vitro assay for kappa-opioid receptors (rabbit vas deferens preparation). 3. Systemically-applied [14C]-EMD 61753 is found in high concentrations in the lungs, liver, adrenal glands and kidneys. Considerably less radioactivity is detected in the whole brain, and this radioactivity is concentrated in the region of the cerebral ventricles in the choroid plexuses. EMD 61753 penetrates only poorly into the CNS. 4. EMD 61753 was weakly effective in pharmacological tests of central activity. This compound reversed haloperidolol-induced DOPA accumulation in the nucleus accumbens of the rat only at a dose of 30 mg kg-1, s.c., (doses of 0.1, 1.0 and 10 mg kg-1, s.c., and 1.0, 10 and 100 mg kg-1, p.o., were inactive). Hexobarbitone-induced sleeping in mice was prolonged by EMD 61753 at threshold doses of 10 mg kg-1, s.c., and 100 mg kg-1, p.o., whereas the motor performance of rats in the rotarod test was impaired by EMD 61753 with an ID50 value of 453 mg kg-1, s.c. 5. EMD 61753 produced dose-dependent, naloxone-reversible antinociception in the mouse formalin test (1st phase ID50 1.9 mg kg-1, s.c., and 10.4 mg kg-1, p.o.; 2nd phase ID50 0.26 mg kg-1, s.c., and 3.5 mg kg-1, p.o.) and rodent abdominal constriction test (ID50 mouse 1.75 mg kg-1, s.c., and 8.4 mg kg-1, p.o.; ID50 rat 3.2 mg kg-1, s.c., and 250 mg kg-1, p.o.). EMD 61753 was inactive, or only weakly effective, in the rat pressure test under normalgesic conditions. After the induction of hyperalgesia with carrageenin, however, this compound elicited potent, dose-dependent (ID50 0.08 mg kg-1, s.c., and 6.9 mg kg-1, p.o., after remedial application, and 0.2 mg kg-1, s.c., and 3.1 mg kg-1, p.o., after prophylactic application) and naloxone-reversible antinociception. The antinociceptive action of systemically-applied (50 mg kg-1, p.o.) EMD 61753 in the hyperalgesic pressure test was completely inhibited by injection of the K-opioid antagonist norbinaltorphimine (100 Lg) into the inflamed tissue, a result which indicates that this opioid effect is mediated peripherally.6. Cutaneous plasma protein extravasation produced by antidromic electrical stimulation of the rat saphenous nerve was dose-dependently inhibited by systemically-applied EMD 61753 (ID13 values 3.7 mg kg-1, s.c., and 35.8 mg kg-1, p.o.), and this effect was completely antagonized by intraplantar application of norbinaltorphimine (50 microg). Extravasation elicited by the intraplantar application of substance P (10 microg) was not influenced by the administration of EMD 61753.7. EMD 61753 produced dose-dependent diuresis in non-hydrated rats at doses of and above 1.0 mg kg-1, s.c., and 10 mg kg-1, p.o., and in saline-loaded rats at doses of and above 10 mg kg-1, s.c.,and 30mgkg-1, p.o.8. The prostaglandin-mediated fall in mean arterial blood pressure elicited in anaesthetized rats by i.v.application of arachidonic acid was not inhibited by prior treatment with EMD 61753 (10mg kg-1,p.o.). Thus, a blockade of prostaglandin synthesis via inhibition of cyclo-oxygenase activity does not contribute to the in vivo effects of EMD 61753 and its metabolites.9 The present experiments therefore indicate that EMD 61753 is a potent, selective and orally-effective full ic-opioid receptor agonist which has a limited ability to penetrate the blood-brain barrier and elicit centrally-mediated sedation, putative aversion, diuresis, and antinociception. The inhibitory actions of systemically-applied EMD 61753 against hyperalgesic pressure nociception and neurogenic inflammation are mediated peripherally, probably by opioid receptors on the endings of sensory nerve fibres.

Activity at phencyclidine and mu opioid sites mediates the hyperalgesic and antinociceptive properties of the N-terminus of substance P in a model of visceral pain

Substance P, a putative neurotransmitter or neuromodulator of nociception or pain in the spinal cord, exhibits both antinociceptive and hyperalgesic properties. Investigators have shown that the N-terminal metabolite of substance P, substance P(1–7), produces naloxone-reversible antinociception when given supraspinally and systemically in mice and hyperalgesia when injected intrathecally in rats. The goal of our investigation was to identify the receptors mediating these actions of substance P(1–7) at the initial site of release of substance P i.e. in the spinal cord. Thirty minutes after intrathecal injection, substance P(1–7) produced naloxone-reversible antinociception in a dose-dependent manner in the abdominal stretch assay. When administered with naloxone, substance P(1–7) produced hyperalgesia 5 and 10 min after injection, which was inhibited by dizocilpine (MK-801), a phencyclidine ligand and non-competitive antagonist of N-methyl-d-aspartate. Antinociception was inhibited by the μ-selective opioid antagonist β-funaltrexamine, but not by the μ-selective opioid antagonist naloxonazine or the δ-selective antagonist naltrindole, indicating a μ 2-opioid receptor-mediated effect. These findings suggest that the N-terminal portion of substance P may modulate nociception or pain, as demonstrated in the acetic acid abdominal stretch (writhing) assay, via activation of two different receptor systems. Substance P(1–7)-induced hyperalgesia is mediated by a phencyclidine-sensitive mechanism and antinociception involves activity at μ-opioid, most likely μ 2, receptors.

Central and peripheral actions of the novel kappa-opioid receptor agonist, EMD 60400.

1. The pharmacological characteristics of the kappa-opioid receptor agonist, EMD 60400, have been investigated, with particular reference to its central and peripheral sites of action and its ability to influence nociception. The kappa agonists ICI 197067 and ICI 204448 were tested for purposes of comparison. 2. EMD 60400 and ICI 197067 bind with high affinity (IC50 values of 2.8 and 1.5 nM, respectively) and high selectivity to kappa-opioid receptors. ICI 204448 has a lower binding affinity (IC50 13.0 nM) and selectivity for kappa-opioid receptors. 3. EMD 60400, ICI 197067, and ICI 204448 are full and potent agonists in the rabbit vas deferens in vitro assay for kappa-opioid receptors (IC50 values of 41.8, 15.7 and 15 nM, respectively). 4. Ex vivo binding experiments in mice revealed that EMD 60400 and ICI 197067 were well taken up after s.c. administration. Brain levels of EMD 60400 were lower than those of ICI 197067 at comparable doses, indicating that EMD 60400 does not penetrate into the CNS as well as ICI 197067. 5. Haloperidol-induced DOPA accumulation in the nucleus accumbens of the rat was dose-dependently reversed by s.c. application of EMD 60400 and ICI 197067 at doses of and above 3 and 0.3 mg kg-1, respectively. ICI 204448 had no effect on DOPA accumulation at 30 mg kg-1, s.c. 6. Prolongation of hexobarbitone-induced sleeping time in mice and motor impairment in the rat rotarod test were observed for EMD 60400 at doses above 3 and 2.5 mg kg-1, s.c., respectively, and for ICI 197067 at doses above 0.3 and 0.25 mg kg-1, s.c., respectively. ICI 204448 was inactive in these tests at doses of 30 and 100 mg kg-1, s.c., respectively.7. EMD 60400 applied s.c. produced dose-dependent naloxone-reversible antinociception in the mouse formalin test (1st and 2nd phase ID50 0.44 and 0.47 mg kg-1, respectively) and rodent writhing test (ID50 mouse 0.55 mg kg-1 and rat 0.3mg kg-1). Furthermore, EMD 60400 was considerably more potent in the rat pressure pain test after the induction of inflammation with carrageenin than under normalgesic conditions (ID50 values 0.1 Microg kg-1 and 4.0 mg kg-1, s.c., respectively). The action of EMD 60400 (50 microgkg-1, s.c.) in the hyperalgesic pressure pain test was completely antagonized by injection of the K-opioid antagonist, norbinaltorphimine (100 microg) into the inflamed tissue, thus demonstrating the peripheral opioid nature of this effect.8. EMD 60400 produced dose-dependent inhibition of neurogenic plasma extravasation elicited byantidromic electrical stimulation of the rat saphenous nerve (ID50 value 0.3 mg kg-1, i.v.). This inhibition was completely antagonized by intraplantar injection of norbinaltorphimine (50 microg).9. EMD 60400, ICI 197067, and ICI 204448 have diuretic effects in rats at doses of and above 0.1, 0.01,and 0.3 mg kg-1, s.c., respectively. An antidiuretic action was also observed with ICI 197067 at very low doses (3 and 6 microgkg-1, s.c.).10. Pharmacological and biochemical data therefore indicate that the three K-opioid receptor agonists tested here have different tendencies to elicit centrally-mediated sedation and putative aversion(ICI 197067 > EMD 60400 > ICI 204448) which correspond to their ability to cross the blood-brain barrier. EMD 60400 combines high affinity and selectivity for the K receptor with a degree of peripheral selectivity. The peripheral actions of systemically-applied EMD 60400 against hyperalgesic pressure pain and neurogenic inflammation are very probably mediated by opioid receptors on the endings of sensory nerve fibres.

Phencyclidine (PCP) receptors mediate substance P N-terminus-induced hyperalgesia while mu-type opioid receptors mediate its antinociceptive effect

Substance P (SP) exhibits both antinociceptive and hyperalgesic properties in the central nervous system (CNS). The N-terminus of SP, which has no neurokinin activity, induces naloxone-reversible antinociception in mice (1) and hyperalgesia in rats (2) as demonstrated using the major SP metabolite, SP(1-7). To investigate more specific receptor mechanisms of these SP(1-7)-induced effects in mice, we injected SP(1-7) intrathecally (i.t.), i.e. at the first site of SP release in the CNS in response to a peripheral nociceptive input. Thirty minutes after i.t. injection, SP(1-7) produced naloxone-reversible time- and dose-dependent antinociception in the abdominal stretch assay

Cholestasis in the male rat is associated with naloxone-reversible antinociception

Clinical observations have suggested that cholestasis is associated with increased neurotransmission mediated by the opioid system in the central nervous system. As opiate agonists (e.g. morphine) mediate analgesia, increased opioidergic tone in cholestasis should be associated with a decreased response to pain. To test this hypothesis, the response of rats with acute cholestasis to a nociceptive stimulus was measured by the use of the tail-flick test, an extensively validated assay for measuring opiate-induced antinociception. Five and 7 days after bile-duct resection, the mean tail-flick latency was longer than before surgery (p < 0.05), whereas the corresponding means for unoperated and sham-resected controls were not significantly different from their respective baseline values. The increase in the mean tail-flick latency in the bile-duct resection group was reversed by (-)-naloxone (1 mg/kg subcutaneously), but not by its enantiomer (+)-naloxone (10 mg/kg subcutaneously) (p < 0.001). The stereoselective reversal of antinociception in cholestasis by naloxone indicates that this phenomenon is opioid-receptor mediated. In contrast, prolongation of the mean TFL found in the rat model of thioacetamide-induced acute hepatocellular necrosis was not reversed by (-)-naloxone, indicating that antinociception in this model is not opioid mediated. These findings provide support for the hypothesis that cholestasis is associated with increased opioidergic tone.

L-Tyrosine-induced antinociception in the mouse: involvement of central δ-opioid receptors and bulbo-spinal noradrenergic system

L-Tyrosine methyl ester (L-Tyr-OMe) produces naloxone-reversible antinociception in mice. This effect was characterized in the present study. L-Tyr-OMe administered s.c. at doses of 100–400 mg/kg elicited dose-dependent antinociception, which was antagonized by s.c. and i.c.v. naloxone at 1 mg/kg and 10 ng/mouse, respectively, but not by i.t. injection at 10 ng/mouse. The antinociceptive effect of systemic L-Tyr-OMe was also inhibited by naltrindole (0.1 mg/kg s.c.), a selective δ-opioid receptor antagonist, but was resistant to L-leucyl-L-arginine (3 μg/mouse i.c.v.), a kyotorphin (L-tyrosyl-L-arginine) receptor antagonist. This effect was reduced by i.t., but not by i.c.v., phentolamine at 0.1–1 μg/mouse, while methysergide (10 μg/mouse i.t.) and p-chlorophenylalanine (500 mg/kg s.c.) did not have such an inhibitory effect. L-Tyr-OMe, given i.c.v. and i.t. at 0.1–10 μg/mouse, also produced dose-dependent antinociception, the latter effect being naloxone-reversible but resistant to i.t. phentolamine. These results suggest that L-Tyr-OMe (s.c.)-induced antinociception is mediated by central δ-opioid receptors and by the bulbo-spinal noradrenergic system, but not by the central ‘kyotorphinergic’ system, and that L-Tyr-OMe, when given i.t., produces antinociception via spinal opioidergic, but not noradrenergic systems.

Central, naloxone-reversible antinociception by diclofenac in the rat

The antinociceptive effect of subcutaneously (s.c.), intracerebroventricularly (i.c.v.) or intrathecally (i.t.) administered diclofenac was studied in a series of experiments employing the tail-flick (0.01-10.0 mg/kg body weight i.p., 1-50 micrograms i.c.v., 1-10 micrograms i.t.) and hot-plate (0.01-50 mg/kg body weight i.p., 1-50 micrograms i.c.v., 1-10 micrograms i.t.) models representing somatosensory stimuli and the writhing test (0.001 mg-10 mg s.c., 0.1-200 micrograms i.c.v., 0.1-100 micrograms i.t.) and colorectal distension (1-200 micrograms i.c.v.) models representing noxious visceral stimuli. Diclofenac did not exert any antinociceptive effects in the tail-flick or hot-plate models. In the writhing test, diclofenac dose-dependently inhibited the number of writhings after s.c. administration (0.001-10.0 mg/kg body weight) with an ED50 of 1 mg/kg. A similar dose-dependent action of diclofenac was seen after i.c.v. (0.1-200 micrograms) and i.t. (0.1-100 micrograms) administration with an ED50 of 3 micrograms in both cases. The antinociceptive effect of diclofenac administered s.c., i.c.v. or i.t. was almost completely reversed by pretreatment with naloxone, (1 mg/kg body weight s.c.). In the colorectal distension model, the i.c.v. administration of diclofenac (1-200 micrograms), which attenuated the cardiovascular response to colorectal distension, was reversed by naloxone. The pressor and tachycardia response to a 20 s, 80 mmHg colorectal distension was inhibited by diclofenac 100 micrograms i.c.v. (16.1 +/- 1.7 mmHg or 58% and 39.4 +/- 0.4 bpm or 70% versus control, respectively). It is concluded that diclofenac exerts a central, naloxone-reversible antinociceptive action in experimental animals after noxious visceral stimuli but not after somatosensory stimuli.