Non-selective Opioid Receptor Antagonist Naloxone Research Articles

Roles of nociceptin/orphanin FQ and nociceptin/orphanin FQ peptide receptor in respiratory rhythm generation in the medulla oblongata: an in vitro study

Nociceptin/orphanin FQ (N/OFQ) is the endogenous agonist of the orphan opioid receptor-like receptor (NOP receptor, previously termed ORL1), a novel member of the opioid receptor family. The aim of the present study, using in vitro newborn rat preparations, was to elucidate the roles N/OFQ and the NOP receptor play in medullary generation of respiratory rhythm. The brainstem-spinal cord from 3-day-old Wistar rats was isolated and perfused with artificial cerebrospinal fluid (27.5 degrees C) equilibrated with oxygen 95% and carbon dioxide 5% at pH 7.4. Respiratory activity was recorded from the C4/C5 ventral roots. The effects of N/OFQ (10 nM, 30 nM, 100 nM) on respiratory frequency (fR) (bursts min(-1)) was measured. Drugs were administered through the recording chamber by means of a perfusion system. In addition, the effects of pretreatment with the classical non-selective opioid receptor antagonist naloxone 1 microM, and the selective NOP antagonist CompB 10 microM, were evaluated. Statistical significance was evaluated using ANOVA followed by Dunnett's test (P<0.05). N/OFQ reduced fR in a concentration-dependent manner. Pretreatment with CompB 10 microM prevented the N/OFQ 10 nM-induced fR reduction, whereas CompB itself was inactive. Pretreatment with naloxone did not prevent the N/OFQ-induced fR reduction. N/OFQ acts as a neuromodulator to reduce fR in the respiratory rhythm- generating centre of the medulla oblongata, and this action of N/OFQ is mediated by NOP receptors.

Vagal modulation of spinal nicotine-induced inhibition of the inflammatory response mediated by descending antinociceptive controls

Noxious stimuli activate neuroendocrine axes, inhibiting inflammation, an effect that is powerfully attenuated by ongoing activity in subdiaphragmatic vagal afferents. To evaluate whether this inhibitory effect of vagal afferent activity is mediated by descending antinociceptive control, we tested whether antagonizing descending antinociceptive controls: (i) enhances the inhibition of inflammation produced by spinal nicotine (which stimulates central terminals of nociceptors) and (ii) occludes the enhancing effect of subdiaphragmatic vagotomy, in the rat. Spinal intrathecal co-administration of the α-adrenergic receptor antagonist phentolamine and the non-selective opioid receptor antagonist naloxone, and acute subdiaphragmatic vagotomy each produced enhancement, with similar magnitude, of nicotine-induced inhibition of plasma extravasation, produced by the potent inflammatory mediator, bradykinin. The combination of subdiaphragmatic vagotomy and intrathecal receptor antagonists, however, produced no further enhancement compared to each treatment alone. These findings support the suggestion that activity in descending antinociceptive controls modulates noxious stimulus-induced inhibition of inflammation and the vagal modulation of noxious stimulus-induced inhibition of inflammation is mediated by descending antinociceptive controls.

Noninvasive remote ischemic preconditioning for global protection of skeletal muscle against infarction.

The aim of this study was to investigate the efficacy and mechanism of action of a noninvasive remote ischemic preconditioning (IPC) technique for the protection of multiple distant skeletal muscles against ischemic necrosis (infarction). It was observed in the pig that three cycles of 10-min occlusion and reperfusion in a hindlimb by tourniquet application reduced the infarction of latissimus dorsi (LD), gracilis (GC), and rectus abdominis (RA) muscle flaps by 55%, 60%, and 55%, respectively, compared with their corresponding control (n = 6, P < 0.01) when they were subsequently subjected to 4 h of ischemia and 48 h of reperfusion. This infarct-protective effect of remote IPC in LD muscle flaps was abolished by an intravenous bolus injection of the nonselective opioid receptor antagonist naloxone (3 mg/kg) 10 min before remote IPC and a continuous intravenous infusion (3 mg/kg) during remote IPC and by an intravenous bolus injection of the selective delta 1-opioid receptor antagonist 7-benzylidenealtrexone maleate (3 mg/kg). However, this infarct-protective effect of remote IPC was not affected by an intravenous bolus injection of the ganglionic blocker hexamethonium chloride (20 mg/kg) or the nonspecific adenosine receptor antagonist 8-(p-sulfophenyl)theophylline (10 mg/kg) or by a local intra-arterial injection of the adenosine1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (3 mg/muscle flap) given 10 min before remote IPC. It was also observed that this remote IPC of skeletal muscle against infarction was associated with a slower rate of muscle ATP depletion during the 4 h of sustained ischemia and a reduced muscle neutrophilic myeloperoxidase activity after 1.5 h of reperfusion. These observations led us to speculate that noninvasive remote IPC by brief cycles of occlusion and reperfusion in a pig hindlimb is effective in global protection of skeletal muscle against infarction. This infarct-protective effect is most likely triggered by the activation of opioid receptors in the skeletal muscle, and remote IPC is associated with an energy-sparing effect during sustained ischemia and attenuation of neutrophil accumulation during reperfusion.

Sexual dimorphism in very low dose nalbuphine postoperative analgesia

In recent studies we demonstrated that the analgesic effect of the κ-like opioids is significantly greater in women, that low dose nalbuphine (5 mg) produces profound anti-analgesia (i.e. enhances pain) in men, and that addition of a low dose of the non-selective opioid receptor antagonist naloxone (0.4 mg) to nalbuphine (5 mg) abolishes the sex difference and results in significantly enhanced analgesia in both sexes. To further delineate the dose-dependent analgesic and anti-analgesic effects of nalbuphine, the present study evaluated the effect of a lower dose of nalbuphine (2.5 mg), with and without naloxone, on dental postoperative pain. In women, nalbuphine alone induced modest, short duration analgesia, which was antagonized rather than enhanced by the addition of naloxone (0.4 mg). In men, this dose of nalbuphine alone did not produce analgesia or anti-analgesia, and naloxone (0.4 mg) did not alter the response to nalbuphine. Thus, the anti-analgesic effect of nalbuphine, present in both sexes at the 5 mg dose disappears at the lower dose of nalbuphine. In addition, the mild analgesia in women produced by this lower dose of nalbuphine is antagonized by naloxone.

Lack of Evidence of an Interaction between Leu-Enkephalin and Muscarinic-Like Responses in the Frog Semicircular Canal

Using multiunit recording of action potentials from the whole vestibular nerve, we studied whether opioid peptide leu-enkephalin (Enk) may modulate muscarinic-like responses in semicircular organs of the frog. When acetylcholine (ACh) (0.1–1 mM) was applied with 1 nM Enk, the maximal frequency increase induced by ACh was reduced.However,the frequency decrease of ACh responses under Enk did not differ from the frequency decline in basal spike discharge induced by Enk alone. Administration of atropine (1 µM) left the response to Enk intact and blocked the excitatory effect of ACh. No modification of the ACh response under Enk was observed in the presence of the non-selective opioid receptor antagonist naloxone (10 µM). This study suggests that no interaction exists between the ACh-mediated excitatory action on resting activity in the isolated semicircular canal preparation and the suppressive action of Enk.

Functional interactions between δ- and μ-opioid receptors in rat thermoregulation

The selective δ-opioid receptor agonist deltorphin II (25.0–100.0 μg, i.c.v.) produced biphasic effects on core temperature in rats, in which hypothermia was followed by hyperthermia. Pretreatment with the selective δ-opioid receptor antagonist, naltrindole (25.0 μg, i.c.v.), blocked hypothermia produced by deltorphin II and had a tendency to potentiate the hyperthermic effect of deltorphin II. The non-selective opioid receptor antagonist naloxone (1.5 mg kg −1, s.c.) potentiated hypothermia, and blocked hyperthermia, produced by deltorphin II (100.0 μg). Also, naloxone potentiated hypothermia produced by a lower dose of deltorphin II (25.0 μg), which did not produce hyperthermia. A similar pattern was found for the selective μ-opioid receptor antagonist, β-funaltrexamine (5.0 μg, i.c.v.), which potentiated and blocked deltorphin II-induced hypo- and hyperthermia, respectively. The selective κ-opioid receptor antagonist nor-binaltorphimine (20.0 μg, i.c.v.) had no effects on deltorphin II-induced temperature changes. The present results suggest that deltorphin II produces hypothermia through activation of δ-opioid receptors, whereas the hyperthermic effect of deltorphin II involves activation of μ-opioid receptors. This μ-opioid receptor stimulatory effect of deltorphin II is furthermore more pronounced than was anticipated based on the reported in vitro properties of this compound. The biphasic effect of deltorphin II implies a negative interaction between δ- and μ-opioid receptors in thermoregulation in rats.

Local Injection of a Selective Endothelin-B Receptor Agonist Inhibits Endothelin-1-Induced Pain-Like Behavior and Excitation of Nociceptors in a Naloxone-Sensitive Manner

We showed previously that subcutaneous injection of the injury-associated peptide mediator endothelin-1 (ET-1) into the rat plantar hindpaw produces pain behavior and selective excitation of nociceptors, both through activation of ET(A) receptors likely on nociceptive terminals. The potential role of ET(B) receptor activation in these actions of ET-1-has not been examined. Therefore, in these experiments, we studied the effect of blocking or activating ET(B) receptors on ET-1-induced hindpaw flinching and excitation of nociceptors in rats. An ET(B) receptor-selective antagonist, BQ-788 (3 mm), coinjected with ET-1 (200 microm) reduced the time-to-peak of flinching and significantly enhanced the average maximal flinch frequency (MFF). In contrast, coinjection of an ET(B) receptor selective agonist, IRL-1620 (100 or 200 microm), with ET-1 reduced the average MFF and the average total number of flinches. Interestingly, this unexpected inhibitory effect of IRL-1620 was prevented by the nonselective opioid receptor antagonist naloxone (2.75 mm). To confirm these inhibitory actions, we studied the effects of IRL-1620 on ET-1-induced spike responses in single, physiologically characterized nociceptive C-fibers. IRL-1620 suppressed spike responses to ET-1 in all (n = 12) C-units, with mean and maximum response frequencies of 0.08 +/- 0.02 and 1.5 +/- 0.4 impulses/sec versus 0.32 +/- 0.07 and 4.17 +/- 0.17 impulses/sec for ET-1 alone. In additional support of the behavioral results, coinjection of naloxone (2.75 mm) completely prevented this inhibitory action of IRL-1620. These results establish that ET(B) receptor activation inhibits ET-1-induced pain behavior and nociception in a naloxone-sensitive manner and point to a previously unrecognized dual modulation of acute nociceptive signaling by ET(A) and ET(B) receptors in cutaneous tissues.

Effects of opioid antagonists on unconditioned and conditioned hyperactivity to morphine

In a series of experiments, the ability of selective μ- (β-funaltrexamine, β-FNA), δ- (naltrindole, nalt) and κ- (nor-binaltorphimine, nor-BNI) opioid receptor antagonists to attenuate the unconditioned and conditioned hyperactive effects of morphine was examined. For comparison, the nonselective opioid receptor antagonist naloxone (nalx) was also examined. Locomotor activity served as the behavioral measure. Experiment 1 found that doses of 1 and 4, but not 16 mg/kg, of morphine effectively produced conditioned hyperactivity (CH). Experiments 2a–d found that β-FNA, nalt, nor-BNI and nalx, respectively, attenuated unconditioned morphine-induced hyperactivity. Experiments 3a–c, however, found that none of the selective antagonists, given individually, attenuated CH. In contrast, nalx did attenuate CH (Experiment 3d). Collectively results suggest that the unconditioned and conditioned hyperactive responses to morphine are mediated by different receptor systems and that activation of multiple opioid-receptor subtypes mediate expression of CH.

Nociceptin/orphanin FQ receptors modulate glutamate extracellular levels in the substantia nigra pars reticulata. A microdialysis study in the awake freely moving rat

Intracerebral microdialysis was employed in awake freely moving rats to investigate the effects of nociceptin/orphanin FQ receptor ligands on glutamate extracellular levels in the substantia nigra pars reticulata. Nociceptin/orphanin FQ, ineffective at 0.1 μM, induced a prolonged stimulation of nigral glutamate levels at 1 and 10 μM (mean effect of 137±9 and 167±13%, respectively, of basal values). These effects were prevented by the novel nociceptin/orphanin FQ receptor antagonist [Nphe 1]nociceptin/orphanin FQ(1-13)NH 2 (100 and 300 μM, respectively) but not by the non-selective opioid receptor antagonist naloxone (10 μM). [Nphe 1]nociceptin/orphanin FQ(1-13)NH 2 (100 μM) inhibited by itself glutamate outflow (maximal reduction to 71±4%) while naloxone was ineffective. The nociceptin/orphanin FQ receptor ligand [Phe 1ψ(CH 2-NH)Gly 2]nociceptin/orphanin FQ(1-13)NH 2 also facilitated glutamate outflow at 10 μM (mean effect of 145±10%). Intranigral perfusion with tetrodotoxin (1 μM) or with the dopamine D 2 receptor antagonist raclopride (1 μM), failed to affect basal glutamate output and prevented the facilitatory effect of nociceptin/orphanin FQ (10 μM). However, perfusion with the GABA A receptor antagonist bicuculline (10 μM) increased local glutamate extracellular levels by itself and attenuated the effect of the peptide. Our data suggest that nociceptin/orphanin FQ increases glutamate extracellular levels in the substantia nigra pars reticulata via activation of nociceptin/orphanin FQ receptors located on non-glutamatergic, possibly dopaminergic and GABAergic, neuronal elements.

Differential antinociceptive effects induced by intrathecally administered endomorphin-1 and endomorphin-2 in the mouse

Two highly selective μ-opioid receptor agonists, endomorphin-1 and endomorphin-2, have been identified and postulated to be endogenous ligands for μ-opioid receptors. Intrathecal (i.t.) administration of endomorphin-1 and endomorphin-2 at doses from 0.039 to 5 nmol dose-dependently produced antinociception with the paw-withdrawal test. The paw-withdrawal inhibition rapidly reached its peak at 1 min, rapidly declined and returned to the pre-injection levels in 20 min. The inhibition of the paw-withdrawal responses to endomorphin-1 and endomorphin-2 at a dose of 5 nmol observed at 1 and 5 min after injection was blocked by pretreatment with a non-selective opioid receptor antagonist naloxone (1 mg/kg, s.c.). The antinociceptive effect of endomorphin-2 was more sensitive to the μ 1-opioid receptor antagonist, naloxonazine than that of endomorphin-1. The endomorphin-2-induced paw-withdrawal inhibition at both 1 and 5 min after injection was blocked by pretreatment with κ-opioid receptor antagonist nor-binaltorphimine (10 mg/kg, s.c.) or the δ 2-opioid receptor antagonist naltriben (0.6 mg/kg, s.c.) but not the δ 1-opioid receptor antagonist 7-benzylidine naltrexone (BNTX) (0.6 mg/kg s.c.). In contrast, the paw-withdrawal inhibition induced by endomorphin-1 observed at both 1 and 5 min after injection was not blocked by naloxonazine (35 mg/kg, s.c.), nor-binaltorphimine (10 mg/kg, s.c.), naltriben (0.6 mg/kg, s.c.) or BNTX (0.6 mg/kg s.c.). The endomorphin-2-induced paw-withdrawal inhibition was blocked by the pretreatment with an antiserum against dynorphin A-(1-17) or [Met 5]enkephalin, but not by antiserum against dynorphin B-(1-13). Pretreatment with these antisera did not affect the endomorphin-1-induced paw-withdrawal inhibition. Our results indicate that endomorphin-2 given i.t. produces its antinociceptive effects via the stimulation of μ 1-opioid receptors (naloxonazine-sensitive site) in the spinal cord. The antinociception induced by endomophin-2 contains additional components, which are mediated by the release of dynorphin A-(1-17) and [Met 5]enkephalin which subsequently act on κ-opioid receptors and δ 2-opioid receptors to produce antinociception.

Inhibitory activity of nociceptin/orphanin FQ on capsaicin-induced bronchoconstriction in the guinea-pig

In vivo studies were conducted in the guinea-pig to investigate the activity of the selective ORL1 receptor agonist nociceptin/orphanin FQ against capsaicin-induced bronchoconstriction, a response mediated by the release of tachykinins from pulmonary sensory nerves. Anesthetized guinea-pigs were ventilated with a rodent ventilator and placed in a whole-body plethysmograph, and pulmonary resistance (R L) and dynamic lung compliance (C Dyn) were monitored. Intravenous administration of nociceptin/orphanin FQ (0.3 mg/kg) inhibited the capsaicin-induced bronchoconstriction. The new nonpeptide ORL1 receptor antagonist 1-[(3 R,4 R)-1-cyclooctylmethyl-3-hydroxymethyl-4-piperidyl]-3-ethyl-1,3-dihydro-2 H-benzimidazol-2-one (J-113397) administered intravenously (1 mg/kg) produced a significant blockade of the inhibitory effect of nociceptin/orphanin FQ (0.3 mg/kg) on capsaicin-induced bronchoconstriction, whereas the nonselective opioid receptor antagonist naloxone (1 mg/kg) had no effect. Nociceptin/orphanin FQ (0.3 mg/kg) did not affect the bronchoconstriction induced exogenously by the tachykinin NK 2 receptor agonist [β-ala 8]-neurokinin A (4–10). We conclude that nociceptin inhibits in vivo capsaicin-evoked tachykinin release from sensory nerve terminals in the guinea-pig by a prejunctional mechanism. This inhibitory action does not involve activation of opioid receptors.

Nociceptin inhibits capsaicin-induced bronchoconstriction in isolated guinea pig lung

The isolated perfused guinea pig lung was used to investigate the effect of nociceptin against bronchoconstriction elicited by endogenous and exogenous tachykinins. The opioid receptor-like 1 (ORL1) receptor agonist, nociceptin/orphanin FQ (0.001–1 μM) produced a dose-related inhibition of the capsaicin-induced bronchoconstriction (10 −5–10 3 μg) in isolated guinea pig lung ( P<0.05), a response mediated by the release of endogenous tachykinins from lung sensory nerves. The new ORL1 receptor antagonist 1-[(3 R,4 R)-1-Cyclooctylmethyl-3-hydroxymethyl-4-piperidyl]-3-ethyl-1,3-dihydro-2 H-benzimidazol-2-one (J-113397) (0.3 μM) significantly blocked the inhibitory effect of nociceptin/orphanin FQ (0.01 μM) on capsaicin-induced bronchoconstriction, whereas the non-selective opioid receptor antagonist naloxone (1 μM) had no effect. Nociceptin/orphanin FQ (1 μM) did not affect the bronchoconstriction induced exogenously by the tachykinin NK2 receptor agonist neurokinin A. In conclusion, the present data provide evidence that nociceptin inhibits capsaicin-evoked tachykinin release from sensory nerve terminals in guinea pig lung by a prejunctional mechanism. This inhibitory action occurs independently from activation of opioid receptors. The present study also indicates that J-113397 is a potent ORL1 receptor antagonist.

Involvement of mu-Opioid Receptor in the Salsolinol-Associated Place Preference in Rats Exposed to Conditioned Fear Stress

Salsolinol, a condensation product of dopamine with acetaldehyde (the initial oxidation product of ethanol), has long been discussed as an endogenous opioid system-activating factor contributing to the etiology of alcoholism. Moreover, psychological stress has been considered to play an important role in the development of alcoholism. Male Sprague Dawley rats were subjected to conditioned fear stress (at 24 hr after electric footshock exposure), and then salsolinol or saline was injected intraperitoneally. For conditioning, rats were immediately confined to the nonpreferred compartment after salsolinol injection and to the preferred compartment after saline injection on alternate days. This conditioning session was repeated twice daily (for 8 days). Test session was carried out 1 day after the last conditioning session. Salsolinol [10 mg/kg, but not 1, 3, or 30 mg/kg, intraperitoneally (ip)] without conditioned fear stress induced a slight, but significant, place preference. In contrast, salsolinol (1, 3, and 10 mg/kg, ip) with conditioned fear stress induced a marked and significant place preference. The nonselective opioid receptor antagonist naloxone hydrochloride (which can pass into the brain), but not the nonselective peripheral opioid receptor antagonist naloxone methiodide (which cannot pass into the brain), significantly attenuated the salsolinol (3 mg/kg, ip)-induced place preference under conditioned fear stress. Moreover, the selective mu-opioid receptor antagonist beta-funaltrexamine significantly attenuated the salsolinol-induced place preference. Furthermore, 0.3 mg/kg salsolinol (which produced no significant place preference) combined with the mu-opioid receptor agonist morphine [0.1 mg/kg, subcutaneously (sc)], at the dose which alone produced no significant place preference, produced a marked and significant place preference. Both naloxone hydrochloride and beta-funaltrexamine significantly attenuated the salsolinol plus morphine-induced place preference. Salsolinol may have some rewarding effect, and its rewarding effect may be potentiated by psychological stress. In addition, the rewarding effect of salsolinol especially under psychological stress may involve the endogenous central opioid system, i.e., mu-opioid receptor.

Alpha-2 adrenergic and opioid receptor-mediated gastroprotection

Clonidine inhibited the development of gastric mucosal lesions induced by either acidified ethanol or indomethacin. The ED 50 values were: 7.1 and 5.2 μg.kg –1 orally, respectively. The gastroprotective effect was antagonised by the pre-synaptic alpha-2 antagonist yohimbine, the more selective alpha-2 antagonist Ch-38083 and the pre-synaptic alpha-2B antagonist prazosin. Moreover, the non-selective opioid receptor antagonist naloxone, the delta receptor selective naltrindole also reversed the clonidine-induced mucosal protective action. Clonidine was also effective following intracerebroventricular administration with the ED 50 of 37 ng/rat against ethanol-induced mucosal damage. Our results suggest that: 1) the gastroprotective effect of clonidine is likely to be mediated by alpha-2B adrenoceptor subtype; 2) there is an interaction between pre-synaptic alpha-2 adrenoceptors and opioid system; and 3) clonidine can induce gastroprotection by central mechanism.

Opioid receptor agonistic characteristics of mitragynine pseudoindoxyl in comparison with mitragynine derived from Thai medicinal plant Mitragyna speciosa

We have previously elucidated the opiate-like action of mitragynine, an active principle isolated from the Thai medicinal plant Mitragyna speciosa. In the present study, effects of the related compound, mitragynine pseudoindoxyl on electrically stimulated contraction in guinea pig ileum and mouse vas deferens, and on its binding affinity in the guinea pig brain membranes were studied. Mitragynine pseudoindoxyl inhibited the electrically stimulated ileum and mouse vas deferens contractions in a concentration-dependent manner. In the ileum, the effective concentration is in an nM order, being nearly equivalent to reported concentrations of the μ-opioid receptor agonist [D-Ala 2, Met-Phe 4, Gly-ol 5] enkephalin (DAMGO), and is 100- and 20-fold smaller than those of mitragynine and morphine, respectively. In the vas deferens, it is 35-fold smaller than that of morphine. The inhibitory action of mitragynine pseudoindoxyl in the ileum was antagonized by the non-selective opioid receptor antagonist naloxone and the μ-receptor antagonist naloxonazine. It was also antagonized by the δ-receptor antagonist naltrindole in the vas deferens. Mitragynine pseudoindoxyl showed a similar binding affinity to DAMGO and naltrindole at μ- and δ-receptors, respectively. However, the affinity at κ-receptors was negligible. The present study demonstrates that mitragynine pseudoindoxyl, a novel alkaloid structurally different from other opioid agonists, acts on opioid receptors, leading to a potent inhibition of electrically stimulated contraction in the ileum through the μ-receptors and in mouse vas deferens through δ-receptors.

Fentanyl increases dopamine release in rat nucleus accumbens: involvement of mesolimbic mu- and delta-2-opioid receptors

The effects of the μ-receptor agonist fentanyl on extracellular levels of dopamine in rat nucleus accumbens were studied in awake animals by in vivo brain microdialysis. Fentanyl dose-dependently increased the levels of dopamine when given intravenously (μg/kg) or via a microdialysis probe placed into the ventral tegmental area or the nucleus accumbens (nmol). The effect of fentanyl given into the nucleus accumbens was blocked by systemic administration of the non-selective opioid receptor antagonist naloxone and by accumbens administration of d-Phe-Cys-Tyr-D-Trp-Orn-Thr-Phe-Thr-NH 2 (nmol), a μ-opioid receptor antagonist, and naltrindole (nmol), a non-selective δ-opioid receptor antagonist, in a dose-dependent manner. The δ 2-opioid receptor antagonist, naltriben (nmol), also blocked the effects of fentanyl, whereas the δ 1-opioid receptor antagonist, (E)-7-benzylidenenaltrexone (nmol), was ineffective. When marginally effective doses of d-Phe-Cys-Tyr-D-Trp-Orn-Thr-Phe-Thr-NH 2 and naltriben were given simultaneously, the effect of fentanyl was nearly fully blocked; the pretreatment itself had no effect. Administration of the μ-opioid receptor agonist [ d-Ala 2, N-Me-Phe 4,Gly 5-ol]-enkephalin (nmol), the δ 1-opioid receptor agonist [ d-Pen 2,5]-enkephalin (nmol) or the δ 2-opioid receptor agonist [ d-Ala 2,Glu 4]-deltorphin (nmol) into the nucleus accumbens enhanced the amount of accumbal dopamine. This study provides evidence that not only activation of δ 1- and δ 2-opioid receptors, but also activation of μ-opioid receptors in the nucleus accumbens increases the release of accumbal dopamine in freely moving rats. We suggest that the effect of intra-accumbens administration of fentanyl upon accumbal release of dopamine is either due to the simultaneous activation of μ-opioid receptors and δ 2-opioid receptors or due to activation of μ-opioid receptors that interact with δ 2-opioid receptors in a complex manner.

The nociceptin receptor-mediated inhibition of the rat rostral ventrolateral medulla neurons in vitro

The recently available antagonist selective for novel nociceptin receptor, [Phe 1ψ(CH 2-NH)Gly 2]NC(1–13)NH 2, was utilized in this study to verify specificity of nociceptin receptor in mediating the nociceptin-induced inhibition of electrical activity of neurons in the rostral ventrolateral medulla of rat brain slices. Perfusion of nociceptin (10 nM) considerably reduced spontaneously firing frequency of the medullary neurons. Co-perfusion of [Phe 1ψ(CH 2–NH)Gly 2]NC(1–13)NH 2 (10 μM) completely blocked the nociceptin-induced depression of the neuronal activity. Blocking effect of [Phe 1ψ(CH 2–NH)Gly 2]NC(1–13)NH 2 was concentration-dependent. However, the nociceptin antagonist did not modify basal, and opioid peptide enkephalin-depressed, firing rates of the neurons. In contrast to [Phe 1ψ(CH 2-NH)Gly 2]NC(1–13)NH 2, the non-selective opioid receptor antagonist naloxone (10 μM) failed to affect the nociceptin inhibition even though naloxone at a lower concentration (1 μM) readily blocked enkephalin-induced depression of the neuronal activity. These data indicate that the nociceptin-induced inhibition of spontaneous discharge of the rostral ventrolateral medulla neurons is specifically mediated by [Phe 1ψ(CH 2–NH)Gly 2]NC(1–13)NH 2-sensitive nociceptin receptors distinct from typical naloxone-sensitive opioid receptors.

Time window for the contribution of the delta-opioid receptor to cardioprotection by ischemic preconditioning in the rat heart.

The present study aimed to examine (1) whether the role of the opioid receptor in ischemic preconditioning (PC) is consistent regardless of the duration of ischemic insult and (2) which opioid receptor subtype contributes to PC. In the first series of experiments, the effects of PC, a nonselective opioid receptor antagonist (naloxone), and their combination on the infarct size after various durations of ischemia were assessed. In anesthetized, open-chest rats, the coronary artery was occluded for 20, 30, or 40 minutes to induce infarction and was reperfused for 3 hours, PC was performed with two cycles of 5-minute ischemia followed by 5-minute reperfusion before the sustained ischemia. At 25 minutes before the ischemia, naloxone was injected alone or in combination with subsequent PC. Infarct size was determined by tetrazolium staining and was expressed as a percentage of the risk area size (%IS/RA). In the second series of experiments, the effects of a delta-receptor-selective antagonist, naltrindole (NTI), and a kappa-receptor selective antagonist, nor-binaltrophimine (nor-BNI), on PC before 30-minute coronary occlusion were assessed. In untreated controls, %IS/RA was 53.1 +/- 3.2 after 20 minutes, 67.9 +/- 3.9 after 30 minutes, and 87.8 +/- 2.0 after 40 minutes of ischemia, respectively. PC significantly reduced %IS/RA after 20, 30, and 40 minutes of ischemia to 3.1 +/- 0.8, 12.8 +/- 1.1, and 42.1 +/- 4.3, respectively (P < 0.05 vs. each control). Naloxone (6 mg/kg) partially attenuated the protection afforded by PC when the sustained ischemia was 30 minutes (%IS/RA = 27.4 +/- 4.5; P < 0.05 vs. PC), but this inhibitory effect of naloxone was not detected when the duration of the ischemia was 20 or 40 minutes. NTI (10 mg/kg) also attenuated infarct size limitation by PC after 30 minutes of ischemia (%IS/RA = 25.6 +/- 3.7), but nor-BNI (10 mg/kg) failed to modify infarct size limitation by PC (%IS/RA = 13.3 +/- 3.2). The present results suggest that activation of the opioid delta-receptor partly contributes to preconditioning against infarction in the rat and that there may be a time window (at around 30 minutes after the onset of ischemia) for this opioid receptor-mediated protective mechanism.

The portacaval-shunted rat: a new model for the study of the mechanisms controlling voluntary ethanol consumption and ethanol preference?

Portacaval anastomosis (PCA) is a surgical procedure whereby blood from the portal vein is shunted into the inferior vena cava. PCA in the rat results in a significant increase (from 0.77 +/- 0.26 to 3.51 +/- 0.37 g of ethanol/kg/day) in voluntary ethanol consumption in a free-choice paradigm between water and 5% ethanol solution. After PCA surgery, increased voluntary ethanol consumption starts abruptly at 6 to 7 days and is maintained for > 28 weeks. Voluntary ethanol consumption in rats after PCA results in blood ethanol levels up to 158 mg%. After PCA, the ethanol preference ratio (defined as the percentage of total fluid intake constituted by ethanol) increased from 19 +/- 2% to 78 +/- 2% (P < 0.001). Administration of the nonselective opioid receptor antagonist naloxone (5 mg/kg, sc) resulted in a significant 6-fold attenuation of voluntary ethanol consumption by rats with PCA, an effect that was not mediated by an effect on locomotor activity. These findings, together with previous reports of widespread alterations of the mu- and delta-opioid receptors in the brain after PCA, suggest that increased voluntary ethanol consumption and ethanol preference in PCA rats may result from activation of the endogenous opioid system. Preliminary studies suggest that rats with PCA manifest behavioral signs consistent with the development of dependence. The portacaval-shunted rat may provide a useful preparation for the study of mechanisms, in particular those involving the liver, implicated in the development of increased voluntary ethanol consumption and ethanol preference.

Cholecystokinin-Induced Antinociception is Not Blocked by CCK-A or CCK-B Receptor Antagonists

Williams, C. L., G. C. Rosenfeld and T. F. Burks. Cholecystokinin-induced antinociception is not blocked by CCK-A or CCK-B receptor antagonists. Peptides 18(3) 409–414, 1997.—To determine the relative importance of CCK-A, CCK-B, and opioid receptors in mediating the antinociceptive actions of cholecystokinin, we evaluated the actions of selective agonists and antagonists in the mouse hot plate assay. The agonists used were CCK (1–30 nmol ICV), a CCK-A receptor agonist (SNF9019; 0.3–10 nmol ICV), and a CCK-B receptor agonist (SNF9007; 0.3–10 nmol ICV). The antagonists used were the CCK-A receptor antagonist, L364,718 (12.5 nmol ICV), CCK-B receptor antagonist, L365,260 (2.5–25 nmol ICV), and the nonselective opioid receptor antagonist naloxone (1 mg/kg SC). CCK and its receptor-selective analogues, SNF9019 and SNF9007, resulted in antinociception that was blocked by naloxone, but was not antagonized by L364,718 or L365,260. In contrast, in positive control experiments, the inhibitory effects of CCK, SNF9019, and SNF9007 on gastrointestinal propulsion in mice were antagonized by identical ICV doses of L364,718 and L365,260. We conclude that centrally administered CCK produces antinociception in the mouse hot plate assay via opioid receptors, but independent of CCK-A or CCK-B receptors. It is necessary to speculate that other CCK receptors, not antagonized by currently available selective antagonists, may exist.