Selective Mu-receptor Agonist Research Articles

Functional selectivity of EM-2 analogs at the mu-opioid receptor.

The mu opioid receptor agonists are the most efficacious pain controlling agents but their use is accompanied by severe side effects. More recent developments indicate that some ligands can differentially activate receptor downstream pathways, possibly allowing for dissociation of analgesia mediated through the G protein from the opioid-related side effects mediated by β-arrestin pathway. In an effort to identify such biased ligands, here we present a series of thirteen endomorphin-2 (EM-2) analogs with modifications in positions 1, 2, and/or 3. All obtained analogs behaved as mu receptor selective agonists in calcium mobilization assay carried out on cells expressing opioid receptors and chimeric G proteins. A Bioluminescence Resonance Energy Transfer (BRET) approach was employed to determine the ability of analogs to promote the interaction of the mu opioid receptor with G protein or β-arrestin 2. Nearly half of the developed analogs showed strong bias towards G protein, in addition four compounds were nearly inactive towards β-arrestin 2 recruitment while blocking the propensity of EM-2 to evoke mu-β-arrestin 2 interaction. The data presented here contribute to our understanding of EM-2 interaction with the mu opioid receptor and of the transductional propagation of the signal. In addition, the generation of potent and selective mu receptor agonists strongly biased towards G protein provides the scientific community with novel tools to investigate the in vivo consequences of biased agonism at this receptor.

Evidence for functional delta-opiate receptors in the rat intestine.

The selective delta-opiate agonists D-Ser2, Leu5, Thr6-enkephalin (DSLET), D-Ala2, D-Leu5-enkephalin and D-Pen2, D-Pen5-enkephalin caused inhibition of the cholinergic contraction produced by transmural stimulation of the rat isolated jejunum. Dynorphin A, which is an agonist at both kappa- and delta-opioid receptors also inhibited the cholinergic contraction, as did leu- and met-enkephalin. The selective mu-receptor agonist D-Ala2-NMe-Phe4, Gly-ol5-enkephalin was the least potent of all peptides tested. In general, the order of potency of the peptides was similar to that reported for the delta-receptor-rich mouse vas deferens with potency values similar to those recorded previously for the hamster vas deferens. The selective delta-opioid antagonist naltrindole caused parallel shifts to the concentration-effect curve to DSLET giving a pA2 value of 10.15. The results indicate that the previously identified delta-binding sites in the rat jejunum may correspond to functional delta-opiate receptors involved in attenuating acetylcholine release.

Shell/core differences in mu- and delta-opioid receptor modulation of dopamine efflux in nucleus accumbens

The mu- and delta-opioid receptors located at the terminal level in nucleus accumbens are involved in the opiate modulation of dopamine release in this brain area. However, recent studies suggest that the effects of opioid drugs on the core subregion of nucleus accumbens may completely differ from those observed in the shell. We used in vivo microdialysis to simultaneously apply selective mu- and delta-opioid receptor agonists and to measure extracellular levels of dopamine in three subregions of the accumbens, namely shell, core, and the transition zone between them. The regional analysis of these subregions of the accumbens demonstrated that basal levels of dopamine and its metabolites were higher in the core, and decreased from this subregion to the shell. Retrodialysis application to the core of both the selective mu-receptor agonist ([ d-Ala 2, N-Me-Phe 4, Gly 5-ol]-enkephalin (DAMGO) (1 μmol/L)) and of the selective delta-opioid receptor agonist ([ d-Pen 2, d-Pen 5]-enkephalin (DPDPE) (50 nmol/L)) increased the dialysate levels of dopamine. However, the application of these drugs to the shell significantly reduced the dopamine levels in this subregion. Local application of the same doses of these drugs in the transition zone between the shell and the core did not significantly affect the dopamine levels in dialysates. These results suggest that the opioid circuits modulating dopaminergic activity in the shell could differ from those in the core of the nucleus accumbens.

Melatonin attenuates the development of antinociceptive tolerance to delta-, but not to mu-opioid receptor agonist in mice

The effects of melatonin (Mel) on the development of tolerance to antinociceptive actions induced by mu- and delta-opioid receptor agonists were determined in male Kunming mice. In the mouse tail-flick tests, selective mu and delta receptor agonists were repeatedly administered to mice supraspinally (intracerebroventricularly, i.c.v.) in the absence or presence of melatonin. Administration of endomorphin-1 (EM-1, a mu-opioid receptor agonist) or deltorphin I (del I, a delta-opioid receptor agonist) twice daily for 4 days produced antinociceptive tolerance compared with vehicle controls. Co-administration with melatonin prevented the development of tolerance to deltorphin I analgesia, and this effect was dose dependent. However, melatonin did not affect the development of antinociceptive tolerance to endomorphin-1. Additionally, the attenuation of deltorphin I tolerance by melatonin was reduced by chronic treatment with luzindole (luz), a selective antagonist on the MT 2 receptor subtype. Taken together, these data suggest that melatonin interferes with the neural mechanisms involved in the development of tolerance to delta-opioid agonist analgesia via its receptor.

Intubating Conditions and Hemodynamic Changes during Tracheal Intubation with Propofol Followed by Remifentanil without the Use of Muscle Relaxant

Background: Studies have shown that tracheal intubation can be facilitated safely and effectively after induction of anesthesia with hypnotic and opioid without employing any muscle relaxants. Remifentanil is a new, ultra-short acting, selective mu-receptor agonist. The goal of our present study was to evaluate the appropriate induction dose of remifentanil with propofol for tracheal intubation without employing any muscle relaxants. Methods: We have assessed intubating conditions in three groups of 45 ASA I or II Patients. Each group received intravenous dosage of 2 mg/kg of propofol with remifentanil 1/kg (Group I), 2/kg (Group II) or 3/kg (Group III), respectively. No muscle relaxant was administered. Intubating conditions were assessed based on jaw relaxation, vocal cord position, vocal cord movement, airway reaction, and movement of limbs. Intubating conditions, mean arterial blood pressure and heart rate were measured just before induction (T00, baseline), before intubation (T0), 1 minute after intubation (T1), 2 minutes after intubation (T2) and 3 minutes after intubation (T3). Results: Intubating conditions were clinically acceptable in 26.7, 86.7 and 93.3% of patients, after 1, 2 or 3/kg administration of remifentanil, respectively. Clinically acceptable intubating conditions were significantly more in Group II and III when compared with Group I (P ＜ 0.05). Both HR (heart rate) and MAP (mean arterial pressure) were significantly lower in Group III when compared with Group I before intubation (T0), 1 minute after intubation (T1), 2 minutes after intubation (T2) and 3 minutes after intubation (T3) (P ＜ 0.05). Conclusions: We conclude that 2-3/kg of remifentanil may reliably provide 'good' to 'excellent' conditions for tracheal intubation when administered after administration of 2 mg/kg propofol. The conditions for intubation were significantly better in patients receiving 2 mg/kg of propofol followed by 2-3/kg of remifentanil than those receiving 1/kg of remifentanil.

Tolerance develops in spinal cord, but not in brain with chronic [Dmt1]DALDA treatment

Previously, we reported that H-2',6'-dimethyltyrosine [Dmt(1)]-d-Arg-Phe-Lys-NH(2) (DALDA), an analogue of the naturally occurring opioid peptide dermorphin, is a highly potent and selective mu receptor agonist with low cross-tolerance to morphine. In the present study, we investigated the effect of treating mice chronically with [Dmt(1)]DALDA. The AD(50) of [Dmt(1)]DALDA (s.c.) increased eight-fold in animals given this drug chronically; in contrast, the AD(50) increased two-fold in mice chronically treated with morphine. The AD(50) of morphine (s.c.) in these [Dmt(1)]DALDA-treated animals was increased more than 120 times, while that of the more selective mu agonist [d-Ala(2)-MePhe(4)-Gly-ol(5)]enkephalin (DAMGO) given intrathecally was increased more than 240 times. However, the AD(50) of DAMGO given intracerebroventricularly was essentially the same in animals treated chronically with [Dmt(1)]DALDA as in naive animals. The dose of naloxone required to precipitate withdrawal in [Dmt(1)]DALDA-treated animals was 20 times lower than that in morphine-tolerant animals. Using real-time quantitative PCR, we found that expression of the mu opioid receptor, delta opioid receptor, preproenkephalin and preprodynorphin genes was upregulated in the brain by [Dmt(1)]DALDA treatment. No significant changes in expression of opioid receptor or opioid peptide genes were detected in the spinal cord of [Dmt(1)]DALDA-treated mice, nor in the brain or spinal cord of morphine-treated mice. We conclude that a high degree of tolerance to [Dmt(1)]DALDA develops in the spinal cord but not brain, and cannot be accounted for by changes in expression of opioid receptors or opioid peptides in these tissues.

Activation of mu opioid receptors in the ventrolateral periaqueductal gray inhibits reflex micturition in anesthetized rats

This study tested the hypothesis that morphine and other opiates cause urinary retention by activating mu opioid receptors in the midbrain periaqueductal gray (PAG) region. Selective mu, delta and kappa receptor agonists were microinjected into the PAG of urethane-anesthetized rats and the amplitude and incidence of bladder contractions were recorded during continuous saline infusion. Arterial pressure was monitored through a femoral artery catheter. Microinjection of the mu receptor agonist DAMGO into the ventrolateral PAG (vlPAG) suppressed volume-evoked bladder contractions completely. Bladder contractions ceased within 5 min of DAMGO injection and remained essentially undetectable for the rest of the 20 min recording period. Microinjection of the delta receptor agonist DPDPE into the vlPAG did not significantly affect either the amplitude of bladder contractions or the time interval separating contractions. The kappa receptor agonist U-69593 caused no discernible change in amplitude but increased the interval between bladder contractions significantly. Microinjection of DAMGO, DPDPE or U-69593 into the lateral or dorsolateral PAG columns was ineffective. DAMGO injection into the vlPAG increased arterial pressure whereas DPDPE and U-69593 produced a small but significant depressor response. These data support the hypothesis that mu and kappa receptors in the vlPAG participate in the micturition reflex.

Spinal nociceptive mechanisms: Mu- but Not Delta- Opioid Receptor activation depresses nociceptive transmission in the isolated spinal cord of neonatal rats

The roles and functions of opioid receptors, especially delta-receptors in mediating nociceptive transmission in infant animals are not clear. The present study was designed to investigate the involvements of delta- and mu-opioid receptors in spinal nociceptive transmission in the neonatal rat. Immature rats (7-14 days old) of either sex were deeply anaesthetized with urethane. Hemisected spinal cords with attached dorsal and ventral roots were isolated and perfused continuously with artificial cerebrospinal fluid at room temperature. Dorsal root evoked-population ventral root potentials (DR-VRP) in the ventral root were recorded in response to supermaximal electrical stimulation of the corresponding dorsal root (segments L4-5). Supermaximal electrical shock to a dorsal root evoked a typical DR-VRP, consisting of an initial monosynaptic compound action potential of motoneurones (MSP) superimposed on a population excitatory postsynaptic potential (e.p.s.p.) followed by a prolonged population e.p.s.p. , which lasts for tens of seconds. Bath perfusion of SNC-80, a highly selective non-peptide delta-opioid receptor agonist, at 1(n=2), 10 (n=8) and 100 (n=3) mM did not depress the DR-VRP. In contrast, DAMGO, a highly selective mu-receptor agonist, dose-dependently (3, 10, 30, 100 nM) depressed the nociceptive reflex activity. At 100 nM (n=4), the evoked long during DR-VRPs were depressed by 59 + 4% (area under the curve) of pre-drug control. Steady state depression was reached at 10-15 min following DAMGO perfusion and the depressant effect was reversed by washout. Unlike mu-opioid receptors, the present data indicate that delta-opioid receptors are not actively involved in spinal nociceptive related reflex activity in the immature rat.

The effects of endomorphin-1 on conditioned defeat in Syrian hamsters (Mesocricetus auratus)

The present study examined the effect of endomorphin-1 (EM1), an endogenous opioid with a high affinity for the mu opiate receptor, on conditioned defeat. Conditioned defeat is a phenomenon in which hamsters that have been defeated subsequently fail to exhibit normal territorial aggression and instead display submissive/defensive behaviors even when paired with a non-aggressive intruder. In experiment 1, animals were placed in the home cage of a larger resident for 15 min and were defeated. After 24 h, animals received a 3-μl injection of EM1 (0.0, 0.3, 3.0, or 10 nmol) into the left lateral cerebral ventricle 5 min before a smaller non-aggressive intruder was placed in the home cage of the experimental animal. In experiment 2, animals were infused with EM1 immediately after the initial defeat and were paired with a non-aggressive intruder 24 h later as in experiment 1. EM1 reduced the duration of submissive/defensive behavior in experiment 1 (P<0.05) but not in experiment 2 (P>0.05). These data support the hypothesis that the highly selective mu receptor agonist endomorphin-1 modulates the expression of conditioned defeat, but provides no support for the hypothesis that endomorphin-1 modulates the consolidation of conditioned defeat.

Inhibition of calcium channels by opioid- and adenosine-receptor agonists in neurons of the nucleus accumbens.

The pharmacological effects of opioid- and adenosine-receptor agonists on neural signalling were investigated by measuring drug actions on barium current flowing through calcium channels in acutely-dissociated neurons of the rat nucleus accumbens (NAc). Under whole-cell voltage clamp, opioids acted via mu, but not delta or kappa, receptors to partially inhibit barium current. Mean inhibition was 35+/-2% (+/-s.e.mean, n = 33) for methionine-enkephalin and 37+/-1% (n = 65) for the selective mu receptor agonist DAMGO, both measured at saturating agonist concentrations in neurons with diameter > or = 20 microm. EC(50) for DAMGO was 100 nM. Perfusion of naloxone reversed the current inhibition by DAMGO. Adenosine also partially inhibited barium current in these neurons. Mean inhibition was 28+/-2% (n = 29) for adenosine and 33+/-3% (n = 27) for the selective A1 receptor agonist N(6)CPA, both at saturating concentrations in neurons with diameter > or = 20 microm. EC(50) for N(6)CPA was 34 nM. Adenosine inhibition was reversed by perfusion of an A1 receptor antagonist, 8-cyclopentyl-1,3-dipropylxanthine, while the selective A2A receptor agonist, CGS 21680, had no effect. Inhibition by opioids and adenosine was mutually occlusive, suggesting a converging pathway onto calcium channels. These actions involved a G-protein-coupled mechanism, as demonstrated by the partial relief of inhibition by strong depolarization and by the application of N-ethylmaleimide or GTP-gamma-S. Inhibition of barium current by opioids had their greatest effect in large neurons, that is, in presumed interneurons. In contrast, opioid inhibition in neurons with diameter < or = 15 microm was 11+/-2% (n = 26) for methionine-enkephalin and 11+/-4% (n = 17) for DAMGO, both measured at saturating agonist concentrations. Adenosine inhibition in neurons with diameter < or = 15 microm was 22+/-5% (n = 9). These results implicate the interneurons as a locus for the modulation of the excitability of projection neurons in the NAc during the processes of addiction and withdrawal.

Nociceptin/orphanin FQ exacerbates excitotoxic white-matter lesions in the murine neonatal brain.

Intracerebral administration of the excitotoxin ibotenate to newborn mice induces white-matter lesions, mimicking brain lesions that occur in human preterm infants. Nociceptin (NC), also called orphanin FQ, is the endogenous ligand of the opioid receptor-like 1 (ORL1) receptor and does not bind classical high-affinity opioid receptors. In the present study, administration of NC exacerbated ibotenate-induced white-matter lesions while coadministration of ibotenate with either of two NC antagonists reduced excitotoxic white-matter lesions by up to 64%. Neither ibotenate plus endomorphin I (a selective mu receptor agonist), nor ibotenate plus naloxone (a classical opioid receptor antagonist) modulated the excitotoxic lesion. Pretreatment with antisense oligonucleotides targeting the NC precursor peptide mRNA significantly reduced ibotenate-induced white-matter damage. Finally, high doses of fentanyl, which stimulates both classical mu-opioid receptors and ORL1, exacerbated excitotoxic white-matter lesion. This toxic effect was blocked by inhibiting ORL1 but not classical opioid receptors. Together, these findings show that endogenous or exogenous stimulation of the ORL1 receptor can be neurotoxic and that blocking NC signaling protects the white matter against excitotoxic challenge. These data point to potential new avenues for neuroprotection in human preterm infants at high risk of brain lesions.

Cornea-responsive medullary dorsal horn neurons: modulation by local opioids and projections to thalamus and brain stem.

Previously, it was determined that microinjection of morphine into the caudal portion of subnucleus caudalis mimicked the facilitatory effects of intravenous morphine on cornea-responsive neurons recorded at the subnucleus interpolaris/caudalis (Vi/Vc) transition region. The aim of the present study was to determine the opioid receptor subtype(s) that mediate modulation of corneal units and to determine whether opioid drugs affected unique classes of units. Pulses of CO(2) gas applied to the cornea were used to excite neurons at the Vi/Vc ("rostral" neurons) and the caudalis/upper cervical spinal cord transition region (Vc/C1, "caudal" neurons) in barbiturate-anesthetized male rats. Microinjection of morphine sulfate (2.9-4.8 nmol) or the selective mu receptor agonist D-Ala, N-Me-Phe, Gly-ol-enkephalin (DAMGO; 1.8-15.0 pmol) into the caudal transition region enhanced the response in 7 of 27 (26%) rostral units to CO(2) pulses and depressed that of 10 units (37%). Microinjection of a selective delta ([D-Pen(2,5)] (DPDPE); 24-30 pmol) or kappa receptor agonist (U50488; 1.8-30.0 pmol) into the caudal transition region did not affect the CO(2)-evoked responses of rostral units. Caudal units were inhibited by local DAMGO or DPDPE but were not affected by U50,488H. The effects of DAMGO and DPDPE were reversed by naloxone (0.2 mg/kg iv). Intravenous morphine altered the CO(2)-evoked activity in a direction opposite to that of local DAMGO in 3 of 15 units, in the same direction as local DAMGO but with greater magnitude in 4 units, and in the same direction with equal magnitude as local DAMGO in 8 units. CO(2)-responsive rostral and caudal units projected to either the thalamic posterior nucleus/zona incerta region (PO/ZI) or the superior salivatory/facial nucleus region (SSN/VII). However, rostral units not responsive to CO(2) pulses projected only to SSN/VII and caudal units not responsive to CO(2) projected only to PO/ZI. It was concluded that the circuitry for opioid analgesia in corneal pain involves multiple sites of action: inhibition of neurons at the caudal transition region, by intersubnuclear connections to modulate rostral units, and by supraspinal sites. Local administration of opioid agonists modulated all classes of corneal units. Corneal stimulus modality was predictive of efferent projection status for rostral and caudal units to sensory thalamus and reflex areas of the brain stem.

Involvement of opioid receptors in N-methyl-D-aspartate-induced arterial hypertension in periaqueductal gray matter.

Arterial hypertension induced by microinjections of N-methyl-D-aspartate (NMDA) (2 nmol/rat) into the midbrain periaqueductal gray matter was used to assess the involvement of opioid receptors (mu, delta and kappa) in modulating pressor periaqueductal gray neurons. Groups (n = 5-8) of urethane-anaesthetised rats received, 5 min before NMDA, microinjections of selective opioid receptor antagonists in the periaqueductal gray area and arterial blood pressure was monitored. Pretreatments with naloxone (5 nmol/rat), a non selective mu receptor antagonist, or naltrindole hydrochloride (5 nmol/rat), a selective delta receptor antagonist, significantly (P < 0.05) decreased by 31% and 37%, respectively, NMDA-induced hypertension. The latency for the maximum increase of NMDA-induced hypertension was also significantly (P < 0.05) increased with naloxone. Pretreatment with nor-binaltorphimine (5 nmol/rat), a selective kappa receptor antagonist, only increased the latency of NMDA-induced hypertension. Each opioid antagonist failed per se to alter arterial blood pressure. Microinjection of morphine (13 nmol/rat), a non selective mu receptor agonist, significantly decreased (P < 0.05) by 57.5% NMDA-induced arterial hypertension and this effect was antagonised by naloxone. Combined pretreatments in the periaqueductal gray area with naloxone and the GABAA antagonist bicuculline (2.5 nmol/rat; 5 min before naloxone) antagonised the effect of naloxone on NMDA-induced hypertension. In contrast, bicuculline significantly (P < 0.05) potentiated morphine-induced decrease of NMDA hypertension.(ABSTRACT TRUNCATED AT 250 WORDS)

Voltage-dependent effects of opioid peptides on hippocampal CA3 pyramidal neurons in vitro

Opioid peptides, and especially the dynorphins, have been localized to several circuits in the CA3 hippocampal region, yet electrophysiological studies often find mixed effects of opiates on the excitability of CA3 neurons. Reasoning that these mixed effects might involve voltage-dependent actions, we tested the effect of several opiates on CA3 pyramidal neurons using single-electrode voltage-clamp recording in a slice preparation of rat hippocampus. In most CA3 neurons, the voltage-dependent K+ current known as the M-current (IM) was uniquely sensitive to the opioid peptides, with the direction of response dependent upon the opiate type and concentration. Thus, an opiate selective for kappa receptors, U-50,488H, significantly augmented IM. The kappa-selective agonists dynorphin A and dynorphin B, which exist in mossy fiber afferents to CA3 pyramidal neurons, also markedly augmented IM at low concentrations (20-100 nM). By contrast, dynorphin A at higher concentrations (1-1.5 microM) often reduced IM. Similarly, several opiates [e.g., D-Ala2,D-Leu5-enkephalin: (DADL), [D-Pen2,5]-enkephalin (DPDPE)] known to act on the delta receptor subtypes reduced the M-current, with partial reversal of this effect by naloxone. Neither the selective mu-receptor agonist [D-Ala2, NMe-Phe4, Gly-ol]-enkephalin (DAMGO) nor the nonopioid fragment of dynorphin, des-Tyr-dynorphin, consistently altered IM. These opiate effects on IM were accompanied by changes in conductance and holding current consistent with their respective effects on IM. Dynorphin A did not measurably affect the Q-current, a conductance known to contribute to inward rectification in hippocampal pyramidal neurons. The opiate effects on IM were not altered by pretreatment with Cs+ (which blocks IQ) or Ca2+ channel blockers. The opposing effects of the dynorphins (both A and B) and DADL on IM were antagonized by naloxone (1-3 microM), and the dynorphin-induced augmentations of IM were usually reversed by the kappa receptor antagonist norbinaltorphimine. These results suggest that the opiates can have opposing effects on the same voltage-dependent K+ channel type (the M channel) in the rat CA3 pyramidal neuron, with the direction of the response depending on which receptor subtype is activated. These data not only help explain the mixed effects of opiates seen in other studies, but also suggest a potential postsynaptic function for the endogenous opiates contained in the CA3 mossy fibers.

Effects of met-enkephalin on GABAergic spontaneous miniature IPSPs in organotypic slice cultures of the rat hippocampus

The action of met-enkephalin on GABAergic spontaneous miniature IPSPs (smIPSPs) was investigated in CA1 neurons from hippocampal slice cultures. In the presence of excitatory amino acid blockers (2,3-dihydroxy-6-nitro-7-sulphamoyl-benzo(F)quinoxaline, DL-2-amino-5-phosphonovaleric acid) and TTX, a continuous high-frequency bombardment of smIPSPs was recorded. The smIPSPs were blocked by the GABAA antagonist bicuculline. The occurrence of the smIPSPs was random and their amplitude distribution was skewed toward larger smIPSPs. Met-enkephalin (10-20 microM) reversibly reduced the frequency and changed the amplitude distribution of the smIPSPs. The proportion of "large" smIPSPs was reduced, but a loss of "small" smIPSPs also contributed to the reduction in smIPSP frequency. The selective mu-receptor agonist DAGO mimicked the effect of met-enkephalin and naloxone blocked the effect of DAGO. Hyperpolarization of the neuronal membranes, produced by reducing the extracellular K+ concentration, did not reduce the frequency of the smIPSPs, nor did it block the effect of DAGO. Reduction of the extracellular concentration of Ca2+ combined with an increase in extracellular Mg2+ or the addition of Cd2+ did not reduce the smIPSP frequency, nor did it block the effect of DAGO. These results suggest that CA1 pyramidal cells of hippocampal organotypic cultures are tonically inhibited by spontaneous release of GABA, through a release mechanism that is independent of propagated sodium action potentials. Met-enkephalin and DAGO reduce the tonic inhibition by reducing the frequency of the smIPSPs, through a direct action on the presynaptic GABAergic terminals. The effect was probably not mediated by hyperpolarization of the presynaptic membrane or by modulation of presynaptic Ca2+ currents.

The kappa opioid agonist U-50,488H antagonizes respiratory effects of mu opioid receptor agonists in conscious rats.

The interactive effects of mu and kappa opioid receptor agonists on respiratory function were investigated following their i.c.v. injection into conscious rats. The highly selective mu receptor agonist [D-Ala2,N-Methyl-Phe4,Gly-ol] enkephalin (DAMGO; 1.2-10 nmol) and the relatively selective mu agonist morphine (20 and 30 nmol) significantly decreased arterial pH and PO2, and increased arterial PCO2 and blood pressure. Morphine and a low dose of DAMGO (1.2 nmol) also significantly elevated respiratory rate. Heart rate was decreased by DAMGO and, depending upon dose, was either decreased (20 nmol) or increased (30 nmol) by morphine. The selective kappa opioid agonist U-50,488H (200 nmol i.c.v.), which by itself had no significant effect on either respiration or cardiovascular function, dose-dependently antagonized the acidotic, hypoxemic and hypercapnic effects of both DAMGO (2.5 nmol) and morphine (30 nmol). Furthermore, these mu antagonistic properties of U-50,488H were blocked completely after pretreatment with 25 nmol of the highly selective kappa opioid antagonist nor-binaltorphimine. These results indicate that the antagonism of mu opioid respiratory depressant effects by U-50,488H is kappa opioid receptor mediated.

Roles of delta and mu opioid receptors in mediating the effects of enkephalins on avoidance conditioning.

The effects on one-way active avoidance conditioning of pre-training, systemic administration of the selective mu-receptor agonist [D-Ala2,N-Me-Phe4, Gly-ol]enkephalin (DAGO), and the selective mu-receptor antagonist (D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2 (CTOP), were determined in Swiss-Webster mice. A low dose of DAGO (0.92 micrograms/kg) moderately enhanced avoidance acquisition, whereas a 100 micrograms/kg dose of CTOP more dramatically impaired acquisition. However, the avoidance-enhancing dose of DAGO significantly increased locomotor activity as measured in a separate group of mice in the avoidance chamber, and the avoidance-impairing dose of CTOP significantly decreased activity. Under these same training conditions, earlier studies (Schulteis et al. 1988; Schulteis and Martinez 1990) demonstrated that enkephalins impaired avoidance learning, and selective delta-receptor antagonists such as ICI 174,864 enhanced learning; in contrast to the present study, both of these effects were dissociated from performance effects such as alterations in locomotor activity. Taken together, the results suggested that the effects of enkephalins were mediated by the delta-, but not mu-, class of opioid receptor.

Intrathecal pertussis toxin treatment attenuates opioid antinociception and reduces high-affinity state of opioid receptors.

The effect of pertussis toxin on opioid antinociception was studied in rats. Intrathecal injection of a single dose of pertussis toxin reduced the antinociceptive effect of PL017, a highly selective mu-opioid agonist, in a dose- and time-dependent manner. The maximal effective dose of pertussis toxin was 1 microgram, and the maximal effect was seen at day 3. The effect of the toxin lasted for 2 weeks, and the antinociceptive response recovered partially at the third week. The dose-response curves of the antinociceptive effect of PL017 were shifted to the right with increasing doses of pertussis toxin. Three days after pertussis injection, receptor-binding activity of membranes in the lumbosacral segment of spinal cords decreased to 70% of control as assayed by 125I-FK33824, a highly selective mu-receptor agonist. In experiments using [3H]naloxone as the radiolabeled ligand, displacement curves of FK33824 were shifted to the right after pertussis toxin treatment. The shift also was dose and time dependent. Scatchard analysis of binding data showed that, after pertussis toxin treatment, there was no significant change in the total number of binding sites, but a class of low-affinity binding sites appeared in addition to the high-affinity sites. When spinal membranes were washed in Na+ (100 mM) and guanosine diphosphate (100 microM) and binding was assayed in the presence of Mg2+ (5 mM), all the mu-receptors were in the high-affinity state in control membranes. After the pertussis toxin treatment, the ratio of low-affinity sites to high-affinity sites increased.(ABSTRACT TRUNCATED AT 250 WORDS)

Bilateral injections of a selective mu-receptor agonist (morphiceptin) into the medial preoptic nucleus produces a marked delay in the initiation of sexual behavior in the male rat.

We examined the putative functional role of the medial preoptic nucleus mu-receptor population in the expression of male copulatory behavior in sexually vigorous Long-Evans rats. In the first experiment, three doses of morphiceptin (10, 500, and 1000 ng) a selective mu-receptor agonist injected bilaterally into the medial preoptic nucleus, produced a marked delay in the initiation of male copulatory behavior compared to saline injected controls. These injections significantly lengthened intromission and mount latencies while having no appreciable effect on any other parameter of male copulatory behavior. In a separate experiment, the transient inhibition of the expression of male copulatory behavior was completely abolished following pretreatment of naloxone 20 minutes prior to bilateral injections of morphiceptin (1000 ng) into the medial preoptic nucleus. Collectively, these results suggest that the delay in the initiation of copulation that is commonly observed following peripheral or central injections of opioids is mediated at least in part by mu receptors located within the medial preoptic nucleus.

Effect of opioid peptides on circular muscle of canine duodenum.

1. The effects of opioid peptides on inhibitory transmission in the circular muscle layer of canine duodenum were investigated in vitro using simultaneous mechanical and intracellular electrical recording techniques. 2. Exogenously added [Met5]enkephalin, [Leu5]enkephalin and dynorphin (1-13) decreased the amplitude of non-adrenergic, non-cholinergic inhibitory junction potentials (IJPs) evoked by transmural nerve stimulation. 3. A selective delta-receptor agonist, DPDPE ([D-Pen2, D-Pen5]enkephalin), and a selective mu-receptor agonist, PL017 (Try-Pro-NMePhe-D-Pro-NH2), decreased the amplitude of IJPs whereas a selective kappa-receptor agonist, U-50,488H ([trans-3,4-dichloro-N-methyl-N-(2-91-pyrolidinyl)-cyclohexyl]- benzeneacetamide methanesulphonate), in large doses (1 microM) produced only a small reduction. 4. A selective delta-receptor antagonist, ICI-174,864, blocked the effect of DPDPE but not that of PL017 suggesting the presence of distinct delta- and mu-opioid receptors on inhibitory motor nerves. 5. Exogenously added dynorphin (1-13) decreased the amplitude of IJPs. delta-Opioid receptors appeared to be involved because ICI-174,864, a selective delta-antagonist, blocked the inhibitory effect of exogenously added dynorphin (1-13). 6. The inhibitory effect of the opioid peptides was still observed in preparations of circular muscle devoid of myenteric and submucosal plexuses, indicating that the site of action was on inhibitory motor nerve fibres located within the circular muscle layer and not on neuronal cell bodies in the enteric plexuses. 7. It was concluded that in the canine small intestine, opioid peptides could modulate release of inhibitory transmitter(s) at or near nerve terminals of inhibitory motor nerves innervating circular muscle cells.