Blockade effect of mu and kappa opioid antagonists on the anti-nociception induced by intra-periaqueductal grey injection of oxytocin in rats

Intra-periaqueductal grey (PAG) injection of 1 nmol of oxytocin induced significant increases in hindpaw withdrawal latency (HWL) to thermal and mechanical stimulation in rats. The anti-nociceptive effect of oxytocin was attenuated significantly by subsequent intra-PAG injection of the mu opioid antagonist beta-funaltrexamine (beta-FNA) and the kappa opioid antagonist nor-binaltorphimine (nor-BNI), but not by the delta antagonist naltrindole. The results demonstrated that mu and kappa opioid receptors, not delta receptors, were involved in the oxytocin-induced anti-nociception in PAG of rats.     

Further evidence for the interaction of mu- and delta-opioid receptors in the antinociceptive effects of the dual inhibitor of enkephalin catabolism,RB101(S). A spinal c-Fos protein study in the rat under carrageenin inflammation

We have previously shown that RB101, a dual inhibitor of enkephalin-degrading enzymes, decreased carrageenin-evoked c-Fos protein expression at the spinal cord level in awake rats. Moreover, we have also shown that c-Fos expression is a useful marker of the possible direct or indirect interactions between neural pathways, such as opioid and cholecystokinin systems. We now investigated the respective roles of the three main types of opioid receptors (mu, delta, or kappa) and their possible interactions, in the depressive effects of RB101 in inflammatory nociceptive conditions induced by intraplantar carrageenin (6 mg/150 microl of saline). We used beta-funaltrexamine (beta-FNA), naltrindole (NTI), and nor-binaltorphimine (BNI) as specific antagonists for mu, delta- and kappa-opioid receptors, respectively. c-Fos protein-immunoreactivity (c-Fos-IR) was evaluated as the number of c-Fos-IR nuclei in the lumbar spinal cord 90 min after carrageenin. c-Fos-IR nuclei were preferentially located in the superficial (I-II) and deep (V-VI) laminae of segments L4-L5 (areas containing numerous neurons responding exclusively, or not, to nociceptive stimuli). RB101(S) (30 mg/kg, i.v.) significantly reduced the total number of carrageenin-evoked c-Fos-IR nuclei (30% reduction, P<0.01). This effect was completely blocked by beta-FNA (10 mg/kg, i.v.), or NTI (1 mg/kg, i.v.). In contrast, BNI (2.5 mg/kg, i.v.) did not reverse the reducing effects of RB101(S) on carrageenin-evoked c-Fos protein expression. These results suggest that functional interactions occur between mu- and delta-opioid receptors in enkephalin-induced antinociceptive effects.     

Role of mu, delta and kappa opioid receptors in ethanol-reinforced operant responding in infant rats

We recently observed that naloxone, a non-specific opioid antagonist, attenuated operant responding to ethanol in infant rats. Through the use of an operant conditioning technique, we aimed to analyze the specific participation of mu, delta, and kappa opioid receptors on ethanol reinforcement during the second postnatal week. In Experiment 1, infant rats (PDs 14-17) were trained to obtain 5, 7.5, 10, or 15% ethanol, by operant nose-poking. Experiment 2 tested blood ethanol levels (BELs) attained by operant behavior. In Experiment 3, at PDs 16-18, rats received CTOP (mu antagonist: 0.1 or 1.0mg/kg), naltrindole (delta antagonist: 1.0 or 5.0mg/kg) or saline before training. In Experiment 4, rats received nor-binaltorphimine (kappa antagonist: 10.0 or 30.0mg/kg, a single injection after completion of PD 15 operant training), spiradoline mesylate (kappa agonist: 1.0 or 5.0mg/kg; at PDs 16-18) or saline (PDs 16-18), before the conditioning. Experiments 5 and 6 assessed possible side effects of opioid drugs in locomotor activity (LA) and conditioned taste aversion (CTA). Ethanol at 7.5 and 10% promoted the highest levels of operant responding. BELs were 12-15mg/dl. In Experiment 3 naltrindole (dose-response effect) and CTOP (the lowest dose) were effective in decreasing operant responding. Nor-binaltorphimine at 10.0mg/kg and spiradoline at 5.0mg/kg also blocked ethanol responding. The effects of opioid drugs on ethanol reinforcement cannot be explained by effects on LA or CTA. Even though particular aspects of each opioid receptor require further testing, a fully functional opioid system seems to be necessary for ethanol reinforcement, during early ontogeny.     

Inhibition of trigemino-hypoglossal reflex in rats by oxytocin is mediated by mu and kappa opioid receptors

Recent studies showed that oxytocin plays an important role in the modulation of pain at different levels of the central nervous system. The present study was undertaken to investigate the effect of oxytocin on trigemino-hypoglossal reflex in rats. With the experimental settings used in this study, we have demonstrated that oxytocin showed significant analgesic effect after intracerebroventricular administration in rats, as assayed by the amplitude of the retractory movements of the tongue after tooth pulp stimulation. Antinociceptive effect of oxytocin was inhibited by subsequent perfusion of cerebral ventricles with oxytocin antagonist, [deamino-Cys1-D-Tyr(OEt)2-Thr4-Orn8]-oxytocin, atosiban. An involvement of opioid system in the oxytocin-induced analgesia was studied after intracerebroventricular administration of different opioid antagonists: non-selective naloxone, mu-selective beta-funaltrexamine, delta-selective naltrindole, and kappa-selective nor-binaltorphimine. It was shown that inhibition of antinociceptive effects was mediated through mu and kappa opioid receptors, indicating that there is a synergy between oxytocin and opioid systems in transmitting and modulating pain stimuli. Co-administration of oxytocin and a mu-selective endogenous opioid ligand endomorphin-2 did not significantly increase the antinociceptive activity of endomorphin-2.     

c-Fos and peptide immunoreactivities in the central extended amygdala of morphine-dependent rats after naloxone-precipitated withdrawal

The central extended amygdala, a forebrain macrostructure, may represent a common substrate for acute drug reward and the dysphoric effects of drug withdrawal. To test its involvement during opiate withdrawal, we studied the distribution of c-Fos immunoreactive neurons, in relation to their neuropeptide content, in brain sections from morphine-dependent or naive rats, killed 90 min after naloxone or saline intraperitoneal injection. Naloxone treatment in naive rats induced a slight increase in c-Fos immunoreactivity in the central amygdaloid nucleus, the lateral bed nucleus of the stria terminalis and the interstitial nucleus of the posterior limb of the anterior commissure. In morphine-dependent rats, naloxone injection significantly increased the number of c-Fos-positive neurons in these structures as well as in the majority of the other central extended amygdala components. Double immunocytochemistry was used to determine the neurochemical nature of c-Fos-positive neurons in the central extended amygdala. Corticotropin-releasing factor- and methionine-enkephakin-immunoreactive neurons displayed c-Fos immunoreactivity in naive rats after naloxone injection, whereas only enkephalinergic neurons were found to be c-Fos positive in morphine-dependent rats after naloxone injection. The possible involvement of the corticotropin-releasing factor system during withdrawal is discussed. These results suggest that the whole central extended amygdala is activated during opiate withdrawal, with a lateral to medial decreasing gradient, and emphasize the role of peptidergic systems in this morphofunctional continuum.     

The development of tolerance to locomotor effects of morphine and the effect of various opioid receptor antagonists in rats chronically treated with morphine

Behavioural measures are considered to be highly sensitive indices of opioid withdrawal. Opioids, depending on dose and time protocols may induce both reduction and enhancement of locomotor activity and chronic opioid treatment results in tolerance and sensitisation to these effects. In the present study the locomotor activity as experimental model was used to assess the development of tolerance to subcutaneous morphine challenge at different time points following morphine withdrawal in rats exposed to gradually increasing subcutaneous doses of morphine for 11 days. Tolerance developed to the inhibitory action of morphine (10 mg/kg) was observed even 8 weeks after morphine withdrawal, while tolerance to its locomotor activity enhancing effect (3 mg/kg) was detected 18 h after withdrawal, but not 3 weeks later. In the other series of experiments the locomotor activity of animals exposed to chronic morphine treatment was tested 18 h after spontaneous or subcutaneously administrated opioid antagonists precipitated withdrawal. Spontaneous withdrawal resulted in a moderate decrease of locomotion. Both the non-selective antagonist naloxone in low, mu opioid-receptor selective doses and the delta opioid-receptor selective naltrindole induced marked reduction of locomotor activity. The results provide further evidence that both mu and delta opioid-receptors might be affected during chronic morphine treatment.     

Morphine can produce analgesia via spinal kappa opioid receptors in the absence of mu opioid receptors

Previous studies have demonstrated the virtual lack of analgesia in mu opioid receptor knockout mice after systemic administration of morphine. Thus, it has been suggested that analgesic actions of morphine are produced via the mu opioid receptor, despite its ability to bind to kappa and delta receptors in vitro. However, it is not clear whether the results of these studies reflect the effect of morphine in the spinal cord. In the present study, we report study of the analgesic actions of spinally-administered morphine and other opioid receptor agonists in mu opioid receptor knockout and wild type mice. Morphine produced a dose-dependent antinociceptive effect in the tail flick test in the knockout mice, although higher doses were needed to produce antinociception than in wild type mice. The antinociceptive effect of morphine was completely blocked by naloxone (a non-selective opioid antagonist) and nor-binaltorphimine (nor-BNI, a selective kappa-opioid receptor antagonist), but not by naltrindole (a selective delta-opioid receptor antagonist). U-50,488H (a selective kappa-opioid receptor agonist) also produced a dose-dependent antinociceptive effect in knockout mice but presented lower analgesic potency in knockout mice than in wild type mice. Analgesic effects of [d-Pen2,d-Pen5]enkephalin (DPDPE, a selective delta-opioid receptor agonist) were observed in wild type mice but abolished in knockout mice. SNC80 (a selective delta-opioid receptor agonist) was not antinociceptive even in wild type mice. The present study demonstrated that morphine can produce thermal antinociception via the kappa opioid receptor in the spinal cord in the absence of the mu opioid receptor. Lower potency of U50,488H in mu opioid receptor knockout mice suggests interaction between kappa and mu opioid receptors at the spinal level.     

Involvement of opioid receptors in electroacupuncture-produced anti-hyperalgesia in rats with peripheral inflammation

Our previous study showed that electroacupuncture (EA) significantly attenuated inflammatory hyperalgesia. It has also been reported that EA analgesia in uninjured animals is mediated by mu and delta opioid receptors at 2-15 Hz and by kappa opioid receptor at 100 Hz. Because persistent pain changes neural response to external stimulation, we hypothesized that (1) the mechanisms of EA anti-hyperalgesia may be different under conditions of persistent pain and that (2) combining EA with a sub-effective dose of morphine could enhance EA anti-hyperalgesia. Hyperalgesia, decreased paw withdrawal latency (PWL) to a noxious thermal stimulus, was induced by subcutaneously injecting complete Freund's adjuvant (CFA) into the hind paws of rats. Selective antagonists against mu (D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-ThrNH2, CTOP), delta (naltrinodole, NTI) and kappa (nor-binaltorphimine, BNI) opioid receptors were administered intrathecally 10 min before each of two EA treatments at acupoint Huantiao (GB30), one immediately post and the other 2 h post-CFA. Morphine was given (i.p.) 40 min before the second EA treatment. PWL was measured before and 2.5 and 5 h post-CFA. Both 10 and 100 Hz EA-produced anti-hyperalgesia were blocked spinally by mu- and delta- but not kappa-receptor antagonists. EA combined with a sub-threshold dose of morphine (2.5 mg/kg) enhanced anti-hyperalgesia additively (10 Hz EA) or synergistically (100 Hz EA) compared to that produced by each component alone. These results suggest selective involvement of mu and delta, but not kappa, receptors in EA-produced anti-hyperalgesia in rats. A combined EA and opioid drug protocol may provide an improved treatment strategy for inflammatory pain.     

Sigma opioid receptor; SKF-10,047 update

We found that (-)-SKF-10,047 blocks EEG and behavioral effects of morphine in the naive rat, precipitates withdrawal in morphine-dependent rats, produces physical dependence as evidenced by naloxone-induced withdrawal, and displaces [3H] dihydromorphine from brain homogenates. (+)-SKF-10,047 did not produce dependence upon chronic treatment, and it did not displace [3H] dihydromorphine from brain homogenates. Such pharmacodynamic dissociation of SKF-10,047 effects suggests an association of sigma receptors with psychotogenic, but not opioid characteristics. The latter are most likely mediated by mu or kappa receptors.     

U-50,488, a selective kappa opioid agonist: Comparison to other reputed kappa agonists

1. U-50,488 is a structurally novel, non-mu opioid. In the present experiments it was compared to the reputed kappa opioid agonists, ketazocine, ethylketocyclazocine and bremazocine as regards analgesic cross tolerance to morphine and U-50,488, antagonism of analgesia by naloxone and MR-2266 (in vivo pA2 determination), and narcotic antagonist properties (antagonism of morphine analgesia and precipitation of abstinence in morphine-dependent mice). 2. The analgesic mechanism of bremazocine was similar to that of U-50,488 but the former compound had, in addition, considerable mu-antagonist activity. The analgesic mechanisms of the ketazocines were less selective; both shared both mu and kappa agonist properties. U-50, 488, however, had no such mu agonist or antagonist effects and thus is a more selective kappa agonist. 3. This compound and its congeners may prove useful in the elucidation of the functions of kappa receptors in the central nervous system.     

Mercaptoacetate induces feeding through central opioid-mediated mechanisms in rats

The endogenous opioid system has been implicated in the mediation of food intake elicited by such regulatory challenges as glucoprivation induced by 2-deoxy-D-glucose (2DG) or food deprivation in rodents. Administration of the free fatty acid oxidation inhibitor, mercaptoacetate (MA), produces a potent short-term increase in feeding in rats, the mechanisms of which have been dissociated from that elicited by 2DG. The present study evaluated whether MA-induced feeding in rats was mediated by the endogenous opioid system through systemic administration of the general opioid antagonist, naltrexone, through central administration of either general, mu, mu(1), kappa(1) or delta opioid antagonists, and through central administration of antisense oligodeoxynucleotide (AS ODN) probes directed against specific exons of either the mu (MOR-1), kappa (KOR-1), kappa(3) (KOR-3/ORL-1) or delta (DOR-1) opioid receptor clones. MA-induced feeding was significantly and dose-dependently reduced by systemic naltrexone (0.005-5 mg/kg); these ingestive effects were quite selective since neither total, ambulatory nor stereotypic activity was affected by either MA itself or MA paired with naltrexone. MA-induced feeding was significantly reduced by central pretreatment with either naltrexone (0.1-20 microgram) or mu-selective (beta-funaltrexamine, 0.1-20 microgram), mu(1)-selective (naloxonazine, 1-20 microgram), kappa(1)-selective (nor-binaltorphamine, 0.1-20 microgram), or delta-selective (naltrindole, 1-20 microgram) opioid receptor antagonists. MA-induced feeding was significantly reduced by AS ODN probes directed against either exons 1, 2 or 3, but not exon 4 of the MOR-1 clone, exon 3, but not exons 1 or 2 of the KOR-1 clone, exons 1 or 2, but not exon 3 of the KOR-3/ORL-1 clone, and exon 1, but not exons 2 or 3 of the DOR-1 clone. These data are discussed in terms of opioid mediation of ingestive responses related to fat, and in terms of potential central sites of action at which lipoprivic ingestive responses might act.     

Effects of morphine and morphine withdrawal on adrenergic neurons of the rat rostral ventrolateral medulla

In urethane anesthetized rats, iontophoretic application of morphine or α-methylnoradrenaline (α-MNE) inhibited (80–100%) the discharges of all putative adrenergic (C1) cells of the rostral ventrolateral medulla (RVLM). The effect of morphine was blocked selectively by naloxone while that of α-MNE was blocked selectively by theα₂-adrenergic antagonist idazoxan. Putative C1 cells were inhibited (75–100%) by low i.v. doses of clonidine (10–15 μg/kg). Most cells (7/10) were also inhibited by morphine i.v. (81% at 7 mg/kg). Two cells were slightly excited at doses below 2 mg/kg and inhibited at higher doses. Three cells were excited only. All effects of morphine i.v. were reversed by naloxone (1 mg/kg, i.v.). Intravenous administration of naloxone to morphine-dependent rats increased significantly the firing rate of all putative C1 adrenergic cells (from 5.8 ± 0.9 spikes/s to 12.3 ± 1.5 spikes/s;n = 8). During withdrawal these cells could still be inhibited (80–100%) by i.v. injection of clonidine (15 μg/kg). C-Fos expression induced by naltrexone-precipitated withdrawal was examined in the brainstem of freely moving morphine-dependent rats pretreated with clonidine or saline before injection of the opioid antagonist. The locus coeruleus (LC) of the same rats was examined for comparison. Morphine withdrawal without clonidine treatment significantly increased the number of Fos-like-immunoreactive (Fos-LIR) cells in the RVLM and LC. Clonidine pretreatment (1 mg/kg, i.p.) reduced the number of withdrawal-activated Fos-LIR cells in LC by 81%. In the RVLM this reduction averaged 37% for all cell types and 48% for C1 adrenregic cells. Further, a very large proportion of RVLM neurons that expressed c-Fos during morphine withdrawal (83%) were immunoreactive forα₂A-adrenergic receptors. This study suggests that, like noradrenergic cells of the LC, C1 adrenergic neurons of the RVLM are: (i) inhibited by both opiate andα₂-adrenergic receptor agonists; and (ii) activated during naloxone-precipitated morphine withdrawal, Since C1 cells are considered essential to sympathetic tone generation, their inhibition by morphine may contribute to the hypotensive effects of this opioid agonist in non-dependent individuals. Their excitation during opiate withdrawal may also contribute to the autonomic activation that characterizes this syndrome. Finally, inhibition of C1 cells by clonidine may contribute to the clinically recognized efficacy of this drug to attenuate autonomic signs of opiate withdrawal.     

Infusion of beta-FNA into the thalamus attenuates morphine-induced c-Fos induction in the rat caudate putamen.

The medial thalamus contains mu opioid receptors and sends a glutamatergic projection to the caudate putamen (CPu) in rat. Morphine-induced c-Fos expression in the CPu has been shown to be blocked by pretreatment with antagonists to N-methyl-D-aspartate receptors, indicating the involvement of glutamate in this morphine-induced response. The importance of the glutamatergic projections from the thalamus was assessed by infusing the mu opioid receptor antagonist, beta-funaltrexamine (beta-FNA), prior to systemic morphine injection. Infusion of beta-FNA near specific medial thalamic nuclei attenuated morphine-induced c-Fos expression in the CPu.     

Dependence and withdrawal following intracerebroventricular and systemic morphine administration: functional anatomy and behavior

Regional cerebral glucose utilization (RCGU) and behavior during precipitated morphine withdrawal were studied in rats made dependent by either intracerebroventricular (i.c.v.) or subcutaneous (s.c.) administration of morphine. [14C]2-deoxy-D-glucose autoradiography revealed that RCGU increased in an anatomically related group of limbic and brainstem structures in rats that were in morphine withdrawal precipitated by naloxone administration compared to morphine-dependent controls that were not in precipitated withdrawal. Correlation of RCGU for 24 brain structures comparing i.c.v. vs s.c. morphine-treated rats was highly significant for groups in withdrawal and for controls (r values, 0.958 and 0.971, respectively). Withdrawal behaviors including autonomic signs of withdrawal, withdrawal jumping, and incidence of diarrhea were not different between the two groups in withdrawal (i.c.v. and s.c.). Weight loss during withdrawal increased (P less than 0.05) in rats made dependent by s.c. morphine administration compared to rats that received morphine by the i.c.v. route. Taken together, these results indicate that RCGU changes during morphine withdrawal result solely from effects of chronic morphine in the central nervous system, not in peripheral sites. The increased weight loss of s.c.-treated, morphine-dependent rats in withdrawal suggests an independent peripheral effect perhaps mediated by visceral opiate receptors.     

SoRI 9409, a non-peptide opioid mu receptor agonist/delta receptor antagonist, fails to stimulate [35S]-GTP-gamma-S binding at cloned opioid receptors.

Recent work suggests that opioids which combine mu agonist and delta antagonist activity may be non-addicting antinociceptive agents. SoRI 9409 (5'-(4-Chlorophenyl)-17-(cyclopropylmethyl)-6,7-didehydro-3,14-dihydroxy-4,5alpha-epoxypyrido-[2',3':6,7]morphinan) is a naltrexone-derived non-peptide ligand which demonstrates partial mu and kappa agonist activity and antagonist activity at delta receptors. Chronic administration of SoRI 9409 to mice failed to produce tolerance to its antinociceptive effect and SoRI 9409 produced less withdrawal signs than naloxone in acute and chronic morphine dependence models. To further characterize SoRI 9409 we determined its effects in the guanosine 5'-O-(3-[35S]thio)-triphosphate binding assay. SoRI 9409 demonstrated no agonist activity at cloned mu delta, or kappa receptors. Other experiments demonstrated that SoRI 9409 was a potent and selective delta antagonist (K(i) = 0.08 nM) which acted also as an antagonist at mu and kappa receptors. Its profile of activity resembled that of naltrindole (NTI). Viewed collectively, the in vitro data reported here predict that SoRI 9409 should be a mu antagonist in vivo, which is not observed. Resolving these discrepant findings will require additional research.     

Mapping of c-fos gene expression in the brain during morphine dependence and precipitated withdrawal, and phenotypic identification of the striatal neurons involved

The c-fos gene is expressed in the central nervous system in response to various neuronal stimuli. Using in situ hybridization, we examined the effects of chronic morphine treatment and withdrawal on c-fos mRNA in the rat brain, and particularly within identified striatal neurons. Morphine dependence was induced by subcutaneous implantation of two pellets of morphine for 6 days and withdrawal was precipitated by administration of naltrexone. Placebo animals and morphine-dependent rats showed a very weak c-fos mRNA expression in all the structures studied. Our study emphasized the spatial variations in c-fos mRNA expression, and also revealed a peak expression of c-fos mRNA at 1 h after naltrexone-precipitated withdrawal in the projection areas of dopaminergic neurons, noradrenergic neurons and in several regions expressing opiate receptors. Interestingly, morphine withdrawal induces c-fos mRNA expression in the two efferent populations of the striatum (i.e. striatonigral and striatopallidal neurons) both in the caudate putamen and nucleus accumbens. Moreover, the proportions of activated neurons during morphine withdrawal are different in the caudate putamen (mostly in striatopallidal neurons) and in the shell and core parts of the nucleus accumbens (mostly in striatonigral neurons). The activation of striatopallidal neurons suggests a predominant dopaminergic regulation on c-fos gene expression in the striatum during withdrawal. On the contrary, c-fos induction in striatonigral neurons during withdrawal seems to involve a more complex regulation like opioid-dopamine interactions via the mu opioid receptor and the D1 dopamine receptor coexpressed on this neuronal population or the implication of other neurotransmitter systems.     

Selective opioid receptor antagonist effects upon intake of a high-fat diet in rats

Short-term (2 h) intake of a high-fat diet in rats was significantly inhibited by intravenous (0.1-10 mg/kg: 39-67%) and central (1-5 micrograms, i.c.v.: 51%) naloxone. The irreversible mu opioid antagonist, beta-funaltrexamine (10 micrograms, i.c.v.: 37%), but not the irreversible mu 1 antagonist, naloxonazine (10 mg/kg, i.v.) inhibited intake, suggesting mu 2 receptor mediation. The delta antagonist, ICI 174864 (1-10 micrograms, i.c.v.: 41%) inhibited high-fat diet intake only at doses that also produced motor dysfunction.     

The role of opioid receptors in morphine withdrawal in the infant rat

Exposure to opiates such as morphine can lead to psychological and physical dependence in both adult and infant humans. Infant rats experience opiate withdrawal behaviors that are qualitatively different from the withdrawal behaviors displayed by adult rats. In the adult, withdrawal is largely mediated by the mu-opioid receptor. We sought to understand more about what role each opioid receptor (mu, kappa, and delta) plays in the display of the physical withdrawal in the infant rat. Beginning on postnatal day 1, infant rats were injected with morphine sulfate twice a day for 6.5 days. On the afternoon of the seventh day the infant rats were given an i.c. injection of a vehicle, the mu-opioid receptor antagonist CTOP, the kappa-opioid receptor antagonist nor-BNI, or the delta-opioid receptor antagonist naltrindole. CTOP precipitated withdrawal behaviors in the 7-day-old rat in a dose-dependent manner. Neither nor-BNI nor naltrindole induced any significant changes in the frequency of the withdrawal behaviors. These data suggest that in the infant rat control of certain behavioral withdrawal signs is modulated primarily by the mu-opioid receptor, as is the case in the adult rat.     

Delta(2)-opioid receptor mediation of morphine-induced CCK release in the frontal cortex of the freely moving rat.

Numerous pharmacological data have been accumulated in support of the existence of physiological interactions between cholecystokinin (CCK) and opioids in the central nervous system. With the aim of further characterizing these interactions, an in vivo microdialysis approach was used to directly assess the possible influence of opioids on the extracellular levels of CCK-like material (CCKLM) in the frontal cortex of the awake, freely moving rat. Systemic administration of a high dose of morphine (10 mg/kg i.p.) produced a marked increase (up to +200%) of cortical CCKLM outflow, and this effect could be completely prevented by systemic (1.5 mg/kg i.p.) as well as intracortical (10 microM) administration of the opioid receptor antagonist naloxone. The opioid receptors activated by morphine appeared to be of the delta type because the intracortical infusion of naltrindole (10 microM) also prevented the effect of morphine, whereas CTOP (10 microM), a selective mu-opioid receptor antagonist, and nor-binaltorphimine (10 microM), a selective kappa-opioid receptor antagonist, were inactive. In addition, naltriben (10 microM), which acts selectively at the delta(2) subtype, also abolished the stimulatory effect of morphine on cortical CCKLM outflow, whereas 7-benzylidenenaltrexone (10 microM), a selective delta(1)-opioid receptor antagonist (10 microM), did not alter the morphine effect. Conversely, the direct stimulation of cortical delta(2)-opioid receptors by local infusion of [D-Ala(2)] deltorphin II mimicked the stimulatory effect of systemic morphine on CCKLM outflow. These data indicate that delta(2)-opioid receptors play a key role in opioid-CCK interactions in the rat frontal cortex.