Dopaminergic modulation of kappa opioid-mediated ultrasonic vocalization, antinociception, and locomotor activity in the preweanling rat.

The purpose of this study was to determine whether dopamine (DA) systems modulate kappa opioid-mediated ultrasonic vocalizations (USVs), antinociception, and locomotion in young rats. Seventeen-day-old rats were injected with the kappa agonist U-50,488 (0.0-7.5 mg/kg) and saline, the D?-like receptor agonist R(-)-propylnorapomorphine (NPA; 0. 1 or 1.0 mg/kg), the indirect DA agonist cocaine (10 or 20 mg/kg), or the DA antagonist flupenthixol (0.25 or 0.5 mg/kg). USVs and locomotion were measured for 6 min, with antinociception being assessed with a tail-flick test. Kappa receptor stimulation produced analgesia and increased USVs and locomotion. U-50,488-induced analgesia was potentiated by NPA, whereas U-50,488-induced USVs were attenuated by both DA agonists. NPA and flupenthixol depressed U-50,488's locomotor effects. These results show that DA systems interact with kappa opioid systems to modulate USVs, antinociception, and locomotion in preweanling rats. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

No evidence for presynaptic opioid receptors on cholinergic, but presence of kappa-receptors on dopaminergic neurons in the rabbit caudate nucleus: involvement of endogenous opioids

The effects of various opioid receptor agonists and antagonists were studied in rabbit caudate nucleus slices preincubated with either [3H]dopamine or [3H]choline, superfused with medium (containing in most experiments the D2 receptor antagonist domperidone) and subjected to electrical field stimulation. The stimulation-evoked [3H]overflow from slices prelabeled with [3H]dopamine (evoked [3H]dopamine release) was significantly reduced by preferential kappa-opioid receptor agonists, like U-50,488 H, but not by mu- or delta-opioid receptor selective drugs. Opioid receptor antagonists shifted the concentration/response curve of U-50,488 H to the right (apparent pA2-value of the kappa-selective antagonist nor-binaltorphimine: 10.1) and enhanced the evoked dopamine release in the presence of a mixture of peptidase inhibitors. On the other hand, the [3H]overflow from rabbit caudate nucleus slices prelabeled with [3H]choline (evoked acetylcholine release) remained almost unaffected by any opioid receptor agonist, as long as the presynaptic D2 heteroreceptor was blocked with domperidone: in the absence of domperidone, U-50,488 H exhibited facilitatory effects. For comparison, the effects of the preferential delta-opioid receptor agonist DPDPE was also studied in slices of the rat striatum, where it clearly inhibited the evoked acetylcholine release. From our data we conclude that in the rabbit caudate nucleus the evoked dopamine release is inhibited by both exogenous and endogenous opioids via presynaptic kappa-opioid receptors, whereas the evoked release of acetylcholine is not, or only indirectly (via released dopamine) affected by opioids.     

Cross-tolerance to acute administration of mu and kappa opioid agonists at the spinal cord level in the rat

Development of tolerance and cross-tolerance after acute administration of the mu agonist morphine and the kappa agonist U-50,488H was assessed in rats, through recording of a C-fiber-evoked spinal nociceptive reflex. Rats rendered tolerant to morphine (a single dose of 1 mg/kg i.p.) showed, after a 5-hour period, tolerance to morphine and cross-tolerance to the kappa-opioid receptor agonist U-50,488H, as revealed by depressed C-reflex responsiveness. In contrast, pretreatment with U-50,488H (a single dose of 1 mg/kg i.p.) rendered tolerant the rats to U-50,488H, but the animals did not develop cross-tolerance to morphine. Results indicate that acute administration of mu and kappa ligands leads to development of unidirectional cross-tolerance in rat spinal cord. This points to limitations in using alternated mu and kappa opioid agonists to bypass the problem of development of opioid tolerance in chronic pain complaints.     

Effect of 7-nitroindazole on tolerance to morphine, U-50,488H and [D-Pen2, D-Pen5] enkephalin in mice

The effects of 7-nitroindazole (7-NI), an inhibitor of the neuronal nitric oxide synthase (nNOS) which does not increase blood pressure, on tolerance to the antinociceptive activity of mu-(morphine), kappa-(U-50,488H) and delta-([D-Pen2, D-Pen5]enkephalin, DPDPE) opioid receptor agonists were determined in mice. Male Swiss-Webster mice were made tolerant by twice daily injections of morphine (20 mg/kg, s.c.), U-50,488H (25 mg/kg, i.p.) or DPDPE (20 micrograms/mouse, i.c.v.) for 4 days. When tested on day 5, tolerance to their antinociceptive activity was evidenced by decreased response in chronic drug treated mice in comparison to vehicle-injected mice. Concurrent administration of 7-NI (20, 40 or 80 mg/kg, i.p.) with DPDPE did not modify the development of tolerance to the antinociceptive action of DPDPE. However, 7-NI (40 or 80 mg/kg, i.p.) inhibited the development of tolerance to the antinociceptive activity of morphine and U-50,488H but the lower dose of 7-NI (20 mg/kg, i.p.) was not effective. Chronic administration of 7-NI by itself did not modify the acute response to morphine, U-50,488H or DPDPE. It is concluded that a specific inhibitor of nNOS can inhibit tolerance to the antinociceptive activity of mu- and kappa- but not of delta-opioid receptor agonists in mice.     

Establishing morphine and U-50,488H as discriminative stimuli in a three-choice assay with pigeons.

Pigeons were trained in a 3-choice assay to discriminate among injections of 5.6 mg/kg U-50,488H, 5.6 mg/kg morphine, and vehicle solution. In dose-response tests, subjects rarely responded on the U-50,488H-appropriate key when morphine was administered or on the morphine-appropriate key when they received U-50,488H. A high dose of naltrexone (1.0 mg/kg) completely blocked the morphine cue but failed to block completely the U-50,488H cue. In generalization tests, d-amphetamine primarily engendered saline-appropriate responding. Ethylketazocine produced mixed results, in that moderate doses produced responding on both the morphine- and U-50,488H-appropriate keys, but 3.2 mg/kg engendered primarily morphine-appropriate responding. These results demonstrate the feasibility, but not necessarily the value, of 3-choice discrimination procedures involving mu and kappa agonists and vehicle. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Ligand binding profiles of U-69, 593-sensitive and-insensitive sites in human cerebral cortex membranes: evidence of kappa opioid receptors heterogeneity

Receptor binding studies were performed to characterize the properties of subtypes of kappa opioid receptors in membrane preparations of human cerebral cortex. [3H]U69,593 ([3H]U69), a selective kappa 1-agonist, and [3H]diprenorphine ([3H]DIP), a non-selective opioid antagonist, in the presence of 1 microM each of DAMGO, DPDPE and U-69 to block mu-, delta-, and kappa 1-sites, labeled single population of binding sites, respectively. [3H]U-69 binding sites (KD = 3.8 +/- 0.2 nM, Bmax = 6.3 +/- 0.2 fmol/mg protein) had a binding profile that correspond to kappa 1-receptor. That is, dynorphin A (1-13) (Dyn A), bremazocine (BZC), U50,488H (U50), (-)ethylketocyclazocine (EKC) and nor-binaltorphimine (nor-BNI) bound to this site with high affinities. [3H]DIP labeled binding sites (Kd = 7.3 +/- 0.2 nM, Bmax = 102 +/- 9 fmol/mg protein) that were not sensitive to U-50, but to BZC, EKC and nor-BNI. These results indicate that kappa 1 and Kappa 2 opioid receptors exist in human cerebral cortex with different ligand binding profiles.     

Uncertainty analysis for wheelchair propulsion dynamics

Nefiracetam is undergoing preclinical and clinical tests as a cognition-enhancing drug in Alzheimer's disease (AD). Nicotinic cholinergic receptors are lost in AD, and nicotinic as well as muscarinic cholinergic receptors are involved in the modulation of eyeblink conditioning. Experiments were carried out using young rabbits to examine the effect of nefiracetam on cholinergic antagonists to nicotinic (mecamylamine) and muscarinic (scopolamine) receptors. Rabbits were tested for 15 days in the 750 ms delay eyeblink classical conditioning paradigm in paired and explicitly unpaired conditions. Nefiracetam at a dose of 15 mg/kg significantly ameliorated the effects of 0.5 mg/kg mecamylamine, and nefiracetam at a dose of 10 mg/kg significantly ameliorated the effect of 1.5 mg/kg scopolamine. The vehicle alone and nefiracetam alone groups performed similarly to the groups treated with mecamylamine or scopolamine and nefiracetam. Reversal by nefiracetam of a nicotinic as well as a muscarinic cholinergic antagonist indicates that the drug may affect deficits specific to AD.     

Genesis and significance of hypertension in autosomal dominant polycystic kidney disease

The aim of the present study was to determine whether U-50,488H and U-62,066E, kappa-opioid receptor agonists cause a neuroprotective action against hypoxia/hypoglycemia-induced reduction in 2-deoxyglucose (2-DG) uptake of hippocampal slices from U-50,488H-tolerant rats. Both U-50,488H and U-62,066E exhibited an attenuating effect on hypoxia/hypoglycemia-induced reduction in 2-DG uptake of hippocampal slices. Hypoxia/hypoglycemia-induced deficit of 2-DG uptake was prevented by cotreatment with naloxone, an opioid receptor antagonist, but potentiated by cotreatment with morphine, a mu-opioid receptor agonist. Chronic administration of U-50,488H resulted in the development of tolerance to the analgesic effect as well as the neuroprotective effect whereas this treatment affected neither basal- nor hypoxia/hypoglycemia-induced decreases in 2-DG uptake. Chronic administration of U-50,488H did not modify naloxone-induced attenuation of 2-DG uptake deficit but slightly potentiated the morphine-induced exacerbation. These findings suggest that the tolerance to kappa-opioid receptors does not affect the mu-opioid receptor-mediated neuroprotective or neurotoxic action.     

Expression of interstitial collagenase (matrix metalloproteinase-1) is related to the activity of human endometriotic lesions

The effects of the competitive antagonist of the N-methyl-D-aspartate (NMDA) receptor, LY235959, were determined on the analgesic and hypothermic effects as well as on the development of tolerance to these effects of U-50,488H, a kappa-opioid receptor agonist in mice and rats. In the mouse, a single injection of LY235959 given 10 min prior to U-50,488H did not modify the analgesic action of the latter. Similarly, chronic administration of LY235959 twice a day for 4 days did not modify U-50,488H-induced analgesia in mice. Repeated pretreatment of mice with LY235959 dose-dependently attenuated the development of tolerance to the analgesic actions of U-50,488H. In the rat, LY235959 by itself produced a significant analgesia and prior treatment of rats with LY235959 enhanced the analgesic action of U-50,488H. Similar effects were seen with the hypothermic action. Pretreatment of rats with LY235959 attenuated the development of tolerance to the analgesic but not to the hypothermic action of U-50,488H. These results provide evidence that LY235959 produces differential actions on nociception and thermic responses by itself and when given acutely with U-50,488H in mice and rats. However, when the animals are pretreated with LY235959, similar inhibitory effects are observed on the development of tolerance to the analgesic action of U-50,488H in both the species. These studies demonstrate an involvement of the NMDA receptor in the development of kappa-opioid tolerance and suggest that the biochemical consequences of an opioid's interaction with the opioid receptor are not the only factors that contribute to the acute and chronic actions of opioid analgesic drugs.     

Effects of kappa opioid agonists on the discriminative stimulus effects of cocaine in rhesus monkeys.

The study examined the effects of the kappa opioid agonists U50,488 and ethylketocyclazocine (EKC) on cocaine discrimination in rhesus monkeys trained to discriminate cocaine (0.4 mg/kg) from saline. Administration of U50,488 and EKC alone produced primarily saline-appropriate responding. Kappa agonist pretreatments produced variable effects on cocaine discrimination across monkeys, attenuating the discriminative stimulus effects of cocaine in some monkeys, but either having no effect on cocaine discrimination or enhancing the discriminative stimulus effects of cocaine in other monkeys. The effects of kappa agonists on cocaine discrimination were reversed by pretreatment with the opioid antagonist naloxone (1.0 mg/kg). These results indicate that kappa agonists do not consistently block the discriminative stimulus effects of cocaine in rhesus monkeys. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Effects of scopolamine infusions into the anterior and posterior cingulate on passive avoidance and water maze navigation

We examined the role of anterior and posterior cingulate cortical muscarinic receptors in water maze spatial learning and passive avoidance. Pretraining and posttraining trial scopolamine (a mixed a muscarinic acetylcholine antagonist) infusions into the anterior cingulate cortex dose dependently (3 no effect; 10 and 30 micrograms impaired) impaired passive avoidance performance. Pretesting infusion into the anterior cingulate had no effect on passive avoidance. Scopolamine infusion into the anterior cingulate did not impair spatial navigation. On the contrary, scopolamine (3 micrograms no effect, 10 and 30 micrograms impaired) infusions into the posterior cingulate before daily training trials impaired water maze navigation to a hidden platform, but did not affect navigation to a visible escape platform or passive avoidance. Posttraining and pretesting infusion into the posterior cingulate did not impair WM spatial navigation. The present results indicate that muscarinic acetylcholine receptor antagonist may modulate passive avoidance performance via cholinergic receptors located in anterior cingulate cortex and the ability to develop a spatial navigation strategy via muscarinic receptors located in posterior cingulate.     

Majonoside-R2, a major constituent of Vietnamese ginseng, attenuates opioid-induced antinociception

The effects of majonoside-R2 on antinociceptive responses caused by the mu-opioid receptor agonist morphine and the selective kappa-opioid receptor agonist U-50, 488H were examined by the tail-pinch test in mice. Intraperitoneal (IP) or intracerebroventricular (ICV) injection of majonoside-R2 (3.1-6.2 mg/kg, IP or 5-10 micrograms/mouse, ICV) and diazepam (0.1-0.5 mg/kg, IP or 0.5-1.0 microgram/mouse, ICV), as well as an opioid receptor antagonist naloxone (2 mg/kg, IP or 5 micrograms/mouse, ICV), dose-dependently attenuated the antinociception caused by subcutaneously administered morphine and U-50,488H. Moreover, when co-administered ICV or intrathecally (IT) with morphine (4 micrograms/mouse) or U-50,488H (60 micrograms/mouse), majonoside-R2 (5-20 micrograms/mouse) also exhibited antagonism against the antinociceptive action of these opioid receptor agonists in the tail-pinch test. The inhibitory effects of majonoside-R2 (10 micrograms/mouse, ICV) and diazepam (1 microgram/mouse, ICV) were reversed by flumazenil (2.5 micrograms/mouse, ICV), a selective benzodiazepine receptor antagonist, and picrotoxin (0.25 microgram/mouse, ICV), a GABA-gated chloride channel blocker. These results suggest that majonoside-R2 attenuates the opioid-induced antinociception by acting at the spinal and supraspinal levels, and that the GABAA receptor complex at the supraspinal level is involved in the effect of ICV administered majonoside-R2.     

Differential effects of cholinergic blockade on performance of rats in the water tank navigation task and in a radial water maze.

Tested the disruptive effect of cholinergic blockade under conditions in which either the working memory or the spatial mapping requirements of the behavioral task were emphasized. In Exp I, 13 male hooded rats were trained in an 8-arm radial water maze to asymptotic performance. When delays of 5, 10, 20, and 40 min were inserted between Choice 4 and Choice 5, incidence of errors in Choices 5–8 increased after pretrial (20 min) intraperitoneal scopolamine (0.2 mg/kg) faster than under control conditions and approached chance level with the 40-min delay. Scopolamine after Choice 4 or pretrial methylscopolamine was ineffective. In Exp II, 30 Ss were trained in a Morris water tank. Acquisition was impaired by pretrial injection (20 min) of 0.1 and 0.2 mg/kg scopolamine, but a higher dose (1.0 mg/kg) was required to impair overtrained performance. In a working memory version of the navigation task, scopolamine administered 20 min before the 1st trial deteriorated retention tested 40 min later at a dose of 1.0 but not at 0.4 and 0.2 mg/kg. It is concluded that the disruptive effect of scopolamine is proportional to the demands on the working memory component of the task, whereas the use of an overtrained mapping strategy is relatively resistant to cholinergic blockade. (35 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Physostigmine, but not 3,4-diaminopyridine, improves radial maze performance in memory-impaired rats

The results of some studies suggest that 3,4-diaminopyridine (3,4-DAP), a drug that enhances the release of acetylcholine, may improve memory. The present study examined the ability of 3,4-DAP to reverse the memory impairment produced by scopolamine and the ability of 3,4-DAP and physostigmine to reverse the memory impairment produced by quinolinic acid lesions of the nucleus basalis magnocellularis (nbm) in rats. Mnemonic functioning was assessed with the use of a partially baited eight-arm radial maze. Entries into arms that were never baited were defined as reference memory errors; entries into baited arms from which the food already had been eaten were defined as working memory errors. In Experiment 1, 0.1 mg/kg scopolamine produced a significant increase in working and reference memory errors. Various doses of 3,4-DAP had no significant ameliorative effect on the mnemonic deficit. In Experiment 2, cholinergic function was impaired using a unilateral intra-nbm injection of quinolinic acid (120 nmol in 1.0 microliter). These lesions reduced the levels of the cholinergic marker, choline acetyltransferase, in the cortex by more than 40%. Results showed that the nbm lesion animals were significantly more impaired on the working than reference memory component of the task. Physostigmine (0.01, 0.05, 0.10, 0.20, 0.50 mg/kg) dose-dependently decreased the number of working but not reference memory errors. 3,4-DAP (10(-8), 10(-6), 10(-4), 10(-2), 10(0) mg/kg) had no reliable effect. It was concluded that physostigmine, but not 3,4-DAP, ameliorates memory impairments following decreases in cholinergic function.     

Possible mechanisms of salt-induced hypertension in Dahl salt-sensitive rats

The effects of acute and chronic administration of cocaine on the antinociception and tolerance to the antinociceptive actions of mu-(morphine), kappa-(U-50,488H), and delta-([D-Pen2,D-Pen5]enkephalin; DPDPE), opioid receptor agonists were determined in male Swiss-Webster mice. Intraperitoneal injection of 40 mg/kg of cocaine by itself produced weak antinociceptive response as measured by the tail-fick test but the lower doses were ineffective. Administration of morphine (10 mg/kg, SC), U-50,488H (25 mg/kg, IP) or DPDPE (10 microg/mouse, ICV) produced antinociception in mice. Cocaine (20 mg/kg) potentiated the antinociceptive action of morphine and DPDPE but had no effect on U-50,488H-induced antinociception. Administration of morphine (20 mg/kg, SC), U-50,488H (25 mg/kg, IP) or DPDPE (20 microg/mouse, ICV) twice a day for 4 days resulted in the development of tolerance to their antinociceptive actions. Tolerance to the antinociceptive actions of morphine and U-50,488H was inhibited by concurrent treatment with 20 or 40 mg/kg doses of cocaine; however, tolerance to the antinociceptive action of DPDPE was not modified by cocaine. It is concluded that cocaine selectively potentiates the antinociceptive action of mu- and delta- but not of the kappa-opioid receptor agonist. On the other hand, cocaine inhibits the development of tolerance to the antinociceptive actions of mu- and kappa- but not of delta-opioid receptor agonists in mice.     

Opioid antagonist profile of SC nor-binaltorphimine in the formalin paw assay

The antinociceptive effects of mu and kappa agonists were examined after the systemic administration of the opioid antagonists nor-binaltorphimine (nor-BNI) and naloxone in the late response or tonic nociceptive phase of the mouse formalin assay. Initially, SC morphine (ED50, 0.97 mg/kg), racemic U-50488H (ED50, 0.79 mg/kg), (-)U-50488 (ED50, 0.41 mg/kg), and another agonist PD 117,302 (ED50, 0.28 mg/kg) were found to produce graded increases in the level of antinociception as measured by this procedure; naloxone, administered immediately before morphine and U-50488H, antagonized their antinociceptive actions. The effects of morphine and U-50488H then were evaluated 10 min to 96 h after the administration of nor-BNI. Subcutaneous nor-BNI at 30.0 mg/kg, but not at 3.0 or 10.0 mg/kg, attenuated the antinociceptive effects of morphine and U-50488H when the interval separating nor-BNI and the agonists was kept constant at 1 h. Time-course analysis of the effects of combinations of nor-BNI with morphine led to irregular findings: 10.0 mg/kg of nor-BNI lessened the effects of morphine (2.0 mg/kg) if the dosing interval was 10 min, whereas 30.0 mg/kg of nor-BNI attenuated the effects of morphine (2.0 mg/kg) if the dosing interval was 1 or 4 h; 10.0 mg/kg of nor-BNI also diminished the antinociceptive effects of U-50488H (1.7 mg/kg) only if the interval spacing the two drugs was 24 h. In comparison, a threefold higher dose of nor-BNI (30.0 mg/kg) reduced the effects of U-50488H (1.7 mg/kg) if the interval was 1 h or more. In these latter experiments, the antagonist effects of SC nor-BNI (30.0 mg/kg) were evident up to 96 h posttreatment. These results show that the mu opioid antagonist activity of nor-BNI is variable and that the kappa opioid antagonist selectivity of nor-BNI is a function of dose and treatment interval and is long-lasting even after systemic administration.     

p-Hydroxybenzyl alcohol attenuates learning deficits in the inhibitory avoidance task: involvement of serotonergic and dopaminergic systems

p-Hydroxybenzyl alcohol (HBA), an aglycone of gastrodin, is an active ingredient of Gastrodia elata BLUME. In this study, we investigated the action of HBA on acquisition of an inhibitory avoidance response in rats and used piracetam as a positive control. The results indicated that scopolamine, a cholinergic receptor antagonist, injected before training impaired retention. HBA did not attenuate the scopolamine-induced impairment, but piracetam did. p-Chloroamphetamine, a serotonin releaser, injected before training impaired retention. HBA at 5 mg/kg and piracetam at 100 mg/kg could counteract the p-chloroamphetamine-induced deficit. Apomorphine, a dopaminergic receptor agonist, also impaired retention. HBA at 5 mg/kg and piracetam at 300 mg/kg could ameliorate the apomorphine-induced amnesia. The above results indicated that HBA, different from piracetam, can attenuate impairments induced by p-chloroamphetamine and apomorphine, but had no effect on impairment induced by scopolamine in an inhibitory avoidance task in rats. Such findings suggest that HBA may act through suppressing dopaminergic and serotonergic activities and thus improves learning.     

Characterization, expression, and immunohistochemical localization of 5 alpha-reductase in human skin

The effect of the mu opioid agonist DAGO, delta opioid agonist DPDPE and kappa opioid agonist U50,488H on 3H-dopamine (3H-DA) uptake was studied in synaptosomes prepared from rat striatum and nucleus accumbens. Over the range of concentrations tested (1 nM-10 microM) DAGO and DPDPE were devoid of effects on 3H-DA uptake in the striatum and the nucleus acumbens. In contrast, U50,488H significantly decreased 3H-DA uptake in both structures. The inhibition of uptake induced by the kappa agonist was not reversed in the presence of the opiate antagonists naloxone (10 microM) or nor-binaltorphimine (0.1 microM). Dynorphin A (1-13) also induced a significant reduction in 3H-DA uptake in both structures at the concentrations of 10 and 30 microM. This inhibitory effect was not reversed by naloxone (10 microM). These data suggest that kappa opioid agonists modulate dopamine uptake in the striatum and the nucleus accumbens and their effects may not be due to an activation of opioid receptors.     

Opioid control of the fetal stretch response: Implications for the first suckling episode.

On their 1st experience with milk, fetal rats express a stretch response that is similar to the postnatal behavior exhibited by infant rats at the nipple. Fetuses also possess a functional opioid system that is activated by prenatal milk exposure. The opioid receptor antagonist naloxone and the specific kappa antagonist nor-binaltorphimine blocked the stretch response and prevented the increase in rearlimb activity that is typically induced by milk. The mu antagonist β-funaltrexamine blocked the stretch while permitting the expression of rearlimb activity. The kappa agonist U50,488 promoted rearlimb activity in the absence of milk, whereas the mu agonist [{d}-Ala–2,NMe-Phe–4,Gly–5-ol]-enkephalin (DAMGO) exerted little influence on fetal behavior. Fetuses pretreated with U50,488 stretched to nonmilk stimuli (saline or lemon), but fetuses pretreated with DAMGO did not. Opioid activation is part of a chain of events that culminates in the fetal stretch response and may be important in promoting milk ingestion during the newborn's 1st suckling episode. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

The kappa opioid receptor expressed on the mouse R1.1 thymoma cell line down-regulates without desensitizing during chronic opioid exposure

The R1.1 mouse thymoma cell line expresses a single class of kappa opioid receptors that is negatively coupled to adenylyl cyclase through a Bordetella pertussis toxin-sensitive inhibitory guanine nucleotide-binding protein. The aim of the present study was to determine whether chronic opioid treatment of R1.1 cells altered either the binding properties or the functional response associated with the kappa opioid receptor. Culturing of R1.1 cells with the kappa-selective agonist (trans)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl] benzeneacetamide methane-sulfonate hydrate (U50,488) for 3 hr and longer, followed by extensive washing of R1.1 cell membranes, produced a concentration- and time-dependent reduction in the binding of the kappa-selective ligand (5 alpha,7 alpha,8 beta)-(-)-N-methyl-N-(7-(1-pyrrolidinyl)-1- oxaspiro(4,5)dec-8-yl) benzeneacetamide ([3H]U69,593). Culturing of R1.1 cells with 100 nM U50,488 for 24 hr produced approximately a 50% reduction in the Bmax value for [3H]U69,593 and [3H]naloxone binding. In contrast to the reduction in binding, there was no change in the inhibition of adenylyl cyclase activity by (-)-U50,488. To determine whether kappa opioid receptor function was maintained by spare receptors after agonist-induced down-regulation, membranes from untreated R1.1 cells were incubated with 400 nM of the irreversible opioid antagonist beta-chlornaltrexamine (beta-CNA) followed by extensive washing. beta-CNA produced a 50% reduction in the [3H]U69,593 binding and a 6-fold increase in the IC50 value for (-)-U50,488 inhibition of adenylyl cyclase activity, with no change in the maximal inhibition of cyclic AMP levels.(ABSTRACT TRUNCATED AT 250 WORDS)