Chronic administration of U50,488H fails to produce hypothalamo-pituitary-adrenal axis tolerance in neonatal rats.

The present study investigated the effect of chronic administration of a kappa opioid receptor agonist on the function of kappa and mu opioid, serotonergic and cholinergic regulation of secretion from the hypothalamo-pituitary-adrenal axis in neonatal rats. After chronic treatment with saline or U50,488H (trans-(+/-)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl]- benzeneacetamide methane sulfonate), a kappa opioid receptor agonist and subsequent pharmacological challenge, corticosterone (CS) in serum was determined. Kappa tolerance did not develop in pups treated on postnatal days 5-9 with increasing doses of U50,488H (0.5-2.5 mg/kg). When the rats were treated with the same chronic regimen of U50,488H at different stages of development from birth through weaning, only weanling rats became tolerant to U50,488H. In the absence of measurable kappa tolerance, the responses of corticosterone in serum to morphine, quipazine, a serotonin receptor agonist and physostigmine, an inhibitor of acetylcholinesterase, were attenuated in neonatal rats, treated with U50,488H. These studies suggest that kappa tolerance is more difficult to induce in developing rats than in adults and that regulation of the function of the hypothalamo-pituitary-adrenal axis by other neurotransmitter systems is altered by treatment with kappa opioid receptor agonists, even in the apparent absence of tolerance.     

Gastric acid secretion by central injection of dynorphin A-(1-17), an endogenous ligand of kappa-opioid receptor, in urethane-anesthetized rats.

Gastric acid secretion has been proposed to be regulated by opioid receptors in the central nervous system (CNS). Previously, we reported that central injection of synthetic agonists of kappa-opioid receptors stimulated gastric acid secretion in rats, and the secretion by the agonists was inhibited by norbinaltorphimine (an antagonist of kappa-opioid receptor). In the present study, we investigated the effect of dynorphin A-(1-17), an endogenous ligand of kappa-opioid receptor on the gastric acid secretion in the perfused stomach of urethane-anesthetized rats. Injection of dynorphin A-(1-17) (0.1-1 microg per rat) into the lateral cerebroventricle (LV) stimulated the secretion in a dose-dependent manner. The effect of dynorphin A-(1-17) was almost completely inhibited by the LV injection of norbinaltorphimine (10 microg) and in vagotomized rats. Although some studies of dynorphin A-(1-17) after central injection showed non-opioid effects such as the involvement of N-methyl-D-aspartate (NMDA) receptor, the effect of dynorphin A-(1-17) was not inhibited by a selective antagonist of the NMDA receptor ((+/-)-3-(2-carboxypiperazin-4-yl)-1-propylphosphonic acid, 10 microg). The LV injection of naloxone benzoylhydrazone (a kappa3-opioid receptor agonist, 100 microg) also stimulated the secretion in norbinaltorphimine-sensitive manner. These findings showed that both an endogenous ligand dynorphin A-(1-17) and a synthetic kappa3-opioid receptor agonist stimulated gastric acid secretion via kappa-opioid receptors in the CNS of rats in vivo.     

Centrally mediated opioid induced depression of hepatic glutathione: effects of intracerebroventricular administration of mu, kappa, sigma and delta agonists

It has recently been demonstrated that morphine produces a loss of hepatocellular glutathione in mice by virtue of its action within the central nervous system. The ability of opioid receptor antagonists to abolish morphine's effect on hepatic glutathione suggests that this action is opioid-receptor mediated. The involvement of opioid receptors in this phenomenon is confirmed in the present study in mice by the ability of naltrexone, 100 micrograms administered intracerebroventricularly (i.c.v.), to completely block the decrease in hepatic glutathione induced by an i.c.v. injection of 100 micrograms of morphine. Intracerebroventricular administration of the selective mu (mu) opioid receptor agonist, (D-Ala2,N-MePhe4,Gly-ol5)enkephalin (DAGO; 25-50 micrograms), or the selective delta (delta) opioid agonist, [D-Pen2,D-Pen5]enkephalin (DPDPE; 3-50 micrograms), like morphine, produced significant decreases in hepatic glutathione 3 h after administration. The selective kappa (kappa) opioid receptor agonists, ethylketocyclazocine (1-30 micrograms) and trans-(+/-)3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl] benzeneacetamide-methane sulfonate (U50 488; 10-300 micrograms), as well as the selective sigma (sigma) opioid agonists, phencyclidine (PCP; 50-300 micrograms) and N-allylnormetazocine (SKF 10,047; 1-30 micrograms), had no effect on the concentrations of glutathione in the liver. It appears from these data that stimulation of mu- or delta-, but not kappa- or sigma-opioid receptors within the central nervous system results in a loss of hepatocellular glutathione.     

Sex-related differences in mechanical nociception and antinociception produced by mu- and kappa-opioid receptor agonists in rats

Previous studies indicate that in antinociceptive procedures employing thermal, chemical and electrical stimuli, opioids are generally more potent in male than female rodents. The purpose of the present study was to examine nociception and opioid antinociception in male and female rats using a mechanical nociceptive stimulus. Results indicated that males had a higher threshold for nociception, and in tests in which a constant pressure was applied to the hindpaw, the paw withdrawal latencies were consistently longer in males. Opioids with activity at the mu receptor, including levorphanol, morphine, dezocine, buprenorphine, butorphanol and nalbuphine, were generally more potent and/or effective in males. In contrast, sex differences were not consistently observed with the kappa-opioid receptor agonists spiradoline, (5,7,8b)-N-methyl-N[2-1(1-pyrrolidinyl),1-oxaspiro[4,5]dec-8-yl benzeneacetamide (U69593), trans-(+/-)-3,4-dichloro-N-methyl-[2-(1-pyrrolidinyl)-cyclohexyl]benzeneacetamide (U50488), enadoline, ethylketocyclazocine, and nalorphine. These findings suggest that males and females differ in their responsiveness to mechanical nociception and that sex differences in sensitivity to kappa-, but not mu-, opioid receptor agonists are specific to certain nociceptive stimulus modalities.     

The kappa agonists PD117302 and U50488 produce a biphasic effect on 24 hour food intake in the rat

The effects of the selective kappa opioid receptor agonists PD117302 [(+/-)trans-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl-]benzo-[b]- thiophene-4-acetamide] and U50488 (trans-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]- benzeneacetamide) were investigated on food intake in non-deprived rats over a 24 hr period following subcutaneous administration. At doses of 2.5 mg/kg and 5 mg/kg, both PD117302 and U50488 initially increased food intake. Between 6-24 hr, however, PD117302 (2.5-10 mg/kg) and U50488 (2.5 mg/kg and 5 mg/kg) caused a reduction in food intake. The largest doses of PD117302 and U50488 also reduced total 24 hr food intake. The reduction in food intake produced by PD117302 (5 mg/kg) was selective, since the intake of water was not affected. It is concluded that the kappa agonists PD117302 and U50488 produce a biphasic effect on 24 hr food intake, with an initial hyperphagia followed by a decrease in food intake.     

Nonspecific effects of the selective kappa-opioid receptor agonist U-50,488H on dopamine uptake and release in PC12 cells

kappa-Opioid receptor agonists decrease the levels of extracellular dopamine in vivo and in vitro. However, the mechanism(s) underlying these actions are unclear. The objective of this study was to distinguish between an effect of the selective kappa-opioid receptor agonist U-50,488H ((trans-(+/-)-3,4-dichloro-N-methyl-N-(2-[1-pyrrolidinyl]cyclohexyl)benzeneacetamide methanesulfonate) on secretion and reuptake of dopamine by PC12 cells. The data show that U-50,488H has both a modest effect to increase dopamine release and a more pronounced effect to inhibit dopamine uptake. Neither effect was sensitive to nor-binaltorphimine or naloxone, suggesting that they are not mediated through an opioid receptor.     

Opioid activity profiles indicate similarities between the nociceptin/orphanin FQ and opioid receptors

Nociceptin (orphanin FQ) is the recently discovered peptide agonist for the orphan receptor opioid receptor-like 1 (ORL1). Despite the high sequence homology between ORL1 and the opioid receptors, most opioids lack affinity for the nociceptin receptor. The affinity and functional profile of opioids possessing activity at the nociceptin receptor was determined using [3H]nociceptin and nociceptin-stimulated [35S]GTPgammaS binding. The mu-opioid receptor-selective agonist lofentanil potently and competitively displaced [3H]nociceptin at rat brain receptors (IC(50) 62 nM). Lofentanil exhibited full agonism for enhancement of [35S]GTPgammaS binding to human recombinant ORL1 receptors (EC(50) 50 nM). The related piperidines ohmefentanyl and sufentanil and the nonselective opioid receptor agonist etorphine were less potent nociceptin receptor agonists. The kappa(1)+kappa(3)-opioid receptor agonist/mu-opioid receptor antagonist naloxone benzoylhydrazone was a pure antagonist at both rat brain and human ORL1 receptors. The nonselective opioid receptor partial agonist buprenorphine and the nonselective opioid receptor antagonist (-)-quadazocine exhibited pure antagonism at rat brain receptors, but displayed partial agonism at human ORL1 receptors. Thus, opioids displaying full agonism at the nociceptin receptor are also opioid receptor agonists, whereas opioids that are antagonists or partial agonists at the nociceptin receptor show antagonism or partial agonism at opioid receptors. In addition, the stereospecificity required at opioid receptors appears to be retained at the nociceptin receptor, since (+)-quadazocine is inactive at both receptors. These findings illustrate the structural and functional homology of the opioid recognition site on these two receptor classes and suggest that opioids may provide leads for the design of nonpeptide nociceptin receptor agonists and antagonists lacking affinity for the classical opioid receptors.     

Up-regulation of RGS4 mRNA by opioid receptor agonists in PC12 cells expressing cloned mu- or kappa-opioid receptors.

The regulators of G-protein signaling (RGS) proteins have been shown to modulate the function of some heterotrimeric G-proteins by stimulating the GTPase activity of G-protein alpha subunits. In this study, by northern blotting analysis, we investigated the regulation of RGS4 mRNA by opioid receptor agonists in PC12 cells stably expressing either cloned mu- or kappa-opioid receptors. Treatment with respective opioid receptor agonists (mu: morphine) and [D-Ala(2), MePhe(4), Gly(ol)(5)] enkephalin (DAMGO), kappa: (+)-(5 alpha,7 alpha,8 beta)-N-methyl-N-[7-(1-pyrrolidinyl)-1-oxaspiro-(4,5)dec-8-y1]benzeneacetamide (U69,593)) for 0.5-24 h significantly and transiently increased the expression of RGS4 mRNA by 140-170% of the control level in a concentration-dependent manner which peaked when treated for 2 h, while treatment of non-transfected PC12 cells with opioid receptor agonists did not. The up-regulation of RGS4 mRNA was significantly blocked by co-treatment with respective opioid antagonists (mu: naloxone, kappa: norbinaltorphimine) or pretreatment with pertussis toxin. These results suggest that the activation of mu- or kappa-opioid receptors increases RGS4 mRNA level, which might contribute to opioid desentilization.     

Characterization of the antinociceptive effects of TRK-820 in the rat

We have already reported that TRK-820, (-)-17-cyclopropylmethyl-3, 14b-dihydroxy-4, 5a-epoxy-6b-[N-methyl-trans-3-(3-furyl)acrylamido]morphinan hydrochloride, a new selective kappa-opioid receptor agonist, has affinity for kappa-subtype opioid receptors other than the kappa(1)-opioid receptor. It would be of interest to examine whether the different kappa-opioid receptor subtype properties of TRK-820 participate in its antinociceptive action in the inflamed paw test and the formalin test. TRK-820 produced a potent antinociceptive effect, which was inhibited by the selective kappa-opioid receptor antagonist nor-binaltorphimine, but not by the mu-opioid receptor antagonist naloxone in the mechanical paw pressure test. TRK-820 also produced a potent antinociceptive effect in rats with adjuvant-induced arthritis. TRK-820 and morphine, a prototype mu-opioid receptor agonist, were equally effective in inhibiting the nociceptive responses in the arthritic rats and in the normal rats, while ICI-199441, 2-(3, 4-dichlorophenyl)-N-methyl-N-[(1S)-1-phenyl-2-(1-pyrrolidinyl)ethyl]- acetamide, a kappa-opioid receptor agonist, was about 5-fold less potent in the arthritic rats than in the normal rats. In the formalin test TRK-820 had a very similar antinociceptive potency to that of ICI-199441, unlike in the arthritic rats in which TRK-820 was 2.5 times more potent than ICI-199441. It is concluded that TRK-820 produced a potent antinociceptive action via the stimulation of kappa-opioid receptors in rats. TRK-820 has a unique antinociceptive profile different from that of the other kappa-opioid receptor agonists such as ICI-199441 in arthritic rats.     

Effects of kappa opioid agonists alone and in combination with cocaine on heart rate and blood pressure in conscious squirrel monkeys

As kappa agonists have been proposed as treatments for cocaine abuse, the cardiovascular effects of the kappa opioid receptor agonists ethylketocyclazocine (EKC) and enadoline were investigated in conscious squirrel monkeys. Both EKC and enadoline increased heart rate with little effect on blood pressure. This effect appeared to be specific for kappa receptors as the mu opioid agonist morphine did not mimic the effects of the kappa agonists. The opioid antagonist naltrexone, at a dose of 1.0 mg/kg, blocked the effect of EKC. An action at both central and peripheral receptors may be responsible for the heart rate increase following kappa agonist treatment. The ganglionic blocker chlorisondamine partially antagonized the effect of EKC on heart rate, suggesting central involvement, while the peripherally-acting agonist ICI 204,448 ((+/-)-1-[2,3- (Dihydro-7-methyl-1H-inden-4-yl)oxy]-3-[(1-methylethyl)amino]-2-butanol hydrochloride) also increased heart rate, supporting a peripheral site of action. When given in combination with cocaine, EKC produced effects that were sub-additive, suggesting that the kappa agonists may be used safely as cocaine abuse treatments.     

Spinal dynorphin A (1-17): possible mediator of antianalgesic action

Earlier studies from this laboratory indicated that intracerebroventricular administration of physostigmine and clonidine activated both a spinal descending analgesic and antianalgesic system. It was proposed that the latter was mediated spinally by dynorphin A (1-17), because small intrathecal doses (fmol) of dynorphin A (1-17) antagonized analgesia, while intrathecal administration of naloxone and nor-binaltorphimine (at doses which had no effect on spinal mu and kappa receptors) enhanced analgesia by attenuating the antianalgesic component. In the present studies in mice, using the tail-flick response, intrathecal administration of dynorphin antibody (antiserum to dynorphin) enhanced the analgesic effect of (10 min) physostigmine and clonidine given intraventricularly. Peak effect for the antiserum was at 1 hr. Inhibition of the tail-flick response, induced by DAMGO (Tyr-D-Ala2-Gly-NMePhe4-Gly-ol5, a mu agonist), U50, 488 H (trans-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]- benzeneacetamide methanesulfonate hydrate, a kappa agonist) and morphine was also enhanced by intrathecal administration of dynorphin antiserum. Thus, a variety of analgesic agonists appear to activate a dynorphin-mediated antianalgesic system. Such a system appears not to be activated by intraventricular administration of beta-endorphin and DPDPE (D-Pen2-D-Pen5-enkephalin, a delta agonist) because neither beta-endorphin- nor DPDPE-induced analgesia was enhanced by intrathecal administration of antiserum. The results of the experiments with the antibody provide further evidence to support the role of dynorphin A (1-17), as a putative endogenous opioid, which mediates an antianalgesic descending system in the spinal cord.     

Highly selective kappa opioid analgesics. 4. Synthesis of some conformationally restricted naphthalene derivatives with high receptor affinity and selectivity.

This paper describes the synthesis and kappa and mu opioid receptor binding affinity of some conformationally restrained derivatives of the arylacetamide group in the selective kappa opioid receptor agonist (+/-)-trans-N-methyl-N-[2-(1-pyrrolidinyl) cyclohexyl]benzo [b]thiophene-4-acetamide monohydrochloride (1,PD117302), which is an analogue of U-50, 488. The methyl-substituted derivatives (+/-)-trans-N, alpha-dimethyl-N-[2-(1-pyrrolidinyl)cyclohexyl]benzo-[b] thiophene-4-acetamide monohydrochloride (6a,b) possess significantly weaker affinity than 1 for the kappa opioid receptor (Ki = 172 and 3.7 nM, respectively). It is proposed that this is due to the conformational restriction imposed by the methyl group of 6. In order to test this proposal the acenaphthene derivative and the 4,5-dihydro-3H-naphtho [1,8-bc]thiophene derivative were prepared. The acenaphthene derivative (+)-N-methyl-N-[7-(1-pyrrolidinyl)-1-oxaspiro [4.5]dec-8-yl]acenaphthenecarboxamide monohydrochloride (9) was found to have high kappa opioid receptor affinity and selectivity (kappa Ki = 0.37 +/- 0.05 nM, mu/kappa = 659, delta/kappa = 1562) and is 100 times more potent than morphine as an analgesic in the rat paw pressure test for analgesia after intravenous administration (MPE50 = 0.014 and 1.4 mg/kg, respectively). The 4,5-dihydro-3H-naphtho[1,8-bc]thiophene derivative (-)-4,5-dihydro-N-methyl-N-[7-(1-pyrrolidinyl)-1-oxaspiro [4.5]dec-8-yl]-3H-naphthol[1,8-bc]thiophene-5-carboxamide p-toluenesulfonate (17) also has high kappa opioid receptor affinity and selectivity (kappa Ki = 4.65 nM, mu/kappa = 109).     

Interaction of bivalent ligand KDN21 with heterodimeric delta-kappa opioid receptors in human embryonic kidney 293 cells

KDN21 is a bivalent ligand that contains delta and kappa opioid antagonist pharmacophores linked through a 21-atom spacer. It has been reported that KDN21 bridges delta and kappa receptors that are organized as heterodimers. We have shown previously that when using [(3)H]diprenorphine as radioligand, KDN21 displayed greatly enhanced affinity in this series for coexpressed delta and kappa opioid receptors (CDK). The present study used in vitro expression systems to investigate interactions of members of the KDN series with delta-kappa heterodimers through competition binding using selective ligands and the mitogen-activated protein kinase (MAPK) assay. In this regard, the use of the selective radioligands [(3)H]naltrindole and [(3)H]norbinaltorphimine (nor-BNI) in competition binding studies revealed that KDN21 has much higher affinity than other KDN members for CDK and bound to CDK more selectively relative to mixed delta and kappa opioid receptors or singly expressed delta and kappa opioid receptors. Other experiments revealed that the binding of naltrindole to delta opioid receptors could increase the binding of nor-BNI to kappa opioid receptors and vice versa, suggesting reciprocal allosteric modulation of receptors in the heterodimer. Regarding the selectivity of KDN21 for phenotypic delta and kappa opioid receptors, we investigated the effect of KDN21 on the activation of MAPKs [extracellular signal-regulated kinases 1 and 2 (ERK1/2)] by delta- or kappa-selective agonists. KDN21 inhibited the activation of ERK1/2 by [D-Pen(2),D-Pen(5)]-enkephalin (delta(1)) and bremazocine (kappa(2)) but had no effect on the activation by deltorphin II (delta(2)) and (+)-(5alpha,7alpha,8beta)-N-methyl-N-[7-(1-pyrrolidinyl)-1-oxaspiro[4.5]dec-8-yl]benzeneacetamide (U69593, kappa(1)). 7-Benzylidenenaltrexone (delta(1)) and bremazocine (kappa(2)) significantly reduced the binding of KDN21 to CDK, whereas naltriben (delta(2)) and U69593 produced no such change. Taken together, these data support the idea that the organization of delta and kappa receptors as heterodimers gives rise to delta(1) and kappa(2) phenotypes.     

ATP-gated K(+) channel openers enhance opioid antinociception: indirect evidence for the release of endogenous opioid peptides

The ATP-gated K(+) channel openers - diazoxide, levcromakalim and morphine - enhance K(+) efflux by opening ATP-gated K(+) channels, thereby inducing cell hyperpolarization. Hyperpolarization decreases intracellular Ca(2+) levels, which leads to a decrease in neurotransmitter release contributing to the antinociceptive effects of the drugs. Previous findings implicate the release of endogenous opioids as the mediator of the antinociceptive effects of ATP-gated K(+) channel openers. Diazoxide and levcromakalim, administered intracerebroventricularly (i.c.v.), produced dose-dependent antinociception as determined by the tail-flick method ?ED(50) 44 microg/mouse [95% confidence limits (CLs) from 28 to 68 microg/mouse] for diazoxide?. Glyburide (10 microg/mouse), an ATP-gated K(+) channel antagonist, attenuated the effects of diazoxide, levcromakalim and morphine. Diazoxide- and levcromakalim-induced antinociception were both antagonized by CTOP (D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr amide), a mu-opioid receptor selective antagonist, and ICI 174,864 (N, N-diallyl-Tyr-Aib-Aib-Phe-Leu), a delta-opioid receptor antagonist, but were differentially attenuated by the kappa-opioid receptor antagonist, nor-Binaltorphimine. Combinations of inactive doses of the K(+) channel openers and opioid receptor agonists produced significant antinociceptive enhancement. Diazoxide (2 microg/mouse) shifted morphine's dose-response curve 47-fold, while levcromakalim (0.1 microg/mouse) shifted the curve 27-fold. The dose-response curve of kappa-opioid receptor agonist U50,488H (trans-(+/-)-3, 4 Dichloro-N-[2-(1-pyrrolidinyl)-cyclohexyl] benzeneacetamide methane sulfonate) was shifted 106-fold by diazoxide in a parallel manner, while levcromakalim administration increased the potency of U50,488H by 15-fold. Diazoxide shifted the dose-response curve of the delta-opioid receptor agonist, DPDPE [(D-Pen(2,5))-enkephalin], leftward in a non-parallel manner, while DPDPE was 6-fold more potent when combined with levcromakalim. We hypothesize that endogenous opioids mediate ATP-gated K(+) channel opener-induced antinociception and enhancement of opioids.     

Absence of G-protein activation by mu-opioid receptor agonists in the spinal cord of mu-opioid receptor knockout mice

1. The ability of mu-opioid receptor agonists to activate G-proteins in the spinal cord of mu-opioid receptor knockout mice was examined by monitoring the binding to membranes of the non-hydrolyzable analogue of GTP, guanosine-5'-O-(3-[35S]thio)triphosphate ([35S]GTPgammaS). 2. In the receptor binding study, Scatchard analysis of [3H][D-Ala2,NHPhe4,Gly-ol]enkephalin ([3H]DAMGO; mu-opioid receptor ligand) binding revealed that the heterozygous mu-knockout mice displayed approximately 40% reduction in the number of mu-receptors as compared to the wild-type mice. The homozygous mu-knockout mice showed no detectable mu-binding sites. 3. The newly isolated mu-opioid peptides endomorphin-1 and -2, the synthetic selective mu-opioid receptor agonist DAMGO and the prototype of mu-opioid receptor agonist morphine each produced concentration-dependent increases in [35S]GTPgammaS binding in wild-type mice. This stimulation was reduced by 55-70% of the wild-type level in heterozygous, and virtually eliminated in homozygous knockout mice. 4. No differences in the [35S]GTPgammaS binding stimulated by specific delta1- ([D-Pen2,5]enkephalin), delta2-([D-Ala2]deltorphin II) or kappa1-(U50,488H) opioid receptor agonists were noted in mice of any of the three genotypes. 5. The data clearly indicate that mu-opioid receptor gene products play a key role in G-protein activation by endomorphins, DAMGO and morphine in the mouse spinal cord. They support the idea that mu-opioid receptor densities could be rate-limiting steps in the G-protein activation by mu-opioid receptor agonists in the spinal cord. These thus indicate a limited physiological mu-receptor reserve. Furthermore, little change in delta1-, delta2- or kappa1-opioid receptor-G-protein complex appears to accompany mu-opioid receptor gene deletions in this region.     

kappa-Opioid inhibition of [3H]dopamine release from rat ventral mesencephalic dissociated cell cultures.

The present study investigated the effect of opioids on [3H]dopamine release from mixed neuronal-glial cell cultures of embryonic rat ventral mesencephalon. Each of the major morphological types of dopaminergic cell was represented in these cultures. These cells exhibited specific uptake of [3H]dopamine, which was subsequently released, in a calcium-dependent manner, in response to a double pulse of elevated extracellular potassium. Spontaneous and potassium-evoked [3H]dopamine release was inhibited by kappa- but not mu- or delta-opioid agonists. The selective kappa 1 agonist (5 alpha, 7 alpha, 8 beta)-(-)-N-methyl-N-[7-(1-pyrollidinyl)-1-oxaspiro(4,5)- dec-8-yl]benzeneacetamide (U69593) produced a dose-dependent inhibition of dopamine release. The effect of U69593 was blocked by the nonselective opiate antagonist naloxone and the selective kappa opioid antagonist norbinaltorphimine. kappa-Opioid inhibition of potassium-evoked [3H]dopamine release was maintained in the presence of tetrodotoxin. These results suggest that functional kappa receptors, modulating dopamine release, are localized on the terminals of dopaminergic neurons.     

Dexamethasone modulates hypotension induced by opioids in anaesthetised rats.

The effect of dexamethasone on hypotension induced by mu-, kappa- and delta-opioid receptor agonists was investigated in pentobarbital-anaesthetised rats. Morphine (nonselective opioid receptor agonist), DAGO (D-Ala2-N-methyl-[Phe4-Gly5-ol]enkephalin; mu-opioid receptor-selective agonist), U50-488H (trans(+/-)-3,4-dichloro-N-methyl-N-(2[1pyrrolidynyl]cyclohexyl)-benzeneacetamide; kappa-opioid receptor-selective agonist) and deltorphin II (delta-opioid receptor-selective agonist), given intravenously, 5 micromol/kg, induced hypotension in rats. This hypotension was characterised by a fall in mean arterial blood pressure in 1-2 min that recovered in 30 min for morphine and U50-488H and in 5 or 20 min for DAGO and deltorphin II, respectively. Dexamethasone per se at a dose of 7.5 micromol/kg, i.v. did not significantly modify the mean arterial blood pressure of animals. Dexamethasone administration 90 min, but not 30 or 60 min, before the opioid agonists injection, prevented the hypotension induced by morphine or U50-488H, but not that induced by DAGO or deltorphin II. Pretreatment with RU-38486 (mifepristone; 7.5 micromol/kg, i.v.), a glucocorticoid receptor antagonist, 15 min before the steroid, prevented dexamethasone inhibition of hypotension induced by morphine and U50-488H. Furthermore, pretreatment with cycloheximide, a protein synthesis inhibitor (3.5 micromol/kg, i.v.), was also able to abolish the effects of dexamethasone on morphine- and U50-488H-induced hypotension. Results of the present study indicate that dexamethasone inhibited kappa-opioid receptor-mediated hypotension in rats, indicating a further important functional interaction between corticosteroids and the opioid system at kappa receptors. The ability of cycloheximide and RU-38486 to block dexamethasone effects indicates that steroid interference with kappa-opioid receptor-mediated hypotension involves a protein synthesis-dependent mechanism via glucocorticoid receptors.     

Involvement of glutamate and gamma-amino-butyric acid receptor systems on gastric acid secretion induced by activation of kappa-opioid receptors in the central nervous system in rats

1. Various neurotransmitters in the brain regulate gastric acid secretion. Previously, we reported that the central injection of kappa-opioid receptor agonists stimulated this secretion in rats. Although the existence of kappa(1)-kappa(3)-opioid receptor subtypes has been proposed, the character is not defined. We investigated the interactions between kappa-opioid receptor subtypes and glutamate, gamma-amino-butyric acid (GABA) or 5-hydroxy tryptamine (5-HT) receptors in the rat brain. 2. Gastric acid secretion induced by the injection of U69593 (8.41 nmol, a putative kappa(1)-opioid receptor agonist) into the lateral cerebroventricle was completely inhibited by the central injection of 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10.9 nmol, an antagonist for non-N-methyl-D-aspartate (non-NMDA) receptors) and by bicuculline infusion (222 micro g kg(-1) per 10 min, i.v., GABA(A) receptor antagonist). The secretion induced by bremazocine (8.52 nmol, a putative kappa(2)-opioid receptor agonist) was inhibited by bicuculline infusion, but not by CNQX. The secretion induced by naloxone benzoylhydrazone (224 nmol, a putative kappa(3)-opioid receptor agonist) was slightly and partially inhibited by CNQX and bicuculline. 3. Treatment with CNQX and bicuculline inhibited gastric acid secretion induced by the injection of dynorphin A-(1-17) into the lateral, but not the fourth, cerebroventricle. Antagonists for NMDA, GABA(B) and 5-HT(2/1C) receptors did not inhibit the secretions by kappa-opioid receptor agonists. 4. In rat brain regions close to the lateral cerebroventricle, kappa-opioid receptor systems (kappa(1)>kappa(3)>kappa(2)) are regulated by the non-NMDA type of glutamate receptor system, and kappa(1)- and kappa(2)-opioid receptor systems are regulated by the GABA(A) receptor system. The present findings show pharmacological evidence for kappa-opioid receptor subtypes that regulate gastric acid secretion in the rat brain.     

Studies on the pharmacology of central opioid-induced increases in plasma catecholamines in conscious rats

Centrally-administered opioid peptides produce elevations in levels of catecholamines in plasma in conscious rats. To investigate the opioid receptor involved in mediating this response, morphine (3.5-105 nmol), [D-Ala2-D-Leu5]enkephalin (DADL, 1.05-35 nmol) and U50, 488H (35-1160 nmol), relatively selective ligands for the mu, delta and kappa receptors respectively, were injected intracerebroventricularly into conscious rats and the levels of catecholamines in plasma determined at the peak of the response. Each agonist produced dose-dependent elevations in levels of noradrenaline and adrenaline, the order of potency being DADL approximately equal to morphine greater than U50,488H. These results exclude involvement of the kappa receptor but do not distinguish between an effect mediated by a mu or delta receptor. The responses to morphine were blocked in the presence of 300 nmol naloxone but were not altered by 41.5 nmol RX 781094, a selective alpha 2-adrenoceptor antagonist. The elevation in levels of adrenaline in plasma produced by small doses of the agonists was not related to behavioural changes.     

Synthesis and evaluation of N-substituted cis-N-methyl-2-(1-pyrrolidinyl)cyclohexylamines as high affinity sigma receptor ligands. Identification of a new class of highly potent and selective sigma receptor probes

Certain benzeneacetamides [(-)- and (+)-cis-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl) cyclohexyl]benzeneacetamide] were recently reported to be potent sigma receptor ligands. In order to determine whether efficacy for the sigma receptor could be improved, a series of compounds related to the benzeneacetamides, N-substituted cis-2-(1-pyrrolidinyl)-N-methylcyclohexylamines, were synthesized and their structure-activity requirements were determined. The compounds were synthesized by starting with the previously reported (+/-)-, 1S,2R-(+)-, and 1R,2S-(-)-cis-2-(1-pyrrolidinyl)-N-methylcyclohexylamines. Analysis of sigma ([3H](+)-3-PPP), kappa ([3H]bremazocine and [3H]U69,593), dopamine-d2 ([3H](-)-sulpiride), and phencyclidine (PCP) ([3H]TCP) receptor binding in guinea pig brain revealed a number of highly potent and selective sigma receptor ligands. Notably, 1S,2R-cis-(-)-N-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl]-(2-naphthyl) acetamide [(-)-29] (Ki = 8.66 +/- 0.35 nM), (+/-)-cis-2-amino-4,5-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl] benzeneacetamide [(+/-)-17] (Ki = 11 +/- 3 nM), 1S,2R-(-)-cis-N-methyl-N-[2-(3,4-dichlorophenyl)ethyl]-2-(1-pyrrolidinyl ) cyclohexylamine [(-)-44] (Ki = 1.3 +/- 0.3 nM), and 1R,2S-(+)-cis-N-methyl-N-[2-(3,4-dichlorophenyl)ethyl]-2-(1-pyrrolidinyl ) cyclohexylamine. [(+)-44] (Ki = 6 +/- 3 nM) exhibited very high affinity at sigma receptors, by displacement of [3H]-(+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine [( 3H]-(+)-3-PPP). These compounds showed insignificant affinity for kappa, dopamine, or PCP receptors, making them valuable tools for the study of sigma receptors. Furthermore, these compounds also exhibited enantioselectivity ranging from 5-fold for (+)- and (-)-44 to 160-fold for (+)- and (-)-29. Several other compounds showed equivalent selectivity but displayed lower sigma receptor affinity.