Pharmacological studies with a nonpeptidic, delta-opioid (-)-(1R,5R,9R)-5,9-dimethyl-2'-hydroxy-2-(6-hydroxyhexyl)-6,7-benzomorphan hydrochloride ((-)-NIH 11082).

In the search for a selective delta-opioid receptor agonist, (-)-(1R,5R,9R)-5,9-dimethyl-2'-hydroxy-2-(6-hydroxyhexyl)-6,7-benzomorphan hydrochloride ((-)-NIH 11082) and the (+)-enantiomer were synthesized and tested. (-)-NIH 11082 displayed antinociceptive activity in the paraphenylquinone test (PPQ test) in male ICR mice [ED50=1.9 (0.7-5.3) mg/kg, s.c.] and showed little, if any, activity in the tail-flick and hot-plate assays. The (+)-enantiomer was essentially inactive indicating stereoselectivity. Opioid receptor subtype characterization studies indicated that naltrindole, a delta-opioid receptor antagonist, was potent versus the ED80 of (-)-NIH 11082 in the PPQ test [AD50=0.75 (0.26-2.20) mg/kg, s.c]. beta-Funaltrexamine and nor-binaltorphimine, selective mu- and kappa-receptor antagonists, respectively, were inactive versus the ED80 of (-)-NIH 11082. In rats with inflammation-induced pain, (-)-NIH 11082 produced antihyperalgesic effects that were attenuated by naltrindole. In morphine-dependent rhesus monkeys of both sexes, (-)-NIH 11082 neither substituted for morphine nor exacerbated withdrawal signs in the dose range of 4.0 to 32.0 mg/kg, s.c. Neither convulsions nor other overt behavioral signs were observed in any of the species tested. The results indicate that (-)-NIH 11082 has delta-opioid receptor properties.

Cannabimimetic properties of ajulemic acid.

Side effects of marijuana-based drugs and synthetic analogs of Delta(9)-tetrahydrocannabinol (Delta(9)-THC), including sedation and dysphoria, have limited their therapeutic application. Ajulemic acid (AJA), a side-chain synthetic analog of Delta(8)-THC-11-oic acid, has been reported to have anti-inflammatory properties without producing undesired psychoactive effects. Moreover, it has been suggested that AJA does not interact with cannabinoid receptors to produce its pharmacological effects. The aim of the present study was to conduct a thorough evaluation of the pharmacological effects of AJA then to determine whether actions at cannabinoid receptor (CB)(1) mediated these effects. This study evaluated the psychoactive and analgesic effects of AJA by examining its cannabimimetic properties in ICR mice (i.e., antinociception, catalepsy, hypothermia, and hypomobility), its discriminative stimulus effects in Long Evans rats trained in a two-lever Delta(9)-THC (3.0 mg/kg) versus vehicle drug discrimination procedure, and its antihyperalgesia effects in a rat model of inflammatory pain [complete Freund's adjuvant (CFA)-induced mechanical hyperalgesia]. Lastly, antagonism tests with SR 141716A [N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboximide hydrochloride], CB(1) receptor antagonist, were conducted. These studies demonstrated that AJA shares a number of CB(1)-mediated pharmacological properties with Delta(9)-THC, including cannabimimetic, discriminative stimulus, and antihyperalgesic effects. Furthermore, a separation between doses that produced antinociception and those that produced the other pharmacological effects in mice was not observed. Moreover, AJA showed nearly equipotency for therapeutic efficacy in the CFA model and for substitution in Delta(9)-THC discrimination. In summary, this study shows that AJA, like Delta(9)-THC, exhibits psychoactive and therapeutic effects at nearly equal doses in preclinical models, suggesting similar limitations in their putative therapeutic profiles.

Effects of combining ethanol (EtOH) with gamma-hydroxybutyrate (GHB) on the discriminative stimulus, locomotor, and motor-impairing functions of GHB in mice.

RATIONALE: Gamma-hydroxybutyrate (GHB) is a drug of abuse that is often coabused with ethanol (EtOH) and has been implicated as a date rape agent in conjunction with EtOH. Much information is lacking regarding the manner in which GHB interacts with EtOH. OBJECTIVE: This study was designed to further characterize the behavioral effects of GHB alone and in combination with EtOH in male Swiss-Webster mice. METHODS: The effects of GHB (0.1-1.0 g/kg) and EtOH (2.0-5.0 g/kg) alone, as well as the effects of GHB in combination with EtOH, were examined using an automated locomotor activity procedure, a functional observational battery (FOB) and a GHB drug discrimination procedure. RESULTS: GHB decreased, whereas EtOH had little effect on locomotor activity. In the FOB, EtOH dose-dependently decreased activity in combination with 0.3 g/kg GHB. Alone, each drug had little effect on the righting reflex, but combining ineffective doses of GHB and EtOH significantly impaired righting. GHB and EtOH decreased forelimb grip strength. Combinations of ineffective doses of GHB and EtOH decreased forelimb grip strength when given together. GHB and EtOH impaired inverted screen performance, and EtOH increased the impairing effects of low, but not high, doses of GHB. GHB and EtOH increased hind limb splay, and EtOH increased the effects of 0.1 and 0.3 g/kg GHB on splay. GHB and EtOH decreased body temperature, and EtOH augmented the temperature-decreasing effects of GHB. EtOH produced less than 50% GHB-like discriminative stimulus effects, and GHB failed to alter the GHB-like discriminative stimulus effects of EtOH. CONCLUSIONS: Overall, EtOH increased the effects of GHB on several gross measures of behavior and only partially occasioned the discriminative stimulus properties of GHB.

Effects of sex, hindpaw injection site and stimulus modality on nociceptive sensitivity in arthritic rats.

Injection of Complete Freund's adjuvant (CFA) into the hindpaw produces inflammation and alterations in nociceptive sensitivity. The present study was designed to compare the effects of CFA injection into the dorsal and plantar surfaces of the hindpaw on nociceptive sensitivity of the hindpaw to mechanical pressure, warm-water and a hotplate stimulus in male and female rats. CFA or vehicle (VEH) was injected into the dorsal or plantar surface of the right hindpaw on day 0 and tests were conducted on days 4, 6, 8, 10, 11 and 18. Up until day 11 the inflammation was confined to the injected hindpaw (monoarthritic state), whereas by day 18 both hindpaws were inflamed (polyarthritic state). The site of the CFA injection had minimal effects on thermal or mechanical sensitivity with the following exceptions. On days 11 and 18 males had higher hotplate latencies when injected in the dorsal as compared to the plantar surface. For both males and females, warm-water paw withdrawal latencies were longer in those rats injected in the dorsal versus the plantar surface on day 18. No sex differences in paw pressure thresholds were observed on days 11 and 18 in CFA-treated rats. In the warm-water paw withdrawal test CFA-treated males exhibited longer latencies than CFA-treated females on day 11, but similar latencies on day 18. In the hotplate test CFA-treated females exhibited shorter latencies than CFA-treated males on days 11 and 18. The present results demonstrate that nociceptive sensitivity is the result of the interplay among sex, CFA injection site (plantar vs. dorsal), arthritic state (mono- vs. polyarthritic) and stimulus modality (mechanical vs. thermal).

Nociceptive sensitivity and opioid antinociception and antihyperalgesia in Freund's adjuvant-induced arthritic male and female rats.

The present study was designed to examine sex differences in complete Freund's adjuvant (CFA)-induced mechanical hyperalgesia and sex differences in opioid antinociception and anti-hyperalgesia. Female rats developed inflammation and hyperalgesia faster and exhibited greater peak hyperalgesia than male rats. In arthritic (CFA-treated) rats, lower thresholds were observed during estrus and proestrus, and in nonarthritic (vehicle-treated) rats, lower thresholds were observed during proestrus. Morphine and oxycodone were more potent in male than female arthritic rats, and butorphanol was more potent and effective in male than female arthritic rats. The potency of morphine was increased in arthritic rats, although to a greater magnitude in males. The potency of oxycodone was increased in male but not female arthritic rats. The potency of butorphanol was increased in arthritic male rats and the maximal antinociceptive effect of butorphanol was increased in arthritic female rats, but it did not result in greater than 20% antinociception. Morphine, oxycodone, and butorphanol all produced antihyperalgesic effects (returning thresholds of arthritic rats to the thresholds of nonarthritic rats) with greater potency in males than females. The peripherally acting opioid agonist loperamide produced intermediate levels of antinociception in male and female arthritic rats and no antinociception in nonarthritic rats. Loperamide was more potent in male than female arthritic rats at producing antihyperalgesia. These data demonstrate sex differences in arthritis-induced hyperalgesia and responsiveness to opioid analgesics. In arthritic rats, the antinociceptive effects of opioid agonists are most probably mediated by both central and peripheral opioid receptors, whereas their antihyperalgesic effects are mediated primarily by actions at peripheral opioid receptors.

Further structurally constrained analogues of cis-(6-benzhydrylpiperidin-3-yl)benzylamine with elucidation of bioactive conformation: discovery of 1,4-diazabicyclo[3.3.1]nonane derivatives and evaluation of their biological properties for the monoamine transporters.

Our structure-activity relationship (SAR) study on piperidine analogues for monoamine transporters led to the development of a series of 3,6-disubstituted piperidine derivatives, structurally constrained versions of flexible piperidine analogues, with preferential affinity for the dopamine transporter (DAT). In our attempt to further rigidify this structure to study influence of rigidity on binding and in vivo activity, we have developed a series of 4,8-disubstituted 1,4-diazabicyclo[3.3.1]nonane derivatives. All synthesized derivatives were tested for their affinity at the DAT, serotonin transporter (SERT), and norepinephrine transporter (NET) in the brain by measuring their potency in competing for the binding of [(3)H]WIN 35, 428, [(3)H]citalopram, and [(3)H]nisoxetine, respectively. Selected compounds were also tested for their ability to inhibit uptake of [(3)H]DA. The SAR study led to the discovery of a potent lead compound (-)-S,S-10c which exhibited high affinity and selectivity for the DAT (IC(50) = 22.5 nM; SERT/DAT = 384 and NET/DAT > 444). It is interesting to note that both (-)-10c and the lead compound from the 3,6-disubstituted series (-)-2 exhibited highest activity in their (S,S) isomer indicating similar requirement of regiospecificity for maximum interaction. Overall, our current SAR results corresponded well with the results from less constrained 3,6-disubstituted versions of these molecules albeit the former class exhibited more stringent requirement in molecular structure for activity. However, the potent compounds in the current series exhibited greater selectivity for the DAT compared to their corresponding lesser constrained 3,6-disubstituted versions indicating an effect of rigidity in selective interaction with the transporter proteins. In an effort to elucidate the bioactive conformational structure of the lead molecules in the current and the 3,6-disubstituted series, a preliminary molecular modeling study was carried out where the most rigid derivative (-)-10c was used as a template structure. Compounds (-)-2 and (-)-10c exhibited stimulant activity in locomotor tests in mice in which (-)-2 exhibited a slower onset and longer duration of action compared to (-)-10c. Both compounds occasioned complete cocaine-like responding in mice trained to discriminate 10 mg/kg ip cocaine from vehicle.

Discriminative stimulus, reinforcing, physical dependence, and antinociceptive effects of oxycodone in mice, rats, and rhesus monkeys.

Despite oxycodone's (4,5-epoxy-14-hydroxy-3-methoxy-17-methylmorphinan-6-one) history of clinical use and the attention it has received as a drug of abuse, few reports have documented its pharmacology's relevance to its abuse or its mechanism of action. The purposes of the present study were to further characterize the analgesic effects of oxycodone, its mechanism of action, and its effects in terms of its relevance to its abuse liability. The results indicate that oxycodone had potent antinociceptive effects in the mouse paraphenylquinone writhing, hot-plate, and tail-flick assays, in which it appeared to be acting as a mu-opioid receptor agonist. It generalized to the heroin discriminative stimulus and served as a positive reinforcer in rats and completely suppressed withdrawal signs in morphine-dependent rhesus monkeys. These results suggest that the analgesic and abuse liability effects of oxycodone are likely mediated through mu-opioid receptors and provide the first laboratory report of its discriminative stimulus, reinforcing, and morphine cross-dependency effects.

Modulation of the locomotor activating effects of the noncompetitive NMDA receptor antagonist MK801 by dopamine D2/3 receptor agonists in mice.

The noncompetitive NMDA receptor antagonist MK801 (dizocilpine) produces behavioral stimulation mediated, in part, through indirect activation of the dopamine (DA) system. Previous reports indicate that D2/3 agonists inhibit MK801-induced stereotypies; however, it is unclear if these agonists also attenuate MK801-induced locomotion. As such, the ability of the D2/3 agonists, quinelorane and quinpirole, and the partial D3 agonist, BP897, to attenuate the locomotor activating effects of MK801 was examined in mice. MK801 (0.1-1.0 mg/kg) produced a biphasic effect on total distance traveled with the intermediate dose of 0.3 mg/kg producing the greatest stimulation. The increase in MK801-induced total distance traveled was attenuated by the coadministration of quinelorane and quinpirole at doses that alone had no effect on activity. Similarly, the partial D3 agonist, BP897, blocked the effects of MK801. The D3-preferring antagonist, nafadotride, reversed the attenuation of quinelorane and partially reversed the attenuation of quinpirole. The D2-preferring antagonist, eticlopride, reversed the attenuating effects of quinelorane, but was not effective against quinpirole. Nafadotride and eticlopride were ineffective against the attenuating effects of BP897 on MK801-induced locomotion. Because BP897 is a partial agonist it was tested against quinelorane/MK801 and quinpirole/MK801 combinations. BP897 reversed the attenuating effects of quinelorane, but not those of quinpirole on MK801's effects. These results demonstrate that the DA system, through D2/3 receptor activation, modulates the locomotor activating effects produced by noncompetitive NMDA receptor blockade.

The modulatory actions of dopamine D2/3 agonists and antagonists on the locomotor-activating effects of morphine and caffeine in mice.

Morphine produces robust increases in locomotor activity in mice. Recent data indicate that dopamine (DA) D2/3 agonists attenuate the discriminative stimulus and antinociceptive effects of mu opioid agonists such as morphine. The present study was designed to determine the extent to which D2/3 receptor activation and blockade can modulate morphine-induced locomotion using a novel cumulative dosing procedure in Swiss-Webster mice. The results indicate that morphine-induced locomotion is nonsignificantly attenuated by the D2/3 agonists quinelorane and quinpirole, whereas the D2/3 antagonists eticlopride and nafadotride, as well as the partial D2/3 agonist BP897, significantly reduced morphine-induced locomotion. To determine the specificity of this modulation, these agonists and antagonists were examined in combination with caffeine, a drug that also indirectly alters DAergic activity. Unlike the effects on morphine, caffeine-induced locomotion was unaltered by eticlopride, nafadotride and BP897, but was attenuated by quinelorane and quinpirole. These results indicate that modulation of D2/3 receptors can, in turn, alter the locomotor-activating effects of morphine.

Effect of strain and sex on mu opioid receptor-mediated G-protein activation in rat brain.

Strain and sex differences in mu opioid-mediated antinociception have been reported in rodents. The present studies evaluated mu opioid receptor-mediated G-protein activation in Lewis and Fischer 344 (F344) male and female rats using agonist-stimulated [35S]GTPgammaS binding. Compared to Lewis rats, F344 rats exhibited a 35% higher level of net DAMGO-stimulated [35S]GTPgammaS binding in striatum. Basal [35S]GTPgammaS binding was approximately 30% lower in thalamus of Lewis than F344 rats. Female Lewis rats also exhibited slightly ( approximately 15%) lower basal [35S]GTPgammaS binding in cingulate cortex relative to F344 rats of either sex. The relative efficacies of the mu partial agonists, morphine and buprenorphine, were also examined. Buprenorphine exhibited approximately 40% lower relative efficacy in the periaqueductal gray in Lewis compared to F344 rats, but no other relative efficacy differences were found between strains or sexes. Moreover, regional differences in the relative efficacy of buprenorphine were also detected in Lewis but not F344 rats. In contrast to these results, the only difference found between sexes was the 13% lower basal [35S]GTPgammaS binding in the cingulate cortex of female compared to male Lewis rats. These results suggest that differences in mu opioid receptor-mediated G-protein activation may contribute to strain differences in opioid antinociception, whereas sex differences may result predominantly from other mechanisms.

RTI 113, a 3-phenyltropane analog, produces long-lasting cocaine-like discriminative stimulus effects in rats and squirrel monkeys.

The 3-phenyltropane analog, 3beta-(4-chlorophenyl)tropane-2beta-carboxylic acid phenyl ester hydrochloride (RTI 113), has been shown to decrease cocaine self-administration in rats, squirrel monkeys and rhesus monkeys. Consequently, it has been proposed as a potential agonist pharmacotherapy for cocaine dependence. However, the time course of the discriminative stimulus effects of RTI 113 has not been determined. The present study was designed to compare the time course of the discriminative stimulus effects of RTI 113 with that of cocaine in rats and squirrel monkeys. Tests were conducted using a drug discrimination procedure in which rats and squirrel monkeys were trained to discriminate 10 and 0.3 mg/kg cocaine from saline, respectively. RTI 113 substituted for cocaine in both species. The time course of the discriminative stimulus effects of RTI 113 was approximately five times longer compared to that of cocaine. RTI 113 is a 3-phenyltropane analog that has long acting cocaine-like discriminative stimulus effects in both rats and squirrel monkeys. These results, combined with those indicating that RTI 113 decreases cocaine self-administration, suggest that RTI 113, or analogs of RTI 113, may be useful tools for developing potential agonist therapies for cocaine dependence.

Sex and rat strain determine sensitivity to kappa opioid-induced antinociception.

RATIONALE: Recent studies indicate that sex and rodent strain are determinants of sensitivity to opioid-induced antinociception. OBJECTIVES: The present study examined the influence of sex and rat strain on kappa opioid-induced antinociception using a series of kappa opioids that vary in their relative effectiveness. METHODS: In a warm-water (50, 52 and 55C) tail-withdrawal procedure, the antinociceptive effects of kappa opioids were determined in male and female rats of the F344, Lewis and Sprague-Dawley (SD) strains. RESULTS: In both males and females of each strain, spiradoline produced high levels of antinociception across all nociceptive stimulus intensities, whereas U50,488 produced high levels only at the low and moderate nociceptive stimulus intensities. Sex differences in the potency and effectiveness of these kappa opioids were relatively small and not consistently obtained. Enadoline, bremazocine and nalorphine were less effective than spiradoline in producing antinociception, and at low and moderate nociceptive stimulus intensities these opioids were both more potent and effective in F344 and SD males than their female counterparts. In contrast, in Lewis rats, only bremazocine was more potent and effective in males. In combination tests, bremazocine shifted the spiradoline dose-effect curve leftward and/or upward in males and rightward in females (i.e., antagonized spiradoline). In contrast, in both males and females enadoline shifted the spiradoline dose-effect curve leftward and/or upward. CONCLUSIONS: These data indicate that kappa opioids were generally more potent and effective as antinociceptive agents in males than females. Similar to data obtained with micro opioids, the magnitude of these sex differences was generally larger with the less effective kappa opioids and determined, in part, by rat strain and nociceptive stimulus intensity.

Effects of the putative antagonist NCS382 on the behavioral pharmacological actions of gammahydroxybutyrate in mice.

RATIONALE: Gammahydroxybutyrate (GHB) is an endogenous chemical found in the human brain that when administered systemically readily crosses the blood-brain barrier and produces behavioral effects. Some previously reported observations, including reports of specific antagonism by NCS382 (6,7,8,9-tetrahydro-5-[H]-benzocycloheptene-5- ol-4-ylidene acetic acid), support the hypothesis that GHB is a neurotransmitter with its own receptor system. In addition to its uncertain physiological role, the recent interest in GHB has been engendered by its illicit use and abuse. OBJECTIVES: To further characterize the behavioral effects of GHB and to evaluate NCS382 for its potential antagonistic effects. METHODS: Following the administration of GHB alone and in combination with NCS382, mice were tested in a Functional Observational Battery (FOB) and for their effects on locomotor activity and on schedule-maintained behavior. Additionally, spontaneous and NCS382-precipitated withdrawal in rats chronically treated with GHB was examined. RESULTS: In the FOB, GHB generally produced depressant-like effects that were generally not reversed by NCS382. GHB also dose dependently reduced locomotor activity and rates of operant behavior, which were generally not reversed by co-administrations with NCS382. Neither spontaneous nor NCS382-precipitated signs of physical dependence were observed following chronic GHB administration. CONCLUSION: GHB dose dependently produced depressant-like effects on learned and unlearned behavior. The putative GHB antagonist NCS382 failed to convincingly antagonize these effects. Physical dependence was not evident following spontaneous withdrawal or NCS382 challenge. Taken together, these results suggest that NCS382's ability to antagonize GHB's effects may be very limited.

Expansion of structure-activity studies of piperidine analogues of 1-[2-(diphenylmethoxy)ethyl]-4-(3-phenylpropyl)piperazine (GBR 12935) compounds by altering substitutions in the N-benzyl moiety and behavioral pharmacology of selected molecules.

A series of substituted N-benzyl analogues of the dopamine transporter (DAT) specific compound, 4-[2-(diphenylmethoxy)ethyl]-1-benzylpiperidine were synthesized and biologically characterized. Different 4'-alkyl, 4'-alkenyl, and 4'-alkynyl substituents were introduced in the phenyl ring of the benzyl moiety along with the replacement of the same phenyl ring by the isomeric alpha- and beta-naphthyl groups. Different polar substitutions at the 3'- and 4'-position were also introduced. Novel compounds were tested for their binding affinity at the dopamine, serotonin, and norepinephrine transporter systems in the brain by competing for [(3)H]WIN 35 428, [(3)H]citalopram, and [(3)H]nisoxetine, respectively. Selected compounds were also evaluated for their activity in inhibiting the uptake of [(3)H]dopamine. Binding results demonstrated that alkenyl and alkynyl substitutions at the 4'-position produced potent compounds in which compound 6 with a vinyl substitution was the most potent. In vivo evaluation of three selected compounds indicated that despite their high potency at the DAT, these compounds stimulated locomotor activity (LMA) less than cocaine when tested across similar dose ranges. In a drug discrimination study procedure, none of these three compounds generalized from cocaine in mice trained to discriminate 10 mg/kg cocaine from vehicle. In a 4 h time course LMA experiment, one of our previous lead piperidine derivatives (1a) showed considerable prolonged action. Thus, in this report, we describe a structure-activity relationship study of novel piperidine analogues assessed by both in vitro transporter assays and in vivo behavioral activity measurements.