Previous Drug Discrimination Studies Research Articles

Acute behavioral tolerance to nicotine in the conditioned taste aversion paradigm

AbstractAcute behavioral tolerance to nicotine is a well‐established phenomenon in animals, although previous drug discrimination studies have suggested that subpopulations of rats may exist that fail to exhibit acute tolerance to nicotine. The present study sought to determine whether these findings might extend to the conditioned taste aversion (CTA) paradigm. In experiment 1, rats were administered 0.8 mg/kg s.c. nicotine or vehicle before a pairing session with saccharin, and then administered either 0.4 mg/kg nicotine or vehicle immediately afterward. Rats treated with vehicle pre‐session and then nicotine post‐session developed a significant CTA to nicotine, whereas rats treated with 0.8 mg/kg nicotine s.c. pre‐session failed to develop a CTA to nicotine. In experiment 2, rats were identified as exhibiting or failing to exhibit acute tolerance to nicotine in a drug discrimination paradigm. Rats that produced acute nicotine tolerance failed to develop a CTA after 24 h, but did so after 48 h, while rats that did not produce nicotine tolerance exhibited CTA after 24 h, but not after 48 h. These findings indicate that acute behavioral tolerance to nicotine extends to its aversion effects. Drug Dev Res 68:522–528, 2007. © 2008 Wiley‐Liss, Inc.

Discriminative stimulus properties of the atypical antipsychotic drug clozapine in rats trained to discriminate 1.25 mg/kg clozapine vs. 5.0 mg/kg clozapine vs. vehicle

Clozapine, the prototype for atypical antipsychotic drugs, is used in the drug discrimination paradigm as a model for screening atypical from typical antipsychotic drugs. Previous drug discrimination studies in rats have shown that a 1.25 mg/kg clozapine training dose provides full stimulus generalization (i.e.) >or=80% condition-appropriate responding) to most atypical antipsychotic drugs, although a 5.0 mg/kg clozapine training dose appears necessary to provide stimulus generalization to other atypical antipsychotic drugs. The present study sought to characterize the pharmacological mechanisms that mediate these clozapine training doses. In rats trained to discriminate 1.25 vs. 5.0 mg/kg clozapine vs. vehicle in a three-choice drug discrimination task, various receptor-selective compounds were tested for stimulus generalization. The antidepressant mianserin was also tested. Full stimulus generalization from the 1.25 mg/kg clozapine training dose occurred only to mianserin (98.8%). Partial substitution (i.e. >or=60% and <80% condition-appropriate responding) to the 5.0 mg/kg clozapine training dose occurred for the muscarinic receptor antagonist scopolamine. The combined total percentage of responding on the 1.25 and 5.0 mg/kg clozapine levers, however, was well above the full substitution criteria at the 0.25, 0.5, and 1.0 mg/kg scopolamine doses. The M1 agonist N-desmethylclozapine, the nicotinic antagonist mecamylamine, the D1 antagonist SCH 23390, the D4 antagonist LU 38-012, the 5-HT1A agonist (+)-8-OH-DPAT, the 5-HT1A antagonist WAY 100 635, the 5-HT2A/2B/2C antagonist ritanserin, the 5-HT6 antagonist RO4368554, the alpha1 antagonist prazosin, the alpha2 antagonist yohimbine, and the histamine H1 antagonist pyrilamine all failed to substitute for either the 1.25 or the 5.0 mg/kg clozapine training doses. These results are consistent with previous evidence that antidepressant drugs have a tendency to substitute for clozapine and that muscarinic receptor antagonism may mediate the discriminative stimulus properties of 5.0 mg/kg clozapine. The lack of stimulus generalization from either clozapine training dose to other receptor-selective compounds, however, fails to explain how this model screens atypical from typical antipsychotic drugs and suggests that the discriminative stimulus properties of clozapine consist of a compound cue.

Antagonism of discriminative stimulus effects of Δ9-THC and (R)-methanandamide in rats

In previous drug discrimination studies we observed surmountable antagonism by Delta(9)-tetrahydrocannabinol (THC) in the presence of constant doses of SR-141716 [N-(piperidin-1-yl)-5-(4-chloro-phenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide] (0.3 and 1 mg/kg), but there was only marginal evidence for surmountable antagonism with combinations of SR-141716 and (R)-methanandamide, a chiral analog of the endocannabioid anandamide. Here we examine antagonism where the cannabinoid CB1 receptor agonist [Delta(9)-THC and (R)-methanandamide] dose is held constant (i.e., the training dose) and the antagonist {i.e., SR-141716 and AM-251 [N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide; 2 ml/kg]} dose varied. We also tested the cannabinoid CB2 receptor antagonist SR-144528 {N-[(1S)-endo-1,3,3-trimethylbicyclo(2.2.1)heptan-2-yl]5-(4-chloro-3-methyl-phenyl)-1-(4-methylbenzyl)pyrazole-3-carboxamide}. Different groups of rats were trained to discriminate between vehicle and three different doses of Delta(9)-THC (1.8, 3, and 5.6 mg/kg, presumably reflecting different efficacy demands) as well as 10 mg/kg (R)-methanandamide. Dose-generalization tests involved different doses of the cannabinoid CB1 receptor agonists. Antagonist tests varied the dose of the antagonist (range: 0.1 and 3 mg/kg for SR-141716 and AM-251, and 1 to 10 mg/kg for SR-144528). SR-141716 and AM-251 doses dependently blocked the agonist-induced discriminative stimulus effects. SR-141716 tended to be slightly more potent than AM-251. The effective dose 50 (ED(50)) of SR-141716 was higher in the 5.6 mg/kg Delta(9)-THC-trained group relative to the two other Delta(9)-THC-trained groups. The cannabinoid CB2 receptor antagonist SR-144528 combined with the training dose of 1.8 mg/kg Delta(9)-THC, as well as when combined with the training dose of 10 mg/kg (R)-methanandamide, did not markedly change drug-appropriate (agonist) responses. Data support that the discriminative stimulus effects of (R)-methanandamide and its overlap with the Delta(9)-THC cue are, indeed, CB1 receptor mediated events as revealed in antagonism tests with the selective central CB1 receptor antagonists SR-141716 and AM-251. The activation of cannabinoid CB2 receptors appears to be insignificant for these discriminations.

Evidence for bidirectional cues as a function of time following treatment with amphetamine: implications for understanding tolerance and withdrawal

Rationale Previous drug-discrimination studies have focused on characterizing the cue properties associated with amphetamine's (AMPH) primary effect. Results from recent experiments indicate that equally prominent cues are associated with AMPH withdrawal. Objectives The purpose of the present study was to investigate the extent to which AMPH-induced withdrawal cues, opposite to those associated with AMPH's primary effect are observed. Methods Since dopamine (DA) has been implicated in mediating the AMPH cue, rats were trained to discriminate between 0.25 mg/kg AMPH, an indirect DA agonist, and 0.033 mg/kg haloperidol (HAL), a DA antagonist at the D2 receptor site. Training doses were chosen so that rats responded about equally on both levers when tested on saline (SAL) providing a behavioral baseline sensitive to assessing AMPH-related bidirectional changes in cue state. Following acquisition of the discrimination, rats were tested for choice of responding on the AMPH and HAL levers at intervals from 6 to 72 h following treatment with a single dose of 3.0 mg/AMPH. Also, in order to investigate the relationship between withdrawal and tolerance to AMPH's cue properties, AMPH dose–response curves were determined 24 h following treatment with SAL, 1.5 and 3.0 mg/kg AMPH. Results At short intervals after treatment with 3.0 mg/kg AMPH, rats responded primarily on the AMPH lever followed by a shift to predominant responding on the HAL lever 16–30 h post-treatment, before returning to predrug levels. Treatment with 1.5 and 3.0 mg/kg AMPH produced parallel dose–response curve shifts to the right. Conclusions Following a single dose of AMPH, robust cues associated with AMPH withdrawal were observed that lasted approximately three times longer than the cues associated with the drug's primary effects. Furthermore, results from the tolerance tests indicate that tolerance reflects a baseline shift rather than a loss in drug efficacy.

Discriminative stimulus effects of ethanol: lack of interaction with taurine.

Recent microdialysis studies showed that ethanol administration increases the release of taurine in various rat brain regions, and it was suggested that this increase in extracellular concentrations of taurine might mediate some of the neurochemical effects of ethanol. Previous drug discrimination studies showed that positive modulators of the GABA(A) receptor consistently substituted for ethanol discriminative stimulus effects. Since taurine is also believed to modulate GABA(A) receptor activity, this study addressed the hypothesis that taurine mediates the discriminative stimulus effects of ethanol due to GABA(A) activation. Male Long-Evans rats were trained to discriminate water from either 1 or 2 g/kg ethanol. In a first experiment, various taurine doses (0-500 mg/kg) were tested to investigate whether taurine substitutes for ethanol. In a second experiment, rats were pretreated with either 500 mg/kg taurine or an equivalent volume of saline before testing for ethanol discrimination with various ethanol doses (0-2.0 g/kg). The results showed that taurine does not substitute for ethanol at any tested doses. In addition, taurine pretreatments failed to modify the dose-response curve for ethanol discrimination. These results demonstrate that taurine is not directly involved in mediating the discriminative stimulus effects of ethanol. It is therefore very unlikely that the brain release of taurine observed after ethanol administration is implicated in the major pharmacological effects of ethanol, i.e. positive modulation of GABA(A) receptor, that mediate its discriminative stimulus effects.

5-HT 2A receptor-stimulated phosphoinositide hydrolysis in the stimulus effects of hallucinogens

The role of 5-HT 2A-mediated stimulation of phosphoinositide hydrolysis in the discriminative effects of hallucinogens was investigated in PC12 cells stably expressing the rat 5-HT 2A receptor (PC12-5-HT 2A cells). The hallucinogenic compounds, d-lysergic acid diethylamide (LSD), (−)2,5-dimethoxy-4-methylamphetamine (DOM), psilocybin, N, N-dimethyltryptamine (DMT), 5-methoxy- N, N-dimethyltryptamine (MDMT) and N, N-diethyltryptamine (DET), all caused a concentration-dependent increase in the generation of [ 3H]inositol phosphates. The nonhallucinogenic compounds, 6-fluoro- N, N-diethyltryptamine (6-F-DET), lisuride and quipazine, also displayed significant efficacy in stimulating phosphoinositide hydrolysis, while 2-bromo-lysergic acid diethylamide (BOL), which is not a hallucinogen, did not alter inositol phosphate generation. The β-carbolines, harmaline and harmane, also did not alter phosphoinositide hydrolysis. Comparison of these results with previous drug discrimination studies indicated the apparent lack of correlation between the degree of substitution in LSD- and DOM-trained animals and efficacy in stimulating phosphoinositide hydrolysis. The present study indicates that 5-HT 2A-mediated stimulation of phosphoinositide hydrolysis does not appear to be the sole critical signaling mechanism involved in the discriminative effects of hallucinogens.

Long-lasting rebound cue effects following single doses of nicotine and amphetamine: implications for understanding tolerance.

Previous drug-discrimination studies have, with the exception of nicotine (NIC), demonstrated tolerance to the cue effects of a broad range of drugs of abuse. Barrett et al. have shown that tolerance to a drug's cue properties reflects drug-induced rebound shifts in the discrimination baseline and not a weakened or less salient cue. The objective of the present study was to use a discrimination task sensitive to bidirectional cue changes to characterize the interoceptive cues associated with both the primary and rebound cues produced by nicotine in an attempt to understand why a recent study by Shoaib et al. failed to observe tolerance to the nicotine cue. Since dopamine (DA) has been implicated in mediating the NIC cue, rats were trained to discriminate between 0.25 mg/kg amphetamine (AMPH), an indirect DA agonist, and 0.033 mg/kg haloperidol (HAL), a DA antagonist at the D2 receptor site. Training doses were chosen so that rats responded about equally on both levers when tested on saline (SAL) following acquisition. This procedure provided a behavioral baseline to assess NIC-related changes along a presumed continuum of DA-mediated cues. Following acquisition of the discrimination: (i) NIC substitution tests were conducted, (ii) rats were tested for lever choice at intervals from 2 h to 48 h following treatment with single doses of 0.25 mg/kg and 0.50 mg/kg NIC, and (iii) rats were challenged with test doses of NIC during a period of NIC rebound. (i) NIC substituted for AMPH in a dose- dependent manner. (ii) At short intervals after treatment with 0.25 mg/kg and 0.50 mg/kg NIC, rats responded primarily on the AMPH lever followed by a shift to predominant responding on the HAL lever 16-24 h post-treatment, before returning to predrug levels. (iii) No evidence was observed for acute tolerance to NIC. The robust and long-lasting rebound cues associated with training level doses of NIC suggest that maximal tolerance would likely develop to the NIC cue during the acquisition phase of the conventional NIC-saline discrimination study.

Two-lever drug-drug discrimination with the 5-HT1 receptor agonists flesinoxan and eltoprazine.

Previous drug discrimination studies with the 5-HT1 receptor agonists flesinoxan and eltoprazine showed a clear 5-HT1A receptor-mediated effect for flesinoxan and the involvement of both 5-HT1A and 5-HT1B receptors in eltoprazine. However, there was no clear antagonism of eltoprazine's cue, possibly due to the compound nature of the eltoprazine stimulus. In the present experiments, in order to create a specific 5-HT1A vs. 5-HT1B receptor-mediated discrimination, rats were trained to discriminate between flesinoxan and eltoprazine. All rats learned the discrimination readily (mean = 41.3 sessions to criterion). With training doses of 1.0 mg/kg, p.o. flesinoxan and 1.5 mg/kg, p.o. eltoprazine, saline administration resulted in 50% of the responses made on both levers. Substitution tests showed that the flesinoxan stimulus was mediated by the 5- HT1A receptor (8-OH-DPAT, buspirone) and the eltoprazine stimulus probably mediated by the 5-HT1B receptor (anpirtoline, TFMPP, RU-24969). The selective 5-HT1A receptor antagonist WAY-100635 antagonized the flesinoxan cue, and the discriminative stimulus of eltoprazine could be completely antagonized with the 5-HT1B/1D receptor antagonist GR-127935. When the training doses of both flesinoxan and eltoprazine were administered concurrently, complete substitution for eltoprazine was obtained. We conclude that rats can learn to discriminate between two serotonergic drugs with overlapping stimulus properties and that the flesinoxan stimulus is mediated by 5-HT1A receptors and the eltoprazine stimulus, under these particular training conditions, by 5-HT1B receptors.

Clozapine discrimination with a low training dose distinguishes atypical from typical antipsychotic drugs in rats.

Previous drug discrimination studies with clozapine have not reliably distinguished between atypical and typical antipsychotics. The present study was conducted to determine whether low-dose clozapine drug discrimination could distinguish atypical from typical antipsychotics. Rats were trained to discriminate 1.25 mg/kg clozapine from vehicle in a two-lever drug discrimination procedure. Generalization testing revealed full substitution with the atypical antipsychotics olanzapine (90.3% maximum generalization), sertindole (99.8%), and risperidone (87.1%) and partial substitution for quetiapine (seroquel, 66.4%) and the typical antipsychotics haloperidol (56.8%) and thioridazine (74.3%). Remoxipride (23.1%) and the typical antipsychotics chlorpromazine (27.9%) and fluphenazine (29.5%) did not reliably substitute for clozapine. In contrast to previous clozapine drug discrimination studies with higher training doses, the atypical antipsychotics olanzapine, sertindole, and risperidone reliably substituted for clozapine while typical antipsychotics did not. These results suggest that low-dose clozapine drug discrimination may be a more sensitive assay for distinguishing atypical from typical antipsychotic drugs.

Discriminative stimulus properties of alprazolam

The triazolobenzodiazepine alprazolam has a unique clinical profile compared to most other benzodiazepines (e.g. diazepam, chlordiazepoxide), in that it is used to treat panic disorder and is effective in depression, two disorders that are usually treated with anti-depressants. Previous drug discrimination studies suggested that alprazolam has stimulus properties in common with antidepressants. In the present study, the discriminative stimulus properties of alprazolam were investigated to test more conclusively the role of benzodiazepine receptors and whether alprazolam has stimulus properties in common with antidepressants. Male Wistar rats (n=12) were trained to discriminate between alprazolam (2.0 mg/kg, PO) and vehicle in an operant two-lever drug discrimination procedure under a tandem VI40"-FR10 schedule of reinforcement. Generalization and antagonism tests were carried out under 2 min extinction. In generalization tests, a number of benzodiazepines (alprazolam, chlordiazepoxide, midazolam, lorazepam) and the barbiturate pentobarbital substituted completely, while zolpidem and abecarnil substituted partially for alprazolam. In contrast, no significant degree of generalization to the antidepressants imipramine and fluvoxamine and the putative antidepressants buspirone and flesinoxan was found. In antagonism studies alprazolam could be antagonized (almost) completely by flumazenil, partially by pentylenetetrazole, but not by methyl 6, 7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM), N-methyl-beta-carboline-3-carboxamide (FG-7142) and picrotoxin. These results show that the discriminative stimulus properties of alprazolam are mediated by benzodiazepine receptors and that the finding that antidepressants share discriminative stimulus effects with alprazolam may have limited generality.

Cocaine-opioid interactions in groups of rats trained to discriminate different doses of cocaine.

The growing abuse of cocaine combined with morphine-like opiates ("speedballs") in human addicts has prompted efforts to characterize the roles of different opioid receptor subtypes in mediating their combined effects. Previous drug discrimination studies in rats have been inconsistent in showing significant interactions between cocaine and opioid agonists in subjects trained to discriminate a relatively high dose of cocaine from vehicle. It is known, however, that the training dose of cocaine can play a key role in drug-substitution and drug-interaction profiles and, therefore, training rats to discriminate a relatively low dose of cocaine may influence its interactions with opioid agonists. The objectives of this study were to examine the degree to which a relatively high (10 mg/kg) versus a relatively low (3.0 mg/kg) cocaine training dose influenced the interactions between cocaine and either the mu opioid agonist morphine or the kappa opioid agonist U50,488. Substitution tests with cumulative doses of cocaine, morphine and U50,488 were conducted, as were studies in which selected doses of morphine or U50,488 were administered prior to cumulative doses of cocaine. In substitution tests, cocaine was 2.9 times more potent under the low- than the high-dose training condition. Morphine substituted fully for cocaine in the majority of subjects trained to discriminate the low, but not the high, dose of cocaine. U50,488 engendered mainly saline-lever responses under both training conditions. In pretreatment studies, morphine enhanced and U50,488 attenuated the discriminative stimulus effects of cocaine in low-dose, but not high-dose, trained rats. In low-dose trained rats, cocaine was five- to eightfold more potent after morphine and three- to fourfold less potent after U50,488 pretreatments. The results demonstrate that cocaine-opioid interactions are dependent on the training dose of cocaine in rats and suggest an opposing influence of mu and kappa opioid receptors in modifying the discriminative stimulus effects of cocaine.

Assessment of the multiple discriminative stimulus effects of ethanol using an ethanol-pentobarbital-water discrimination in rats.

Previous drug discrimination studies have elucidated the importance of gamma-aminobutyric acidA (GABAA), N-methyl-D-aspartate (NMDA) glutamate, and serotonin (5-HT) receptor systems in mediating the discriminative stimulus effects of ethanol. The present study used a three-choice operant drug discrimination procedure in an attempt to determine if salient GABAergic effects could be separated from other stimulus effects of 2.0 g/kg ethanol. Adult male Long-Evans rats (n = 7) were trained to discriminate pentobarbital (10.0 mg/kg; intragastrically (i.g.) from ethanol (2.0 g/kg; i.g.) from water (4.7 ml; i.g.) using food reinforcement. Stimulus substitution tests were conducted following the administration of allopregnanolone (1.0-17.0 mg/kg; intraperitoneally (i.p.)), diazepam (0.1-7.3 mg/kg; i.p.), midazolam (0.0056-17.0 mg/kg; i.p.), dizocilpine (0.01-0.56 mg/kg; i.p.), phencyclidine (1.0-5.6 mg/kg; i.p.), CGS 12066B (3-30 mg/kg; i.p.), RU 24969 (0.1-5.6 mg/kg; i.p.) and morphine (1 or 3.0 mg/kg; i.p.). Within the group, allopregnanolone and midazolam completely substituted (> 80%), and diazepam partly substituted (67%) for the discriminative stimulus effects of pentobarbital. Dizocilpine and phencyclidine partly substituted (58 and 57%, respectively) for ethanol without substantial pentobarbital-appropriate responding. RU 24969, CGS 12066B and morphine did not result in complete substitution for either ethanol or pentobarbital, although RU 24969 resulted in partial (68%) pentobarbital substitution. The ability to train the present three-choice discrimination in rats indicates that the discriminative stimulus effects of 10.0 mg/kg pentobarbital were separable from those of 2.0 g/kg ethanol. The results suggest that the pharmacological effects of ethanol, which can control behavior, may seemingly be modified by training conditions (two-versus three-choice discrimination procedures), to the extent that a receptor system prominently linked to the behavioral activity of ethanol (i.e. GABAA) appears no longer to be involved in the interoceptive effects of the drug.

Assessment of the mixed discriminative stimulus effects of ethanol in a three-choice ethanol-dizocilpine-water discrimination in rats

Previous drug discrimination studies have elucidated the importance of the NMDA, GABA(A) and 5-HT(1) receptor systems in mediating the discriminative stimulus effects of ethanol. The present study used a three-choice drug discrimination paradigm in an attempt to determine whether the salient NMDA antagonistic effects were separable from other stimulus effects of ethanol. Adult Long-Evans rats (n = 7) were trained to discriminate ethanol (1.5g/kg, intragastric (i.g.)), the uncompetitive NMDA antagonist dizocilpine (0.17mg/kg, i.g.) or water (3.5ml, i.g.) under a food-reinforced fixed-ratio 15 (FR15) schedule of reinforcement. Following training, substitution tests were conducted with the GABA(A)/benzodiazepine (GABA(A)/BDZ) positive modulator pentobarbital (PB, 5.6-17mg/kg, i.g.), the uncompetitive NMDA antagonist phenycldine (PCP, 0.1-5.6mg/kg, i.p.) and the 5-HT(1) agonist RU 24969 (0.1-3.0mg/kg, i.p.). Complete substitution of PCP (ED(50), 0.9mg/kg) for dizocilpine was found in all animals. Conversely, PB (ED(50), 10mg/kg) substituted fully for ethanol in five of seven animals, whereas RU 24969 (ED(50), 1.4mg/kg) completely substituted for ethanol in only three of seven animals tested. The result demonstrate that a three-choice discrimination using dizocilpine, ethanol and water as training conditions can be established in rats. By contrasting the discriminative stimulus effects of an uncompetitive NMDA antagonist to ethanol, the ethanol-like effects of pentobarbital and RU 24969 are attenuated compared to previous studies of two-choice ethanol water discrimination.