Jonathan L. Katz

The validity of the reinstatement model of craving and relapse to drug use.

RationaleThe reinstatement procedure has been used increasingly as a laboratory model of craving and relapse to drug abuse. With the number of reports involving this procedure growing, its validity as a model of relapse merits discussion.ObjectivesThe present commentary addresses the validity of the reinstatement procedure in relation to the following three types of models: 1) formal equivalence models, which are assessed on the basis of how well they resemble some phenomenon outside the laboratory (i.e. face validity); 2) correlational models, which are assessed on the basis of how well they predict outcomes of various interventions (such as drug administration or environmental change) when effected outside the laboratory (i.e. predictive validity); and 3) functional equivalence models, which are assessed on the basis of whether the laboratory phenomenon is mechanistically identical or reasonably similar to the phenomenon outside the laboratory (i.e. content validity).MethodsIn order to evaluate the reinstatement model, we briefly examined its various forms and uses, and compared preclinical outcomes to what is known about relapse from the clinical literature.ResultsIn its most general form, the reinstatement model has reasonable face validity; that is, there is a general agreement in appearance or form of the behavior in the model and the clinical target, relapse. This face validity is generally absent for the procedure when it is used as a model of craving. The predictive validity of the model has not been established. Evidence from studies of treatments for drug relapse have not supported the validity of the model, however from studies of the effects of the presentation of various types of stimuli (e.g. drug priming) there is mixed evidence supporting predictive validity. With regard to functional equivalence, there is reasonable evidence supporting functional commonalities between drug self-administration in laboratory animals and human drug abusers, which lends support to the validity of the reinstatement model. However, there are several specific areas of departure between the methods and results using the model and clinical practices and observations about relapse, suggesting a lack of functional equivalence.ConclusionsThere is reasonable evidence to support the face validity of the model, but at this time, neither its predictive validity nor functional equivalence has been fully established, which underscores the need for caution in generalizing results from the model to the clinical condition.

Relationship between conformational changes in the dopamine transporter and cocaine-like subjective effects of uptake inhibitors.

Cocaine exerts its stimulatory effect by inhibiting the dopamine transporter (DAT). However, novel benztropine- and rimcazole-based inhibitors show reduced stimulant effects compared with cocaine, despite higher affinity and selectivity for DAT. To investigate possible mechanisms, we compared the subjective effects of different inhibitors with their molecular mode of interaction at the DAT. We determined how different inhibitors affected accessibility of the sulfhydryl-reactive reagent [2-(trimethylammonium)ethyl]-methanethiosulfonate to an inserted cysteine (I159C), which is accessible when the extracellular transporter gate is open but inaccessible when it is closed. The data indicated that cocaine analogs bind an open conformation, whereas benztropine and rimcazole analogs bind a closed conformation. Next, we investigated the changes in inhibition potency of [3H]dopamine uptake of the compounds at a mutant DAT (Y335A) characterized by a global change in the conformational equilibrium. We observed a close relationship between the decrease in potencies of inhibitors at this mutant and cocaine-like responding in rats trained to discriminate cocaine from saline injections. Our data suggest that chemically different DAT inhibitors stabilize distinct transporter conformations and that this in turn affects the cocaine-like subjective effects of these compounds in vivo.

Cocaine-induced locomotor activity and cocaine discrimination in dopamine D4 receptor mutant mice

RationalePrevious studies have found a role for dopamine D2-like receptors in many of the behavioral effects of cocaine, including its stimulation of locomotor activity and interoceptive discriminative-stimulus effects. However, given the lack of selectivity of most of the available pharmacological tools among D2, D3 and D4 dopamine receptors, the roles of these specific receptors remain unclear.ObjectivesThe roles of specific dopamine D4 receptors in the behavioral effects of cocaine, including its locomotor stimulant and interoceptive discriminative-stimulus effects were investigated using dopamine D4 receptor knockout (DA D4R KO) and wild-type (WT) mice.MethodsThe mice were trained in daily sessions to discriminate IP injections of saline from cocaine (10xa0mg/kg). Responses on one of two response keys intermittently produced a food pellet; one response was reinforced in sessions following cocaine injection (10xa0mg/kg), and the other response was reinforced in sessions following saline injection. Each 20th response produced a food pellet (fixed-ratio, or FR20 schedule of reinforcement). The dose-effects of cocaine and its interaction with the D2-like antagonist, raclopride, were assessed. Horizontal locomotor activity was also assessed in each genotype.ResultsAs previously shown), cocaine was a more potent stimulant of locomotor activity in the DA D4R KO mice compared to WT littermate mice. In addition, cocaine was more potent in producing discriminative-stimulus effects in DA D4R KO mice (ED50 value=0.50xa0mg/kg) compared to their WT littermates (ED50 value=2.6xa0mg/kg). Raclopride shifted the cocaine dose-effect curve in both DA D4R KO and WT mice, though the shift was greater for the DA D4R KO mice.ConclusionsThe present results on the stimulation of activity and interoceptive/subjective effects of cocaine are consistent with the previously reported disregulation of dopamine synthesis in DA D4R KO mice, and further suggest a role of the DA D4R in vulnerability to stimulant abuse.

Assessment of reinforcing effects of benztropine analogs and their effects on cocaine self-administration in rats: comparisons with monoamine uptake inhibitors.

Benztropine (BZT) analogs inhibit dopamine uptake but are less effective than cocaine in producing behavioral effects predicting abuse liability. The present study compared reinforcing effects of intravenous BZT analogs with those of standard monoamine uptake inhibitors and the effects of their oral pretreatment on cocaine self-administration. Responding of rats was maintained by cocaine [0.032–1.0 mg/kg/injection (inj)] or food reinforcement under fixed-ratio five-response schedules. Maximal rates of responding were maintained by 0.32 mg/kg/inj cocaine or substituted methylphenidate, with lower rates maintained at lower and higher doses. The N-methyl BZT analog, AHN 1-055 (3α-[bis(4′-fluorophenyl)methoxy]-tropane), also maintained responding (0.1 mg/kg/inj), although maximal rates were less than those with cocaine. Responding was not maintained above vehicle levels by the N-allyl, AHN 2-005 (N-allyl-3α-[bis(4′-fluorophenyl)methoxy]-tropane), and N-butyl, JHW 007 [N-(n-butyl)-3α-[bis(4′-fluorophenyl)methoxy]-tropane], BZT analogs, and it was not maintained with nisoxetine or citalopram. Presession treatment with methylphenidate (3.2–32 mg/kg) dose-dependently shifted the cocaine self-administration dose-effect curve leftward, whereas nisoxetine and citalopram effects were not significant. An intermediate dose of AHN 1-055 (32 mg/kg) increased responding maintained by low cocaine doses and decreased responding maintained by higher doses. A higher dose of AHN 1-055 completely suppressed cocaine-maintained responding. Both AHN 2-005 and JHW 007 dose-dependently (10–32 mg/kg) decreased cocaine self-administration, shifting its dose-effect curve down. Decreases in cocaine-maintained responding occurred at doses of methylphenidate and BZT analogs that left food-maintained responding unchanged. During a component in which injections were not available, methylphenidate and AHN 1-055, but not AHN 2-005 or JHW 007, increased response rates. These findings further support the low abuse liability of BZT analogs and their potential development as medications for cocaine abuse.

Reinforcing Effects of σ-Receptor Agonists in Rats Trained to Self-Administer Cocaine

σ-Receptor (σR) antagonists have been reported to block certain effects of psychostimulant drugs. The present study examined the effects of σR ligands in rats trained to self-administer cocaine (0.032–1.0 mg/kg/inj i.v.) under fixed-ratio 5-response schedules of reinforcement. Maximal rates of responding were maintained by 0.32 mg/kg/inj cocaine, or by the σR agonists, 1,3-di-(2-tolyl)guanidine (DTG; 1.0 mg/kg/inj) or 2-(4-morpholinethyl) 1-phenylcyclohexane-1-carboxylate hydrochloride (PRE-084; 0.32 mg/kg/inj), when substituted for cocaine. Lower response rates were maintained at higher and lower doses of the compounds. No dose of the σR antagonists [N-[2-(3,4-dichlorophenyl)ethyl]-N-methyl-2-(1-pyrrolidinyl)ethylamine (BD 1008), N-[2-(3,4-dichlorophenyl)ethyl]-N-methyl-2-(dimethylamino)ethylamine (BD 1047), N-[2-(3,4-dichlorophenyl)ethyl]-4-methylpiperazine (BD 1063)] maintained responding appreciably above levels obtained when responding had no consequences. Presession treatment with σR agonists dose-dependently shifted the cocaine self-administration dose-effect curve leftward. The dopamine-uptake inhibitor, (−)-2β-carbomethoxy-3β-(4-fluorophenyl)tropane (WIN 35,428), dose-dependently shifted the DTG and PRE-084 self-administration dose-effect curves leftward. Treatment with the σR antagonists dose-dependently decreased response rates maintained by DTG or PRE-084, but did not affect cocaine self-administration. Response rates maintained by maximally effective DTG or PRE-084 doses were decreased by σR antagonists at lower doses than those that decreased response rates maintained by food reinforcement. Although σR antagonists block some cocaine-induced effects, the lack of effect on cocaine self-administration suggests that the primary reinforcing effects of cocaine do not involve direct effects at σRs. However, the self-administration of σR agonists in cocaine-trained subjects, facilitation of cocaine self-administration by σR-agonist pretreatment, and the facilitation of σR-agonist self-administration by WIN 35,428, together suggest enhanced abuse-related effects resulting from concomitant dopaminergically mediated actions and σR-mediated actions of the drugs.

Variability of drug self-administration in rats

RationaleAlthough temporal patterns of drug self-administration in animals are known to be highly regular, this regularity has rarely been quantified or systematically compared across reinforcers.ObjectivesOver a range of unit doses, this study assessed: (1) the within-subject variability of inter-infusion intervals (latencies); (2) the estimated whole-body drug level at the time of self-infusion; (3) the within-subject variability of these drug levels; and (4) the statistical dependence between successive latencies, to determine whether regularity is maintained by compensatory, moment-to-moment adjustment of latencies.MethodsRats were trained with cocaine (10–1000xa0μg/kg per infusion, IV), remifentanil (an ultra-short acting opioid; 0.25–32xa0μg/kg per infusion, IV), or food (20–180xa0mg/delivery).ResultsWithin subjects, latencies were most consistent from infusion to infusion at unit doses on the descending limb of the dose-response curve. However, the drug level at the time an infusion was initiated was actually least consistent at these doses. Sequential latencies showed only a weak autocorrelation for both drugs.ConclusionThese results suggest that highly consistent response patterns are not simply a product of precise titration of drug levels. The weak autocorrelation between sequential latencies suggests that temporal regularity of responding is not maintained through compensatory adjustments of post-infusion pauses.

Relationships among dopamine transporter affinities and cocaine-like discriminative-stimulus effects

Abstractu2005 Rationale: The discriminative-stimulus effects of cocaine have been reported to be mediated by indirect agonist actions initiated by the blockade of dopamine uptake, and the potencies of drugs that have discriminative-stimulus effects like cocaine are directly related to their dopamine transporter binding affinities. The binding to the dopamine transporter by cocaine and many of its analogs has been reported to fit better using a two-site model than a one-site model. Objectives: The present study examined the relationship among binding affinities of dopamine uptake inhibitors at these two sites and their potencies to produce discriminative-stimulus effects. Methods: The inhibition constants (Ki values) were derived for unlabeled dopamine uptake inhibitors for displacement of [3H]WIN 35,428 from rat caudate putamen membranes. These Ki values were related to the ED50 values obtained in rats trained to discriminate 10 mg/kg cocaine from saline injections under a fixed-ratio 20 schedule of food reinforcement. Results: Among the dopamine uptake inhibitors studied, the binding data for eight compounds (WIN 35,428, nomifensine, WIN 35,981, WIN 35,065-2, methylphenidate, cocaine, cocaethylene, and bupropion) were better fit by a two-site model than a one-site model. The data for the remaining eleven compounds (RTI-31, RTI-55, RTI-121, RTI-32, LU19-005, BTCP, GBR12909, GBR12935, mazindol, LU17-133, and EXP561) were better fit by a one-site model. Of the drugs that were fit best by a two-site model, there was a higher correlation among the Ki values for the high-affinity site and the ED50 values (R2=0.655; P=0.015) than there was for the low-affinity site (R2=0.543; P=0.037). Of the remaining drugs, there was a high correlation among the Ki values and the ED50 values for the discriminative-stimulus effects (R2=0.523; P=0.012). Conclusions: These data suggest that the discriminative-stimulus effects of cocaine are more closely related to actions mediated by high-affinity binding to the dopamine transporter than they are to actions mediated by the low-affinity site. The further assessment of the respective contributions of high- and low-affinity binding to the behavioral effects of cocaine will be greatly enhanced with the development of pharmacological tools that have a high degree of selectivity for one of these components.

Decreases in cocaine self-administration with dual inhibition of the dopamine transporter and σ receptors.

Sigma receptor (σR) antagonists attenuate many behavioral effects of cocaine but typically not its reinforcing effects in self-administration procedures. However, the σR antagonist rimcazole and its N-propylphenyl analogs, [3-(cis-3,5-dimethyl-4-[3-phenylpropyl]-1-piperazinyl)-propyl]diphenylamine hydrochloride (SH 3-24) and 9-[3-(cis-3,5-dimethyl-4-[3-phenylpropyl]-1-piperazinyl)-propyl]carbazole hydrobromide (SH 3-28), dose-dependently decreased the maximal rates of cocaine self-administration without affecting comparable responding maintained by food reinforcement. In contrast, a variety of σR antagonists [N-phenethylpiperidine oxalate (AC927), N-[2-(3,4-dichlorophenyl)ethyl]-N-methyl-2-(1-pyrrolidinyl)ethylamine dihydrobromide (BD 1008), N-[2-(3,4-dichlorophenyl)ethyl]-N-methyl-2-(dimethylamino) ethylamine dihydrobromide (BD 1047), N-[2-(3,4-dichlorophenyl) ethyl]-4-methylpiperazine dihydrochloride (BD 1063), and N,N-dipropyl-2-[4-methoxy-3-(2-phenylethoxy)phenyl]-ethylamine monohydrochloride (NE-100)] had no effect on cocaine self-administration across the range of doses that decreased rates of food-maintained responding. Rimcazole analogs differed from selective σR antagonists in their dual affinities for σRs and the dopamine transporter (DAT) assessed with radioligand binding. Selective DAT inhibitors and σR antagonists were studied alone and in combination on cocaine self-administration to determine whether actions at both σRs and the DAT were sufficient to reproduce the effects of rimcazole analogs. Typical DAT inhibitors [2β-carbomethoxy-3β-(4-fluorophenyl)tropane (WIN 35,428), methylphenidate, and nomifensine] dose-dependently shifted the cocaine dose-effect curve leftward. Combinations of DAT inhibitor and σR antagonist doses that were behaviorally inactive alone decreased cocaine self-administration without effects on food-maintained responding. In addition, whereas the DAT inhibitors were self-administered at rates similar to those of cocaine, neither rimcazole analogs nor typical σR antagonists (NE-100 and AC927) maintained responding above control levels across a wide range of doses. These findings suggest that the unique effects of rimcazole analogs are due to dual actions at the DAT and σRs and that a combined target approach may have utility in development of medical treatments for cocaine abuse.

Human cocaine-seeking behavior and its control by drug-associated stimuli in the laboratory.

Second-order schedules of drug self-administration were developed to incorporate the effects of drug-related environmental stimuli into an animal model of drug abuse, making it more similar to human situations. Ironically, little is known about how human subjects behave under these schedules. In this study, human volunteers with a history of cocaine use worked on a second-order schedule in which every 100th lever response produced an auditory–visual brief stimulus (2 s). The first stimulus produced after 1 h was extended to 10 s and paired with an intravenous injection of cocaine (25 mg). Up to three injections were allowed per session. In different phases of the experiment, presentation of the brief stimulus was discontinued and/or saline solution (placebo) was injected instead of cocaine. Injections of cocaine were found to maintain responding even when the brief stimulus was not presented. Placebo injections alone did not maintain responding. In contrast, the brief stimulus maintained high levels of responding under placebo conditions, even though self-reports indicated that subjects could clearly discriminate that they were not receiving cocaine. These results demonstrate that drug-related environmental stimuli can maintain persistent drug seeking during periods of drug unavailability. As this procedure directly measures the effects of stimuli on drug seeking, it may provide a valuable complement to indirect measures, such as self-reports of craving, that are often used with human subjects. The similarity of the response patterns in humans and animals also supports the use of second-order schedules in animals as a valid model of human drug seeking.

Dopamine transporter binding without cocaine-like behavioral effects: synthesis and evaluation of benztropine analogs alone and in combination with cocaine in rodents.

Abstract. Rationale: Previous SAR studies demonstrated that small halogen substitutions on the diphenylether system of benztropine (BZT), such as a para-Cl group, retained high affinity at the cocaine binding site on the dopamine transporter. Despite this high affinity, the compounds generally had behavioral effects different from those of cocaine. However, compounds with meta-Cl substitutions had effects more similar to those of cocaine. Objectives: A series of phenyl-ring analogs of benztropine (BZT) substituted with 3′-, 4′-, 3′,4′′- and 4′,4′′-position Cl-groups were synthesized and their pharmacology was evaluated in order to assess more fully the contributions to pharmacological activity of substituents in these positions. Methods: Compounds were synthesized and their pharmacological activity was assessed by examining radioligand binding and behavioral techniques. Results: All of the compounds displaced [3H]WIN 35,428 binding with affinities ranging from 20 to 32.5xa0nM. Affinities at norepinephrine ([3H]nisoxetine) and serotonin ([3H]citalopram) transporters, respectively, ranged from 259 to 5120 and 451 to 2980xa0nM. Each of the compounds also inhibited [3H]pirenzepine binding to muscarinic M1 receptors, with affinities ranging from 0.98 to 47.9xa0nM. Cocaine and the BZT analogs produced dose-related increases in locomotor activity in mice. However, maximal effects of the BZT analogs were uniformly less than those produced by cocaine, and were obtained 2–3xa0h after injection compared to the relatively rapid onset (within 30xa0min) of cocaine effects. In rats trained to discriminate IP saline from 29xa0µmol/kg cocaine (10xa0mg/kg), cocaine produced a dose-related increase in responding on the cocaine lever, reaching 100% at the training dose; however, none of the BZT analogs fully substituted for cocaine, with maximum cocaine responding from 20 to 69%. Despite their reduced efficacy compared to cocaine in cocaine discrimination, none of the analogs antagonized the effects of cocaine. As has been reported previously for 4′-Cl-BZT, the cocaine discriminative-stimulus effects were shifted leftward by co-administration of the present BZT analogs. Conclusions: The present results indicate that although the BZT analogs bind with relatively high affinity and selectivity at the dopamine transporter, their behavioral profile is distinct from that of cocaine. The present results suggest that analogs of BZT may be useful as treatments for cocaine abuse in situations in which an agonist treatment is indicated. These compounds possess features such as reduced efficacy compared to cocaine and a long duration of action that may render them particularly useful leads for the development of therapeutics for cocaine abusers.