Differential modulation of the discriminative stimulus effects of methamphetamine and cocaine by alprazolam and oxazepam in male and female rats.

Drug users often combine benzodiazepines with psychostimulants, such as methamphetamine. However, very little research has been conducted on this type of polydrug use, particularly in female subjects. The present study was therefore designed to examine the effects of two benzodiazepines, alprazolam and oxazepam, on the discriminative stimulus effects of methamphetamine and cocaine in both male and female rats. Rats were trained to discriminate methamphetamine (1.0 mg/kg, ip) or cocaine (10 mg/kg, ip) from saline using a two-lever operant, food-reinforced, drug discrimination design. Pretreatment with oxazepam (5, 10 and 20 mg/kg, ip) significantly attenuated methamphetamine discrimination in both male and female rats. In contrast, however, the high dose of alprazolam (4 mg/kg, ip) actually augmented the subjective effects of lower doses of methamphetamine (0.125 and 0.25 mg/kg, ip). Oxazepam produced similar effects on the subjective effects of cocaine as with methamphetamine, significantly reducing cocaine discrimination in both male and female rats. However, neither the high nor low dose of alprazolam (2 and 4 mg/kg, ip) produced any apparent effect on cocaine discrimination. Finally, while similar results were observed in both male and female rats across these experiments, methamphetamine and cocaine discrimination were more sensitive to oxazepam in female subjects. The results of these experiments suggest that alprazolam and oxazepam can differentially affect the subjective effects of methamphetamine and cocaine. These results also demonstrate that alprazolam can differentially affect the discriminative stimulus effects of methamphetamine and cocaine.

Effects of saline substitution on responding and plasma corticosterone in rats trained to self-administer different doses of cocaine.

Research from our laboratory has explored the role of the hypothalamo-pituitary-adrenal (HPA) axis in cocaine reinforcement. These experiments were designed to determine the involvement of the HPA axis in extinction. Male Wistar rats were trained to self-administer cocaine [0.125, 0.25, or 0.5 mg/kg/infusion (inf)] and food pellets (45 mg) under a multiple, alternating schedule of reinforcement. When self-administration was stable, saline was substituted for cocaine. Blood samples were taken at the end of the sessions following cocaine self-administration, the first exposure to saline substitution (first); and once the criteria for extinction were met (final). Plasma corticosterone was measured using radioimmunoassays. Although there was a significant increase in the number of infusions obtained during the first saline substitution test by rats trained with 0.5 mg/kg/inf of cocaine, there was a decrease in infusions received when 0.125 mg/kg/inf of cocaine was tested. Following repeated exposure to the extinction conditions, responding by rats trained to self-administer all three doses of cocaine was decreased to similar levels. In addition, there were significant differences in plasma corticosterone in rats trained with different doses of cocaine. Lever-pressing behavior and plasma corticosterone varied during extinction in relation to the training dose of cocaine and according to whether the rats had been exposed to single or repeated extinction testing. These data are discussed in terms of the potential difficulties involved in interpreting the effects of compounds intended to reduce drug reinforcement.

Effects of the CRH receptor antagonist CP-154,526 on intravenous cocaine self-administration in rats.

The role for corticotropin-releasing hormone (CRH) receptors in the maintenance of intravenous cocaine self-administration in rats was investigated using the centrally active, small molecule CRH1 receptor antagonist CP-154,526. In these experiments, adult male Wistar rats were allowed alternating 15-min periods of access to food reinforcement and cocaine self-administration (0.125, 0.25 or 0. 5 mg/kg/infusion) during daily 2-h sessions. A 1-min timeout separated access to the two reinforcers. Pretreatment with CP-154, 526 produced dose-related decreases in cocaine self-administration without affecting food-reinforced responding, suggesting a specific effect of the antagonist on cocaine-maintained behavior. Drug intake was decreased across several doses of cocaine, with the dose-response curve for cocaine self-administration shifted downward and flattened, suggesting that CP-154,526 decreased cocaine reinforcement. Furthermore, responding on the cocaine lever following CP-154,526 pretreatment was significantly suppressed, even during the first 15 min of the session, a time when rats typically sample the cocaine lever during extinction, suggesting that CRH receptors may also be involved in some of the conditioned effects of cocaine as well. These data are discussed in terms of the role for CRH in the neurobehavioral effects of cocaine.

Ketoconazole reduces low dose cocaine self-administration in rats.

Ketoconazole is an oral antimycotic agent approved by the FDA for the treatment of fungal disease which also blocks the synthesis of adrenocorticosteroids and functions as a glucocorticoid receptor antagonist. In these experiments, adult male Wistar rats were allowed alternating 15-min periods of access to food reinforcement and cocaine self-administration (0.125, 0.25 or 0.5 mg/kg per infusion) during daily 2-h sessions. A 1-min timeout separated access to the two reinforcers. Pretreatment with ketoconazole (25 mg/kg, i.p.) significantly decreased plasma corticosterone and reduced low dose (i.e. 0.125-0.25 mg/kg per infusion) cocaine self-administration without affecting food-reinforced responding. In fact, pretreatment with ketoconazole resulted in rates and patterns of self-administration at these doses that were indistinguishable from those observed during cocaine extinction. However, cocaine self-administration at the highest dose tested in these experiments (i.e. 0.5 mg/kg per infusion) was not significantly affected by ketoconazole. These data suggest the potential utility of ketoconazole or related drugs as adjuncts in the treatment of cocaine abuse and further underscore the role for corticosterone in cocaine reinforcement.

Tolerance and sensitization to the behavioral effects of cocaine in rats: relationship to benzodiazepine receptors.

Tolerance and sensitization to the behavioral effects of cocaine were investigated in rats responding under a fixed-consecutive-number eight schedule of food reinforcement. The development of tolerance or sensitization was induced by delivering the drug either immediately before or after each behavioral session during chronic administration. Chronic cocaine administered before each session resulted in tolerance, as indicated by the shift to the right in the cocaine dose response curve. This tolerance was more likely to develop in the presence of an external discriminative stimulus. On the other hand, when cocaine was delivered after each session, the injections did not disrupt responding and sensitization or increased sensitivity rather than tolerance developed. This sensitization was more likely to occur when the external discriminative stimulus was not present. These data suggest that either tolerance or sensitization to the behavioral effects of cocaine can occur following the same number of chronic injections, with the effect dependent on the context under which the drug is delivered. Significant differences in benzodiazepine receptor binding measured autoradiographically using [3H]flumazenil were observed between rats that received cocaine before or after each session, suggesting that the development of tolerance and sensitization may be mediated through changes in benzodiazepine receptors in discrete brain regions.

Role of corticosterone in intravenous cocaine self-administration in rats.

The role of corticosterone in cocaine reinforcement was investigated in rats exposed to either response-contingent electric footshock, noncontingent shock or no shock prior to the initiation of testing for intravenous cocaine self-administration. Although rats from the two shock groups were consistently more sensitive to cocaine, plasma corticosterone was always significantly higher in all rats that subsequently self-administered low doses of cocaine compared to the rats that did not, regardless of the treatment condition. In fact, plasma corticosterone was always greater than approximately 150 ng/ml for rats that self-administered low doses of cocaine, suggesting that this stress hormone must be increased above a critical threshold for stable low-dose cocaine self-administration to subsequently occur. Plasma corticosterone was also measured following exposure to cocaine and was significantly elevated in rats from all three treatment groups during cocaine self-administration, provided that doses of cocaine that would maintain responding were tested. When the dose would not maintain self-administration, plasma corticosterone was markedly lower, suggesting that at higher concentrations, the cocaine injections alone were sufficient to increase plasma corticosterone above a critical threshold, even for rats which had low precocaine levels of the hormone. These data suggest a significant role for corticosterone in both the acquisition and maintenance of cocaine reinforcement in rats.

Effects of surgical and pharmacological adrenalectomy on the initiation and maintenance of intravenous cocaine self-administration in rats.

Previous research has suggested the potential involvement of the hypothalamic-pituitary-adrenal (HPA) axis in psychostimulant reinforcement. In particular, we have found significant correlations between electric footshock-induced increases in plasma corticosterone and the acquisition, or lack thereof, of intravenous cocaine self-administration in rats. The experiments presented here were designed to further determine the role for corticosterone in cocaine reinforcement in rats by decreasing plasma levels of the hormone with surgical and pharmacological adrenalectomies. Bilateral adrenalectomy completely abolished the acquisition of intravenous cocaine self-administration over a wide range of doses (0.03125 to 1.0 mg/kg/infusion) without affecting food maintained responding. This suppression of self-administration was partially reversed by adding corticosterone (100 micrograms/ml) to the rats' drinking water. In another group of rats, pretreatment with metyrapone, which blocks the synthesis of corticosterone, resulted in dose-related decreases in ongoing cocaine self-administration. These data suggest that corticosterone is not only important, but may also be necessary for both the acquisition and maintenance of cocaine reinforcement in rats.

Non-contingent electric footshock facilitates the acquisition of intravenous cocaine self-administration in rats.

The experiments described below were designed to investigate whether contingent versus non-contingent electric footshock would affect the acquisition of intravenous cocaine self-administration in rats. During the first component of a multiple schedule, triads of rats were trained to respond under a discrete-trial, fixed-ratio 10 schedule of food reinforcement. Random footshock presentation (0.6 mA) for the first and second rats from each triad was yoked to food lever responding by the rats in the first group only, while the third group of rats was never shocked. When stable baselines of food-reinforced responding were obtained, all three rats from each triad were allowed to self-administer increasing doses of cocaine (0.031-0.5 mg/kg per infusion) during the second component. Rats from the second group, receiving non-contingent footshock presentation, self-administered cocaine (0.125 mg/kg per infusion) at higher rates and at one-half the dose which maintained responding in rats from the other two treatment groups. Plasma corticosterone, measured before the acquisition of cocaine self-administration, was highly correlated with drug intake at this concentration for all three groups of rats. These data demonstrate that non-contingent experimental stress facilities the acquisition of intravenous cocaine self-administration in rats.

Effects of alprazolam on intravenous cocaine self-administration in rats.

The specificity of benzodiazepine pretreatment on the reinforcing efficacy of cocaine was investigated using a multiple schedule of cocaine and food presentation. Cocaine was available under a fixed-ratio 4 schedule of reinforcement during 1 h of the session, while food was delivered under a discrete-trial, fixed-ratio 10 schedule during the other. Following initial exposure to alprazolam, responding maintained by both cocaine and food was significantly reduced. However, tolerance quickly developed to the sedative effects of alprazolam on food-maintained responding, while no reduction in the effects of the drug on cocaine self-administration was observed. Alprazolam (0.5 to 4.0 mg/kg, IP) significantly reduced cocaine intake without affecting food-maintained responding following subsequent testing with the drug. These data suggest a potential specific effect (e.g., anxiolytic) of alprazolam in cocaine reinforcement.

Lack of an effect of 6-hydroxydopamine lesions of the nucleus accumbens on intravenous morphine self-administration.

The neurotoxin, 6-hydroxydopamine (6-OHDA), has been used to selectively destroy dopamine containing neurons in discrete brain regions. Lesions of the nucleus accumbens with this neurotoxin decrease or eliminate cocaine and amphetamine self-administration and either increase or do not affect opiate self-administration in rats with unrestricted access to food and water. This study reports the effects of 6-OHDA lesions of the nucleus accumbens on responding maintained by food, water or morphine (3.3 mg/infusion). Six male rats with continuous access to three response levers were trained on a concurrent chained, fixed-ratio 1, fixed-ratio 9 schedule of reinforcer presentation. After stable patterns of responding were maintained by the three reinforcers, dose-effect curves for morphine were determined by substituting other doses of morphine or vehicle for 24-hour periods. Bilateral sham vehicle or 6-OHDA lesions of the nucleus accumbens were then completed and the effects of the lesion on food, water and morphine intake determined. Dose-effect evaluations were repeated after the lesion. The 6-OHDA lesions did not significantly affect responding maintained by food, water or morphine. The absence of an effect is most likely not the result of an insensitive baseline since other neurotoxin lesions produce long-term and selective decrements in morphine self-administration without affecting food and water responding. Like so many other manipulations, the magnitude of the effect that a neurotoxin lesion can exert on behavior may depend on the specific procedures that are used to maintain responding.

Kainic acid lesions of the nucleus accumbens selectively attenuate morphine self-administration.

The influence of kainic acid lesions of intrinsic and efferent neurons of the central medial nucleus accumbens on responding simultaneously maintained by food, water and morphine self-administration was assessed. Rats were trained on a multioperant baseline to respond on three different levers that resulted in either a food pellet, the presentation of a water dipper or an infusion of morphine. While responding on the morphine lever was related to dose (0.83-13.2 mg/infusion), increasing concentrations of the drug had little or no effect on responding maintained by food and water before the lesion. Bilateral infusions of the neurotoxin into the nucleus accumbens decreased morphine self-administration but did not appreciably alter food or water intake. Food extinction probes before the lesion produced significant increases in drug intake and decreases in responding on the water lever, but the neurotoxin lesion attenuated the food extinction induced decrease in water intake. These data suggested that kainic acid lesions of the nucleus accumbens decrease the reinforcing efficacy of morphine but do not alter the reinforcing properties of food and water. The neuronal systems potentially involved in mediating the reinforcing effects of environmental events are discussed.

Effects of 6-OHDA lesions of the central medial nucleus accumbens on rat intravenous morphine self-administration.

The function of dopaminergic innervations of the central medial nucleus accumbens in the processes maintaining intravenous morphine self-administration was assessed by lesioning with 6-OHDA and comparing drug intake with sham-vehicle treated littermates. Localized bilateral lesions of this structure resulted in significant increases in morphine intake shifting the dose-effect relationship to the right with twice the dose necessary to maintain prelesion rates of self-administration. Content of dopamine and dihydroxyphenylacetic acid was decreased in the nucleus accumbens after the lesion, but unchanged in the adjacent pyriform cortex and anterior caudate nucleus-putamen, while serotonin was significantly decreased in the pyriform cortex. High affinity uptake measurements also suggested nucleus accumbens dopaminergic and pyriform cortex serotonergic innervations to be affected by the lesion. The shift to the right in the dose effect relationship after the lesion suggests these neuronal systems to be excitatory to the processes mediating self-administration.

Reinforcing stimulus properties of endocoids.

Intracranial self-administration methodologies can be used to directly assess the neurobiological substrates of the reinforcing stimulus properties of drugs and endogenous ligands that may bind to receptors initiating these neuronal events. The reinforcing properties of the intracranial infusion of methionine enkephalin into the nucleus accumbens where it is released from nerve endings were demonstrated. The mechanisms of the reinforcing stimulus resulting from the intracranial self-administration of cocaine into the medial prefrontal cortex were shown to require presynaptic dopaminergic intervations and postsynaptic dopaminergic D2 receptors. The endogenous ligand that competes for cocaine binding may initate a reinforcing stimulus through similar mechanisms which could be directly assessed with intracranial self-administration procedures.

Reinforcer interactions under concurrent schedules of food, water, and intravenous morphine.

Responding by six rats was maintained under a concurrent chained fixed-ratio 1, fixed-ratio 9 schedule (conc chain FR1 FR9 ) of food, water, and morphine presentations. The subjects had continuous access to the schedule contingencies on a reversed 12-h light-dark cycle. Local rates and temporal patterns were very similar for responding maintained by the three reinforcers with food and water intake occurring predominantly during the dark cycle, while morphine infusions were evenly distributed. Food and water extinction (24-h duration) decreased the number of ratios completed on both the food and water levers. Moreover, food extinction resulted in a large increase in I.V. morphine self-administration. Morphine extinction increased responding on the morphine lever while almost eliminating responding on the water lever. Changes in the dose of morphine (2.5-40 mg/kg/injection) did not significantly affect food and water intake, but were inversely related to responding on the morphine lever. Saline substitutions resulted in effects similar to those observed during morphine extinction. The schedule used in this study provides a method for examining the specificity of a number of pharmacological and neurochemical manipulations.

Changes in biogenic amine and benzodiazepine receptors correlated with conditioned emotional response and its reversal by diazepam.

Groups of littermate rats were trained to respond for food reinforcement on a variable interval one-min (VI 1) schedule, after which they were classically conditioned to associate a conditioned stimulus (CS) with footshock (conditioned emotional response; conditioned suppression; CER). Two control groups received yoked footshock (no CS) or the visual-auditory stimulus only (no footshock). On test day, a group of the CER conditioned animals received injections of either vehicle or diazepam prior to exposure to the VI 1 food-reinforced schedule. After 30 min of the VI 1 schedule, the CS was presented continuously for 15 min, after which the animals were decapitated, the brains removed, membranes prepared and in vitro receptor binding evaluated. During the CS, the CER animals suppressed responding and exhibited conditioned fear (emotional) behavior, while the control groups, and animals given acute diazepam, maintained normal responding. [3H]Diazepam binding was reduced in the CER animals, yet acute benzodiazepine administration did not effect this binding. [3H]QNB binding was reduced by CER and increased by diazepam administration. Adrenergic, serotonergic and dopaminergic systems were also evaluated. Traditional biogenic amine systems may respond to CER and diazepam administration in some compensatory manner.