Tony M. Kerr

Dopamine activation of endogenous cannabinoid signaling in dorsal striatum

We measured endogenous cannabinoid release in dorsal striatum of freely moving rats by microdialysis and gas chromatography/mass spectrometry. Neural activity stimulated the release of anandamide, but not of other endogenous cannabinoids such as 2-arachidonylglycerol. Moreover, anandamide release was increased eightfold over baseline after local administration of the D2-like (D2, D3, D4) dopamine receptor agonist quinpirole, a response that was prevented by the D2-like receptor antagonist raclopride. Administration of the D1-like (D1, D5) receptor agonist SKF38393 had no such effect. These results suggest that functional interactions between endocannabinoid and dopaminergic systems may contribute to striatal signaling. In agreement with this hypothesis, pretreatment with the cannabinoid antagonist SR141716A enhanced the stimulation of motor behavior elicited by systemic administration of quinpirole. The endocannabinoid system therefore may act as an inhibitory feedback mechanism countering dopamine-induced facilitation of motor activity.

Enduring Resistance to Extinction of Cocaine-Seeking Behavior Induced by Drug-Related Cues

The conditioning of cocaines pharmacological actions with environmental stimuli is thought to be a critical factor in long-lasting relapse risk associated with cocaine addiction. To study the significance of environmental stimuli in enduring vulnerability to relapse, the resistance to extinction of drug-seeking behavior elicited by a cocaine-related stimulus was examined. Male Wistar rats were trained to associate discriminative stimuli (SD) with the availability of intravenous cocaine (S+) vs. the availability of non-rewarding (S−) saline solution, and then placed on extinction conditions during which intravenous solutions and SD were withheld. The rats were then presented with the S+ or S− alone in 60-min reinstatement sessions conducted at 3-day intervals. To examine the long-term persistence of the motivating effects of the cocaine S+, a subgroup of rats was re-tested following an additional three months of abstinence during which time the rats remained confined to their home cages. Re-exposure to the cocaine S+ selectively elicited robust responding at the previously active lever. The efficacy and selectivity of this stimulus to elicit responding remained unaltered throughout a 34-day phase of repeated testing as well as following the additional extended abstinence period. In pharmacological tests, conducted in a separate group of rats, the dopamine (DA) D1 antagonist SCH 39166 (10 μg/kg), the D2/3 antagonist nafadotride (1 mg/kg), and the D2/3 agonist PD 128907 (0.3 mg/kg) suppressed the cue-induced response reinstatement while the D1 agonist SKF 81297 (1.0 mg/kg) produced a variable behavioral profile attenuating cue-induced responding in some rats while exacerbating this behavior in others. The results suggest that the motivating effects of cocaine-related stimuli are highly resistant to extinction. The undiminished efficacy of the cocaine S+ to induce drug-seeking behavior both with repeated testing and following long-term abstinence parallels the long-lasting nature of conditioned cue reactivity and cue-induced cocaine craving in humans, and confirms a significant role of learning factors in long-lasting vulnerability to relapse associated with cocaine addiction. Finally, the results support a role of DA neurotransmission in cue-induced cocaine-seeking behavior.

5-HT1A receptor mutant mice exhibit enhanced tonic, stress-induced and fluoxetine-induced serotonergic neurotransmission

Mutant mice that lack serotonin1A receptors exhibit enhanced anxiety‐related behaviors, a phenotype that is hypothesized to result from impaired autoinhibitory control of midbrain serotonergic neuronal firing. Here we examined the impact of serotonin1A receptor deletion on forebrain serotonin neurotransmission using in vivo microdialysis in the frontal cortex and ventral hippocampus of serotonin1A receptor mutant and wild‐type mice. Baseline dialysate serotonin levels were significantly elevated in mutant animals as compared with wild‐types both in frontal cortex (mutant = 0.44 ± 0.05 n m; wild‐type = 0.28 ± 0.03 n m) and hippocampus (mutant = 0.46 ± 0.07 n m; wild‐type = 0.27 ± 0.04 n m). A stressor known to elicit enhanced anxiety‐like behaviors in serotonin1A receptor mutants increased dialysate 5‐HT levels in the frontal cortex of mutant mice by 144% while producing no alteration in cortical 5‐HT in wild‐type mice. There was no phenotypic difference in the effect of this stressor on serotonin levels in the hippocampus. Fluoxetine produced significantly greater increases in dialysate 5‐HT content in serotonin1A receptor mutants as compared with wild‐types, with two‐ and three‐fold greater responses being observed in the hippocampus and frontal cortex, respectively. This phenotypic effect was mimicked in wild‐types by pretreatment with the serotonin1A antagonist 4‐iodo‐N‐[2‐[4‐(methoxyphenyl)‐1‐piperazinyl]ethyl]‐N‐2‐pyridinyl‐benzamide (p‐MPPI). These results indicate that deletion of central serotonin1A receptors results in a tonic disinhibition of central serotonin neurotransmission, with a greater dysregulation of serotonin release in the frontal cortex than ventral hippocampus under conditions of stress or increased interstitial serotonin levels.

Ethanol anticipation enhances dopamine efflux in the nucleus accumbens of alcohol-preferring (P) but not Wistar rats.

It has been argued that the anticipation of ethanol consumption can activate reinforcement substrates involved in alcohol-seeking behavior. To test this hypothesis nucleus accumbens (NAc) dopamine (DA) efflux was monitored in alcohol-preferring (P) and Wistar rats. Rats from each line were divided into two groups: one trained to self-administer 0.05% saccharin and the other trained to self-administer 10% (w/v) ethanol. On the test day dopamine efflux was monitored in all animals during saccharin self-administration. In ethanol-expecting P rats, self-administration of saccharin produced a significant elevation in extracellular DA (150% of baseline within the first 15-20min). In contrast, a significant increase in extracellular DA was only observed during the final 30min of the test session in ethanol-expecting Wistar rats self-administering saccharin. The self-administration of saccharin in animals expecting the sweetener failed to elevate extracellular DA in both strains. Overall, these results suggest that the mere expectation of ethanol availability enhances the efflux of DA in the NAc of the P, but not the Wistar rat, which may play a role in the initiation or maintenance of ethanol seeking behavior in the P line.

Simple microbore high-performance liquid chromatographic method for the determination of dopamine and cocaine from a single in vivo brain microdialysis sample.

A microbore chromatographic method for the analysis of both dopamine and cocaine from in vivo brain microdialysis samples is described. To eliminate the need for separate chromatographic systems for each analyte, post-column electrochemical and ultraviolet detection systems were arranged in series. The limit of quantitation for dopamine (5 fmol) was well within range for detecting dialysate concentrations of this neurotransmitter in rats which were in a baseline, drug-free state. The limit of quantitation for cocaine (0.5 pmol) was sufficient to detect brain cocaine levels following the peripheral administration of a low dose of this psychostimulant (5 mg/kg, i.p.). Estimates of dialysate dopamine and cocaine concentrations after 5, 10 and 20 mg/kg cocaine (i.p.) were in agreement with reports which utilized separate HPLC analyses for each analyte.

Orexin-A/Hypocretin-1 Mediates Cocaine-Seeking Behavior in the Posterior Paraventricular Nucleus of the Thalamus via Orexin/Hypocretin Receptor-2

Orexin/hypocretin (Orx/Hcrt) projections from the lateral hypothalamus to the paraventricular nucleus of the thalamus (PVT) are implicated in drug addiction. Specifically, the posterior section of the PVT (pPVT) innervates brain structures that modulate motivated behavior. This study investigated the role of pPVT-Orx/Hcrt transmission in cocaine-seeking behavior. Because the effects of Orx/Hcrt are mediated by two Orx/Hcrt receptors (Hcrt-r1 and Hcrt-r2), we examined the extent to which Hcrt-r1 and Hcrt-r2 are involved in Orx/Hcrt-induced cocaine seeking. Male Wistar rats were made cocaine dependent by self-administering cocaine 6 hours/day (long access) for 21 days. After self-administration training, the rats underwent daily extinction training, during which cocaine was withheld. After extinction, the rats were injected into the pPVT with Orx-A/Hcrt-1 (0–2 µg) alone or, using a single dose of 0.5 µg, in combination with an Hcrt-r1 antagonist (SB334867; 0–15 µg) or an Hcrt-r2 antagonist (TCSOX229; 0–15 µg). Orx-A/Hcrt-1 alone reinstated (primed) cocaine seeking. Unexpectedly, coadministration of Orx-A/Hcrt-1 with SB334867 did not have any effects on Orx-A/Hcrt-1-induced reinstatement, whereas when coadministered with Orx-A/Hcrt-1, TCSOX229 prevented cocaine-seeking behavior. These results indicate that Hcrt-r2 in the pPVT mediates the reinstating effect of Orx-A/Hcrt-1 in animals with a history of cocaine dependence and further identify Hcrt-r2 as a possible molecular target that can guide future therapeutic approaches for the prevention of drug-seeking behavior.

The paraventricular nucleus of the thalamus is differentially recruited by stimuli conditioned to the availability of cocaine versus palatable food.

The paraventricular nucleus of the thalamus (PVT) is not traditionally considered part of the brain addiction neurocircuitry but has received growing attention with regard to a role in the modulation of drug‐seeking behavior. This study sought to establish the pattern of neural activation induced by a response‐reinstating discriminative stimulus (SD) conditioned to either cocaine (COC) or a conventional reinforcer using a palatable food substance, sweetened condensed milk (SCM). Male Wistar rats were trained to associate one SD (S+; COC or SCM availability) and a distinctly different SD (S−; non‐reward; i.e. the availability of saline or the absence of SCM). Following extinction of COC‐ and SCM‐reinforced responding, rats were presented with the respective S+ or S− alone and tested for the reinstatement of reward seeking. The COC S+ and SCM S+ elicited identical reinstatement, whereas the non‐reward S− was behaviorally ineffective. PVT sections were obtained following completion of the reinstatement tests and labeled for Fos. The number of Fos+ neurons was compared among rats that were presented with the COC S+, SCM S+ or S−. Rats that were presented with the COC S+ exhibited a significant increase in Fos expression compared with rats that were presented with the S−. Moreover, Fos expression was significantly correlated with the number of reinstatement responses that were induced by the COC S+. In contrast, the SCM S+ and S− produced identical increases in Fos expression, without behaviorally relevant correlations. The findings implicate the PVT as an important site that is selectively recruited during COC‐seeking behavior.

Distinct memory engrams in the infralimbic cortex of rats control opposing environmental actions on a learned behavior

Conflicting evidence exists regarding the role of infralimbic cortex (IL) in the environmental control of appetitive behavior. Inhibition of IL, irrespective of its intrinsic neural activity, attenuates not only the ability of environmental cues predictive of reward availability to promote reward seeking, but also the ability of environmental cues predictive of reward omission to suppress this behavior. Here we report that such bidirectional behavioral modulation in rats is mediated by functionally distinct units of neurons (neural ensembles) that are concurrently localized within the same IL brain area but selectively reactive to different environmental cues. Ensemble-specific neural activity is thought to function as a memory engram representing a learned association between environment and behavior. Our findings establish the causal evidence for the concurrent existence of two distinct engrams within a single brain site, each mediating opposing environmental actions on a learned behavior. DOI: http://dx.doi.org/10.7554/eLife.21920.001

Differential role of hypothalamic orexin/hypocretin neurons in reward seeking motivated by cocaine versus palatable food

Hypothalamic orexin/hypocretin (Orx/Hcrt) neurons are thought to mediate both food‐reinforced behaviors and behavior motivated by drugs of abuse. However, the relative role of the Orx/Hcrt system in behavior motivated by food versus drugs of abuse remains unclear. This investigation addressed this question by contrasting hypothalamic Orx/Hcrt neuronal activation associated with reinstatement of reward seeking induced by stimuli conditioned to cocaine (COC) versus highly palatable food reward, sweetened condensed milk (SCM). Orx/Hcrt neuronal activation in the lateral hypothalamus, dorsomedial hypothalamus and perifornical area, determined by dual c‐fos/orx immunocytochemistry, was quantified in rat brains, following reinstatement of reward seeking induced by a discriminative stimulus (S+) conditioned to COC or SCM. The COC S+ and SCM S+ initially produced the same magnitude of reward seeking. However, over four subsequent tests, behavior induced by the SCM S+ decreased to extinction levels, whereas reinstatement induced by the COC S+ perseverated at undiminished levels. Following both the first and fourth tests, the percentage of Orx/Hcrt cells expressing Fos was significantly increased in all hypothalamic subregions in rats tested with the COC S+ but not rats tested with the SCM S+. These findings point toward a role for the Orx/Hcrt system in perseverating, compulsive‐like COC seeking but not behavior motivated by palatable food. Moreover, analysis of the Orx/Hcrt recruitment patterns suggests that failure of Orx/Hcrt neurons in the lateral hypothalamus to respond to inhibitory inputs from Orx/Hcrt neurons in the dorsomedial hypothalamus/perifornical area may contribute to the perseverating nature of COC seeking.

Unique treatment potential of cannabidiol for the prevention of relapse to drug use: preclinical proof of principle

Cannabidiol (CBD), the major non-psychoactive constituent of Cannabis sativa, has received attention for therapeutic potential in treating neurologic and psychiatric disorders. Recently, CBD has also been explored for potential in treating drug addiction. Substance use disorders are chronically relapsing conditions and relapse risk persists for multiple reasons including craving induced by drug contexts, susceptibility to stress, elevated anxiety, and impaired impulse control. Here, we evaluated the “anti-relapse” potential of a transdermal CBD preparation in animal models of drug seeking, anxiety and impulsivity. Rats with alcohol or cocaine self-administration histories received transdermal CBD at 24 h intervals for 7 days and were tested for context and stress-induced reinstatement, as well as experimental anxiety on the elevated plus maze. Effects on impulsive behavior were established using a delay-discounting task following recovery from a 7-day dependence-inducing alcohol intoxication regimen. CBD attenuated context-induced and stress-induced drug seeking without tolerance, sedative effects, or interference with normal motivated behavior. Following treatment termination, reinstatement remained attenuated up to ≈5 months although plasma and brain CBD levels remained detectable only for 3 days. CBD also reduced experimental anxiety and prevented the development of high impulsivity in rats with an alcohol dependence history. The results provide proof of principle supporting potential of CBD in relapse prevention along two dimensions: beneficial actions across several vulnerability states and long-lasting effects with only brief treatment. The findings also inform the ongoing medical marijuana debate concerning medical benefits of non-psychoactive cannabinoids and their promise for development and use as therapeutics.