Cue-induced Reinstatement Tests Research Articles

Estrogen receptor beta signaling enhances extinction memory recall for heroin-conditioned cues in a sex- and region-specific manner

Return to use, or relapse, is a major challenge in the treatment of opioid use disorder (OUD). Relapse can be precipitated by several factors, including exposure to drug-conditioned cues. Identifying successful treatments to mitigate cue-induced relapse has been challenging, perhaps due to extinction memory recall (EMR) deficits. Previously, inhibition of estradiol (E2) signaling in the basolateral amygdala (BLA) impaired heroin-cue EMR. This effect was recapitulated by antagonism of BLA estrogen receptors (ER) in a sex-specific manner such that blocking ERα in males, but ERβ in females, impaired EMR. However, it is unclear whether increased E2 signaling, in the BLA or systemically, enhances heroin-cue EMR. We hypothesized that ERβ agonism would enhance heroin-cue EMR in a sex- and region-specific manner. To determine the capacity of E2 signaling to improve EMR, we pharmacologically manipulated ERβ across several translationally designed experiments. First, male and female rats acquired heroin or sucrose self-administration. Next, during a cued extinction session, we administered diarylpropionitrile (DPN, an ERβ agonist) and tested anxiety-like behavior on an open field. Subsequently, we assessed EMR in a cue-induced reinstatement test and, finally, measured ERβ expression in several brain regions. Across all experiments, females took more heroin and sucrose than males and had greater responses during heroin-cued extinction. Administration of DPN in the BLA enhanced EMR in females only, driven by ERβ’s impacts on memory consolidation. Interestingly, however, systemic DPN administration improved EMR for heroin cues in both sexes across several different tests, but did not impact sucrose-cue EMR. Immunohistochemical analysis of ERβ expression across several different brain regions showed that females only had greater expression of ERβ in the basal nucleus of the BLA. Here, in several preclinical experiments, we demonstrated that ERβ agonism enhances heroin-cue EMR and has potential utility in combatting cue-induced relapse.

Exploring ketamine’s reinforcement, cue-induced reinstatement, and nucleus accumbens cFos activation in male and female long evans rats

Ketamine (KET), a non-competitive N-methyl-d-aspartate (NMDA) receptor antagonist, has rapid onset of antidepressant effects in Treatment-Resistant Depression patients and repeated infusions are required to sustain its antidepressant properties. However, KET is an addictive drug, and so more preclinical and clinical research is needed to assess the safety of recurring treatments in both sexes. Thus, the aim of this study was to investigate the reinforcing properties of various doses of KET (0-, 0.125-, 0.25-, 0.5 mg/kg/infusion) and assess KET’s cue-induced reinstatement and neuronal activation in both sexes of Long Evans rats. Neuronal activation was assessed using the protein expression of the immediate early gene cFos in the nucleus accumbens (Nac), an important brain area implicated in reward, reinforcement and reinstatement to most drug-related cues. Our findings show that KET has reinforcing effects in both male and female rats, albeit exclusively at the highest two doses (0.25 and 0.5 mg/kg/infusion). Furthermore, we noted sex differences, particularly at the highest dose of ketamine, with female rats displaying a higher rate of self-administration. Interestingly, all groups that self-administered KET reinstated to drug-cues. Following drug cue-induced reinstatement test in rats exposed to KET (0.25 mg/kg/infusion) or saline, there was higher cFos protein expression in KET-treated animals compared to saline controls, and higher cFos expression in the core compared to the shell subregions of the Nac. As for reinstatement, there were no notable sex differences reported for cFos expression in the Nac. These findings reveal some sex and dose dependent effects in KET’s reinforcing properties and that KET at all doses induced similar reinstatement in both sexes. This study also demonstrated that cues associated with ketamine induce comparable neuronal activation in the Nac of both male and female rats. This work warrants further research into the potential addictive properties of KET, especially when administered at lower doses which are now being used in the clinic for treating various psychopathologies.

Pharmacological activation of the amygdala, but not single prolonged footshock-induced acute stress, interferes with cue-induced motivation toward food rewards in rats.

In the face of threats, animals adapt their behaviors to cope with the situation. Under such circumstances, irrelevant behaviors are usually suppressed. In this study, we examined whether food-seeking motivation would decrease under activation of the amygdala, an important nucleus in the regulation of stress response in the central nervous system, or after a physical acute stress session. In Experiment 1, we pharmacologically activated the basolateral nucleus (BLA) or the central nucleus of the amygdala (CeA) before a cue-induced reinstatement test in rats. Our results showed that activation of the BLA or the CeA abolished cue-induced motivation toward food rewards, while locomotor activity and free food intake were not affected. In Experiments 2 and 3, we further assessed anxiety and despair levels, as well as cue-induced reinstatement, after a single prolonged footshock-induced acute stress in rats. Behaviorally, acute stress did not affect anxiety level, despair level, or cue-induced motivation toward food rewards. Physiologically, there was no difference in cellular activities of the amygdala immediately after acute stress. To conclude, our results suggested that pharmacological activation of the amygdala decreased cue-induced motivation toward food reward. However, physiological acute stress did not immediately interfere with the negative emotions, motivation, or amygdala activities of the animals.

Inefficacy of N-acetylcysteine in mitigating cue-induced amphetamine-seeking

Glutamatergic imbalances are characteristic of SUDs. Astrocytic and neuronal transporters help regulate glutamate homeostasis and disruptions in this homeostasis engender SUD. The cysteine-glutamate exchanger (xCT) is primarily localized on astrocytes and maintains glutamate concentrations. This process is disrupted by cocaine use, and the therapeutic N-acetylcysteine (NAC) lowers cue-induced relapse to cocaine by restoring xCT function. However, little research has shown how these effects extend to other psychostimulants, such as amphetamine (AMP). Here, we assessed xCT expression following relapse to AMP cues, and if NAC can attenuate relapse via changes to astrocyte and xCT expression. We administered NAC (100 mg/kg ip) daily during a 14-day abstinence period following AMP (0.1 mg/kg/infusion; 2 h sessions) self-administration. Relapse was tested following one (WD 1) or 14 days (WD 14) of withdrawal. The overall number of astrocytes was also quantified within the medial prefrontal cortex (mPFC) and nucleus accumbens (ACb). NAC failed to lower cue-induced AMP craving via cue-induced relapse and reinstatement testing. Cue-induced craving did not increase from WD 1 to WD 14. AMP-exposed rats had greater astrocyte counts in the mPFC and ACb when compared AMP-naïve rats. Repeated injection with NAC decreased xCT expression within the mPFC and ACb. Overall, these results suggest that NAC may be an ineffective treatment option for lowering cue-induced relapse to AMP. Further, the results suggest that stimulating xCT via NAC may not be an effective therapeutic approach for decreasing cue-seeking for AMP.

Effects of sex and estrous cycle on intravenous oxycodone self-administration and the reinstatement of oxycodone-seeking behavior in rats.

The increasing misuse of both prescription and illicit opioids has culminated in a national healthcare crisis in the United States. Oxycodone is among the most widely prescribed and misused opioid pain relievers and has been associated with a high risk for transition to compulsive opioid use. Here, we sought to examine potential sex differences and estrous cycle-dependent effects on the reinforcing efficacy of oxycodone, as well as on stress-induced or cue-induced oxycodone-seeking behavior, using intravenous (IV) oxycodone self-administration and reinstatement procedures. In experiment 1, adult male and female Long-Evans rats were trained to self-administer 0.03 mg/kg/inf oxycodone according to a fixed-ratio 1 schedule of reinforcement in daily 2-h sessions, and a dose-response function was subsequently determined (0.003-0.03 mg/kg/inf). In experiment 2, a separate group of adult male and female Long-Evans rats were trained to self-administer 0.03 mg/kg/inf oxycodone for 8 sessions, followed by 0.01 mg/kg/inf oxycodone for 10 sessions. Responding was then extinguished, followed by sequential footshock-induced and cue-induced reinstatement tests. In the dose-response experiment, oxycodone produced a typical inverted U-shape function with 0.01 mg/kg/inf representing the maximally effective dose in both sexes. No sex differences were detected in the reinforcing efficacy of oxycodone. In the second experiment, the reinforcing effects of 0.01-0.03 mg//kg/inf oxycodone were significantly attenuated in females during proestrus/estrus as compared to metestrus/diestrus phases of the estrous cycle. Neither males nor females displayed significant footshock-induced reinstatement of oxycodone seeking, but both sexes exhibited significant cue-induced reinstatement of oxycodone seeking at magnitudes that did not differ either by sex or by estrous cycle phase. These results confirm and extend previous work suggesting that sex does not robustly influence the primary reinforcing effects of oxycodone nor the reinstatement of oxycodone-seeking behavior. However, our findings reveal for the first time that the reinforcing efficacy of IV oxycodone varies across the estrous cycle in female rats.

Medial orbitofrontal cortical regulation of different aspects of Pavlovian and instrumental reward seeking.

The medial subregion of the orbitofrontal cortex (mOFC) is thought to play an important role representing the expected outcome of a given course of action, as lesioning or inactivating this cortical region results in the adoption of choice strategies based more on observable (rather than previously learned) information. Despite this, its role in mediating basic associative learning remains to be fully clarified. The present series of experiments examined the role of the mOFC in (1) Pavlovian conditioned approach, (2) conditioned reinforcement, (3) extinction, and (4) cue-induced reinstatement of food-seeking behavior. Separate cohorts of rats went through Pavlovian or instrumental training. Intra-mOFC infusions of either saline or GABA agonists (to temporarily inactivate neural activity) were given prior to Pavlovian approach, conditioned reinforcement, first or second day of instrumental extinction training, or cue-induced reinstatement test days. mOFC inactivation increased lever-CS contacts in Pavlovian conditioned approach and (2) had no effect on conditioned reinforcement. These manipulations (3) accelerated within-session instrumental extinction during the initial extinction session, but impaired subsequent extinction learning on drug-free days. (4) mOFC inactivation induced differential effects on reinstatement that depended on baseline performance. mOFC inactivation abolished reinstatement in "Reinstater" rats (who displayed robust responding under control conditions) and robustly increased reinstatement in "Non-Reinstater" rats (who showed little reinstatement under control conditions) suggesting that individual differences in reinstatement may be supported by differences in mOFC mediated representations of expected outcomes. These findings have important implications for understanding how the mOFC uses stimulus-outcome and action-outcome expectancies to guide behavior, and how dysfunction within this region may contribute to pathological patterns of reward seeking.

Interleukin-1 receptor-associated kinase 4 (IRAK4) in the nucleus accumbens regulates opioid-seeking behavior in male rats

Opioid addiction remains a severe health problem. While substantial insights underlying opioid addiction have been yielded from neuron-centric studies, the contribution of non-neuronal mechanisms to opioid-related behavioral adaptations has begun to be recognized. Toll-like receptor 4 (TLR4), a pattern recognition receptor, has been widely suggested in opioid-related behaviors. Interleukin-1 receptor-associated kinase 4 (IRAK4) is a kinase essential for TLR4 responses, However, the potential role of IRAK4 in opioid-related responses has not been examined. Here, we explored the role of IRAK4 in cue-induced opioid-seeking behavior in male rats. We found that morphine self-administration increased the phosphorylation level of IRAK4 in the nucleus accumbens (NAc) in rats; the IRAK4 signaling remained activated after morphine extinction and cue-induced reinstatement test. Both systemic and local inhibition of IRAK4 in the NAc core attenuated cue-induced morphine-seeking behavior without affecting the locomotor activity and cue-induced sucrose-seeking. In addition, inhibition of IRAK4 also reduced the cue-induced reinstatement of fentanyl-seeking. Our findings suggest an important role of IRAK4 in opioid relapse-like behaviors and provide novel evidence in the association between innate immunity and drug addiction.

Network-Based Discovery of Opioid Use Vulnerability in Rats Using the Bayesian Stochastic Block Model.

Opioid use disorder is a psychological condition that affects over 200,000 people per year in the U.S., causing the Centers for Disease Control and Prevention to label the crisis as a rapidly spreading public health epidemic. The behavioral relationship between opioid exposure and development of opioid use disorder (OUD) varies greatly between individuals, implying existence of sup-populations with varying degrees of opioid vulnerability. However, effective pre-clinical identification of these sub-populations remains challenging due to the complex multivariate measurements employed in animal models of OUD. In this study, we propose a novel non-linear network-based data analysis workflow that employs seven behavioral traits to identify opioid use sub-populations and assesses contributions of behavioral variables to opioid vulnerability and resiliency. Through this analysis workflow we determined how behavioral variables across heroin taking, refraining and seeking interact with one another to identify potentially heroin resilient and vulnerable behavioral sub-populations. Data were collected from over 400 heterogeneous stock rats in two geographically distinct locations. Rats underwent heroin self-administration training, followed by a progressive ratio and heroin-primed reinstatement test. Next, rats underwent extinction training and a cue-induced reinstatement test. To enter the analysis workflow, we integrated data from different cohorts of rats and removed possible batch effects. We then constructed a rat-rat similarity network based on their behavioral patterns and implemented community detection on this similarity network using a Bayesian degree-corrected stochastic block model to uncover sub-populations of rats with differing levels of opioid vulnerability. We identified three statistically distinct clusters corresponding to distinct behavioral sub-populations, vulnerable, resilient and intermediate for heroin use, refraining and seeking. We implement this analysis workflow as an open source R package, named mlsbm.

Environmental enrichment reduces heroin seeking following incubation of craving in both male and female rats

BackgroundContemporary treatments for heroin use disorder demonstrate only limited efficacy when the goals are long term abstinence and prevention of relapse. We have demonstrated that environmental enrichment (EE) reduces cue-induced heroin reinstatement in male rats. The present study is an attempt to extend the “anti-relapse” effects of EE to female rats and to periods where incubation of craving is hypothesized to occur. MethodsThis experiment implemented a 3-phase procedure. In Phase 1, male and female rats were trained to self-administer heroin for 15 days. Phase 2 consisted of a 3- or 15-day forced abstinence (FA) period. In Phase 3 half of the rats were placed into EE and the other half in non-EE housing and subsequently tested for responding in extinction (no heroin or cues) for 15 days followed by a cue-induced reinstatement test. ResultsWe found that rats in the 15 days FA condition showed significantly enhanced drug seeking during extinction, irrespective of sex. We also found that EE significantly reduced this effect. During reinstatement, EE significantly reduced drug seeking in male and female rats and in both 3- and 15-day FA groups. ConclusionsEE, with or without prolonged FA, effectively reduced heroin seeking in male and female rats. These findings indicate that EE can reduce drug-seeking in males and females and when putative incubation of craving (i.e., prolonged abstinence period) has occurred and suggest that it may aid in the development of future long-term behavioral treatments for individuals at risk for heroin relapse.

Cell Adhesion Factors in the Orbitofrontal Cortex Control Cue-Induced Reinstatement of Cocaine Seeking and Amygdala-Dependent Goal Seeking.

Repeated cocaine exposure causes dendritic spine loss in the orbitofrontal cortex, which might contribute to poor orbitofrontal cortical function following drug exposure. One challenge, however, has been verifying links between neuronal structural plasticity and behavior, if any. Here we report that cocaine self-administration triggers the loss of dendritic spines on excitatory neurons in the orbitofrontal cortex of male and female mice (as has been reported in rats). To understand functional consequences, we locally ablated neuronal β1-integrins, cell adhesion receptors that adhere cells to the extracellular matrix and thus support dendritic spine stability. Degradation of β1-integrin tone: (1) caused dendritic spine loss, (2) exaggerated cocaine-seeking responses in a cue-induced reinstatement test, and (3) impaired the ability of mice to integrate new learning into familiar routines, a key function of the orbitofrontal cortex. Stimulating Abl-related gene kinase, overexpressing Proline-rich tyrosine kinase, and inhibiting Rho-associated coiled-coil containing kinase corrected response strategies, uncovering a β1-integrin-mediated signaling axis that controls orbitofrontal cortical function. Finally, use of a combinatorial gene silencing/chemogenetic strategy revealed that β1-integrins support the ability of mice to integrate new information into established behaviors by sustaining orbitofrontal cortical connections with the basolateral amygdala.SIGNIFICANCE STATEMENT Cocaine degenerates dendritic spines in the orbitofrontal cortex, a region of the brain involved in interlacing new information into established behaviors. One challenge has been verifying links between cellular structural stability and behavior, if any. In this second of two related investigations, we study integrin family receptors, which adhere cells to the extracellular matrix and thereby stabilize dendritic spines (see also DePoy et al., 2019). We reveal that β1-integrins in the orbitofrontal cortex control food- and cocaine-seeking behaviors. For instance, β1-integrin loss amplifies cocaine-seeking behavior and impairs the ability of mice to integrate new learning into familiar routines. We identify likely intracellular signaling partners by which β1-integrins support orbitofrontal cortical function and connectivity with the basolateral amygdala.

P.046 Expression of the adult ADHD-associated gene ADGRL3 is dysregulated by genetic risk variants and environmental risk factors

Attention-deficit/hyperactivity disorder (ADHD) is often comorbid with cocaine abuse. Controversy exists regarding long-term consequences of ADHD medications on cocaine abuse liability. Whereas childhood methylphenidate treatment may be preventative, methylphenidate in teens appears to further increase later cocaine abuse risk. In rodents, adolescent methylphenidate treatment further increases adult cocaine self-administration in the Spontaneously Hypertensive Rat (SHR) model of ADHD, whereas adolescent atomoxetine treatment does not. Effects of ADHD medications on cocaine cue reactivity, a critical component of addiction, are unknown.To investigate this, SHR, Wistar–Kyoto (inbred control) and Wistar (outbred control) rats received therapeutically relevant doses of methylphenidate (1.5 mg/kg, oral) and atomoxetine (0.3 mg/kg, intraperitoneal), or respective vehicles from post-natal day 28–55. Cocaine seeking, reflecting cue reactivity, was measured in adulthood during self-administration maintenance and cue-induced reinstatement tests conducted under a second-order schedule.Compared to control strains, SHR earned more cocaine infusions, emitted more cocaine-seeking responses during maintenance and reinstatement testing, and required more sessions to reach the extinction criterion. Compared to vehicle, adolescent methylphenidate, but not atomoxetine, further increased cocaine intake during maintenance testing in SHR. Adolescent atomoxetine, but not methylphenidate, decreased cocaine seeking during reinstatement testing in SHR. Neither medication had effects on cocaine intake or cue reactivity in control strains.The SHR successfully model ADHD and cocaine abuse comorbidity and show differential effects of adolescent ADHD medications on cocaine intake and cue reactivity during adulthood. Thus, SHR have heuristic value for assessing neurobiology underlying the ADHD phenotype and for evaluating pharmacotherapeutics for ADHD.

Effects of chronic stress on reinstatement of palatable food seeking: Sex differences and relationship to trait anxiety

Previous research in our lab has established a causal role for chronic stress exposure in subsequent increases in relapse-like behaviors in male rats with a history of palatable food self-administration. Given that many of the neurobehavioral consequences of stress are sex dependent, we aimed to determine whether sex differences exist with regard to the effects of chronic stress on relapse. Additionally, because high trait anxiety confers vulnerability to stress-related disorders, we examined whether individual differences in trait anxiety were related to differences in relapse-like behavior after chronic stress exposure. Following elevated plus maze testing for classification into high- or low-anxiety phenotypes, male and female rats responded for highly palatable food pellets. During subsequent extinction training, stress was manipulated (0 or 90 min restraint/day for 7 days). Rats were then tested for cue- and pellet priming-induced reinstatement of palatable food seeking. Results showed that female rats displayed higher levels of responding during cue-induced reinstatement tests compared to males, and that a history of chronic stress caused an attenuation of cue-induced reinstatement in female, but not male, rats. Regarding pellet priming-induced reinstatement, there was a three-way interaction such that neither stress history nor anxiety phenotype was related to reinstatement in females, but a history of stress in males caused increased and decreased responding in low- and high-anxiety rats, respectively. These results suggest that biological sex and trait anxiety level may help to explain differences in vulnerability to relapse among individuals exposed to chronic stress. Such information may be useful in designing more personalized and effective treatments for obesity and eating disorders.

Regulation of Garcinol on Histone Acetylation in the Amygdala and on the Reconsolidation of a Cocaine-Associated Memory.

Exposure to drug-related cues often disrupts abstinence from cocaine use by triggering memories of drug effects, leading to craving and possible relapse. One prospective method of treatment is weakening cocaine-associated memories via impairment of memory reconsolidation. Previous experiments have shown that systemic injection of the amnestic agent garcinol impairs the reconsolidation of cocaine-cue memories in a temporally constrained, cue-specific, and persistent manner. Here, we investigated garcinol’s effect on cocaine-cue memory reconsolidation when administered to the lateral nucleus of the amygdala (LA), as well as its epigenetic activity following systemic garcinol administration and also when given in conjunction with trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor. Rats received 12 days of cocaine self-administration training during which time an active lever press resulted in an i.v. cocaine infusion that was concurrently paired with the presentation of a light/tone cue. After 8 days of lever extinction, rats received a memory reactivation session followed by a cue-induced reinstatement test. Intra-LA garcinol following memory reactivation significantly impaired reconsolidation only if the memory was reactivated. Additional studies revealed a significant reduction in histone H3 K27 acetylation and reduced expression of the immediate-early genes Arc and Egr-1 in the LA. When administered alone, TSA enhanced the reinstatement of a cocaine-cue memory, an effect that was prevented when garcinol was concurrently administered. These data indicate the LA is a key structure responsive to garcinol, suggest that one of garcinol’s mechanisms of action is through the reduction of memory-related gene expression in the LA, implicate changes in histone acetylation in memory reconsolidation, and support garcinol as a potential therapeutic tool for sustaining abstinence.

Tamoxifen Blocks the Development of Motivational Features of an Addiction-Like Phenotype in Female Rats.

Women become addicted sooner after initiating cocaine use as compared to men. Preclinical studies reveal a similar vulnerability in females, with findings from ovariectomized rats suggesting that estradiol mediates the enhanced vulnerability. However, since ovariectomy depletes not only estradiol, but all ovarian hormones, its role in a physiological context is not clear. Thus, the goal of this study was to determine the role of estradiol in the development of an addiction-like phenotype in ovary-intact females treated chronically with the selective estrogen receptor (ER) modulator tamoxifen. We hypothesized that tamoxifen, by antagonizing ERs, would block the development of an addiction-like phenotype as defined by an enhanced motivation for cocaine (assessed under a progressive-ratio schedule), and a heightened vulnerability to relapse (assessed under an extinction/cue-induced reinstatement procedure). Effects were examined following extended access cocaine self-administration (24-h/day; 4-discrete trials/h; 1.5 mg/kg/infusion) and 14-days of abstinence, conditions optimized for inducing an addiction-like phenotype. As predicted, motivation for cocaine was increased following extended-access self-administration and protracted abstinence in the vehicle (sesame oil) and no-injection control groups, but not in the tamoxifen group indicating that ER signaling is critical for the development of this feature of an addiction-like phenotype. Surprisingly, the increase in motivation for cocaine following abstinence was also attenuated in the vehicle group as compared to no-injection controls suggesting that oil/injections also affected its development. Contrary to our hypothesis, tamoxifen did not decrease vulnerability to relapse as this group responded at similar levels during initial extinction sessions and cue-induced reinstatement testing as compared to controls. Tamoxifen did, however, impair extinction learning as this group took longer to extinguish as compared to controls. Taken together, these findings indicate that estradiol is critical for the extinction of drug-associated cues and the development of motivational features of addiction.

(-)-Stepholidine reduces cue-induced reinstatement of cocaine seeking and cocaine self-administration in rats

BackgroundDopamine receptors are implicated in cocaine reward and seeking. We hypothesize that (-)-stepholidine, a dopamine D1/D2/D3 multi-receptor agent, would be effective in reducing cocaine reward and seeking in an animal model. We investigated the effects of (-)-stepholidine in cue-induced reinstatement of cocaine seeking and cocaine self-administration (reward). MethodsCue-induced reinstatement experiment: Rats were trained to press a lever reinforced by cocaine (1 mg/kg/injection) for 15 consecutive daily sessions, after which the response was extinguished by withholding cocaine and cocaine-paired cues (light and pump activation). This was followed by a cue-induced reinstatement test where subjects were exposed to two cocaine cue presentations and presses on the active lever produced cues. Subjects were treated with one of four (-)-stepholidine doses prior to the reinstatement test. Cocaine self-administration (reward) experiment: Rats were trained to self-administer cocaine under a progressive ratio schedule of reinforcement. After stable breakpoints were established, rats were injected with four doses of (-)-stepholidine prior to testing; each dose was injected prior to a separate test session with no-treatment sessions intervening to re-establish break points. Results(-)-Stepholidine significantly reduced cue-induced reinstatement of cocaine seeking in a dose-related manner. Additionally, (-)-stepholidine significantly reduced break points for cocaine reward. (-)-Stepholidine did not significantly affect locomotor activity. Conclusions(-)-Stepholidine reduces cue-induced reinstatement of cocaine seeking and cocaine reward, suggesting that it may be useful in treating relapse in cocaine addiction.

Melatonin reduces motivation for cocaine self-administration and prevents relapse-like behavior in rats.

Melatonin is a hormone involved in the entrainment of circadian rhythms, which appears dysregulated in drug users. Further, it has been demonstrated that melatonin can modulate the reinforcing effects of several drugs of abuse and may therefore play a role in drug addiction. Here, we investigated whether administration of melatonin reduces relapse-like behavior and the motivation to seek cocaine in rats. Male Sprague-Dawley rats were submitted to long-term cocaine self-administration training. Thereafter, melatonin effects were assessed on: (1) the motivation to work for cocaine in the break point test, (2) the relapse-like behavior in the cue-induced reinstatement test, (3) the distance traveled in the open field test, and (4) sucrose preference in a two-bottle choice paradigm. Melatonin, 25 or 50mg/kg, was injected 3-4h after the dark phase onset, 30min prior to each test. Both doses of melatonin decreased the number of active pokes in both break point and cue-induced reinstatement tests, demonstrating that melatonin can reduce the cocaine-seeking behavior and the motivation to work for cocaine. Administration of the higher dose of this hormone, however, significantly reduced the number of inactive pokes during the cue-induced reinstatement test and tended to reduce animals' locomotor activity in the open field test. Sucrose preference was unchanged in both vehicle- and melatonin-treated animal groups. Our data suggest that melatonin administration may lower the risk of relapse triggered by cues in cocaine-experienced animals.

Impact of Environmental Enrichment on Perineuronal Nets in the Prefrontal Cortex following Early and Late Abstinence from Sucrose Self-Administration in Rats.

Perineuronal nets (PNNs) are aggregates of extracellular matrix that form structures surrounding a subset of GABAergic interneurons. The staining intensity of PNNs appears to be related to plasticity. Environmental enrichment (EE) influences plasticity during adulthood: EE decreases the rewarding effects of drugs of abuse and diminishes both drug- and sucrose-seeking behavior. We determined the impact of EE on PNN intensity in the medial prefrontal cortex (mPFC) in rats trained to self-administer sucrose. We examined the number and intensity of PNNs within the prelimbic (PL), infralimbic (IL), and orbitofrontal (OF) regions of the mPFC of adult Long-Evans rats that were trained for sucrose self-administration followed by acute or chronic EE during abstinence and a cue-induced reinstatement test. Rats exposed to EE prior to a cue-induced reinstatement of sucrose seeking had an increase in PNN staining compared with rats in standard housing. Conversely, naïve rats given 1 day of EE had a decrease in PNN intensity in the PL, no change in the IL, and an increase in the OF. Our findings demonstrate that EE increases PNN intensity in the mPFC after sucrose training, suggesting that training enhances the ability of EE to increase PNN intensity. We further demonstrate an interaction between time of abstinence, duration of EE exposure, and cue-induced reinstatement. Our results suggest that increased PNN intensity after EE may alter the excitatory/inhibitory balance of mPFC neurons such that rats are less responsive to a sucrose cue.

The tendency to sign-track predicts cue-induced reinstatement during nicotine self-administration, and is enhanced by nicotine but not ethanol.

Some individuals are particularly responsive to reward-associated stimuli ("cues"), including the effects of these cues on craving and relapse to drug-seeking behavior. In the cases of nicotine and alcohol, cues may acquire these abilities via the incentive-enhancing properties of the drug. To determine the interaction between cue-responsivity and nicotine reinforcement, we studied the patterns of nicotine self-administration in rats categorized based on their tendency to approach a food-predictive cue ("sign-trackers") or a reward-delivery location ("goal-trackers"). In a second experiment, we determined whether nicotine and ethanol altered the incentive value of a food cue. Rats were classified as sign- or goal-trackers during a Pavlovian conditioned approach paradigm. Rats then self-administered intravenous nicotine (0.03mg/kg infusions) followed by extinction and cue-induced reinstatement tests. We also tested the effects of nicotine (0.4mg/kg base s.c.) or ethanol (0.7g/kg i.p.) on the approach to, and reinforcing efficacy of, a food cue. Sign-trackers showed greater reinstatement in response to a nicotine cue. Further, nicotine enhanced sign-tracking but not goal-tracking to a food cue and also enhanced responding for the food cue during the conditioned reinforcement test. Conversely, ethanol reduced sign-tracking and increased goal-tracking, but had no effect on conditioned reinforcement. Our studies demonstrate that the tendency to attribute incentive value to a food cue predicts enhanced cue-induced reinstatement. Additionally, the incentive value of food cues is differentially modulated by nicotine and ethanol, which may be related to the reinforcing effects of these drugs.

Effects of oxytocin on methamphetamine-seeking exacerbated by predator odor pre-exposure in rats.

The endogenous oxytocin system has emerged as an inhibitor of drug-seeking and stress in preclinical models. The goal of this study was to examine whether systemic oxytocin administration attenuated methamphetamine (METH)-seeking in rats pre-exposed to a predator odor threat. In Experiment 1, rats were exposed for 5 days to the predator odor, 2,5-dihydro-2,4,5-trimethylthiazoline (TMT), or saline before METH self-administration began. After extinction training, rats were injected with 1 mg/kg, ip oxytocin (OXT) or saline 30 min before a cue-induced reinstatement test followed by re-extinction and a TMT-induced reinstatement test. In Experiment 2, TMT pre-exposure was followed by 10 days of 1 mg/kg OXT or saline injections before METH self-administration, extinction, and a TMT-induced reinstatement test. In Experiment 1, TMT pre-exposed rats that were injected with saline 30 min before reinstatement exhibited greater drug-seeking induced by conditioned cues or TMT than that exhibited by saline pre-exposed rats. A single injection of OXT 30 min before reinstatement suppressed METH-seeking in both saline- and TMT pre-exposed rats. In Experiment 2, TMT pre-exposed rats that received saline injections for 10 days prior to METH self-administration exhibited enhanced drug-seeking induced by TMT during stress-induced reinstatement. OXT injections for 10 days prior to METH self-administration blocked only the stress-induced exacerbation of drug-seeking in TMT pre-exposed rats. These results support further research on the development of oxytocin as a novel therapeutic drug that has enduring effects on drug-seeking exacerbated by stress.

The selective dopamine D3 receptor antagonist, SR 21502, reduces cue-induced reinstatement of heroin seeking and heroin conditioned place preference in rats

BackgroundBecause the role of dopamine (DA) D3 receptors has been investigated primarily in relation to cocaine-related behaviors little is known of the role of these receptors in heroin seeking. PurposesTo investigate the effect of the selective DA D3 receptor antagonist, SR 21502, on cue-induced reinstatement of heroin seeking and heroin conditioned place preference (CPP). MethodsIn experiment 1, rats were trained to self-administer intravenous heroin for 15 days followed by extinction. Following extinction animals were treated with one of several SR 21502 doses (0, 7.5, 10 or 15mg/kg) and a cue-induced reinstatement test was conducted. In experiment 2, animals were conditioned to experience heroin in one compartment of a CPP apparatus and saline in the other. On the test day animals were treated with 0, 3.75, 7.5, 10 or 15mg/kg of SR 21502 and tested for their CPP. ResultsThe results from experiment 1 showed a significant dose-related reduction in cue-induced reinstatement of active lever pressing in the 7.5 and 10mg groups and an absence of the reinstatement effect in the 15mg group. In experiment 2, animals treated with vehicle or 3.75mg of SR 21502 showed significant heroin place preferences but those treated with the higher doses showed no CPP. ConclusionsOur findings suggest that DA D3 receptors play a significant role in heroin approach behaviors driven by conditioned stimuli. As such, we propose that SR 21502 holds potential as an effective pharmacotherapeutic agent for relapse prevention and should be studied further.