Augmented Cocaine Seeking in Response to Stress or CRF Delivered into the Ventral Tegmental Area Following Long-Access Self-Administration Is Mediated by CRF Receptor Type 1 But Not CRF Receptor Type 2

Stressful events are determinants of relapse in recovering cocaine addicts. Excessive cocaine use may increase susceptibility to stressor-induced relapse through alterations in brain corticotropin-releasing factor (CRF) regulation of neurocircuitry involved in drug seeking. We previously reported that the reinstatement of cocaine seeking by a stressor (footshock) is CRF dependent and is augmented in rats that self-administered cocaine under long-access (LgA; 6 h daily) conditions for 14 d when compared with rats provided shorter daily cocaine access [short access (ShA) rats; 2 h daily]. Further, we have demonstrated that reinstatement in response to intracerebroventricular CRF administration is heightened in LgA rats. This study examined the role of altered ventral tegmental area (VTA) responsiveness to CRF in intake-dependent increases in CRF- and stress-induced cocaine seeking. Bilateral intra-VTA administration of CRF (250 or 500 ng/side) produced reinstatement in LgA but not ShA rats. In LgA rats, intra-VTA CRF-induced reinstatement was blocked by administration of the CRF-receptor type 1 (CRF-R1) antagonist antalarmin (500 ng/side) or CP-376395 (500 ng/side), but not the CRF-R2 antagonist astressin-2B (500 ng or 1 μg/side) or antisauvagine-30(ASV-30; 500 ng/side) into the VTA. Likewise, intra-VTA antalarmin, but not astressin-2B, blocked footshock-induced reinstatement in LgA rats. By contrast, neither intra-VTA antalarmin nor CP-376395 altered food-reinforced lever pressing. Intra-VTA injection of the CRF-R1-selective agonist cortagine (100 ng/side) but not the CRF-R2-selective agonist rat urocortin II (rUCN II; 250 ng/side) produced reinstatement. These findings reveal that excessive cocaine use increases susceptibility to stressor-induced relapse in part by augmenting CRF-R1-dependent regulation of addiction-related neurocircuitry in the VTA

Corticosterone-Induced Potentiation of Cocaine Seeking: A Potential Role for Organic Cation Transporter 3

While it is known that stress plays a role in the relapse of cocaine-seeking behavior, recent studies demonstrate that stress may be acting as a stage setter rather than directly triggering further cocaine use. This model suggests stimuli that do not normally evoke relapse under stress-free conditions may result in drug seeking when the exposure occurs under stressful conditions. In this study, we examined the corticosterone-dependent potentiation of cocaine-induced reinstatement by a stressor, electric footshock (EFS), in rats following cocaine self-administration and extinction. We found that in rats with a history of drug exposure under low intake conditions, footshock alone did not reinstate cocaine seeking, but did result in a potentiation of reinstatement in response to a subthreshold dose of cocaine (2.5 mg/kg, ip.). This effect was abolished in adrenalectomized rats and reproduced in intact animals receiving a physiologically relevant dose of corticosterone (2.0 mg/kg, ip.). Administration of the glucocorticoid receptor (GR) antagonist RU-486 did not block these effects, suggesting a rapid, non-GR mediated mechanism. In order to determine the site of action, we performed in vivo microdialysis to measure dopamine levels in the nucleus accumbens (NAc) and found that, similar to its actions on cocaine-seeking behavior, corticosterone potentiated cocaine-induced increases in dopamine. In support of this evidence, direct administration of corticosterone into the NAc or prefrontal cortex (PFC) potentiated reinstatement to a subthreshold dose of cocaine, an effect that was blocked with pretreatment of the dopamine antagonist, fluphenazine. Through immunoflourescence studies, we have shown the presence of a high capacity, corticosterone sensitive, monoamine transporter, organic cation transporter 3 (OCT3), located in the NAc adjacent to tyrosine hydroxylase terminals, suggesting a potential interaction with dopamine clearance. Based on this, we hypothesize the mechanism involves corticosterone inhibition of OCT3-mediated dopamine clearance. In support of this mechanism, pretreatment of rats with normetanephrine, a non-glucocorticoid OCT3 inhibitor mimics the effect of corticosterone on reinstatement. These results suggest a novel mechanism through which stress may modulate dopaminergic signaling and promote drug-seeking behavior

Oral Administration of Levo-Tetrahydropalmatine Attenuates Reinstatement of Extinguished Cocaine Seeking by Cocaine, Stress or Drug-Associated Cues in Rats

Cocaine addiction is characterized by a persistently heightened susceptibility to drug relapse. For this reason, the identification of medications that prevent drug relapse is a critical goal of drug abuse research. Drug re-exposure, the onset of stressful life events, and exposure to cues previously associated with drug use have been identified as determinants of relapse in humans and have been found to reinstate extinguished cocaine seeking in rats. This study examined the effects of acute oral (gavage) administration of levo-tetrahydropalmatine (l-THP), a tetrahydroprotoberberine isoquinoline with a pharmacological profile that includes antagonism of D1, D2 and D3 dopamine receptors, on the reinstatement of extinguished cocaine seeking by a cocaine challenge (10 mg/kg, ip), a stressor (uncontrollable electric footshock [EFS]) or response-contingent exposure to a stimulus (tone and light complex) previously associated with drug delivery in male Sprague–Dawley rats. Extinguished drug seeking was reinstated by ip cocaine, EFS, or response-contingent presentation of drug-associated cues in vehicle-pretreated rats following extinction of iv cocaine self-adminisration. Oral administration of either 3.0 or 10.0 mg/kg l-THP 1 h prior to reinstatement testing significantly attenuated reinstatement by each of the stimuli. Food-reinforced responding and baseline post-extinction responding were significantly attenuated at the 10.0, but not the 3.0 mg/kg, l-THP dose, indicating that the effects of 3 mg/kg l-THP on reinstatement were likely independent of non-specific motor impairment. These findings further suggest that l-THP may have utility for the treatment of cocaine addiction

Corticosterone Acts in the Nucleus Accumbens to Enhance Dopamine Signaling and Potentiate Reinstatement of Cocaine Seeking

Stressful life events are important contributors to relapse in recovering cocaine addicts, but the mechanisms by which they influence motivational systems are poorly understood. Studies suggest that stress may “set the stage” for relapse by increasing the sensitivity of brain reward circuits to drug-associated stimuli. We examined the effects of stress and corticosterone on behavioral and neurochemical responses of rats to a cocaine prime after cocaine self-administration and extinction. Exposure of rats to acute electric footshock stress did not by itself reinstate drug-seeking behavior but potentiated reinstatement in response to a subthreshold dose of cocaine. This effect of stress was not observed in adrenalectomized animals, and was reproduced in nonstressed animals by administration of corticosterone at a dose that reproduced stress-induced plasma levels. Pretreatment with the glucocorticoid receptor antagonist RU38486 did not block the corticosterone effect. Corticosterone potentiated cocaine-induced increases in extracellular dopamine in the nucleus accumbens (NAc), and pharmacological blockade of NAc dopamine receptors blocked corticosterone-induced potentiation of reinstatement. Intra-accumbens administration of corticosterone reproduced the behavioral effects of stress and systemic corticosterone. Corticosterone treatment acutely decreased NAc dopamine clearance measured by fast-scan cyclic voltammetry, suggesting that inhibition of uptake2-mediated dopamine clearance may underlie corticosterone effects. Consistent with this hypothesis, intra-accumbens administration of the uptake2 inhibitor normetanephrine potentiated cocaine-induced reinstatement. Expression of organic cation transporter 3, a corticosterone-sensitive uptake2 transporter, was detected on NAc neurons. These findings reveal a novel mechanism by which stress hormones can rapidly regulate dopamine signaling and contribute to the impact of stress on drug intake

Stress Promotes Drug Seeking Through Glucocorticoid-Dependent Endocannabinoid Mobilization in the Prelimbic Cortex

Background Clinical reports suggest that rather than directly driving cocaine use, stress may create a biological context within which other triggers for drug use become more potent. We hypothesize that stress-induced increases in corticosterone “set the stage” for relapse by promoting endocannabinoid-induced attenuation of inhibitory transmission in the prelimbic cortex (PL). Methods We have established a rat model for these stage-setting effects of stress. In this model, neither a stressor (electric footshock) nor stress-level corticosterone treatment alone reinstates cocaine seeking following self-administration and extinction, but each treatment potentiates reinstatement in response to an otherwise subthreshold cocaine priming dose (2.5 mg/kg, intraperitoneal). The contributions of endocannabinoid signaling in the PL to the effects of stress-level corticosterone on PL neurotransmission and cocaine seeking were determined using intra-PL microinfusions. Endocannabinoid-dependent effects of corticosterone on inhibitory synaptic transmission in the rat PL were determined using whole-cell recordings in layer V pyramidal neurons. Results Corticosterone application attenuated inhibitory synaptic transmission in the PL via cannabinoid receptor type 1 (CB1R)– and 2-arachidonoylglycerol–dependent inhibition of gamma-aminobutyric acid release without altering postsynaptic responses. The ability of systemic stress-level corticosterone treatment to potentiate cocaine-primed reinstatement was recapitulated by intra-PL injection of corticosterone, the CB1R agonist WIN 55,212-2, or the monoacylglycerol lipase inhibitor URB602. Corticosterone effects on reinstatement were attenuated by intra-PL injections of either the CB1R antagonist, AM251, or the diacylglycerol lipase inhibitor, DO34. Conclusions These findings suggest that stress-induced increases in corticosterone promote cocaine seeking by mobilizing 2-arachidonoylglycerol in the PL, resulting in CB1R-mediated attenuation of inhibitory transmission in this brain region

Selective Induction of Cell Death in Melanoma Cell Lines through Targeting of Mcl-1 and A1

Melanoma is an often fatal form of skin cancer which is remarkably resistant against radio- and chemotherapy. Even new strategies that target RAS/RAF signaling and display unprecedented efficacy are characterized by resistance mechanisms. The targeting of survival pathways would be an attractive alternative strategy, if tumor-specific cell death can be achieved. Bcl-2 proteins play a central role in regulating survival of tumor cells. In this study, we systematically investigated the relevance of antiapoptotic Bcl-2 proteins, i.e., Bcl-2, Bcl-xL, Bcl-w, Mcl-1, and A1, in melanoma cell lines and non-malignant cells using RNAi. We found that melanoma cells required the presence of specific antiapoptotic Bcl-2 proteins: Inhibition of Mcl-1 and A1 strongly induced cell death in some melanoma cell lines, whereas non-malignant cells, i.e., primary human fibroblasts or keratinocytes were not affected. This specific sensitivity of melanoma cells was further enhanced by the combined inhibition of Mcl-1 and A1 and resulted in 60% to 80% cell death in all melanoma cell lines tested. This treatment was successfully combined with chemotherapy, which killed a substantial proportion of cells that survived Mcl-1 and A1 inhibition. Together, these results identify antiapoptotic proteins on which specifically melanoma cells rely on and, thus, provide a basis for the development of new Bcl-2 protein-targeting therapies

Intergenerational and parent of origin effects of maternal calorie restriction on Igf2 expression in the adult rat hippocampus

Insulin-like growth factor 2 (Igf2) regulates development, memory and adult neurogenesis in the hippocampus. Calorie restriction (CR) is known to modulate non-neuronal Igf2 expression intergenerationally, but its effect has not been evaluated on brain Igf2. Here, Sprague-Dawley (S) dams underwent moderate CR between gestational days 8–21. To identify parent of origin expression pattern of the imprinted Igf2 gene, their offspring (SS F1) were mated with naïve male or female Brown Norway (B) rats to obtain the second generation (BS and SB F2) progeny. CR did not affect adult hippocampal Igf2 transcript levels in SS F1 males or their BS F2 progeny, but increased it in SS F1 females and their SB F2 offspring. The preferentially maternal Igf2 expression in the SB F2 control male hippocampus relaxed to biallelic with CR, with no effect of grandmaternal diet in any other groups. Thus, allele-specific and total expression of hippocampal Igf2 is affected by maternal, grandmaternal CR in a strain and sex-specific manner