Cocaine Cocaine Conditioned Place Preference Research Articles

Sex differences in hippocampal β-adrenergic receptor subtypes drive retrieval, retention, and learning of cocaine-associated memories.

Drug seeking behavior occurs in response to environmental contexts and drug-associated cues. The presence of these pervasive stimuli impedes abstinence success. β-adrenergic receptors (β-ARs) have a long-standing historical implication in driving processes associated with contextual memories, including drug-associated memories in substance use disorders. However, sex differences in the role of β-adrenergic receptors in drug memories remain unknown. Prior reports indicate a selective role for β2-ARs in retrieval and retention of contextual drug memories in males, and substantial sex differences exist in the expression of β-ARs of male and female rats. Therefore, we hypothesized that there are sex differences in selective recruitment of β-ARs during different stages of memory encoding and retrieval. The role of β-ARs in driving retrieval and learning of contextual cocaine memories was investigated using cocaine conditioned place preference (CPP) in adult male and female Sprague-Dawley rats. Rats were infused directly to the dorsal hippocampus with Propranolol (β1 and β2) or ICI-118,551 (β1) and/or Betaxolol (β2), immediately prior to testing (retrieval), or paired to each cocaine (10 mg/kp, IP) conditioning session (learning). In males, administration of either β1, β2, or combined β1 and β2-ARs before the initial CPP testing reduced the expression of a CPP compared to vehicle administration. In females, β2-ARs transiently decreased CPP memories, whereas β1 had long lasting but not immediate effects to decrease CPP memories. Additionally, β1 and combined β1 and β2-ARs had immediate and persistent effects to decrease CPP memory expression. DG Fos + neurons predicted cocaine CPP expression in males, whereas CA1 and CA3 Fos + neurons predicted cocaine CPP expression in females. There are significant sex differences in the role of dorsal hippocampus β-ARs in the encoding and expression of cocaine conditioned place preference. Furthermore, sub regions of the dorsal hippocampus appear to activate differently between male and female rats during CPP. Therefore DG, CA3, and CA1 may have separate region- and sex-specific impacts on driving drug- associated, or context-associated cues.

Cannabidiol (CBD) facilitates cocaine extinction and ameliorates cocaine-induced changes to the gut microbiome in male C57BL/6JArc mice

Cocaine use disorder (CUD) is a global health problem with no approved medications. One potential treatment target is the gut microbiome, but it is unknown if cocaine induces long-lasting effects on gut microbes. A novel therapeutic candidate for CUD, cannabidiol (CBD), can improve gut function in rodent models. It is possible that protective effects of CBD against cocaine use are mediated by improving gut health. We examined this question in this experiment. Cocaine conditioned place preference (CPP) was conducted in adult male C57BL/6JArc mice. Mice were treated with vehicle or 20 mg/kg CBD prior to all cocaine CPP sessions (N = 11–13/group). Mice were tested drug free 1, 14 and 28 days after cessation of cocaine and CBD treatment. Fecal samples were collected prior to drug treatment and after each test session. Gut microbiome analyses were conducted using 16 s rRNA sequencing and correlated with behavioural parameters. We found a persistent preference for a cocaine-environment in mice, and long-lasting changes to gut microbe alpha diversity. Cocaine caused persistent changes to beta diversity which lasted for 4 weeks. CBD treatment reduced cocaine-environment preference during abstinence from cocaine and returned gut beta diversity measures to control levels. CBD treatment increased the relative abundance of Firmicutes phyla and Oscillospira genus, but decreased Bacteroidetes phyla and Bacteroides acidifaciens species. Preference score in cocaine-treated mice was positively correlated with abundance of Actinobacteria, whereas in mice treated with CBD and cocaine, the preference score was negatively correlated with Tenericutes abundance. Here we show that CBD facilitates cocaine extinction memory and reverses persistent cocaine-induced changes to gut microbe diversity. Furthermore, CBD increases the abundance of gut microbes which have anti-inflammatory properties. This suggests that CBD may act via the gut to reduce the memory of cocaine reward. Our data suggest that improving gut health and using CBD could limit cocaine abuse.

Estradiol reduction through aromatase inhibition impairs cocaine seeking in male rats.

Clinical and preclinical research on cocaine use disorder (CUD) has shown that sex differences in drug seeking are influenced by hormonal fluctuations. Estradiol (E2), a sex steroid hormone, has been linked to female drug effects, vulnerability to use/abuse, and psychosocial factors. Preclinical studies show that estradiol in females facilitates the extinction of cocaine-seeking behavior indicating a possible role in regulating extinction learning. Similar to females, males' brains contain the aromatase enzyme which converts testosterone to estradiol. However, it is unclear whether estradiol plays a role in male extinction learning as it does in females. Furthermore, how endogenously aromatized estradiol affects drug addiction in males is unknown. Therefore, this study investigated whether endogenous estradiol regulates cocaine seeking in male rats. We hypothesized that decreased aromatase enzyme activity, resulting in decreased estradiol synthesis in male brains, will impair extinction learning leading to increased cocaine-seeking behavior. This hypothesis was tested using cocaine-conditioned place preference (CPP), and short access self-administration (SA), followed by extinction and reinstatement. Before each extinction session for CPP or SA, male rats received an injection of either 1 (low dose) or 2.5 mg/kg (high dose) of the aromatase inhibitor Fadrozole (FAD), or vehicle. FAD groups showed dose-dependent effects on cocaine-seeking behavior compared to the vehicle group during CPP extinction. Specifically, low dose FAD facilitated extinction of cocaine CPP, whereas high dose FAD impaired it. In contrast, neither dose of FAD had any effects on the extinction of cocaine SA. Interestingly, only the low dose FAD group had decreased active lever pressing during cue- and cocaine-primed reinstatement compared to the vehicle group. Neither dose of FAD had an effect on sucrose extinction or reinstatement of sucrose seeking. These results from CPP experiments suggest that estradiol may impact extinction learning, as a low dose of FAD may strengthen the formation of cocaine extinction memory. Additionally, in male rats undergoing cocaine SA, the same low dose of aromatase inhibitor effectively reduced reinstatement of cocaine-seeking behavior. Thus, estradiol impacts cocaine seeking and extinction in both males and females, and it may also influence the development of sex-specific treatment strategies for CUD.

Sex difference in the effect of environmental enrichment on food restriction-induced persistence of cocaine conditioned place preference and mechanistic underpinnings

Psychosocial and environmental factors, including loss of natural reward, contribute to the risk of drug abuse. Reward loss has been modeled in animals by removal from social or sexual contact, transfer from enriched to impoverished housing, or restriction of food. We previously showed that food restriction increases the unconditioned rewarding effects of abused drugs and the conditioned incentive effects of drug-paired environments. Mechanistic studies provided evidence of decreased basal dopamine (DA) transmission, adaptive upregulation of signaling downstream of D1 DA receptor stimulation, synaptic upscaling and incorporation of calcium-permeable AMPA receptors (CP-AMPARs) in medium spiny neurons (MSNs) of nucleus accumbens (NAc). These findings align with the still evolving ‘reward deficiency’ hypothesis of drug abuse. The present study tested whether a compound natural reward that is known to increase DA utilization, environmental enrichment, would prevent the persistent expression of cocaine conditioned place preference (CPP) otherwise observed in food restricted rats, along with the mechanistic underpinnings. Because nearly all prior investigations of both food restriction and environmental enrichment effects on cocaine CPP were conducted in male rodents, both sexes were included in the present study. Results indicate that environmental enrichment curtailed the persistence of CPP expression, decreased signaling downstream of the D1R, and decreased the amplitude and frequency of spontaneous excitatory postsynaptic currents (EPSCs) in NAc MSNs of food restricted male, but not female, rats. The failure of environmental enrichment to significantly decrease food restriction-induced synaptic insertion of CP-AMPARs, and how this may accord with previous pharmacological findings that blockade of CP-AMPARs reverses behavioral effects of food restriction is discussed. In addition, it is speculated that estrous cycle-dependent fluctuations in DA release, receptor density and MSN excitability may obscure the effect of increased DA signaling during environmental enrichment, thereby interfering with development of the cellular and behavioral effects that enrichment produced in males.

Social defeat stress enhances the rewarding effects of cocaine through α1A adrenoceptors in the medial prefrontal cortex of mice

Various stressors potentiate the rewarding effects of cocaine and contribute to cocaine cravings. However, it remains unclear whether psychosocial stress enhances the rewarding effects of cocaine. Accordingly, this study employed a cocaine-conditioned place preference (CPP) paradigm combined with social defeat (SD) exposure to investigate the effects of acute SD stress on cocaine reward in male mice. We found that SD stress immediately before the posttest significantly increased cocaine CPP, and systemic blockade of α1 adrenoceptors, but not β adrenoceptors, suppressed this increase. Fiber photometry recordings with GRABNE1m sensors revealed increased noradrenaline (NA) levels in the medial prefrontal cortex (mPFC) in test mice in response to attacks by aggressor mice during SD. Moreover, the SD stress-induced enhancement of CPP was effectively suppressed by intra-mPFC infusion of an α1 adrenoceptor antagonist. In vitro whole-cell recordings demonstrated that silodosin, an α1A, but not α1B or α1D, adrenoceptor antagonist, inhibited NA-induced depolarizing currents and facilitation of excitatory synaptic transmissions. Consistently, intra-mPFC silodosin infusion significantly suppressed the SD stress-induced CPP enhancement. Conversely, intra-mPFC infusion of α1A adrenoceptor agonist augmented cocaine CPP in the absence of stress exposure. Additionally, intranasal silodosin administration attenuated the SD stress-induced enhancement of CPP, and chemogenetic inhibition of mPFC excitatory neurons also suppressed the SD stress-induced CPP enhancement. Together, these findings suggest that NA stimulation of α1A adrenoceptors and the subsequent activation of mPFC pyramidal cells may contribute to SD stress-induced amplification of the rewarding effects of cocaine, and intranasal silodosin administration may hold therapeutic potential for mitigating stress-associated cocaine craving.

Endogenous opioid system modulates conditioned cocaine reward in a sex-dependent manner.

Cocaine predictive cues and contexts exert powerful control over behaviour and can incite cocaine seeking and taking. This type of conditioned behaviour is encoded within striatal circuits, and these circuits and behaviours are, in part, regulated by opioid peptides and receptors expressed in striatal medium spiny neurons. We previously showed that augmenting levels of the opioid peptide enkephalin in the striatum facilitates acquisition of cocaine conditioned place preference (CPP), while opioid receptor antagonists attenuate expression of cocaine CPP. However, whether striatal enkephalin is necessary for acquisition of cocaine CPP and maintenance during extinction remains unknown. To address this, we generated mice with a targeted deletion of enkephalin from dopamine D2-receptor expressing medium spiny neurons and tested them in a cocaine CPP paradigm. Low striatal enkephalin levels did not attenuate acquisition of CPP. However, expression of preference, assessed after acute administration of the opioid receptor antagonist naloxone, was blocked in females, regardless of genotype. When saline was paired with the cocaine context during extinction sessions, females, regardless of genotype, extinguished preference faster than males, and this was prevented by naloxone when paired with the cocaine context. We conclude that while striatal enkephalin is not necessary for acquisition, expression, or extinction of cocaine CPP, expression and extinction of cocaine preference in females is mediated by an opioid peptide other than striatal enkephalin. The unique sensitivity of females to opioid antagonists suggests sex should be a consideration when using these compounds in the treatment of cocaine use disorder.

Identification of miRNA-mediated gene regulatory networks in L-methionine exposure counteracts cocaine-conditioned place preference in mice.

Background and Aims: Methionine has been proven to inhibit addictive behaviors of cocaine dependence. This study aimed to identify the potential mechanisms of MET relating to its inhibitory effects on cocaine induced cellular and behavioral changes. Methods: MRNA and miRNA high-throughput sequencing of the prefrontal cortex in a mouse model of cocaine conditioned place preference (CPP) combined with L-methionine was performed. Differentially expressed miRNAs (DE-miRNAs) and differentially expressed genes (DEGs) regulated by cocaine and inhibited by L-methionine were identified. DEGs were mapped to STRING database to construct a protein-protein interaction (PPI) network. Then, the identified DEGs were subjected to the DAVID webserver for functional annotation. Finally, miRNA-mRNA regulatory network and miRNA-mRNA-TF regulatory networks were established to screen key DE-miRNAs and coregulation network in Cytoscape. Results: Sequencing data analysis showed that L-methionine reversely regulated genes and miRNAs affected by cocaine. Pathways associated with drug addiction only enriched in CS-down with MC-up genes targeted by DE-miRNAs including GABAergic synapse, Glutamatergic synapse, Circadian entrainment, Axon guidance and Calcium signaling pathway. Drug addiction associated network was formed of 22 DEGs including calcium channel (Cacna1c, Cacna1e, Cacna1g and Cacng8), ephrin receptor genes (Ephb6 and Epha8) and ryanodine receptor genes (Ryr1 and Ryr2). Calcium channel gene network were identified as a core gene network modulated by L-methionine in response to cocaine dependence. Moreover, it was predicted that Grin1 and Fosb presented in TF-miRNA-mRNA coregulation network with a high degree of interaction as hub genes and interacted calcium channels. Conclusion: These identified key genes, miRNA and coregulation network demonstrated the efficacy of L-methionine in counteracting the effects of cocaine CPP. To a certain degree, it may provide some hints to better understand the underlying mechanism on L-methionine in response to cocaine abuse.

Cav1.3 L-type Ca2+ channel-activated CaMKII/ERK2 pathway in the ventral tegmental area is required for cocaine conditioned place preference

We have previously demonstrated that pharmacological blockade of ventral tegmental area (VTA) Cav1.3 L-type calcium channels (LTCCs) using Cav1.2 dihydropyridine insensitive (Cav1.2DHP−/−) mutant mice attenuates cocaine conditioned place preference (CPP). However, the molecular mechanisms by which Cav1.3 channels mediate the effects of cocaine in the VTA remain largely unknown. In this study using Cav1.2DHP−/− male mice, we find that cocaine place preference increases CaM kinase IIα, ERK2, and CREB phosphorylation in the VTA, proteins strongly linked to cocaine behaviors. To further explore the causal role of these intracellular signaling proteins in cocaine preference, the CaM kinase II inhibitor, KN93 was directly injected into the VTA of male mice before each cocaine conditioning session. We found that KN93 attenuates conditioned preference for cocaine compared to vehicle treated mice and decreased VTA ERK2 and CREB phosphorylation. Additionally, blockade of the ERK pathway with the MEK inhibitor, U0126 or knockdown of ERK2 using siRNA, attenuated cocaine preference and VTA CREB phosphorylation but not CaMKIIα phosphorylation, suggesting that ERK is activated downstream of CaMKIIα. Examination of postsynaptic density (PSD) GluA1 subunit of AMPA receptors in the nucleus accumbens (NAc) that we have previously shown to be upregulated following long withdrawal periods, was blunted by KN93, U0126 and ERK2 siRNA when examined 30 days following cocaine CPP. Taken together, these findings demonstrate that Cav1.3 channels in the VTA are required for cocaine reward behavior and activation of the CaMKIIα/ERK/CREB signaling pathway in the VTA is necessary for long-lasting changes in the NAc.This article is part of the Special Issue on 'L-type calcium channel mechanisms in neuropsychiatric disorders'.

The Impact of HIV Infection in Cocaine Seeking Behaviors

BackgroundSubstance use disorders (SUDs) are characterized by high propensity to relapse and are highly comorbid with HIV infection. The underlying neurobiology of this relationship is poorly understood. Preclinical research on the neurobiobehavioral outcomes of progressive HIV infections may yield crucial information to improve SUD prognosis and reduce risk of relapse in people living with HIV.MethodsTo model progressive HIV, adult male and female C57BL6J mice were inoculated with EcoHIV, a chimeric HIV‐1 which infects murine cells by replacing envelope glycoprotein gp120 with gp80. Five weeks after inoculation mice were trained in a cocaine conditioned place preference (CPP) paradigm (10mg/kg, i.p.) followed by extinction training. Stress‐induced reinstatement was assessed following administration of yohimbine, an α2 adrenergic antagonist, to determine EcoHIV effects. Neuronal activation in the prefrontal cortex (PFC) and nucleus accumbens (NAc) by cocaine exposure was assessed by quantitative analysis the expression of the immediate early gene, cFos.ResultsEcoHIV and sham‐inoculated mice exhibited similar cocaine CPP and extinction. However, EcoHIV mice showed significantly increased yohimbine‐induced reinstatement rate (p<0.05). cFos analysis revealed an elevated neural activation in PFC and NAc in response to cocaine. However, only neurons within infralimbic PFC (IL) and NAc shell subregions showed further increases in activity in cocaine exposed mice infected with EcoHIV vs shams.ConclusionEcoHIV infected mice escalated stress‐induced reinstatement of cocaine seeking compared to controls, suggesting that people living with HIV may be at elevated risk for stress‐induced relapse. Progressive EcoHIV infection may dysregulate IL and NAcShell activity to facilitate the cocaine seeking and reinstatement. Ongoing research will characterize how EcoHIV infection dysregulate IL‐NAcShell pathway specific activity during cocaine reward learning and stress‐induced relapse.

Optogenetic stimulation of dynorphinergic neurons within the dorsal raphe activate kappa opioid receptors in the ventral tegmental area and ablation of dorsal raphe prodynorphin or kappa receptors in dopamine neurons blocks stress potentiation of cocaine reward

Behavioral stress exposure increases the risk of drug-taking in individuals with substance use disorders by mechanisms involving the dynorphins, which are the endogenous neuropeptides for the kappa opioid receptor (KOR). KOR agonists have been shown to encode dysphoria, aversion, and changes in reward valuation, and kappa opioid antagonists are in clinical development for treating substance use disorders. In this study, we confirmed that KORs were expressed in dopaminergic neurons in the ventral tegmental area (VTA) of male C57BL6/J mice. Genetic ablation of KORs from dopamine neurons blocked the potentiating effects of repeated forced swim stress on cocaine conditioned place preference (CPP). KOR activation inhibited dopamine neuron GCaMP6m calcium activity in VTA during swim stress and caused a rebound enhancement during the period after stress exposure. Transient optogenetic inhibition of VTA dopamine neurons with AAV5-DIO-SwiChR was acutely aversive in a real time place preference assay and blunted cocaine CPP when inhibition was administered concurrently with cocaine conditioning. However, when inhibition preceded cocaine conditioning by 30 min, cocaine CPP was enhanced. Retrograde tracing with CAV2-DIO-ZsGreen identified a population of prodynorphinCre neurons in the dorsal raphe nucleus (DRN) projecting to the VTA. Optogenetic stimulation of dynorphinergic neurons within the DRN by Channelrhodopsin2 activated KOR in VTA and ablation of prodynorphin blocked stress potentiation of cocaine CPP. Together, these studies demonstrate the presence of a dynorphin/KOR midbrain circuit that projects from the DRN to VTA and is involved in altering the dynamic response of dopamine neuron activity to enhance drug reward learning.

Distinct Role of Dopamine in the PFC and NAc During Exposure to Cocaine-Associated Cues.

BackgroundDopamine neurotransmission plays a critical role in reward in drug abuse and drug addiction. However, the role of dopamine in the recognition of drug-associated environmental stimuli, retrieval of drug-associated memory, and drug-seeking behaviors is not fully understood. MethodsRoles of dopamine neurotransmission in the prefrontal cortex (PFC) and nucleus accumbens (NAc) in the cocaine-conditioned place preference (CPP) paradigm were evaluated using in vivo microdialysis. ResultsIn mice that had acquired cocaine CPP, dopamine levels in the PFC, but not in the NAc, increased in response to cocaine-associated cues when mice were placed in the cocaine chamber of an apparatus with 2 separated chambers. The induction of the dopamine response and the development of cocaine CPP were mediated through activation of glutamate NMDA (N-methyl-D-aspartate)/AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor signaling in the PFC during conditioning. Activation of dopamine D1 or D2 receptor signaling in the PFC was required for cocaine-induced locomotion, but not for the induction of the dopamine response or the development of cocaine CPP. Interestingly, dopamine levels in the NAc increased in response to cocaine-associated cues when mice were placed at the center of an apparatus with 2 connected chambers, which requires motivated exploration associated with cocaine reward. ConclusionsDopamine neurotransmission in the PFC is activated by the exposure to the cocaine-associated cues, whereas dopamine neurotransmission in the NAc is activated in a process of motivated exploration of cues associated with cocaine reward. Furthermore, the glutamate signaling cascade in the PFC is suggested to be a potential therapeutic target to prevent the progression of drug addiction.

Stress decreases serotonin tone in the nucleus accumbens in male mice to promote aversion and potentiate cocaine preference via decreased stimulation of 5-HT1B receptors.

Stress-induced release of dynorphins (Dyn) activates kappa opioid receptors (KOR) in serotonergic neurons to produce dysphoria and potentiate drug reward; however, the circuit mechanisms responsible for this effect are not known. In male mice, we found that conditional deletion of KOR from Slc6a4 (SERT)-expressing neurons blocked stress-induced potentiation of cocaine conditioned place preference (CPP). Within the dorsal raphe nucleus (DRN), two overlapping populations of KOR-expressing neurons: Slc17a8 (VGluT3) and SERT, were distinguished functionally and anatomically. Optogenetic inhibition of these SERT+ neurons potentiated subsequent cocaine CPP, whereas optical inhibition of the VGluT3+ neurons blocked subsequent cocaine CPP. SERT+/VGluT3- expressing neurons were concentrated in the lateral aspect of the DRN. SERT projections from the DRN were observed in the medial nucleus accumbens (mNAc), but VGluT3 projections were not. Optical inhibition of SERT+ neurons produced place aversion, whereas optical stimulation of SERT+ terminals in the mNAc attenuated stress-induced increases in forced swim immobility and subsequent cocaine CPP. KOR neurons projecting to mNAc were confined to the lateral aspect of the DRN, and the principal source of dynorphinergic (Pdyn) afferents in the mNAc was from local neurons. Excision of Pdyn from the mNAc blocked stress-potentiation of cocaine CPP. Prior studies suggested that stress-induced dynorphin release within the mNAc activates KOR to potentiate cocaine preference by a reduction in 5-HT tone. Consistent with this hypothesis, a transient pharmacological blockade of mNAc 5-HT1B receptors potentiated subsequent cocaine CPP. 5-HT1B is known to be expressed on 5-HT terminals in NAc, and 5-HT1B transcript was also detected in Pdyn+, Adora2a+ and ChAT+ (markers for direct pathway, indirect pathway, and cholinergic interneurons, respectively). Following stress exposure, 5-HT1B transcript was selectively elevated in Pdyn+ cells of the mNAc. These findings suggest that Dyn/KOR regulates serotonin activation of 5HT1B receptors within the mNAc and dynamically controls stress response, affect, and drug reward.

The association between risky decision making and cocaine conditioned place preference is moderated by sex

BackgroundExcessive risk taking is a characteristic trait of several psychiatric conditions, including substance use disorders. High risk-taking (HiR) rats self-administer more cocaine compared to low risk-taking (LoR) rats. However, research has not determined if risk taking is associated with enhanced cocaine conditioned place preference (CPP). MethodsMale and female Sprague Dawley rats (n = 48 each sex) were first tested in the risky decision task (RDT), in which a response on one lever resulted in safe delivery of one food pellet, and a response on a different lever resulted in delivery of two pellets and probabilistic delivery of foot shock. Following RDT training, rats were tested for cocaine CPP. The first session was a pretest that measured rats’ preference for three compartments that provided different visual and tactile cues. Rats then learned to associate one compartment with cocaine (either 10.0 mg/kg or 20.0 mg/kg; i.p.) and one compartment with saline (1.0 ml/kg; i.p.) across eight conditioning sessions. Finally, rats explored all three compartments in a drug-free state. ResultsSex significantly moderated the association between risky decision making and cocaine CPP. While increased risk aversion was somewhat positively associated with cocaine CPP in males, increased risk taking was positively correlated with cocaine CPP in females. ConclusionsThese results highlight the moderating role of sex on the relationship between risky decision making and cocaine reward.

Serum- and glucocorticoid-inducible kinase 1 activity in ventral tegmental area dopamine neurons regulates cocaine conditioned place preference but not cocaine self-administration.

Drugs of abuse regulate the activity of the mesolimbic dopamine (DA) system, and drug-induced changes in ventral tegmental area (VTA) cellular activity and gene regulation are linked to behavioral outputs associated with addiction. Previous work from our lab determined that VTA serum- and glucocorticoid-inducible kinase 1 (SGK1) transcription and catalytic activity were increased by repeated cocaine administration; however, it was unknown if these biochemical changes contributed to cocaine-elicited behaviors. Using transgenic and viral-mediated manipulations, we investigated the role of VTA SGK1 catalytic activity in regulating cocaine conditioned place preference and self-administration. We showed intra-VTA infusion of a catalytically inactive SGK1 mutant (K127Q) significantly decreased cocaine conditioned place preference (CPP). Further, we found that K127Q expression in VTA DA neurons significantly decreased cocaine CPP, while this same manipulation in VTA GABA neurons had no effect. However, blunted VTA DA SGK1 catalytic activity did not alter cocaine self-administration. Altogether, these studies identify the specific VTA cells critical for SGK1-mediated effects on cocaine CPP but not self-administration.

Role of 5-HT1A receptor-mediated serotonergic transmission in the medial prefrontal cortex in acute restraint stress-induced augmentation of rewarding memory of cocaine in mice

Stress enhances cocaine craving. We recently reported that acute restraint stress increases cocaine conditioned place preference (CPP) in mice; however, the underlying mechanisms remain unclear. This study aimed to examine the role of serotonergic transmission in the medial prefrontal cortex (mPFC) in cocaine CPP enhancement by acute restraint stress, which increases extracellular serotonin (5-HT) levels in the mPFC. Intra-mPFC infusion of the selective serotonin reuptake inhibitor (S)-citalopram prior to the test session significantly increased the cocaine CPP score under non-stressed conditions. This is indicative of the substantial role of increased mPFC 5-HT levels in cocaine CPP enhancement. Moreover, intra-mPFC and systemic administration of the 5-HT1A receptor antagonist WAY100635 immediately before restraint stress exposure significantly attenuated stress-induced cocaine CPP enhancement. Our findings suggest that enhanced serotonergic transmission via 5-HT1A receptors in the mPFC is involved in acute stress-induced augmentation of rewarding memory of cocaine; moreover, the 5-HT1A receptor could be a therapeutic target for stress-induced cocaine craving.

Sleep Deprivation Enhances Cocaine Conditioned Place Preference in an Orexin Receptor-Modulated Manner.

Drug addiction and withdrawal are characterized by sleep disruption, but the effects of sleep disruption on these states are not well characterized. Sleep deprivation (SD) immediately before the cocaine conditioning trials enhanced cocaine conditioned place preference (CPP) in a dose-dependent manner (3, 8 mg/kg but not 15 mg/kg) in mice. SD immediately before the postconditioning test also enhanced cocaine CPP preference in a dose-dependent manner (8 mg/kg, but not 3, 15 mg/kg). Exposure to orexin-receptor antagonism (1 mg/kg SB 334867, an orexin 1 receptor antagonist; OX1R) just before cocaine-conditioning trials or the postconditioning test attenuated SD-enhanced preference. This suggests a potential therapeutic role for the manipulation of the orexin system to mitigate drug seeking, especially in the context of sleep loss before drug exposure.

Stress induces reinstatement of extinguished cocaine conditioned place preference by a sequential signaling via neuropeptide S, orexin, and endocannabinoid.

Neurons containing neuropeptide S (NPS) and orexins are activated during stress. Previously, we reported that orexins released during stress, via orexin OX1 receptors (OX1 Rs), contribute to the reinstatement of cocaine seeking through endocannabinoid/CB1 receptor (CB1 R)-mediated dopaminergic disinhibition in the ventral tegmental area (VTA). Here, we further demonstrated that NPS released during stress is an up-stream activator of this orexin-endocannabinoid cascade in the VTA, leading to the reinstatement of cocaine seeking. Mice were trained to acquire cocaine conditioned place preference (CPP) by context-pairing cocaine injections followed by the extinction training with context-pairing saline injections. Interestingly, the extinguished cocaine CPP in mice was significantly reinstated by intracerebroventricular injection (i.c.v.) of NPS (1 nmol) in a manner prevented by intraperitoneal injection (i.p.) of SHA68 (50 mg/kg), an NPS receptor antagonist. This NPS-induced cocaine reinstatement was prevented by either i.p. or intra-VTA microinjection (i.vta.) of SB-334867 (15 mg/kg, i.p. or 15 nmol, i.vta.) and AM 251 (1.1 mg/kg, i.p. or 30 nmol, i.vta.), antagonists of OX1 Rs and CB1 Rs, respectively. Besides, NPS (1 nmol, i.c.v.) increased the number of c-Fos-containing orexin neurons in the lateral hypothalamus (LH) and increased orexin-A level in the VTA. The latter effect was blocked by SHA68. Furthermore, a 30-min restraint stress in mice reinstated extinguished cocaine CPP and was prevented by SHA68. These results suggest that NPS is released upon stress and subsequently activates LH orexin neurons to release orexins in the VTA. The released orexins then reinstate extinguished cocaine CPP via an OX1 R- and endocannabinoid-CB1 R-mediated signaling in the VTA.

(-)-Stepholidine blocks expression, but not development, of cocaine conditioned place preference in rats

The purpose of this study was to investigate the effects of (-)-stepholidine (SPD), a compound with dopamine D1 partial agonist and D2/D3 antagonist properties, on the development and expression of cocaine conditioned place preference (CPP). Subjects (N = 65; male Long Evans rats) were tested using a CPP procedure consisting of 3 phases: (1) a 15-min pre-exposure session where animals could explore each compartment freely, (2) eight 30-min conditioning sessions where animals were restricted to one side or the other with cocaine (10 mg/kg) or saline, respectively, on alternating days and (3) a 15-minute preference test session where animals could explore each compartment freely. To test the effects of SPD on expression of cocaine CPP, rats were administered vehicle (distilled water with 20 % DMSO), 10, 15 or 20 mg/kg SPD (intraperitoneally) 30 min prior to the test session. We found that 20 mg/kg of SPD significantly blocked the expression of cocaine CPP. To test the effects of SPD on the development of CPP, 0 (vehicle), 10, 15 or 20mg/kg SPD were administered 30 min prior to each cocaine conditioning session and vehicle before each saline conditioning session; no treatment was given prior to the test session. A preference test showed that each SPD group maintained a CPP similar to the vehicle group. These data indicate that SPD can block the expression of a cocaine CPP but has no effect on its development, suggesting that it inhibits the effects of cocaine cues on cocaine incentive motivated behavior. These results suggest that SPD may be a potential treatment for cue-driven aspects of cocaine use disorder.

α2A-adrenergic heteroreceptors are required for stress-induced reinstatement of cocaine conditioned place preference.

The α2a-adrenergic receptor (α2a-AR) agonist guanfacine has been investigated as a potential treatment for substance use disorders. While decreasing stress-induced reinstatement of cocaine seeking in animal models and stress-induced craving in human studies, guanfacine has not been reported to decrease relapse rates. Although guanfacine engages α2a-AR autoreceptors, it also activates excitatory Gi-coupled heteroreceptors in the bed nucleus of the stria terminalis (BNST), a key brain region in driving stress-induced relapse. Thus, BNST α2a-AR heteroreceptor signaling might decrease the beneficial efficacy of guanfacine. We aimed to determine the role of α2a-AR heteroreceptors and BNST Gi-GPCR signaling in stress-induced reinstatement of cocaine conditioned place preference (CPP) and the effects of low dose guanfacine on BNST activity and stress-induced reinstatement. We used a genetic deletion strategy and the cocaine CPP procedure to first define the contributions of α2a-AR heteroreceptors to stress-induced reinstatement. Next, we mimicked BNST Gi-coupled α2a-AR heteroreceptor signaling using a Gi-coupled designer receptor exclusively activated by designer drug (Gi-DREADD) approach. Finally, we evaluated the effects of low-dose guanfacine on BNST cFOS immunoreactivity and stress-induced reinstatement. We show that α2a-AR heteroreceptor deletion disrupts stress-induced reinstatement and that BNST Gi-DREADD activation is sufficient to induce reinstatement. Importantly, we found that low-dose guanfacine does not increase BNST activity, but prevents stress-induced reinstatement. Our findings demonstrate a role for α2a-AR heteroreceptors and BNST Gi-GPCR signaling in stress-induced reinstatement of cocaine CPP and provide insight into the impact of dose on the efficacy of guanfacine as a treatment for stress-induced relapse of cocaine use.

Contribution of D1R-expressing neurons of the dorsal dentate gyrus and Cav1.2 channels in extinction of cocaine conditioned place preference.

Cocaine-associated contextual cues can trigger relapse behavior by recruiting the hippocampus. Extinction of cocaine-associated contextual memories can reduce cocaine-seeking behavior, however the molecular mechanisms within the hippocampus that underlie contextual extinction behavior and subsequent reinstatement remain poorly understood. Here, we extend our previous findings for a role of Cav1.2 L-type Ca2+ channels in dopamine 1 receptor (D1R)-expressing cells in extinction of cocaine conditioned place preference (CPP) in adult male mice. We report that attenuated cocaine CPP extinction in mice lacking Cav1.2 channels in D1R-expressing cells (D1cre, Cav1.2fl/fl) can be rescued through chemogenetic activation of D1R-expressing cells within the dorsal dentate gyrus (dDG), but not the dorsal CA1 (dCA1). This is supported by the finding that Cav1.2 channels are required in excitatory cells of the dDG, but not in the dCA1, for cocaine CPP extinction. Examination of the role of S1928 phosphorylation of Cav1.2, a protein kinase A (PKA) site using S1928A Cav1.2 phosphomutant mice revealed no extinction deficit, likely due to homeostatic scaling up of extinction-dependent S845 GluA1 phosphorylation in the dDG. However, phosphomutant mice failed to show cocaine-primed reinstatement which can be reversed by chemogenetic manipulation of excitatory cells in the dDG during extinction training. These findings outline an essential role for the interaction between D1R, Cav1.2, and GluA1 signaling in the dDG for extinction of cocaine-associated contextual memories.