Esketamine Inhibits Cocaine-Seeking Behaviour Subsequent to Various Abstinence Conditions in Rats.

BACKGROUND: Cocaine use disorder (CUD) is a relapsing brain disease caused by a chronic drug intake that involves neural mechanisms and psychological processes, including depression. Preclinical and clinical studies have demonstrated the promise of pharmacological drugs in controlling the reinstatement of cocaine by targeting the N-methyl-D-aspartate (NMDA) receptor. Recent evidence has revealed that esketamine, a (S) enantiomer of ketamine, shows a high affinity to NMDA receptors and has been used in clinical trials to treat moderate-to-severe depression. METHODS: In the present paper, we investigated the effects of esketamine in regulating cocaine-seeking behaviour induced through the use of cocaine (10 mg/kg) or the cocaine-associated conditioned cue after a short (10 days)-lasting period of drug abstinence with extinction training, home cage or enrichment environment conditions in male rats. Furthermore, we investigated the acute effects of esketamine on locomotor activity in drug-naïve animals. RESULTS: Esketamine (2.5-10 mg/kg) administered peripherally attenuated the reinstatement induced with cocaine priming or the drug-associated conditioned cue after different conditions of abstinence. CONCLUSIONS: These results seem to support esketamine as a candidate for the pharmacological management of cocaine-seeking and relapse prevention; however, further preclinical and clinical research is needed to better clarify esketamine's actions in CUD.

Combined treatment with Sigma1R and A2AR agonists fails to inhibit cocaine self-administration despite causing strong antagonistic accumbal A2AR-D2R complex interactions: the potential role of astrocytes.

Previous studies have indicated that acute treatment with the monoamine stabilizer OSU-6162 (5 mg/kg), which has a high affinity for Sigma1R, significantly increased the density of accumbal shell D2R-Sigma1R and A2AR-D2R heteroreceptor complexes following cocaine self-administration. Ex vivo studies using the A2AR agonist CGS21680 also suggested the existence of enhanced antagonistic accumbal A2AR-D2R allosteric interactions after treatment with OSU-6162 during cocaine self-administration. However, a 3-day treatment with OSU-6162 (5 mg/kg) failed to alter the behavioral effects of cocaine self-administration. To test these results and the relevance of OSU-6162 (2.5 mg/kg) and/or A2AR (0.05 mg/kg) agonist interactions, we administered low doses of receptor agonists during cocaine self-administration and assessed their neurochemical and behavioral effects. No effects were observed on cocaine self-administration; however, marked and highly significant increases using the proximity ligation assay (PLA) were induced by the co-treatment on the density of the A2AR-D2R heterocomplexes in the nucleus accumbens shell. Significant decreases in the affinity of the D2R high- and low-affinity agonist binding sites were also observed. Thus, in low doses, the highly significant neurochemical effects observed upon cotreatment with an A2AR agonist and a Sigma1R ligand on the A2AR-D2R heterocomplexes and their enhancement of allosteric inhibition of D2R high-affinity binding are not linked to the modulation of cocaine self-administration. The explanation may be related to an increased release of ATP and adenosine from astrocytes in the nucleus accumbens shell in cocaine self-administration. This can lead to increased activation of the A1R protomer in a putative A1R-A2AR-D2R complex that modulates glutamate release in the presynaptic glutamate synapse. We hypothesized that the integration of changes in presynaptic glutamate release and postjunctional heteroreceptor complex signaling, where D2R plays a key role, result in no changes in the firing of the GABA anti-reward neurons, resulting in no reduction in cocaine self-administration in the present experiments.

Maternal monosaccharide diets evoke cognitive, locomotor, and emotional disturbances in adolescent and young adult offspring rats.

Anxiety and depression are the most common mental disorders affecting people worldwide. Recent studies have highlighted that a maternal high-sugar diet (HSD) could be a risk factor for neurobehavioural dysregulations, including mood disorders. Increased consumption of added sugar in food such as refined fructose/glucose can increase the risk of metabolic disorders and impact susceptibility to mental disorders. Furthermore, a few papers have reported disabilities in learning and memory among offspring after maternal HSD, thus suggesting a relationship between maternal nutrition and offspring neurogenesis. In this study, we evaluated the impact of maternal monosaccharide consumption based on a glucose (GLU) or fructose (FRU) diet during pregnancy and lactation in adolescent and young adult offspring rats of both sexes on cognitive, locomotor, and emotional disturbances. Locomotor activity, short-term memory, anxiety-like and depressive-like behavior were evaluated in the offspring. We report for the first time that the maternal GLU or FRU diet is sufficient to evoke anxiety-like behavior among adolescent and young adult offspring. Moreover, we found that maternal monosaccharide diets lead to hyperactivity and depressive-like behavior in male adolescent rats. We also noticed that a maternal FRU diet significantly enhanced novelty-seeking behavior only in young adult male rats. Our novel findings indicated that the maternal monosaccharide diet, especially a diet enriched in FRU, resulted in strong behavioral alterations in offspring rats at early life stages. This study also revealed that male rats were more susceptible to hyperactivity and anxiety- and depressive-like phenotypes than female rats. These results suggest that maternal monosaccharide consumption during pregnancy and lactation is an important factor affecting the emotional status of offspring.

Novel GPR18 Ligands in Rodent Pharmacological Tests: Effects on Mood, Pain, and Eating Disorders.

The lack of selective pharmacological tools has limited the full unraveling of G protein-coupled receptor 18 (GPR18) functions. The present study was aimed at discovering the activities of three novel preferential or selective GPR18 ligands, one agonist (PSB-KK-1415) and two antagonists (PSB-CB-5 and PSB-CB-27). We investigated these ligands in several screening tests, considering the relationship between GPR18 and the cannabinoid (CB) receptor system, and the control of endoCB signaling over emotions, food intake, pain sensation, and thermoregulation. We also assessed whether the novel compounds could modulate the subjective effects evoked by Δ9-tetrahydrocannabinol (THC). Male mice or rats were pretreated with the GPR18 ligands, and locomotor activity, depression- and anxiety-like symptoms, pain threshold, core temperature, food intake, and THC-vehicle discrimination were measured. Our screening analyses indicated that GPR18 activation partly results in effects that are similar to those of CB receptor activation, considering the impact on emotional behavior, food intake, and pain activity. Thus, the orphan GPR18 may provide a novel therapeutic target for mood, pain, and/or eating disorders, and further investigation is warranted to better discern its function.

5-HT2C Receptor Stimulation in Obesity Treatment: Orthosteric Agonists vs. Allosteric Modulators.

Obesity is a substantial health and economic issue, and serotonin (5-hydroxytryptamine, 5-HT) is an important neurotransmitter system involved in the regulation of body weight. The 5-HT2C receptors (5-HT2CRs), one of 16 of the 5-HT receptor (5-HTRs) subtypes, play a significant role in food intake and body weight control. In this review, we focused on the 5-HTR agonists, such as fenfluramines, sibutramine, and lorcaserin, which act directly or indirectly at 5-HT2CRs and have been introduced into the clinic as antiobesity medications. Due to their unwanted effects, they were withdrawn from the market. The 5-HT2CR positive allosteric modulators (PAMs) can be potentially safer active drugs than 5-HT2CR agonists. However, more in vivo validation of PAMs is required to fully determine if these drugs will be effective in obesity prevention and antiobesity pharmacology treatment. Methodology strategy: This review focuses on the role of 5-HT2CR agonism in obesity treatment, such as food intake regulation and weight gain. The literature was reviewed according to the review topic. We searched the PubMed and Scopus databases and Multidisciplinary Digital Publishing Institute open-access scientific journals using the following keyword search strategy depending on the chapter phrases: (1) "5-HT2C receptor" AND "food intake", and (2) "5-HT2C receptor" AND "obesity" AND "respective agonists", and (3) "5-HT2C receptor" AND "PAM". We included preclinical studies (only present the weight loss effects) and double-blind, placebo-controlled, randomized clinical trials published since the 1975s (mostly related to antiobesity treatment), and excluded the pay-walled articles. After the search process, the authors selected, carefully screened, and reviewed appropriate papers. In total, 136 articles were included in this review.

Impact of Mephedrone on Fear Memory in Adolescent Rats: Involvement of Matrix Metalloproteinase-9 (MMP-9) and N-Methyl-D-aspartate (NMDA) Receptor.

Treatment of Post-Traumatic Stress Disorder (PTSD) is complicated by the presence of drug use disorder comorbidity. Here, we examine whether conditioned fear (PTSD model) modifies the rewarding effect of mephedrone and if repeated mephedrone injections have impact on trauma-related behaviors (fear sensitization, extinction, and recall of the fear reaction). We also analyzed whether these trauma-induced changes were associated with exacerbation in metalloproteinase-9 (MMP-9) and the GluN2A and GluN2B subunits of N-methyl-D-aspartate (NMDA) glutamate receptor expression in such brain structures as the hippocampus and basolateral amygdala. Male adolescent rats underwent trauma exposure (1.5 mA footshock), followed 7 days later by a conditioned place preference training with mephedrone. Next, the post-conditioning test was performed. Fear sensitization, conditioned fear, anxiety-like behavior, extinction acquisition and relapse were then assessed to evaluate behavioral changes. MMP-9, GluN2A and GluN2B were subsequently measured. Trauma-exposed rats subjected to mephedrone treatment acquired a strong place preference and exhibited impairment in fear extinction and reinstatement. Mephedrone had no effect on trauma-induced MMP-9 level in the basolateral amygdala, but decreased it in the hippocampus. GluN2B expression was decreased in the hippocampus, but increased in the basolateral amygdala of mephedrone-treated stressed rats. These data suggest that the modification of the hippocampus and basolateral amygdala due to mephedrone use can induce fear memory impairment and drug seeking behavior in adolescent male rats.

A High-Sugar Diet Consumption, Metabolism and Health Impacts with a Focus on the Development of Substance Use Disorder: A Narrative Review.

Carbohydrates are important macronutrients in human and rodent diet patterns that play a key role in crucial metabolic pathways and provide the necessary energy for proper body functioning. Sugar homeostasis and intake require complex hormonal and nervous control to proper body energy balance. Added sugar in processed food results in metabolic, cardiovascular, and nervous disorders. Epidemiological reports have shown enhanced consumption of sweet products in children and adults, especially in reproductive age and in pregnant women, which can lead to the susceptibility of offspring's health to diseases in early life or in adulthood and proneness to mental disorders. In this review, we discuss the impacts of high-sugar diet (HSD) or sugar intake during the perinatal and/or postnatal periods on neural and behavioural disturbances as well as on the development of substance use disorder (SUD). Since several emotional behavioural disturbances are recognized as predictors of SUD, we also present how HSD enhances impulsive behaviour, stress, anxiety and depression. Apart from the influence of HSD on these mood disturbances, added sugar can render food addiction. Both food and addictive substances change the sensitivity of the brain rewarding neurotransmission signalling. The results of the collected studies could be important in assessing sugar intake, especially via maternal dietary patterns, from the clinical perspective of SUD prevention or pre-existing emotional disorders. Methodology: This narrative review focuses on the roles of a high-sugar diet (HSD) and added sugar in foods and on the impacts of glucose and fructose on the development of substance use disorder (SUD) and on the behavioural predictors of drugs abuse. The literature was reviewed by two authors independently according to the topic of the review. We searched the PubMed and Scopus databases and Multidisciplinary Digital Publishing Institute open access scientific journals using the following keyword search strategy depending on the theme of the chapter: "high-sugar diet" OR "high-carbohydrate diet" OR "sugar" OR "glucose" OR "fructose" OR "added sugar" AND keywords. We excluded inaccessible or pay-walled articles, abstracts, conference papers, editorials, letters, commentary, and short notes. Reviews, experimental studies, and epidemiological data, published since 1990s, were searched and collected depending on the chapter structure. After the search, all duplicates are thrown out and full texts were read, and findings were rescreened. After the selection process, appropriate papers were included to present in this review.

Increased density and antagonistic allosteric interactions in A2AR-D2R heterocomplexes in extinction from cocaine use, lost in cue induced reinstatement of cocaine seeking.

Neurochemical studies were previously performed on the effects of a 10 day extinction learning from cocaine self-administration on D2R and A2AR recognition and D2R Gi/o coupling in the ventral striatum. In the present study biochemical receptor binding and proximity ligation assay were used to study possible changes in the allosteric receptor-receptor interactions and the density of the A2AR-D2R heterocomplexes in the ventral striatum (nucleus accumbens shell) in extinction from cocaine self-administration including cue induced reinstatement of cocaine seeking. A significant and clear-cut reduction of active lever pressing was observed in extinction on day 10 from cocaine use. In cue induced reinstatement of cocaine self-administration a significant return in active lever presses developed. In extinction, significant increases in the density of A2AR-D2R and D2R-Sigma1R heterocomplexes were observed in nucleus accumbens shell. In contrast, cue-induced reinstatement of cocaine seeking produced no significant changes in these heteroreceptor complexes of the nucleus accumbens shell. In the 3H raclopride/quinpirole competition binding experiments, the extinction led to a significant increase in the D2R Ki, High dissociation constant in the ventral striatum upon ex vivo exposure to CGS 21680 (100 nM), compared to the same exposure performed in membrane preparations from yoked saline rats. No significant changes in D2R Ki, High values were observed in membrane preparations from rats after cue-induced reinstatement of cocaine-seeking undergoing the same exposure ex vivo to CGS 21680 when compared with membrane preparations from yoked saline rats undergoing the same procedures. It seems likely that increased formation of A2AR-D2R and putative A2AR-D2R-Sigma1R heterocomplexes in the nucleus accumbens shell is part of the mechanism for the enhanced antagonistic allosteric A2AR-D2R interactions developed in extinction learning from cocaine. It reduces cocaine reward through reduced D2R function, and these inhibitory mechanisms are no longer in operation in cue induced reinstatement of cocaine seeking.

The effects of cocaine exposure in adolescence: Behavioural effects and neuroplastic mechanisms in experimental models.

Drug addiction is a devastating disorder with a huge economic and social burden for modern society. Although an individual may slip into drug abuse throughout his/her life, adolescents are at higher risk, but, so far, only a few studies have attempted to elucidate the underlying cellular and molecular bases of such vulnerability. Indeed, preclinical evidence indicates that psychostimulants and adolescence interact and contribute to promoting a dysfunctional brain. In this review, we have focused our attention primarily on changes in neuroplasticity brought about by cocaine, taking into account that there is much less evidence from exposure to cocaine in adolescence, compared with that from adults. This review clearly shows that exposure to cocaine during adolescence, acute or chronic, as well as contingent or non-contingent, confers a vulnerable endophenotype, primarily, by causing changes in neuroplasticity. Given the close relationship between drug abuse and psychiatric disorders, we also discuss the translational implications providing an interpretative framework for clinical studies involving addictive as well as affective or psychotic behaviours. LINKED ARTICLES: This article is part of a themed issue on New discoveries and perspectives in mental and pain disorders. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v179.17/issuetoc.

Intravenous administration of Tat-NR2B9c peptide, a PSD95 inhibitor, attenuates reinstatement of cocaine-seeking behavior in rats.

Cocaine use disorder is a serious, chronic and relapsing disease of the nervous system, for which effective treatments do not yet exist. Recently, the role of the N-methyl-d-aspartate (NMDA) receptor subunit GluN2B has been highlighted in cocaine abstinence followed by extinction training. Since the GluN2B subunit is stabilized at synaptic level by the interaction with its scaffolding protein PSD95, in this study we aimed at investigating efficacy of Tat-NR2B9c peptide, a PSD95 inhibitor, which disrupts the interaction of PSD95 with GluN2B, in the attenuation of cocaine seeking-behavior or cue-induced reinstatement. We found that Tat-NR2B9c, administered intravenously, attenuated the reinstatement of active lever presses induced by a priming dose of cocaine or by drug-associated conditioned stimuli. At the same time, the GluN2B/PSD95 complex levels were decreased in the ventral hippocampus of rats that previously self-administered cocaine injected with Tat-NR2B9c during cocaine- or cue-induced reinstatement. In conclusion, we here provide the first evidence showing that the disruption of the GluN2B/PSD95 complexes during cocaine abstinence followed by extinction training may represent a useful strategy to reduce reinstatement of cocaine-seeking behavior.

Enhancement of the GluN2B subunit of glutamatergic NMDA receptors in rat brain areas after cocaine abstinence.

BACKGROUND: Cocaine use disorder is associated with compulsive drug-seeking and drug-taking, whereas relapse may be induced by several factors, including stress, drug-related places, people, and cues. Recent observations strongly support the involvement of the N-methyl-D-aspartate (NMDA) receptors in cocaine use disorders and abstinence, whereas withdrawal in different environments may affect the intensification of relapse. METHODS: The aim of this study was to examine the GluN2B subunit expression and its association with the postsynaptic density protein 95 (PSD95) in several brain structures in rats with a history of cocaine self-administration and housed either in an enriched environment or in an isolated condition. Furthermore, a selective antagonist of the GluN2B subunit-CP 101,606 (10 and 20 mg/kg) administered during exposure to cocaine or a drug-associated conditional stimulus (a cue) was used to evaluate seeking behavior in rats. RESULTS: In rats previously self-administering cocaine, we observed an increase in the GluN2B expression in the total homogenate from the dorsal hippocampus under both enriched environment and isolation. Cocaine abstinence under isolation conditions increased the GluN2B and GluN2B/PSD95 complex levels in the PSD fraction of the prelimbic cortex in rats previously self-administering cocaine. Administration of CP 101,606 attenuated cue-induced cocaine-seeking behavior only in isolation-housed rats. CONCLUSION: In summary, in this study we showed region-specific changes in both the expression of GluN2B subunit and NMDA receptor trafficking during cocaine abstinence under different housing conditions. Furthermore, we showed that the pharmacological blockade of the GluN2B subunit may be useful in attenuating cocaine-seeking behavior.

The coming together of allosteric and phosphorylation mechanisms in the molecular integration of A2A heteroreceptor complexes in the dorsal and ventral striatal-pallidal GABA neurons.

The role of adenosine A2A receptor (A2AR) and striatal-enriched protein tyrosine phosphatase (STEP) interactions in the striatal-pallidal GABA neurons was recently discussed in relation to A2AR overexpression and cocaine-induced increases of brain adenosine levels. As to phosphorylation, combined activation of A2AR and metabotropic glutamate receptor 5 (mGluR5) in the striatal-pallidal GABA neurons appears necessary for phosphorylation of the GluA1 unit of the AMPA receptor to take place. Robert Yasuda (J Neurochem 152: 270-272, 2020) focused on finding a general mechanism by which STEP activation is enhanced by increased A2AR transmission in striatal-pallidal GABA neurons expressing A2AR and dopamine D2 receptor. In his Editorial, he summarized in a clear way the significant effects of A2AR activation on STEP in the dorsal striatal-pallidal GABA neurons which involves a rise of intracellular levels of calcium causing STEP activation through its dephosphorylation. However, the presence of the A2AR in an A2AR-fibroblast growth factor receptor 1 (FGFR1) heteroreceptor complex can be required in the dorsal striatal-pallidal GABA neurons for the STEP activation. Furthermore, Won et al. (Proc Natl Acad Sci USA 116: 8028-8037, 2019) found in mass spectrometry experiments that the STEP splice variant STEP61 can bind to mGluR5 and inactivate it. In addition, A2AR overexpression can lead to increased formation of A2AR-mGluR5 heterocomplexes in ventral striatal-pallidal GABA neurons. It involves enhanced facilitatory allosteric interactions leading to increased Gq-mediated mGluR5 signaling activating STEP. The involvement of both A2AR and STEP in the actions of cocaine on synaptic downregulation was also demonstrated. The enhancement of mGluR5 protomer activity by the A2AR protomer in A2AR-mGluR5 heterocomplexes in the nucleus accumbens shell appears to have a novel significant role in STEP mechanisms by both enhancing the activation of STEP and being a target for STEP61.

The Balance of MU-Opioid, Dopamine D2 and Adenosine A2A Heteroreceptor Complexes in the Ventral Striatal-Pallidal GABA Antireward Neurons May Have a Significant Role in Morphine and Cocaine Use Disorders.

The widespread distribution of heteroreceptor complexes with allosteric receptor-receptor interactions in the CNS represents a novel integrative molecular mechanism in the plasma membrane of neurons and glial cells. It was proposed that they form the molecular basis for learning and short-and long-term memories. This is also true for drug memories formed during the development of substance use disorders like morphine and cocaine use disorders. In cocaine use disorder it was found that irreversible A2AR-D2R complexes with an allosteric brake on D2R recognition and signaling are formed in increased densities in the ventral enkephalin positive striatal-pallidal GABA antireward neurons. In this perspective article we discuss and propose how an increase in opioid heteroreceptor complexes, containing MOR-DOR, MOR-MOR and MOR-D2R, and their balance with each other and A2AR-D2R complexes in the striatal-pallidal enkephalin positive GABA antireward neurons, may represent markers for development of morphine use disorders. We suggest that increased formation of MOR-DOR complexes takes place in the striatal-pallidal enkephalin positive GABA antireward neurons after chronic morphine treatment in part through recruitment of MOR from the MOR-D2R complexes due to the possibility that MOR upon morphine treatment can develop a higher affinity for DOR. As a result, increased numbers of D2R monomers/homomers in these neurons become free to interact with the A2A receptors found in high densities within such neurons. Increased numbers of A2AR-D2R heteroreceptor complexes are formed and contribute to enhanced firing of these antireward neurons due to loss of inhibitory D2R protomer signaling which finally leads to the development of morphine use disorder. Development of cocaine use disorder may instead be reduced through enkephalin induced activation of the MOR-DOR complex inhibiting the activity of the enkephalin positive GABA antireward neurons. Altogether, we propose that these altered complexes could be pharmacological targets to modulate the reward and the development of substance use disorders.

The NMDA Receptor Subunit (GluN1 and GluN2A) Modulation Following Different Conditions of Cocaine Abstinence in Rat Brain Structures.

Different neuronal alterations within glutamatergic system seem to be crucial for developing of cocaine-seeking behavior. Cocaine exposure provokes a modulation of the NMDA receptor subunit expression in rodents, which probably contributes to cocaine-induced behavioral alterations. The aim of this study was to examine the composition of the NMDA receptor subunits in the brain structures in rats with the history of cocaine self-administration after cocaine abstinence (i) in an enriched environment, (ii) in an isolated condition, (iii) with extinction training, or (iv) without instrumental task, as well as the Grin1 (encoding GluN1) and Grin2A (encoding GluN2A) gene expression were evaluated after 10-day extinction training in rat brain structures. In the present study, we observed changes only following cocaine abstinence with extinction training, when the increased GluN2A subunit levels were seen in the postsynaptic density fraction but not in the whole homogenate of the prelimbic cortex (PLC) and dorsal hippocampus (dHIP) in rats previously self-administered cocaine. At the same time, extinction training did not change the Grin1 and Grin2A gene expression in these structures. In conclusion, NMDA receptor subunit modulation observed following cocaine abstinence with extinction training may represent a potential target in cocaine-seeking behavior.

Cocaine abstinence modulates NMDA receptor subunit expression: An analysis of the GluN2B subunit in cocaine-seeking behavior.

Cocaine use disorder develops in part due to the strong associations formed between drugs and the stimuli associated with drug use. Recently, treatment strategies including manipulations of drug-associated memories have been investigated, and the possibility of interfering with N-methyl-d-aspartate (NMDA)-mediated neurotransmission may represent an important option. The aim of this study was to examine the significance of the NMDA receptor subunit GluN2B at the molecular level (the expression of the GluN2B subunit, the Grin2B gene and the association of GluN2B with postsynaptic density protein 95 (PSD95)) in the brain structures of rats with a history of cocaine self-administration after i) cocaine abstinence with extinction training or ii) cocaine abstinence without instrumental tasks, as well as at the pharmacological level (peripheral or intracranial administration of CP 101,606, a GluN2B subunit antagonist during the cocaine- or cue-induced reinstatement). The GluN2B subunit levels and the GluN2B/PSD95 complex levels were either increased in the ventral hippocampus (vHIP) with higher levels of Grin2B gene expression in the HIP or decreased in the dorsal striatum (dSTR) after cocaine abstinence with extinction training. Moreover, CP 101,606, a GluN2B subunit antagonist, administered peripherally, attenuated the reinstatement of active lever presses induced by a priming dose of cocaine or by drug-associated conditioned stimuli, while injection into the vHIP reduced the cocaine- or cue with the subthreshold dose of cocaine-induced reinstatement. In cocaine abstinence without instrumental tasks, an increase in the GluN2B subunit levels and the level of the GluN2B/PSD95 complex in the dSTR was observed in rats that had previously self-administered cocaine. In conclusion, cocaine abstinence with extinction training seems to be associated with the up-regulation of the hippocampal GluN2B subunits, which seems to control cocaine-seeking behavior.

Extinction Training after Cocaine Self-Administration Influences the Epigenetic and Genetic Machinery Responsible for Glutamatergic Transporter Gene Expression in Male Rat Brain.

Glutamate is a key excitatory neurotransmitter in the central nervous system. The balance of glutamatergic transporter proteins allows long-term maintenance of glutamate homeostasis in the brain, which is impaired during cocaine use disorder. The aim of this study was to investigate changes in the gene expression of SLC1A2 (encoding GLT-1), and SLC7A11 (encoding xCT), in rat brain structures after short-term (3 days) and long-term (10 days) extinction training using microarray analysis and quantitative real-time PCR. Furthermore, we analyzed the expression of genes encoding transcription factors, i.e., NFKB1 and NFKB2 (encoding NF-κB), PAX6, (encoding Pax6), and NFE2L2 (encoding Nrf2), to verify the correlation between changes in glutamatergic transporters and changes in their transcriptional factors and microRNAs (miRNAs; miR-124a, miR-543-3p and miR-342-3p) and confirm the epigenetic mechanism. We found reduced GLT-1 transcript and mRNA level in the prefrontal cortex (PFCTX) and dorsal striatum (DSTR) in rats that had previously self-administered cocaine after 3 days of extinction training, which was associated with downregulation of PAX6 (transcript and mRNA) and NFKB2 (mRNA) level in the PFCTX and with upregulation of miR-543-3p and miR-342-3p in the DSTR. The xCT mRNA level was reduced in the PFCTX and DSTR, and NFE2L2 transcript level in the PFCTX was decreased on the 3rd day of extinction training. In conclusion, 3-day drug-free period modulates GLT-1 and xCT gene expression through genetic and epigenetic mechanisms, and such changes in expression seem to be potential molecular targets for developing a treatment for cocaine-seeking behavior.

Cocaine Self-Administration and Abstinence Modulate NMDA Receptor Subunits and Active Zone Proteins in the Rat Nucleus Accumbens.

Cocaine-induced plasticity in the glutamatergic transmission and its N-methyl-d-aspartate (NMDA) receptors are critically involved in the development of substance use disorder. The presynaptic active zone proteins control structural synaptic plasticity; however, we are still far from understanding the molecular determinants important for cocaine seeking behavior. The aim of this study was to investigate the effect of cocaine self-administration and different conditions of cocaine forced abstinence on the composition of the NMDA receptor subunits and on the levels of active zone proteins, i.e., Ras-related protein 3A (Rab3A), Rab3 interacting molecules 1 (RIM1) and mammalian uncoordinated protein 13 (Munc13) in the rat nucleus accumbens. We found an up-regulation of the accumbal levels of GluN1 and GluN2A following cocaine self-administration that was paralleled by an increase of Munc13 and RIM1 levels. At the same time, we also demonstrated that different conditions of cocaine abstinence abolished changes in NMDA receptor subunits (except for higher GluN1 levels after cocaine abstinence with extinction training), while an increase in the Munc13 concentration was shown in rats housed in an enriched environment. In conclusion, cocaine self-administration is associated with the specific up-regulation of the NMDA receptor subunit composition and is related with new presynaptic targets controlling neurotransmitter release. Moreover, changes observed in cocaine abstinence with extinction training and in an enriched environment in the levels of NMDA receptor subunit and in the active zone protein, respectively, may represent a potential regulatory step in cocaine-seeking behavior.

Deleterious Effects of Ethanol, Δ(9)-Tetrahydrocannabinol (THC), and Their Combination on the Spatial Memory and Cognitive Flexibility in Adolescent and Adult Male Rats in the Barnes Maze Task.

Research demonstrates that adolescents differ from adults in their response to drugs of abuse. The aim of the present study was to examine the influence of ethanol, Δ9-tetrahydrocannabinol hydrochloride (THC), and a combination of these drugs given during adolescence on spatial memory in adolescent and adult rats. Thus, adolescent rats (postnatal day (PND) 30) were subjected to the following groups: 0.9% NaCl; 1.5 g/kg ethanol; 1.0 mg/kg THC; 1.5 g/kg ethanol + 1.0 mg/kg THC. Rats received drug injection four times at three-day intervals. One day after the last injection, half of the treated animals were tested in the Barnes maze task, whereas the remaining animals were tested on PND 70. Results show that there was a significant age effect on spatial memory in the Barnes maze task after these drug administrations. Adolescent animals demonstrated more potent deficits in the spatial learning and memory (probe trial) and in cognitive flexibility (reversal learning) than did adults. However, in adult rats that received these drugs in adolescence, memory decline was observed only after ethanol and ethanol + THC administration. Thus, our results are important in understanding the deleterious impact of THC and/or ethanol abuse during adolescence on memory function across the lifespan (adolescent versus adult).

The impact of GABAB receptors and their pharmacological stimulation on cocaine reinforcement and drug-seeking behaviors in a rat model of depression.

Depression and cocaine use disorder represent frequent co-current diagnoses and the GABAB receptors are involved in both conditions. This research involved the application of the animal model of depression (bulbectomy, OBX) and cocaine use disorder (self-administration) to assess the efficiency of GABAB receptor agonists, baclofen and SKF-97541, on cocaine rewarding property and reinforcement of seeking-behaviors in rats with depressive phenotype. Additionally, we applied immunoreactive techniques to determine changes in the expression of GABAB receptor subunit 1 and 2 in rats with depression and cocaine addiction. The results obtained the study illustrate that the GABAB receptor agonists reduced the rewarding property of cocaine in both OBX and control (SHAM) rats. Both agonists significantly reduced cue- and cocaine-induced reinstatement in both groups. This is the first report demonstrating a different impact of cocaine abuse on GABAB receptor levels in depressed animals. It was documented that the expression of GABAB1 subunit in the infralimbic cortex increased during self-administration and extinction training in OBX animals. The lower level of expression for this subunit in addictive SHAM rats during self-administration, and increased in extinguished addictive OBX rats was found in the ventrolateral striatum. The expression of GABAB2 subunit changed only in the case of cocaine self-administration paradigm, as a decline of the subunit level in the nucleus accumbens and ventral hippocampus was observed only in OBX rats. The relevance of GABAB receptors in depression and addiction comorbidity is clearly implicated and can open a new era of drug discovery for individuals with dual diagnosis.