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Neuronal networks are regulated by three-dimensional spatial and structural properties. Despite robust evidence of functional implications in the modulation of cognition, little is known about the three-dimensional internal organization of cholinergic networks in the forebrain. Cholinergic networks in the forebrain primarily occur in subcortical nuclei, specifically the septum, nucleus basalis, globus pallidus, nucleus accumbens, and the caudate-putamen. Therefore, the present investigation analyzed the three-dimensional spatial organization of 14,000 cholinergic neurons that expressed choline acetyltransferase (ChAT) in these subcortical nuclei of the mouse forebrain. Point process theory and graph signal processing techniques identified three topological principles of organization. First, cholinergic interneuronal distance is not uniform across brain regions. Specifically, in the septum, globus pallidus, nucleus accumbens, and the caudate-putamen, the cholinergic neurons were clustered compared with a uniform random distribution. In contrast, in the nucleus basalis, the cholinergic neurons had a spatial distribution of greater regularity than a uniform random distribution. Second, a quarter of the caudate-putamen is composed of axonal bundles, yet the spatial distribution of cholinergic neurons remained clustered when axonal bundles were accounted for. However, comparison with an inhomogeneous Poisson distribution showed that the nucleus basalis and caudate-putamen findings could be explained by density gradients in those structures. Third, the number of cholinergic neurons varies as a function of the volume of a specific brain region but cell body volume is constant across regions. The results of the present investigation provide topographic descriptions of cholinergic somata distribution and axonal conduits, and demonstrate spatial differences in cognitive control networks. The study provides a comprehensive digital database of the total population of ChAT-positive neurons in the reported structures, with the x,y,z coordinates of each neuron at micrometer resolution. This information is important for future digital cellular atlases and computational models of the forebrain cholinergic system enabling models based on actual spatial geometry., (© 2022. The Author(s).)

Sex steroid hormones such as 17β-estradiol (estradiol) regulate neuronal function by binding to estrogen receptors (ERs), including ERα and GPER1, and through differential production via the enzyme aromatase. ERs and aromatase are expressed across the nervous system, including in the striatal brain regions. These regions, comprising the nucleus accumbens core, shell, and caudate-putamen, are instrumental for a wide-range of functions and disorders that show sex differences in phenotype and/or incidence. Sex-specific estrogen action is an integral component for generating these sex differences. A distinctive feature of the striatal regions is that in adulthood neurons exclusively express membrane but not nuclear ERs. This long-standing finding dominates models of estrogen action in striatal regions. However, the developmental etiology of ER and aromatase cellular expression in female and male striatum is unknown. This omission in knowledge is important to address, as developmental stage influences cellular estrogenic mechanisms. Thus, ERα, GPER1, and aromatase cellular immunoreactivity was assessed in perinatal, prepubertal, and adult female and male rats. We tested the hypothesis that ERα, GPER1, and aromatase exhibits sex, region, and age-specific differences, including nuclear expression. ERα exhibits nuclear expression in all three striatal regions before adulthood and disappears in a region- and sex-specific time-course. Cellular GPER1 expression decreases during development in a region- but not sex-specific time-course, resulting in extranuclear expression by adulthood. Somatic aromatase expression presents at prepuberty and increases by adulthood in a region- but not sex-specific time-course. These data indicate that developmental period exerts critical sex-specific influences on striatal cellular estrogenic mechanisms., (© 2020 Wiley Periodicals LLC.)

The smoking incentive in patients with schizophrenia (SCZ) depends on stimulation of nicotinic acetylcholine receptors (nAChRs) in the central nervous system. To detect potential predictor genes for nicotine responses in SCZ, we explored common factor using research data in human and animal samples. In lymphoblastoid cell lines from SCZ, the mRNA expression level of α7 nAChR subunit was decreased. In SCZ-like model mice of phencyclidine (PCP; 10 mg/kg/day, subcutaneously for 14 days)-administered mice, the mRNA expression level of α7 nAChR subunit and protein expression level of α7 or α4 nAChR subunit were significantly decreased in the prefrontal cortex during PCP withdrawal. Protein, but not mRNA, expression levels of α7, α4, and β2 nAChR subunits were significantly increased in the nucleus accumbens. Acute (-)-nicotine [(-)-NIC: 0.3 mg/kg, s.c.] treatment attenuated impairments of social behaviors and visual recognition memory. These effects of (-)-NIC were completely blocked by both methyllycaconitine, a selective α7 nAChR antagonist, and dihydro-β-erythroidine (DHβE), a selective α4β2 nAChR antagonist. (-)-NIC did not induce conditioned place preference, but enhanced sensitivity to methamphetamine-induced hyperactivity. These findings suggest that α7 nAChR is associated with development of disease and is implicated in the therapeutic effect of nicotine in SCZ. The smoking incentive in SCZ might be attributed to treat their own symptoms, rather than a result of (-)-NIC dependence, by stimulating α7 and/or α4β2 nAChRs., Competing Interests: Conflict of interest The authors declare no conflict of interest., (Copyright © 2020 Elsevier B.V. and ECNP. All rights reserved.)

Melanin-concentrating hormone (MCH) is an orexigenic neuropeptide produced in the lateral hypothalamus and zona incerta that increases food intake. The neuronal pathways and behavioral mechanisms mediating the orexigenic effects of MCH are poorly understood, as is the extent to which MCH-mediated feeding outcomes are sex-dependent. Here we investigate the hypothesis that MCH-producing neurons act in the nucleus accumbens shell (ACBsh) to promote feeding behavior and motivation for palatable food in a sex-dependent manner. We utilized ACBsh MCH receptor (MCH1R)-directed pharmacology as well as a dual virus chemogenetic approach to selectively activate MCH neurons that project to the ACBsh. Results reveal that both ACBsh MCH1R activation and activating ACBsh-projecting MCH neurons increase consumption of standard chow and palatable sucrose in male rats without affecting motivated operant responding for sucrose, general activity levels, or anxiety-like behavior. In contrast, food intake was not affected in female rats by either ACBsh MCH1R activation or ACBsh-projecting MCH neuron activation. To determine a mechanism for this sexual dimorphism, we investigated whether the orexigenic effect of ACBsh MCH1R activation is reduced by endogenous estradiol signaling. In ovariectomized female rats on a cyclic regimen of either estradiol (EB) or oil vehicle, ACBsh MCH1R activation increased feeding only in oil-treated rats, suggesting that EB attenuates the ability of ACBsh MCH signaling to promote food intake. Collective results show that MCH ACBsh signaling promotes feeding in an estrogen- and sex-dependent manner, thus identifying novel neurobiological mechanisms through which MCH and female sex hormones interact to influence food intake., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Archaerhodopsin (ArchT)-mediated photoinhibition of axon terminals is commonly used to test the involvement of specific long-range neural projections in behavior. Although sustained activation of this opsin in axon terminals has the unintended consequence of enhancing spontaneous vesicle release, it is unclear whether this desynchronized signaling is consequential for ArchT's behavioral effects. Here, we compare axon terminal and cell body photoinhibition of nucleus accumbens (NAc) afferents to test the utility of these approaches for uncovering pathway-specific contributions of neural circuits to behavior. First, in brain slice recordings we confirmed that ArchT photoinhibition of glutamatergic axons reduces evoked synaptic currents and increases spontaneous transmitter release. A further consequence was increased interneuron activity, which served to broadly suppress glutamate input via presynaptic GABA B receptors. In vivo , axon terminal photoinhibition increased feeding and reward-seeking behavior irrespective of the afferent pathway targeted. These behavioral effects are comparable to those obtained with broad inhibition of NAc neurons. In contrast, cell body inhibition of excitatory NAc afferents revealed a pathway-specific contribution of thalamic input to feeding behavior and amygdala input to reward-seeking under extinction conditions. These findings underscore the off-target behavioral consequences of ArchT-mediated axon terminal inhibition while highlighting cell body inhibition as a valuable alternative for pathway-specific optogenetic silencing., (Copyright © 2020 Lafferty and Britt.)

N-Ethylpentylone (NEP) is a popular synthetic cathinone abused worldwide. To obtain more information about its pharmacokinetics and pharmacodynamics, a rapid, simple and sensitive liquid chromatography-tandem mass spectrometry method was developed for the determination of NEP, two important neurotransmitters, dopamine and serotonin, and their metabolites, including 3,4-dihydroxyphenylacetic acid, 3-methoxytyramine and 5-hydroxyindole-3-acetic acid, in rat brain microdialysate. The analytes were separated on a Phnomenex Polar C 18 column, with a mobile phase of 0.1% formic acid in water (A) and 0.1% formic acid in acetonitrile (B) under gradient elution to shorten the total chromatographic run time. A triple quadruple mass spectrometer coupled with an electrospray ionization source in both positive and negative ion mode was used to detect the analytes. This method showed excellent accuracy (87.4-113.5%) and precision (relative standard deviation <15%) at three quality control levels. The limits of detection were 0.2 ng/mL for NEP and 0.2-50 nm for the others and good linearity was obtained. This study pioneered a method to integrate exogenous drugs and endogenous neurotransmitters as the drugs act on the same determination system, which means that this innovation can provide support for further study of the addictive effects of NEP or other synthetic cathinones on extracellular levels of dopamine and 5-hydroxytryptamine., (© 2019 John Wiley & Sons, Ltd.)

Recent studies suggest that the ventral medial prefrontal cortex (vmPFC) encodes both operant drug self-administration and extinction memories. Here, we examined whether these opposing memories are encoded by distinct neuronal ensembles within the vmPFC with different outputs to the nucleus accumbens (NAc) in male and female rats. Using cocaine self-administration (3 h/d for 14 d) and extinction procedures, we demonstrated that vmPFC was similarly activated (indexed by Fos) during cocaine-seeking tests after 0 (no-extinction) or 7 extinction sessions. Selective Daun02 lesioning of the self-administration ensemble (no-extinction) decreased cocaine seeking, whereas Daun02 lesioning of the extinction ensemble increased cocaine seeking. Retrograde tracing with fluorescent cholera toxin subunit B injected into NAc combined with Fos colabeling in vmPFC indicated that vmPFC self-administration ensembles project to NAc core while extinction ensembles project to NAc shell. Functional disconnection experiments (Daun02 lesioning of vmPFC and acute dopamine D1-receptor blockade with SCH39166 in NAc core or shell) confirm that vmPFC ensembles interact with NAc core versus shell to play dissociable roles in cocaine self-administration versus extinction, respectively. Our results demonstrate that neuronal ensembles mediating cocaine self-administration and extinction comingle in vmPFC but have distinct outputs to the NAc core and shell that promote or inhibit cocaine seeking. SIGNIFICANCE STATEMENT Neuronal ensembles within the vmPFC have recently been shown to play a role in self-administration and extinction of food seeking. Here, we used the Daun02 chemogenetic inactivation procedure, which allows selective inhibition of neuronal ensembles identified by the activity marker Fos, to demonstrate that different ensembles for cocaine self-administration and extinction memories coexist in the ventral mPFC and interact with distinct subregions of the nucleus accumbens., (Copyright © 2019 the authors.)

Exposure to stress increases the risk of developing mood disorders. While a subset of individuals displays vulnerability to stress, others remain resilient, but the molecular basis for these behavioral differences is not well understood. Using a model of chronic social defeat stress, we identified region-specific differences in myelination between mice that displayed social avoidance behavior ('susceptible') and those who escaped the deleterious effect to stress ('resilient'). Myelin protein content in the nucleus accumbens was reduced in all mice exposed to stress, whereas decreased myelin thickness and internodal length were detected only in the medial prefrontal cortex (mPFC) of susceptible mice, with fewer mature oligodendrocytes and decreased heterochromatic histone marks. Focal demyelination in the mPFC was sufficient to decrease social preference, which was restored following new myelin formation. Together these data highlight the functional role of mPFC myelination as critical determinant of the avoidance response to traumatic social experiences., Editorial Note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that all the issues have been addressed (see decision letter)., Competing Interests: VB, KD, MA, MW, AA, JD, GT, HP, NB, PC, JL No competing interests declared, (© 2019, Bonnefil et al.)

Overconsumption of sugars contributes to poor health outcomes. Sugars are often added to commercial foods and beverages in low concentrations and these hidden sugars are consumed unnoticed, continuously. These hidden sugars are suggested to increase the motivation for foodstuffs with higher sugar contents, due to their rewarding properties. This process has been attributed in part, to the activity of both dopaminergic and opioidergic systems in the nucleus accumbens. We asked the question whether prolonged continuous consumption of a low concentration sucrose solution was sufficient to trigger alterations in both dopaminergic and opioidergic systems in the nucleus accumbens of male Sprague-Dawley rats. Rats were given access to either, 1% sucrose and water ad libitum for 3 weeks, or water alone, we then assayed the nucleus accumbens for mRNA and protein expression levels of D1 and D2 dopamine receptors which mediate appetitive motivation and wanting behaviors and for μ-opioid receptors which mediate liking of rewarding stimuli. Our data revealed that rats express a strong preference for 1% sucrose, and showed increased μ-opioid receptor mRNA expression bilaterally in the nucleus accumbens; increased D1 receptor mRNA expression in the left nucleus accumbens; and increased D2 receptor mRNA expression and decreased D2 receptor protein expression in the right nucleus accumbens. We also noted clear individual differences in the volumes of sucrose ingested over this period, however these differences did not correlate with the changes in neurochemistry. Our data show that prolonged ad libitum access to low concentration sucrose alters brain circuits critical for coding reward which may contribute to an enhanced drive for sweet foods and beverages., (Crown Copyright © 2018. Published by Elsevier Inc. All rights reserved.)

Aberrant hippocampal activity is observed in individuals with schizophrenia and is thought to underlie the augmented dopamine system function associated with psychosis. The pathway by which the ventral hippocampus (vHipp) regulates dopamine neuron activity has been demonstrated previously and involves a glutamatergic projection to the nucleus accumbens (NAc). Recent postmortem studies have confirmed glutamatergic abnormalities in the NAc of individuals with schizophrenia. Specifically, an increase in vesicular glutamate transporter 2 (vGlut2) expression was reported. Although projections from the hippocampus do express vGlut2, inputs from the thalamus are more likely to account for this alteration; however, the role of thalamic inputs to the NAc in the regulation of dopamine neuron activity has not been elucidated. Here, using male Sprague Dawley rats, we demonstrate that a subset of NAc medium spiny neurons receive convergent inputs from the vHipp and paraventricular nucleus of the thalamus (PVT), with both regions working synergistically to regulate dopamine neuron activity. Activation of either the vHipp or PVT increases the number of spontaneously active dopamine neurons in the ventral tegmental area. Moreover, this regulation requires simultaneous activity in both regions because PVT inactivation can reverse vHipp-induced increases in dopamine neuron population activity and vHipp inactivation can reverse PVT-induced increases. This is relevant to schizophrenia because inactivation of either the vHipp or PVT is sufficient to reverse aberrant dopamine system function in two distinct rodent models. These data suggest that thalamic abnormalities may contribute to the aberrant dopamine system function observed in schizophrenia and that the PVT represents a novel site of intervention for psychosis. SIGNIFICANCE STATEMENT Current treatments for schizophrenia are far from adequate and a more complete understanding of the pathophysiology underlying this disease is warranted if we are to discover novel therapeutic targets. We have previously demonstrated that the aberrant dopamine system function observed in individuals with schizophrenia and rodent models is driven by increases in hippocampal activity. We now demonstrate that thalamic (paraventricular nucleus, PVT) and ventral hippocampal afferents converge in the nucleus accumbens to regulate dopamine system function. Such information provides a potential site for therapeutic intervention for schizophrenia. Indeed, inactivation of the PVT can effectively reverse aberrant dopamine system function in two distinct rodent models displaying circuit level alterations and corresponding behavioral deficits relevant to schizophrenia., (Copyright © 2018 the authors 0270-6474/18/3810607-12$15.00/0.)

Nicotine addiction and abuse remains a global health issue. To date, the fundamental neurobiological mechanism of nicotine addiction remains incompletely understood. Trace amine-associated receptor 1 (TAAR1) is thought to directly modulate dopaminergic system and are thought to be a neural substrate underlying addictive-like behaviors. We aimed to investigate the role of TAAR1 in nicotine addictive-like behaviors. TAAR1 expression after nicotine treatment was evaluated by western blotting. c-Fos immunofluorescence and in vivo fast-scan cyclic voltammetry were used to examine the activation of brain regions and dopamine release, respectively. We then thoroughly and systematically examined the role of TAAR1 in mediating nicotine-induced sensitization, nicotine discrimination, nicotine self-administration, nicotine demand curve, and the reinstatement of nicotine-seeking. Local pharmacological manipulation was conducted to determine the role of TAAR1 in the nucleus accumbens (NAcs) in the reinstatement of nicotine-seeking. We found that the expression of TAAR1 protein was selectively downregulated in the NAc, with no change in either dorsal striatum or prefrontal cortex. TAAR1 activation was sufficient to block nicotine-induced c-Fos expression in the NAc, while also reducing nicotine-induced dopamine release in the NAc. Systemic administration of TAAR1 agonists attenuated the expression and development of nicotine-induced sensitization, nicotine self-administration, the reinstatement of nicotine-seeking, and increased the elasticity of nicotine demand curve, while intra-NAc infusions of a TAAR1 agonist was sufficient to attenuate nicotine reinstatement. Moreover, TAAR1-knockout rats showed augmented cue-induced and drug-induced reinstatement of nicotine-seeking. These results indicated that modulation of TAAR1 activity regulates nicotine addictive-like behaviors and TAAR1 represents a novel target towards the treatment of nicotine addiction.

Background: There is a need to further anatomically describe the nucleus accumbens (NA), as there is a growing neurosurgical interest in this locus but a limited understanding of its structure. In this study, we evaluated quantitative NA parameters and spatial relationships with adjacent structures found in the telencephalon., Methods: A total of 155 NA specimens from coronal sections and 3 NA specimens from transverse sections were stained using the Mulligan technique as modified by Barnard et al. The distance from the NA to other structures was then measured., Results: The mean radius of the 155 NAs in the coronal sections was 6.23 ± 0.964 mm, averaging 8.99 ± 2.02 mm from midline (coordinate x), 27.09 ± 3.15 mm from the insula, 12.95 ± 3.21 mm from the outer border of the putamen, 10.52 ± 2.66 mm from the upper border of the caudate, and 8.84 ± 2.93 mm from the midline of the lateral ventricle. The mean distance from the NA center of gravity to the middle of the intercommissural line parallel to the midline (coordinate y) was 17.08 ±3.61 mm, and the mean vertical distance from the intercommissural line to the NA was 8.12 ± 1.265 mm., Conclusions: We obtained the stereotactic coordinates of (x, y, z) = (8, 17, -8) for the NA. From this and other delineations of the described position of the NA, it is possible to contribute to stereotactic surgical atlases, improving neurosurgical interventions in this structure., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Extensive rodent literature suggests that the endocannabinoid (eCB) system present in the nucleus accumbens (NAc) modulates dopamine (DA) release in this area. However, expression patterns of the cannabinoid receptor type 1 (CB1R), the synthesizing enzyme N-acyl phosphatidylethanolamine phospholipase D (NAPE-PLD), and the degradation enzyme fatty acid amide hydrolase (FAAH) in the NAc have not yet been described in non-human primates. The goal of this study is therefore to characterize the expression and localization of the eCB system within the NAc of vervet monkeys (Chlorocebus sabaeus) using Western blots and immunohistochemistry. Results show that CB1R, NAPE-PLD, and FAAH are expressed across the NAc rostrocaudal axis, both in the core and shell. CB1R, NAPE-PLD, and FAAH are localized in medium spiny neurons (MSNs) and fast-spiking GABAergic interneurons (FSIs). Dopaminergic projections and astrocytes did not express CB1R, NAPE-PLD, or FAAH. These data show that the eCB system is present in the vervet monkey NAc and supports its role in the primate brain reward circuit.

Contexts exert bi-directional control over relapse to drug seeking. Contexts associated with drug self-administration promote relapse, whereas contexts associated with the absence of self-administration protect against relapse. The nucleus accumbens shell (AcbSh) is a key brain region determining these roles of context. However, the specific cell types, and projections, by which AcbSh serves these dual roles are unknown. Here, we show that contextual control over relapse and abstinence is embedded within distinct output circuits of dopamine 1 receptor (Drd1) expressing AcbSh neurons. We report anatomical and functional segregation of Drd1 AcbSh output pathways during context-induced reinstatement and extinction of alcohol seeking. The AcbSh→ventral tegmental area (VTA) pathway promotes relapse via projections to VTA Gad1 neurons. The AcbSh→lateral hypothalamus (LH) pathway promotes extinction via projections to LH Gad1 neurons. Targeting these opposing AcbSh circuit contributions may reduce propensity to relapse to, and promote abstinence from, drug use., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: We previously reported that a methanol extract of Glycyrrhizae radix (MEGR) blocked methamphetamine-induced locomotor sensitization and conditioned place preference in rats. In the present study, the effects of MEGR on repeated nicotine-induced locomotor sensitization and enhanced extracellular dopamine (DA) release in the nucleus accumbens (Nacc) were evaluated., Methods: Male Sprague-Dawley rats received repeated administrations of nicotine (0.4 mg/kg, subcutaneous) or saline twice a day for 7 d and were challenged with nicotine 4 d after the last daily dosing. During the 4-d withdrawal period, the rats were treated once a day with MEGR (60 or 180 mg/kg/d). Extracellular DA levels were measured by in vivo microdialysis, the malondialdehyde levels and the activities of superoxide dismutase and catalase in the Nacc were biochemically evaluated, and the expression of antioxidant proteins was confirmed by Western blot assays. All data were assessed with analysis of variance tests followed by post-hoc comparison tests and p values <0.05 were considered statistically significant., Results: The expression of repeated nicotine-induced locomotor sensitization was dose-dependently attenuated by MEGR, and 180 mg/kg/d MEGR significantly inhibited augmented accumbal DA release induced by a direct local challenge of nicotine. Moreover, 180 mg/kg/d MEGR reversed increases in malondialdehyde production, decreases in superoxide dismutase and catalase activities, and the reduced expression of nuclear factor erythroid 2-related factor 2 and heme oxygenase 1 in the nicotine-sensitized Nacc., Conclusions: These results suggest that MEGR inhibited nicotine-induced locomotion and dopaminergic sensitization via antioxidant action.

Background: Lateral hypothalamus (LH) involves in modulation of tonic pain. Regarding the direct and indirect neural connections between the LH and nucleus accumbens (NAc), we aimed to examine the pain modulatory role of NAc dopamine receptors in modulation of LH-induced analgesia in the formalin test., Methods: Vehicle-control groups received saline or DMSO into the NAc and saline into the LH. Carbachol-control groups received carbachol (250 nmol/L) into the LH, 5 min after saline or DMSO injection into the NAc. In treatment groups, intra-NAc administration of SCH-23390 or sulpiride (D1-and D2-like dopamine receptor antagonists, respectively) was performed 5 min before carbachol injection. Formalin test was done in all rats 5 min after the second injection., Results: The blockade of NAc dopamine receptors reduced carbachol-induced antinociception during both phases of formalin test and reduction in LH-induced analgesia during the late phase was more than that during the early phase. Furthermore, contribution of D2-like dopamine receptors to mediation of anti-hyperalgesic effect of carbachol was greater than that of D1-like dopamine receptors during the late phase., Conclusions: The findings suggest that LH-VTA-NAc circuit is contributed to the modulation of formalin-induced pain. These findings demonstrate that transmission at D1- and D2-like dopamine receptors mediates the LH-induced analgesia., Significance: Blockade of accumbal dopamine receptors attenuated analgesia induced by carbachol injection into the lateral hypothalamus during both phases of formalin test. Effect of blockade of D1- and D2-like dopamine receptors on reduction in antinociception was more during the late phase. Contribution of D2-like dopamine receptors to mediation of antinociception during the late phase was greater than the early phase., (© 2017 European Pain Federation - EFIC®.)

Loneliness is associated with impaired mental and physical health. Studies of lonely individuals reported differential expression of inflammatory genes in peripheral leukocytes and diminished activation in brain reward regions such as nucleus accumbens, but could not address gene expression in the human brain. Here, we examined genome-wide RNA expression in post-mortem nucleus accumbens from donors (N=26) with known loneliness measures. Loneliness was associated with 1710 differentially expressed transcripts and genes from 1599 genes (DEGs; false discovery rate P<0.05, fold change ⩾|2|, controlling for confounds) previously associated with behavioral processes, neurological disease, psychological disorders, cancer, organismal injury and skeletal and muscular disorders, as well as networks of upstream RNA regulators. Furthermore, a number of DEGs were associated with Alzheimer's disease (AD) genes (that was correlated with loneliness in this sample, although gene expression analyses controlled for AD diagnosis). These results identify novel targets for future mechanistic studies of gene networks in nucleus accumbens and gene regulatory mechanisms across a variety of diseases exacerbated by loneliness.

Background: Prenatal alcohol exposure (PA) restricted to days 8-20 of rat gestation reduces the activity of the dopaminergic neurons (DA) in the ventral tegmental area (VTA). Hyperactivity and impulsivity have been observed under this treatment; however, it is unknown whether DA levels are affected. Decision-making in risk situations, meanwhile, has been associated with impulsive behavior, but because studies of this phenomenon in animal models are limited, we do not yet know whether PA has any effect. The present study investigated the effects of PA on risk-taking behavior and locomotor activity in pre-pubertal male rats. In addition, DA levels in the nucleus accumbens (NAcc) and medial prefrontal cortex (mPFC) in response to methylphenidate (MP) were assessed., Methods: Dams were treated with either two daily 3.0-g/kg doses of alcohol or an isocaloric solution, from days 8-20 of gestation. Locomotor activity and risk-taking in pre-pubertal male rats after PA, and DA levels in the NAcc and mPFC after a single dose of MP (5mg/kg), were analyzed., Results: Hyperactivity and increased risk-taking behavior were observed in the rats treated prenatally with alcohol compared to controls. Methylphenidate increased DA levels in the NAcc regardless of prenatal treatment, but significantly higher DA levels were found in the PA group regardless of postnatal treatment with saline or MP., Conclusion: The presence of hyperactivity and increased risk-taking behavior after prenatal alcohol treatment is supported. Results suggest that higher DA levels in the rats with PA could be due to a down-regulation of postsynaptic receptors, the desensitization of presynaptic receptors, or a compensatory over-activity of DA neurons in the VTA., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The nucleus accumbens (NAc) in the ventral striatum integrates many neurochemical inputs including dopamine and serotonin projections from midbrain nuclei to modulate drug reward. Although D1 and D2 dopamine receptors are differentially expressed in the direct and indirect pathway medium spiny neurons (dMSNs and iMSNs, respectively), 5-HT6 receptors are expressed in both pathways, more strongly than anywhere else in the brain, and are an intriguing target for neuropsychiatric disorders. In the present study, we used viral vectors utilizing dynorphin or enkephalin promoters to drive expression of 5-HT6 receptors or green fluorescent protein (GFP) selectively in the dMSNs or iMSNs of the NAc shell. Rats were then trained to self-administer cocaine. Increased 5-HT6 receptor expression in dMSNs did not change any parameter of cocaine self-administration measured. However, increasing 5-HT6 receptors in iMSNs reduced the amount of cocaine self-administered under fixed-ratio schedules, especially at low doses, increased the time to the first response and the length of the inter-infusion interval, but did not alter motivation as measured by progressive ratio 'break point' analysis. Modeling of cocaine pharmacokinetics in NAc showed that increased 5-HT6 receptors in iMSNs reduced the rat's preferred tissue cocaine concentration at each dose. Finally, increased 5-HT6 receptors in iMSNs facilitated conditioned place preference for a low dose of cocaine. We conclude that 5-HT6 receptors in iMSNs of NAcSh increase the sensitivity to the reinforcing properties of cocaine, particularly at low doses, suggesting that these receptors may be a therapeutic target for the treatment of cocaine addiction.

The neuronal metabolomes in rat striatum (STR), prefrontal cortex (PFC), and nucleus accumbens (NAC) were analyzed by Hadamard transform ion mobility mass spectrometry (HT-IMMS) in order to reveal global and specific metabolic changes induced by cocaine self-administration after 1-day or 3-week withdrawal. Metabolite features were comprehensively separated and detected using HPLC-IMMS within minutes. Global metabolic differences were observed by PCA for comparisons between cocaine and saline treatments at 1-day withdrawal time. Metabolite features that were significantly changed were selected using PCA loadings' plot and unpaired LLL test and then tentatively identified by accurate m/z, yielding a complete profile of metabolic changes induced by cocaine self-administration. The majority of these changes were found at the 1-day withdrawal time, but several of them endured even after 3-week withdrawal from cocaine, and these changes were generally brain region specific. Putatively identified metabolites associated with oxidative stress and energy metabolism were also specifically investigated. We discovered that the dysregulation of creatine/creatinine was different between the STR and NAC, demonstrating that metabolic alterations are brain region specific. Glutathione and adenosine were also changed in their abundance, and the results agreed with previous studies. In general, this study provided a high-throughput analytical platform to perform metabolomics analyses with putative identifications for altered metabolite features induced by cocaine treatment, therefore revealing additional metabolic targets of cocaine-induced changes after early and extended withdrawal times.

Background: The contribution of epigenetic factors, such as histone acetylation and DNA methylation, to the regulation of alcohol-drinking behavior has been increasingly recognized over the last several years. GADD45b is a protein demonstrated to be involved in DNA demethylation at neurotrophic factor gene promoters, including at brain-derived neurotrophic factor (Bdnf) which has been highly implicated in alcohol-drinking behavior., Methods: DNA methyltransferase-1 (Dnmt1), 3a, and 3b, and Gadd45a, b, and g mRNA were measured in the nucleus accumbens (NAc) and ventral tegmental areas of high ethanol (EtOH) consuming C57BL/6J (C57) and low alcohol consuming DBA/2J (DBA) mice using quantitative reverse transcriptase polymerase chain reaction (PCR). In the NAc, GADD45b protein was measured via immunohistochemistry and Bdnf9a mRNA using in situ PCR. Bdnf9a promoter histone H3 acetylated at lysines 9 and 14 (H3K9,K14ac) was measured using chromatin immunoprecipitation, and 5-methylcytosine (5MC) and 5-hydroxymethylcytosine (5HMC) using methylated DNA immunoprecipitation. Alcohol-drinking behavior was evaluated in Gadd45b haplodeficient (+/-) and null mice (-/-) utilizing drinking-in-the-dark (DID) and 2-bottle free-choice paradigms., Results: C57 mice had lower levels of Gadd45b and g mRNA and GADD45b protein in the NAc relative to the DBA strain. C57 mice had lower NAc shell Bdnf9a mRNA levels, Bdnf9a promoter H3K9,K14ac, and higher Bdnf9a promoter 5HMC and 5MC. Acute EtOH increased GADD45b protein, Bdnf9a mRNA, and histone acetylation and decreased 5HMC in C57 mice. Gadd45b +/- mice displayed higher drinking behavior relative to wild-type littermates in both DID and 2-bottle free-choice paradigms., Conclusions: These data indicate the importance of the DNA demethylation pathway and its interactions with histone posttranslational modifications in alcohol-drinking behavior. Further, we suggest that lower DNA demethylation protein GADD45b levels may affect Bdnf expression possibly leading to altered alcohol-drinking behavior., (Copyright © 2016 by the Research Society on Alcoholism.)

The dopamine level in the brain and the mesolimbick dopaminergic system are responsible for nicotine addiction. In the present study, extracellular dopamine in nucleus accumbens was collected by a brain microdialysis technique. Also a sensitive high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method with a gradient elution procedure was developed for a precise determination of dopamine in brain dialysate. The retention time of dopamine was about 11.32 min. The linear range was 20 - 1000 pg/mL. The limit of detection (LOD) and the limit of qualification (LOQ) were 5 and 20 pg/mL, respectively. The recovery ranged from 98.2 to 109.0%, and both the intra-RSD and inter-RSD were below <9%, respectively. The probe recovery for dopamine in this brain microdialysis experiment was about 25%. Finally, the dopamine concentrations in the rat brain microdialysates were determined, and the pharmacokinetics of extracellular dopamine in the brain nucleus accumbens after an intravenous injection of nicotine was successfully evaluated.

It is widely accepted that D1 dopamine receptor-expressing striatal neurons convey their information directly to the output nuclei of the basal ganglia, whereas D2-expressing neurons do so indirectly via pallidal neurons. Combining optogenetics and electrophysiology, we found that this architecture does not apply to mouse nucleus accumbens projections to the ventral pallidum. Thus, current thinking attributing D1 and D2 selectivity to accumbens projections akin to dorsal striatal pathways needs to be reconsidered.

This study investigated by microdialysis the role of response contingency and food-associated cues in the responsiveness of dopamine transmission in the nucleus accumbens shell and core to sucrose feeding. In naive rats, single-trial non-contingent presentation and feeding of sucrose pellets increased dialysate shell dopamine and induced full habituation of dopamine responsiveness to sucrose feeding 24 and 48 h later. In rats trained to respond for sucrose pellets on a fixed ratio 1 (FR1) schedule, dialysate dopamine increased in the shell but not in the core during active responding as well as under extinction in the presence of sucrose cues. In rats yoked to the operant rats, the presentation of sucrose cues also increased dialysate dopamine selectively in the shell. In contrast, non-contingent sucrose presentation and feeding in FR1-trained and in yoked rats increased dialysate dopamine to a similar extent in the shell and core. It is concluded that, whereas non-contingent sucrose feeding activated dopamine transmission in the shell and core, response-contingent feeding activated, without habituation, dopamine transmission selectively in the shell as a result of the action of sucrose conditioned cues. These observations are consistent with a critical role of conditioned cues acquired during training and differential activation of shell vs. core dopamine for response-contingent sucrose feeding., (© 2015 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Rationale: Women are twice as likely as men to develop major depressive disorder. Exposure to chronic stress can induce depression in some vulnerable individuals, while others are resistant to depressive-like symptoms after equivalent levels of chronic stress., Objectives: In female rats, individual differences in saccharin intake during chronic social defeat stress may predict subsequent cocaine self-administration, and may be attributed to alterations in mesolimbic dopamine activity., Methods: Female rats were exposed to 21 days of chronic social defeat stress, during which they were evaluated for their anhedonia-like responses in the form of saccharin intake. After chronic social defeat stress, the rats were tested for behavioral cross-sensitization to cocaine and escalated cocaine self-administration in a 24-h "binge." A separate group of animals underwent in vivo microdialysis of the nucleus accumbens (NAc) shell to assess dopamine (DA) in response to acute cocaine challenge., Results: Cluster analysis revealed two phenotypes among the stressed female rats based on their saccharin intake while being exposed to stress, termed stress-resistant (SR, 28 %) and stress-sensitive (SS, 72 %). The amount of cocaine self-administered during the 24-h "binge" was positively correlated with preceding saccharin intake. The NAc DA response to a cocaine challenge was significantly lower in SR rats than in the SS and non-stressed control rats. No other significant differences were observed in behavioral cross-sensitization or cocaine self-administration prior to the "binge.", Conclusion: Female rats showed individual differences in their anhedonic-like response to chronic social defeat stress, and these differences were reliably associated with subsequent cocaine-taking behavior.

The peripheral administration of lipopolysaccharide (LPS) induces depressive-like behavior. Anhedonia is a core symptom of depression, defined as a loss of the capacity to experience pleasure. The present study used the sucrose preference test to investigate the influence of Ginkgo biloba extract (EGb 761) on LPS-induced anhedonia in male rats. The animals were randomly divided into four groups: (I) vehicle + saline, (II) vehicle + LPS, (III) EGb 761 + saline, and (IV) EGb 761 + LPS. Saline or LPS (100 µg/kg) was administered intraperitoneally 2 h before the sucrose preference test. Sucrose consumption was recorded 2, 4, 6, 13, and 24 h after 100 µg/kg of LPS or saline injection in the dark phase of the light/dark cycle. Dopamine and serotonin levels in the nucleus accumbens were measured. Our results indicated that the vehicle + LPS group exhibited a significant decrease in sucrose intake compared with the vehicle + saline group. The EGb 761 + LPS group showed more sucrose and food consumption than the vehicle + LPS group. Additionally, compared with the EGb 761 + LPS group, the vehicle + LPS group had less dopamine levels in the nucleus accumbens. Treatment with EGb 761 had no effect on water intake. Our results suggest that EGb 761 may be useful for reducing anhedonic depressive-like behavior., (© 2014 TheAuthors. Phytotherapy Research published by John Wiley & Sons, Ltd.)

The signal transduction modulator Rgs9-2 (Regulator of G protein signaling 9-2) plays a key role in dopaminergic and opioidergic transmission in the striatum. Rgs9-2 is a potent modulator of opiate reward and analgesia, but its role in chronic pain remains unknown. Here, we use the spared nerve injury model (SNI), to evaluate the influence of Rgs9-2 in sensory symptoms, as well as in anxiety and depression-like behaviors observed under neuropathic pain conditions. Our data demonstrate that knockout of the Rgs9 gene reduces the intensity of thermal hyperalgesia and mechanical allodynia the first few days after nerve injury. This small, but significant effect is only observed at early time points after nerve injury, whereas after the first week of SNI, Rgs9 knockout (Rgs9KO) and Rgs9 wildtype (Rgs9WT) mice show similar levels of mechanical allodynia and thermal hyperalgesia. Furthermore, Rgs9-2 deletion exacerbates anxiety and depression like behaviors several weeks after the emergence of the neuropathic pain symptoms. Our findings also reveal a temporal and regional regulation of Rgs9-2 protein expression by neuropathic pain, as Rgs9-2 levels are reduced in the spinal cord a few days after nerve injury, whereas decreased Rgs9-2 levels in the Nucleus Accumbens (NAc) are only observed several weeks after nerve injury. Thus, adaptations in Rgs9-2 activity in the spinal cord and in the NAc may contribute to sensory and affective components of neuropathic pain., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Objective: Developmental disorders including cognitive deficit, hyperkinetic disorder, and autistic behaviors are frequently comorbid in epileptic patients with SCN1A mutations. However, the mechanisms underlying these developmental disorders are poorly understood and treatments are currently unavailable. Using a rodent model with an Scn1a mutation, we aimed to elucidate the pathophysiologic basis and potential therapeutic treatments for developmental disorders stemming from Scn1a mutations., Methods: We conducted behavioral analyses on rats with the N1417H-Scn1a mutation. With high-performance liquid chromatography, we measured dopamine and its metabolites in the frontal cortex, striatum, nucleus accumbens, and midbrain. Methylphenidate was administered intraperitoneally to examine its effects on developmental disorder-like behaviors and hyperthermia-induced seizures., Results: Behavioral studies revealed that Scn1a-mutant rats had repetitive behavior, hyperactivity, anxiety-like behavior, spatial learning impairments, and motor imbalance. Dopamine levels in the striatum and nucleus accumbens in Scn1a-mutant rats were significantly lower than those in wild-type rats. In Scn1a-mutant rats, methylphenidate, by increasing dopamine levels in the synaptic cleft, improved hyperactivity, anxiety-like behavior, and spatial learning impairments. Surprisingly, methylphenidate also strongly suppressed hyperthermia-induced seizures., Significance: Dysfunction of the mesolimbic dopamine reward pathway may contribute to the hyperactivity and learning impairments in Scn1a-mutant rats. Methylphenidate was effective for treating hyperactivity, learning impairments, and hyperthermia-induced seizures. We propose that methylphenidate treatment may ameliorate not only developmental disorders but also epileptic seizures in patients with SCN1A mutations., (© 2014 The Authors. Epilepsia published by Wiley Periodicals, Inc. on behalf of International League Against Epilepsy.)

Acetylcholinesterase activity was quantitatively evaluated by cytochemical method in brain structures (layers III and V of the sensorimotor cortex, caudate nucleus, nucleus accumbens, hippocampus CA3 field) of August and Wistar rats demonstrating high and low motor activity in the open field test. In August rats, acetylcholinesterase activity in the analyzed brain structures prevailed in animals with high motor activity in comparison with rats with low motor activity. In Wistar rats, the differences between the animals demonstrating high and low motor activity were less pronounced, but varied depending on the experimental series of studies. Comparisons of August rats with low motor activity and Wistar rats with high motor activity (maximum difference of motor function in these animals) revealed significant excess of acetylcholinesterase activity in layer III of the sensorimotor cortex in August rats and no differences in other brain structures of the examined animals.

Many behavioral, physiological, and anatomical studies utilize animal models to investigate human striatal pathologies. Although commonly used, rodent striatum may not present the optimal animal model for certain studies due to a lesser morphological complexity than that of non-human primates, which are increasingly restricted in research. As an alternative, the tree shrew could provide a beneficial animal model for studies of the striatum. The gross morphology of the tree shrew striatum resembles that of primates, with separation of the caudate and putamen by the internal capsule. The neurochemical anatomy of the ventral striatum, specifically the nucleus accumbens, has never been examined. This major region of the limbic system plays a role in normal physiological functioning and is also an area of interest for human striatal disorders. The current study uses immunohistochemistry of calbindin and tyrosine hydroxylase (TH) to determine the ultrastructural organization of the nucleus accumbens core and shell of the tree shrew (Tupaia glis belangeri). Stereology was used to quantify the ultrastructural localization of TH, which displays weaker immunoreactivity in the core and denser immunoreactivity in the shell. In both regions, synapses with TH-immunoreactive axon terminals were primarily symmetric and showed no preference for targeting dendrites versus dendritic spines. The results were compared to previous ultrastructural studies of TH and dopamine in rat and monkey nucleus accumbens. Tree shrews and monkeys show no preference for the postsynaptic target in the shell, in contrast to rats which show a preference for synapsing with dendrites. Tree shrews have a ratio of asymmetric to symmetric synapses formed by TH-immunoreactive terminals that is intermediate between rats and monkeys. The findings from this study support the tree shrew as an alternative model for studies of human striatal pathologies., (Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.)

Background: Conditioned place preference (CPP) to ethanol (EtOH) is an important addiction-related alteration thought to be mediated by changed neurotransmission in the mesocorticolimbic brain pathway. Stress is a factor of major importance for the initiation, maintenance, and reinstatement of drug abuse and modulates the neurochemical outcomes of drugs. Thus, the aim of this study was to investigate the effects of concomitant exposure to chronic EtOH and stress on CPP to this drug and alterations of dopaminergic and serotonergic neurotransmission in mice., Methods: Male Swiss mice were chronically treated with EtOH via a liquid diet and were exposed to forced swimming stress. After treatment, animals were evaluated for conditioning, extinction, and reinstatement of CPP to EtOH. Also, mice exposed to the same treatment protocol had their prefrontal cortex (PFC), nucleus accumbens (NAc), and amygdala dissected for the quantitation of dopamine, serotonin, and their metabolites content., Results: Data showed that previous chronic exposure to EtOH potentiated EtOH conditioning and increased dopaminergic turnover in PFC. Exposure to stress potentiated EtOH conditioning and decreased dopaminergic turnover in the NAc. However, animals exposed to both chronic EtOH and stress did not display alterations of CPP and showed an elevated content of dopamine in amygdala. No treatment yielded serotonergic changes., Conclusions: The present study indicates that previous EtOH consumption as well as stress exposure induces increased EtOH conditioning, which can be related to dopaminergic alterations in the PFC or NAc. Interestingly, concomitant exposure to both stimuli abolished each other's effect on conditioning and PFC or NAc alterations. This protective outcome can be related to the dopaminergic increase in the amygdala., (Copyright © 2014 by the Research Society on Alcoholism.)

Individual differences in the forced swimming test (FST) could be associated with differential temporal dynamics of gene expression and neurotransmitter activity. We tested juvenile male rats in the FST and classified the animals into those with low and high immobility according to the amount of immobility time recorded in FST. These groups and a control group which did not undergo the FST were sacrificed either 1, 6 or 24 h after the test. We analyzed the expression of the CRF, CRFR1, BDNF and TrkB in the prefrontal cortex, hippocampus and nucleus accumbens as well as norepinephrine, dopamine, serotonin, glutamate, GABA and glutamine in the hippocampus and nucleus accumbens. Animals with low immobility showed significant reductions of BDNF expression across time points in both the prefrontal cortex and the nucleus accumbens when compared with non-swim control. Moreover, rats with high immobility only showed a significant decrease of BDNF expression in the prefrontal cortex 6h after the FST. Regarding neurotransmitters, only accumbal dopamine turnover and hippocampal glutamate content showed an effect of individual differences (i.e. animals with low and high immobility), whereas nearly all parameters showed significant differences across time points. Correlational analyses suggest that immobility in the FST, probably reflecting despair, is related to prefrontal cortical BDNF and to the kinetics observed in several other neurochemical parameters. Taken together, our results suggest that individual differences observed in depression-like behavior can be associated not only with changes in the concentrations of key neurochemical factors but also with differential time courses of such factors., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: Binge ethanol (EtOH) intake during adolescence leads to an array of behavioral and cognitive consequences including elevated intake of EtOH during adulthood, with female mice showing greater susceptibility than males. Administration of the metabotropic glutamate receptor 5 (mGluR5) antagonist 3-((2-Methyl-1,3-thiazol-4-yl)ethynyl)pyridine (MTEP) has been shown to reduce EtOH self-administration in adult male mice, but little is known about its effect on female and adolescent mice., Methods: MTEP (0, 10, 20 mg/kg, i.p.) was repeatedly administered to female and male, adult and adolescent C57BL/6J mice during binge sessions using the scheduled high alcohol consumption paradigm. Next, we assessed whether MTEP administration during binge altered the subsequent 24-hour EtOH intake following a period of abstinence. Finally, we investigated whether MTEP administration during binge followed by an abstinence period altered mRNA of glutamatergic genes within the nucleus accumbens of female mice., Results: MTEP significantly decreased binge EtOH intake in all mice, but only female mice exhibited altered subsequent 24-hour EtOH intake. Interestingly, the alteration in subsequent EtOH intake in female animals was age dependent, with adolescent exposure to MTEP during binge decreasing 24-hour intake and adult exposure to MTEP during binge increasing 24-hour intake. Finally, while there were no effects of MTEP pretreatment on the genes examined, there was a robust age effect found during analysis of mGluR1 ( Grm1 ), mGluR5 ( Grm5 ), the NR2A subunit of the NMDA receptor ( Grin2a ), phosphatidylinositol 3-kinase ( Pik3r1 ), mammalian target of rapamycin ( Mtor ), and extracellular signal-regulated kinase ( Mapk1 ) mRNA, with adolescent female animals having lower expression than their adult counterparts., Conclusions: Collectively, the present findings add to existing evidence implicating the contribution of long-term effects of adolescent binge drinking to enhance alcohol abuse in adulthood, while suggesting that mGluR5 antagonism may not be the best pharmacotherapy to treat binge alcohol consumption in female and adolescent animals.

Background: Females exhibit more rapid escalation of cocaine use and enhanced cocaine-taking behavior as compared to males. While ovarian hormones likely play a role in this increased vulnerability, research has yet to examine the role of estradiol in affecting the behavioral and neurological response to cocaine in a brain region- and sex-specific way., Methods: First, we examined stereotypy and locomotor sensitization after repeated cocaine administration (10 mg/kg i.p.) in intact (SHAM) and castrated (CAST) males, and ovariectomized (OVX) females treated with 5 μg estradiol benzoate (EB) or vehicle (OIL). Next, we used in vivo microdialysis to examine the effects of acute EB treatment on cocaine-induced DA in the regions mediating the display of these behaviors (i.e., the dorsolateral striatum, DLS; and the nucleus accumbens, NAc; respectively)., Results: We find that EB enhances sensitization of cocaine-induced stereotypy in OVX females after 12 days of cocaine treatment, and after a 10-day withdrawal. Similarly, the OVX/EB females show enhanced locomotor sensitization compared to the other three groups on the same days. Using in vivo microdialysis to assess the neurochemical response, we find that EB rapidly enhances cocaine-induced DA in DLS dialysate of OVX females but not CAST males, and has no effect in NAc of either sex., Conclusions: With these experiments, we show that there are sex differences in the effects of estradiol to preferentially enhance the response to cocaine in the DLS over the NAc in females, which may contribute to the preferential sensitization of stereotypy in females., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

Data in animal models and surveys in humans have revealed psychiatric complications of long-term anabolic androgenic steroid abuse. However, the neurobiochemical mechanisms behind the observed behavioral changes are poorly understood. The aim of the present study was to investigate the effects of nandrolone decanoate on emotional behavior and neurochemical brain alterations in gonadally intact male rats. The behavioral reactivity to the elevated plus maze and the social interaction test was used to assess anxiety-related symptoms, and the sucrose preference test was used to evaluate anhedonia. Dopaminergic, serotonergic and noradrenergic transmissions were also evaluated in selected brain areas. The chronic administration of nandrolone, at 5 mg kg(-1) injected daily for 4 weeks, induced the loss of sweet taste preference, a sign of anhedonia and dysfunction of the reward pathway. The behavioral outcomes were accompanied by reductions in the dopamine, serotonin and noradrenaline contents in the nucleus accumbens. Alterations in the time spent in the open arms and in the social interaction test were not found, suggesting that nandrolone did not induce an anxiogenic profile. No differences were revealed between the experimental groups in the amygdala in terms of the neurotransmitters measured. Our data suggest that nandrolone-treated rats have a depressive, but not anxiogenic-like, profile, accompanied by brain region-dependent changes in dopaminergic, serotonergic and noradrenergic neurotransmission. As anabolic androgenic steroid dependence is plausibly the major form of worldwide substance dependence that remains largely unexplored, it should be highlighted that our data could contribute to a better understanding of the altered rewards induced by nandrolone treatment and to the development of appropriate treatments.

Paternal care is necessary for the healthy development of social behavior in monogamous rodents and social recognition underpins social behavior in these animals. The effects of paternal care on the development of social recognition and underlying neuroendocrine mechanisms, especially the involvement of oxytocin and estrogen pathways, remain poorly understood. We investigated the effects of paternal deprivation (PD: father was removed from neonatal pups and mother alone raised the offspring) on social recognition in mandarin voles (Microtus mandarinus), a socially monogamous rodent. Paternal deprivation was found to inhibit the development of social recognition in female and male offspring according to a habituation-dishabituation paradigm. Paternal deprivation resulted in increased inactivity and reduced investigation during new encounters with other animals. Paternal deprivation reduced oxytocin receptor (OTR) and estrogen receptor α (ERα) mRNA expression in the medial amygdala and nucleus accumbens. Paternal deprivation reduced serum oxytocin (OT) concentration in females, but had no effect on males. Our results provide substantial evidence that paternal deprivation inhibits the development of social recognition in female and male mandarin voles and alters social behavior later in life. This is possibly the result of altered expression of central OTR and ERα and serum OT levels caused by paternal deprivation., (Copyright © 2013 Elsevier Inc. All rights reserved.)

The ability to map the functional connectivity of discrete cell types in the intact mammalian brain during behavior is crucial for advancing our understanding of brain function in normal and disease states. We combined designer receptor exclusively activated by designer drug (DREADD) technology and behavioral imaging with μPET and [18F]fluorodeoxyglucose (FDG) to generate whole-brain metabolic maps of cell-specific functional circuits during the awake, freely moving state. We have termed this approach DREADD-assisted metabolic mapping (DREAMM) and documented its ability in rats to map whole-brain functional anatomy. We applied this strategy to evaluating changes in the brain associated with inhibition of prodynorphin-expressing (Pdyn-expressing) and of proenkephalin-expressing (Penk-expressing) medium spiny neurons (MSNs) of the nucleus accumbens shell (NAcSh), which have been implicated in neuropsychiatric disorders. DREAMM revealed discrete behavioral manifestations and concurrent engagement of distinct corticolimbic networks associated with dysregulation of Pdyn and Penk in MSNs of the NAcSh. Furthermore, distinct neuronal networks were recruited in awake versus anesthetized conditions. These data demonstrate that DREAMM is a highly sensitive, molecular, high-resolution quantitative imaging approach.

The nucleus accumbens (NAc) is a critical brain region involved in many reward-related behaviors. The NAc comprises major compartments the core and the shell, which encompass several subterritories. GABAergic medium-sized spiny neurons (MSNs) constitute the output neurons of the NAc core and shell. While the functional organization of the NAc core outputs resembles the one described for the dorsal striatum, a simple classification of the NAc shell neurons has been difficult to define due to the complexity of the compartmental segregation of cells. We used a variety of BAC transgenic mice expressing enhanced green fluorescence (EGFP) or the Cre-recombinase (Cre) under the control of the promoter of dopamine D1, D2, and D3 receptors and of adenosine A2a receptor to dissect the microanatomy of the NAc. Moreover, using various immunological markers we characterized in detail the distribution of MSNs in the mouse NAc. In addition, cell-type specific extracellular signal-regulated kinase (ERK) phosphorylation in the NAc subterritories was analyzed following acute administration of SKF81297 (a D1R-like agonist), quinpirole (a D2 receptors (D2R)-like agonist), apomorphine (a non-selective DA receptor agonist), raclopride (a D2R-like antagonist), and psychostimulant drugs, including cocaine and d-amphetamine. Each drug generated a unique topography and cell-type specific activation of ERK in the NAc. Our results show the existence of marked differences in the receptor expression pattern and functional activation of MSNs within the shell subterritories. This study emphasizes the anatomical and functional heterogeneity of the NAc, which will have to be considered in its further study.

Taste stimuli increase extracellular dopamine (DA) in the nucleus accumbens (NAc) and in the medial prefrontal cortex (mPFC). This effect shows single-trial habituation in NAc shell but not in core or in mPFC. Morphine sensitization abolishes habituation of DA responsiveness in NAc shell but induces it in mPFC. These observations support the hypothesis of an inhibitory influence of mPFC DA on NAc DA. To test this hypothesis, we used in vivo microdialysis to investigate the effect of mPFC 6-hydroxy-dopamine (6-OHDA) lesions on the NAc DA responsiveness to taste stimuli. 6-OHDA was infused bilaterally in the mPFC of rats implanted with guide cannulae. After 1 week, rats were implanted with an intraoral catheter, microdialysis probes were inserted into the guide cannulae, and dialysate DA was monitored in NAc shell/core after intraoral chocolate. 6-OHDA infusion reduced tissue DA in the mPFC by 75%. Tyrosine hydroxylase immunohistochemistry showed that lesions were confined to the mPFC. mPFC 6-OHDA lesion did not affect the NAc shell DA responsiveness to chocolate in naive rats but abolished habituation in rats pre-exposed to the taste. In the NAc core, mPFC lesion potentiated, delayed and prolonged the stimulatory DA response to taste but failed to affect DA in pre-exposed rats. Behavioural taste reactions and motor activity were not affected. The results indicate a top-down control of NAc DA by mPFC and a reciprocal relationship between DA transmission in these two areas. Moreover, habituation of DA responsiveness in the NAc shell is dependent upon an intact DA input to the mPFC., (© 2012 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.)

A rapid and sensitive liquid chromatography/tandem mass spectrometry (LC-MS/MS) method has been developed for the simultaneous measurement of adenosine (ADE), dopamine (DA), acetylcholine (ACh) and 5-hydroxytryptamine (5-HT) in mouse brain microdialysates. High method sensitivity (LLOQ of 0.05nM) was achieved by optimization of chromatographic and mass spectrometric parameters. The method was fully validated for its sensitivity, selectivity, matrix effect and stability. The LC-MS/MS method was successfully applied to evaluate the effect of the systemic administration of cocaine or amphetamine on the extracellular levels of ADE, DA, ACh and 5-HT in the mouse nucleus accumbens by microdialysis., (Copyright © 2012 Elsevier B.V. All rights reserved.)

In our previous study, we first demonstrated a significant effect of dextromethorphan (DM) on morphine-seeking behavior in morphine-dependent rats, when DM was given during morphine withdrawal. Using the same conditioned place preference (CPP) paradigm modified for measuring drug-seeking-related behavior, we further investigated the possible effect of DM on methamphetamine (MA)-seeking in MA-dependent rats. Our data showed that DM could also effectively suppress the drug-seeking behavior for MA, when administered during MA withdrawal. This suggests that DM may possess a pharmacological property to prevent drug-seeking behavior for addictive drugs in general. To examine the action sites of DM in the brain, DM was microinjected into the VTA or the NAc, and tested for its effect on MA-seeking during withdrawal. Both intra-VTA and intra-NAc injections of DM were able to block the MA-seeking, suggesting that DM has a dual action sites. In our neurochemical results, intra-NAc injection of DM showed a clear reduction of DA turnover rate at the NAc and the mPFC in response to MA challenge during withdrawal, which matched with the behavioral results. However, intra-VTA injection of DM reduced the DA turnover rate at the mPFC but did not have effect on the DA turnover rate at the NAc. Although further investigations may be needed to verify the connection between our neurochemical and behavioral results, the present study highlights the therapeutic potential of DM in antidrug-seeking behavior of MA and that the mechanism could be related to its effect on the mesolimbic and mesocortical dopaminergic pathways., (Copyright © 2012 Wiley Periodicals, Inc.)

Altered brain serotonin activity is implicated in schizophrenia. We have previously shown differential involvement of serotonergic projections from the dorsal or median raphe nucleus in phencyclidine-induced hyperlocomotion in rats, a behavioral model of aspects of schizophrenia. Here we further investigated the effects of serotonergic lesions of the raphe nuclei on phencyclidine-induced hyperlocomotion by parallel assessment of Fos-like immunoreactivity (FLI), a marker of neuronal activation in the brain. Male Sprague-Dawley rats were anesthetized with pentobarbitone and stereotaxically microinjected with 5 μg of the serotonergic neurotoxin, 5,7-dihydroxytryptamine (5,7-DHT), into either the dorsal raphe (DRN) or median raphe nucleus (MRN). Two weeks after the surgery, rats with lesions of the MRN, but not those with lesions of the DRN, showed significant enhancement of the hyperlocomotion induced by injection of 2.5 mg/kg of phencyclidine. Rats with MRN lesions also showed significantly higher levels of FLI in the polymorphic layer of the dentate gyrus in the dorsal hippocampus (PoDG) when compared with sham-operated controls. Rats with lesions of the DRN showed significantly higher levels of FLI in the nucleus accumbens (NAcc). These results indicate that FLI in the PoDG, but not the NAcc, correlates with enhanced phencyclidine-induced locomotor hyperactivity in MRN-lesioned rats. These results support our previous studies suggesting a role of serotonergic projections from the MRN to the dorsal hippocampus in some of the symptoms of schizophrenia., (Copyright © 2012 Wiley Periodicals, Inc.)

Extracellular glycine modulates accumbal dopamine levels as well as ethanol-induced dopamine overflow. Glycine availability is also crucial for regulating alcohol consumption and the glycine transporter 1 (GlyT-1) inhibitor Org25935 robustly decreases alcohol intake in rats. To explore whether the alcohol-intake reducing effect of Org25935 is substance bound, we examined the effect of a different selective GlyT-1 inhibitor, Org24598, on ethanol consumption in rats and compared the effect with that of acamprosate, a drug currently in clinical use. We studied the effects of daily Org24598 and acamprosate injections on male Wistar rats with ~60% ethanol preference in a limited access two bottle free-choice model for 12 days, followed by alcohol deprivation for 14 days before a second test period of 10 days. Finally, rats underwent in vivo microdialysis where dopamine, glycine, taurine and β-alanine in n. accumbens were measured. Org24598 profoundly reduced ethanol intake and the effect remained throughout both treatment periods. Acamprosate promptly reduced ethanol intake, but on the third day tolerance developed to this effect and acamprosate failed to influence alcohol consumption during the second test period. Neither Org24598 nor acamprosate reduced water intake. Following the drinking study, the Org24598 group displayed higher basal accumbal dopamine levels compared with acamprosate and vehicle groups. Both Org24598 and acamprosate reduced the ethanol-induced dopamine response in n. accumbens. The study demonstrates a robust anti-alcohol intake effect of the GlyT-1 inhibitor Org24598, supporting the new concept that GlyT-1 inhibition reduces ethanol consumption. GlyT-1 inhibition may represent a new treatment principle for alcoholism that is superior to acamprosate., (© 2011 The Authors, Addiction Biology © 2011 Society for the Study of Addiction.)

Brainstem noradrenergic neurons innervate the mesocorticolimbic reward pathway both directly and indirectly, with norepinephrine facilitating dopamine (DA) neurotransmission via α1-adrenergic receptors (α1ARs). Although α1AR signaling in the prefrontal cortex (PFC) promotes mesolimbic transmission and drug-induced behaviors, the potential contribution of α1ARs in other parts of the pathway, such as the ventral tegmental area (VTA) and nucleus accumbens (NAc), has not been investigated before. We found that local blockade of α1ARs in the medial NAc shell, but not the VTA, attenuates cocaine- and morphine-induced locomotion. To determine the neuronal substrates that could mediate these effects, we analyzed the cellular, subcellular, and subsynaptic localization of α1ARs and characterized the chemical phenotypes of α1AR-containing elements within the mesocorticolimbic system using single and double immunocytochemical methods at the electron microscopic (EM) level. We found that α1ARs are found mainly extra-synaptically in axons and axon terminals in the NAc and are enriched in glutamatergic and dopaminergic elements. α1ARs are also abundant in glutamatergic terminals in the PFC, and in GABA-positive terminals in the VTA. In line with these observations, microdialysis experiments revealed that local blockade of α1ARs attenuated the increase in extracellular DA in the medial NAc shell following administration of cocaine. These data indicate that local α1ARs control DA transmission in the medial NAc shell and behavioral responses to drugs of abuse.

The role of the amygdala in the mediation of fear and anxiety has been extensively investigated. However, how the amygdala functions during the organization of the anxiety-like behaviors generated in the elevated plus maze (EPM) is still under investigation. The basolateral (BLA) and the central (CeA) nuclei are the main input and output stations of the amygdala. In the present study, we ethopharmacologically analyzed the behavior of rats subjected to the EPM and the tissue content of the monoamines dopamine (DA) and serotonin (5-HT) and their metabolites in the nucleus accumbens (NAc), dorsal hippocampus (DH), and dorsal striatum (DS) of animals injected with saline or midazolam (20 and 30 nmol/0.2 µL) into the BLA or CeA. Injections of midazolam into the CeA, but not BLA, caused clear anxiolytic-like effects in the EPM. These treatments did not cause significant changes in 5-HT or DA contents in the NAc, DH, or DS of animals tested in the EPM. The data suggest that the anxiolytic-like effects of midazolam in the EPM also appear to rely on GABA-benzodiazepine mechanisms in the CeA, but not BLA, and do not appear to depend on 5-HT and DA mechanisms prevalent in limbic structures.

The frequent coexistence of depression in epileptic patients raises the issue of simultaneous use of antidepressants along with antiepileptic drugs in the management of such cases. However, it is necessary to evaluate the safety of these antiepileptic/antidepressant drug combinations. The present study investigates the effect of the antidepressant paroxetine (a selective serotonin reuptake inhibitor) administered alone or in combination with the antiepileptic drug sodium valproate on chemoconvulsions induced by picrotoxin (PTX). Seizure score was recorded in vivo, and the levels of thiobarbituric acid-reactive substances and gamma aminobutyric acid (GABA) were measured in the nucleus accumbens of the tested groups of mice. The results show enhancement of seizure severity with significant reduction in GABA levels upon PTX treatment that were reversed by its combination with sodium valproate. On the other hand, paroxetine administered in combination with sodium valproate provided significant protection against PTX-induced convulsions as well as a significant increase in GABA levels in selected brain areas. These results favor their application in management of epilepsy-depression comorbidities.

Postpartum rats that had been previously raised in an artificial rearing (AR) apparatus, without their mothers or siblings during the preweaning period, show altered maternal responses towards their own offspring in adulthood. In mother-reared (MR) rats, nucleus accumbens (NAC) dopamine (DA) responses to pups evoke a robust sustained rise during the postpartum period and following treatment with estrogen/progesterone parturient-like hormones (Afonso et al., 2009). These MR females had siblings that received AR rearing with varying amounts of preweaning tactile stimulation (ARmin; ARmax). The present study examined NACshell DA responses to pup and food stimuli in these AR rats, and statistically compared them to their MR siblings. Microdialysis samples were collected from adult (90 days postnatal) AR females in different parity states (cycling vs. postpartum, Exp. 1), or after ovariectomy with different hormone treatments (sham vs. hormone, Exp. 2. After basal sample collection, pup and then food stimuli were individually presented to the females in the dialysis chamber. As with their MR siblings, basal DA concentrations were lower and pup-evoked DA responses greater in hormonally-primed AR females than in non-primed AR controls. Compared to their postpartum MR sisters (Exp. 1), AR rats had increased basal DA levels, reduced pup related DA elevations, and disrupted maternal behavior. The postpartum AR impairment in pup-evoked DA was reversed by additional pre-weaning tactile stimulation. Exogenous hormones (Exp. 2) eliminated AR impairments on pup-evoked DA responses. Although MR and AR siblings had comparable DA responses to food stimuli, upon reanalyzing MR data it was found that only postpartum dams had DA responses to pups greater than to food. These data suggest that that the hormonally induced suppression of basal DA levels may reflect saliency of pups which was greater in MR than in AR dams. Preweaning tactile stimulation could partially reverse these effects only in naturally cycling or parturient animals., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Attention-deficit hyperactivity disorder (ADHD) is typically characterized as a disorder of inattention and hyperactivity/impulsivity but there is increasing evidence of deficits in motivation. Using positron emission tomography (PET), we showed decreased function in the brain dopamine reward pathway in adults with ADHD, which, we hypothesized, could underlie the motivation deficits in this disorder. To evaluate this hypothesis, we performed secondary analyses to assess the correlation between the PET measures of dopamine D2/D3 receptor and dopamine transporter availability (obtained with [(11)C]raclopride and [(11)C]cocaine, respectively) in the dopamine reward pathway (midbrain and nucleus accumbens) and a surrogate measure of trait motivation (assessed using the Achievement scale on the Multidimensional Personality Questionnaire or MPQ) in 45 ADHD participants and 41 controls. The Achievement scale was lower in ADHD participants than in controls (11±5 vs 14±3, P<0.001) and was significantly correlated with D2/D3 receptors (accumbens: r=0.39, P<0.008; midbrain: r=0.41, P<0.005) and transporters (accumbens: r=0.35, P<0.02) in ADHD participants, but not in controls. ADHD participants also had lower values in the Constraint factor and higher values in the Negative Emotionality factor of the MPQ but did not differ in the Positive Emotionality factor-and none of these were correlated with the dopamine measures. In ADHD participants, scores in the Achievement scale were also negatively correlated with symptoms of inattention (CAARS A, E and SWAN I). These findings provide evidence that disruption of the dopamine reward pathway is associated with motivation deficits in ADHD adults, which may contribute to attention deficits and supports the use of therapeutic interventions to enhance motivation in ADHD.

A sensitive assay method was developed for a parallel, rapid and precise determination of dopamine and its metabolites, homovanillic acid, 3-methoxytyramine and 3,4-dihydroxyphenylacetic acid, from brain microdialysates. The method consisted of a pre-treatment step, freeze-drying (lyophilization), to concentrate dopamine and its metabolites from the microdialysates, and a detection step using liquid chromatography combined with electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). In particular, the reaction monitoring mode was selected for its extremely high degree of selectivity and the stable-isotope-dilution assay for its high precision of quantification. The developed method was characterized by the following parameters: the precision of the developed method was determined as ≥88.6% for dopamine, ≥89.9% for homovanillic acid, ≥86.1% for 3-methoxytyramine and ≥88.1% for 3,4-dihydroxyphenylacetic acid; the mean accuracy was determined as ≥88.2% for dopamine, ≥88.3% for homovanillic acid, ≥85.9% for 3-methoxytyramine and ≥88.6% for 3,4-dihydroxyphenylacetic acid. The developed method was compared to (1) other combinations of pre-treatment methods (solid phase extraction and nitrogen stripping) with LC-MS and (2) another detection method, liquid chromatography, with electrochemical detection. The novel developed method using combination of lyophilization with LC-ESI-MS/MS was tested on real samples obtained from the nucleus accumbens of rat pups after an acute methamphetamine administration. It was proven that the developed assay could be applied to both a simultaneous analysis of all four substrates (dopamine, homovanillic acid, 3-methoxytyramine and 3,4-dihydroxyphenylacetic acid) in microdialysis samples acquired from the rat brain and the monitoring of their slight concentration changes on a picogram level over time following methamphetamine stimulus., (Copyright © 2011 Elsevier B.V. All rights reserved.)

High caloric intake during early postnatal development can have long term consequences for the offspring. We previously reported that the adult offspring of dams fed a high-fat diet during the last week of gestation and throughout lactation display blunted locomotor response to amphetamine (AMP) and reduced sensitization to the drug compared to offspring of control diet dams. Here, we report that the subsensitivity of high-fat offspring to AMP's locomotor stimulant action reflects, at least in part, altered regulation of nucleus accumbens (NAc) dopamine (DA) transmission. When compared to controls, the DA response of high-fat animals to AMP, as measured with microdialysis, was attenuated in the NAc, but unaffected in the prefrontal cortex (PFC). A relatively higher activity of NAc synaptosomal DA transporter sites without changes in vesicular monoamine transporter (VMAT) uptake capacity was also observed in high-fat offspring. Moreover, ventral tegmental area (VTA) D(2) receptor mRNA levels were decreased in high-fat offspring, suggesting a reduction in DA release-regulating D(2) autoreceptors in terminal regions such as the NAc. The magnitude of locomotor response to D(2/3) receptor activation (with quinpirole) was greater in high-fat than in control animals despite having comparable postsynaptic D(2) mRNA levels in the NAc. Finally, while operant responding for a sugar-enriched food reward did not differ between diet groups, high-fat offspring displayed increased operant responding for a fat-enriched reward compared to controls. These findings add to mounting evidence that early life exposure to elevated dietary maternal fat can lead to long lasting changes in DA-mediated behavioral responses to stimulant drugs and fat-enriched foods., (Copyright © 2011 IBRO. Published by Elsevier Ltd. All rights reserved.)