Manipulation of radixin phosphorylation in the nucleus accumbens core modulates risky choice behavior.

Ezrin-Radixin-Moesin (ERM) proteins are actin-binding proteins that contribute to morphological changes in dendritic spines. Despite their significant role in regulating spine structure, the role of ERM proteins in the nucleus accumbnes (NAcc) is not well known, especially in in the context of risk-reward decision-making. Here, we measured the relationship between synaptic excitation and inhibition (E/I ratio) from medium spiny neurons in the NAcc core obtained in the rat after a rat gambling task (rGT). Then, after surgery of a phosphomimetic pseudo-active mutant form of radixin (Rdx-T564D) in the NAcc core, we examined its role in synaptic plasticity and the accompanying risk-choice behavior in rGT. We found that basal E/I ratio in the NAcc core was higher in risk-averse rats than risk-seeking rats. However, it was significantly reduced in risk-averse rats similar to that in risk-seeking rats in the presence of Rdx-T564D in the NAcc core. Furthermore, the head sizes of spines were shifted in risk-averse rats expressing Rdx-T564D in the NAcc core, similar to those observed in risk-seeking rats. The effects of Rdx-T564D in risk-averse rats were again manifested as behavioral changes, with reduced selection of optimal choices and increased selection of disadvantageous ones. In this study, we demonstrated that manipulation of radixin phosphorylation status in the NAcc core can alter glutamatergic synaptic transmission and spine structure at this site, as well as risk choice behaviors in the rGT. These novel findings illustrate that radixin in the NAcc core plays a significant role in determining risk preference during the rGT.

Glycogen Synthase Kinase 3ß Is a Key Regulator in the Inhibitory Effects of Accumbal Cocaine- and Amphetamine-Regulated Transcript Peptide 55-102 on Amphetamine-Induced Locomotor Activity.

Microinjection of cocaine- and amphetamine-regulated transcript (CART) peptide 55-102 into the nucleus accumbens (NAcc) core significantly attenuates psychostimulant-induced locomotor activity. However, the molecular mechanism remains poorly understood. We examined the phosphorylation levels of Akt, glycogen synthase kinase 3ß (GSK3ß), and glutamate receptor 1 (GluA1) in NAcc core tissues obtained 60 min after microinjection of CART peptide 55-102 into this site, followed by systemic injection of amphetamine (AMPH). Phosphorylation levels of Akt at Thr308 and GSK3ß at Ser9 were decreased, while those of GluA1 at Ser845 were increased, by AMPH treatment. These effects returned to basal levels following treatment with CART peptide 55-102. Furthermore, the negative regulatory effects of the CART peptide on AMPH-induced changes in phosphorylation levels and locomotor activity were all abolished by pretreatment with the S9 peptide, an artificially synthesized indirect GSK3ß activator. These results suggest that the CART peptide 55-102 in the NAcc core plays a negative regulatory role in AMPH-induced locomotor activity by normalizing the changes in phosphorylation levels of Akt-GSK3ß, and subsequently GluA1 modified by AMPH at this site. The present findings are the first to reveal GSK3ß as a key regulator of the inhibitory role of the CART peptide in psychomotor stimulant-induced locomotor activity.

Differential transcriptome profile underlying risky choice in a rat gambling task.

Background and aims: Proper measurement of expected risk is important for making rational decisions, and maladaptive decision making may underlie various psychiatric disorders. However, differentially expressed genetic profiling involved in this process is still largely unknown. A rodent version of the gambling task (rGT) has been developed to measure decision-making by adopting the same principle of Iowa Gambling Task in humans. In the present study, we examined using next-generation sequencing (NGS) technique whether there are differences in gene expression profiles in the medial prefrontal cortex (mPFC) and the nucleus accumbens (NAc) when rats make different choices toward risk in rGT. Methods: Rats were trained in a touch screen chamber to learn the relationships between 4 different light signals on the window of the screen and accompanied reward outcomes or punishments set up with different magnitudes and probabilities. Once they showed a stabilized pattern of preference upon free choice, rats were classified into risk-averse or risk-seeking groups. After performing the rGT, rats were decapitated, the mPFC and the NAc was dissected out, and NGS was performed with the total RNA extracted. Results: We found that 477 and 36 genes were differentially expressed (approximately 75 and 83% out of them were downregulated) in the mPFC and the NAc, respectively, in risk-seeking compared to risk-averse rats. Among those, we suggested a few top ranked genes that may contribute to promoting risky choices. Discussion and conclusions: Our findings provide insights into transcriptional components underlying risky choices in rats.

Disruption of amphetamine sensitization by alteration of dendritic thin spines in the nucleus accumbens core.

Repeated injections of psychomotor stimulants like amphetamine (AMPH) to rodents can induce behavioral sensitization, which represents a long-lasting craving that is usually observed in human addicts. Behavioral sensitization is characteristically maintained for a long duration, accompanied by structural plasticity in some brain areas involved in reward circuitry. For example, it increased dendritic spine densities in the nucleus accumbens (NAcc), which is considered to reflect neurophysiological changes at this site, leading to addictive behaviors. The ezrin, radixin, and moesin (ERM) proteins regulate spine maturity by modifying their phosphorylation at the C-terminal region. We previously showed that ERM phosphorylation is reduced by AMPH in the NAcc core, suggesting that ERM-mediated spine changes at this site might be associated with AMPH sensitization. To test this hypothesis, we administered AMPH to rats according to a sensitization development schedule, with lentivirus encoding a phosphomimetic pseudo-active mutant of radixin (Rdx T564D) in the NAcc core, and examined dendritic spines at this site. We found that compared to acute AMPH, AMPH sensitization increased thin spine density with a similar ratio of filopodia-like to mature thin spines. However, with Rdx T564D, the density of thin spines increased, with augmented filopodia-like thin spines, resulting in no AMPH sensitization. These results indicate that Rdx T564D forces thin spines to immaturity and thereby inhibits AMPH sensitization, for which an increase in mature thin spines is normally necessary. These findings provide significant clues to our understanding of the role of dendritic spines in mediating the development of psychomotor stimulant addiction.

Decrease of glycogen synthase kinase 3ß phosphorylation in the rat nucleus accumbens shell is necessary for amphetamineinduced conditioned locomotor activity.

Phosphorylation levels of glycogen synthase kinase 3ß (GSK3ß) negatively correlated with psychomotor stimulant-induced locomotor activity. Locomotor sensitization induced by psychomotor stimulants was previously shown to selectively accompany the decrease of GSK3ß phosphorylation in the nucleus accumbens (NAcc) core, suggesting that intact GSK3ß activity in this region is necessary for psychomotor stimulants to produce locomotor sensitization. Similarly, GSK3ß in the NAcc was also implicated in mediating the conditioned effects formed by the associations of psychomotor stimulants. However, it remains undetermined whether GSK3ß plays a differential role in the two sub-regions (core and shell) of the NAcc in the expression of drug-conditioned behaviors. In the present study, we found that GSK3ß phosphorylation was significantly lower in the NAcc shell obtained from rats expressing amphetamine (AMPH)-induced conditioned locomotor activity. Further, we demonstrated that these effects were normalized by treatment with lithium chloride, a GSK3ß inhibitor. These results suggest that the behavior produced by AMPH itself and a conditioned behavior formed by associations with AMPH are differentially mediated by the two sub-regions of the NAcc.

Soft subdermal implant capable of wireless battery charging and programmable controls for applications in optogenetics.

Optogenetics is a powerful technique that allows target-specific spatiotemporal manipulation of neuronal activity for dissection of neural circuits and therapeutic interventions. Recent advances in wireless optogenetics technologies have enabled investigation of brain circuits in more natural conditions by releasing animals from tethered optical fibers. However, current wireless implants, which are largely based on battery-powered or battery-free designs, still limit the full potential of in vivo optogenetics in freely moving animals by requiring intermittent battery replacement or a special, bulky wireless power transfer system for continuous device operation, respectively. To address these limitations, here we present a wirelessly rechargeable, fully implantable, soft optoelectronic system that can be remotely and selectively controlled using a smartphone. Combining advantageous features of both battery-powered and battery-free designs, this device system enables seamless full implantation into animals, reliable ubiquitous operation, and intervention-free wireless charging, all of which are desired for chronic in vivo optogenetics. Successful demonstration of the unique capabilities of this device in freely behaving rats forecasts its broad and practical utilities in various neuroscience research and clinical applications.

Immunohistochemical detection of GluA1 subunit of AMPA receptor in the rat nucleus accumbens following cocaine exposure.

α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors are differentially regulated in the nucleus accumbens (NAcc) of the brain after cocaine exposure. However, these results are supported only by biochemical and electrophysiological methods, but have not been validated with immunohistochemistry. To overcome the restriction of antigen loss on the postsynaptic target molecules that occurs during perfusion-fixation, we adopted an immersion-fixation method that enabled us to immunohistochemically quantify the expression levels of the AMPA receptor GluA1 subunit in the NAcc. Interestingly, compared to saline exposure, cocaine significantly increased the immunofluorescence intensity of GluA1 in two sub-regions, the core and the shell, of the NAcc on withdrawal day 21 following cocaine exposure, which led to locomotor sensitization. Increases in GluA1 intensity were observed in both the extra-post synaptic density (PSD) and PSD areas in the two sub-regions of the NAcc. These results clearly indicate that AMPA receptor plasticity, as exemplified by GluA1, in the NAcc can be visually detected by immunohistochemistry and confocal imaging. These results expand our understanding of the molecular changes occurring in neuronal synapses by adding a new form of analysis to conventional biochemical and electrophysiological methods.

Ezrin-radixin-moesin proteins are regulated by Akt-GSK3ß signaling in the rat nucleus accumbens core.

The ezrin-radixin-moesin (ERM) proteins are a family of membrane-associated proteins known to play roles in cell-shape determination as well as in signaling pathways. We have previously shown that amphetamine decreases phosphorylation levels of these proteins in the nucleus accumbens (NAcc), an important neuronal substrate mediating rewarding effects of drugs of abuse. In the present study, we further examined what molecular pathways may be involved in this process. By direct microinjection of LY294002, a PI3 kinase inhibitor, or of S9 peptide, a proposed GSK3ß activator, into the NAcc core, we found that phosphorylation levels of ERM as well as of GSK3ß in this site are simultaneously decreased. These results indicate that ERM proteins are under the regulation of Akt-GSK3ß signaling pathway in the NAcc core. The present findings have a significant implication to a novel signal pathway possibly leading to structural plasticity in relation with drug addiction.

Impulsive Action and Impulsive Choice Are Differentially Expressed in Rats Depending on the Age at Exposure to a Gambling Task.

Impulsivity is considered an important feature associated with the development of numerous psychiatric disorders, including addictions. In the behavioral approach, impulsivity can be broadly divided into two distinct subtypes: impulsive action and choice. In the present study, we used a rodent version of the gambling task (rGT) to examine how impulsive action and impulsive choice are differentially influenced by difference in age at exposure (i.e., late adolescents/young adults vs. mature adults) to rGT. Rats were trained in a touch-screen chamber to learn the relationships between 4 light signals on the window of the screen and accompanying reward outcomes or punishments associated with different magnitudes and probabilities. Depending on their stabilized pattern of preference when allowed free choice, rats were categorized into risk-averse or risk-seeking group. While undergoing a series of experimental schemes, including extinction, re-acquisition, and acute cocaine injection, rats were re-tested for their premature response during inter-trial interval and choice preference toward the 4 different windows in rGT. Notably, rats exposed early, compared with those exposed late, to rGT showed increased impulsive action, particularly during re-acquisition period, in all sub-groups. In contrast, rats exposed late, compared with those exposed early, to rGT showed increased impulsive choice after acute cocaine injection, but these results were only obtained in a sub-group pre-categorized as high impulsive and risk-averse. These results suggest that different aspects of impulsivity can be differentially expressed during decision-making, and differentially influenced by the age at exposure to a gambling task.

Leptin in the nucleus accumbens blocks the increase of GluA1 phosphorylation induced by acute cocaine administration.

Leptin in the rat's nucleus accumbens (NAcc) core has previously shown to disrupt the effects of acute administration of cocaine on both locomotor activity and the phosphorylation levels of glycogen synthase kinase 3ß. In the present study, we further measured the phosphorylation levels of GluA1 after bilateral microinjections of leptin in this site followed by acute administration of cocaine. Interestingly, leptin in the NAcc core significantly blocks the increase of GluA1 phosphorylation levels at serine 845 induced by acute administration of cocaine. These effects were not observed for GluA1 phosphorylation levels at serine 831. We have also found additional evidence that glycogen synthase kinase 3ß may play a major role in mediating these processes.

Leptin in the nucleus accumbens core disrupts acute cocaine effects: Implications for GSK3ß connections.

An adipose-derived peptide hormone, leptin, has a regulatory role in reward-related behaviors produced by drugs of abuse. Although it is known that leptin modulates mesolimbic dopaminergic pathways, little is known about its direct role in the nucleus accumbens (NAcc). In the present study, we measured acute cocaine-induced locomotor activity in the rat and the phosphorylation levels of GSK3ß after bilateral microinjections of leptin into the NAcc core. Interestingly, leptin in the NAcc core significantly disrupts acute cocaine's effects on both locomotor activity and signaling molecules. In order to further confirm the role of GSK3ß in these processes, we microinjected S9 peptide, a small synthetic peptide acting as a competitive inhibitor against phosphorylation site of GSK3ß, followed by leptin co-microinjection, and found that leptin's effects on cocaine were all nullified. These results indicate that leptin in the NAcc core has a negative regulatory role in acute cocaine' effects, and suggest that GSK3ß may play a major role in mediating these processes.

Locomotor sensitization is expressed by ghrelin and D1 dopamine receptor agonist in the nucleus accumbens core in amphetamine pre-exposed rat.

Ghrelin modulates mesolimbic dopaminergic pathways in the brain in addition to its role in feeding. We investigated what roles ghrelin in the nucleus accumbens (NAcc) core may play in mediating locomotor activating effects of amphetamine (AMPH). First, when rats were administered with AMPH (1 mg/kg, i.p.) following a bilateral microinjection of ghrelin (0.1 or 0.5 µg/side) into the NAcc core, their locomotor activity was significantly enhanced, while these effects were blocked by co-microinjection of ghrelin receptor antagonist (0.5 µg/side) into this site. Second, we pre-exposed rats to saline or amphetamine (1 mg/kg, i.p.) every 2 to 3 days for a total of four times. After 2 weeks of drug-free withdrawal period, we examined the effect of saline, ghrelin (0.5 µg/side), D1 dopamine receptor agonist, SKF81297 (0.5 µg/side) or ghrelin (0.5 µg/side) + SKF81297 (0.5 µg/side) directly microinjected into the NAcc core on locomotor activity. When we measured rats' locomotor activity for 1 hour immediately following microinjections, only ghrelin + SKF81297 produces sensitized locomotor activity, while all others have no effects. These results suggest that ghrelin may have a distinct role in the NAcc core to provoke the sensitized locomotor activity induced by psychomotor stimulants, and further, it may produce these effects by interaction with D1 dopamine receptors.

Interaction between trait and housing condition produces differential decision-making toward risk choice in a rat gambling task.

Poor decision-making is a core problem in psychiatric disorders such as pathological gambling and substance abuse. Both trait and environmental factors are considerably important to affect decision-making. However, it has not yet been systematically shown how they interact to affect risk preference in animal models evaluating decision-making. Here, we trained rats, housed in pairs or in isolation, in a touch screen chamber to detect the association between four different light signals on the screen and accompanied reward and punishment outcomes arranged with different schedules. Then, the rats were allowed to freely choose from 4 different light signals. Once animals showed a stabilized pattern of preference (risk-averse or risk-seeking), they were injected with saline or cocaine (a single injection per day for 7 days) followed by 2 weeks of withdrawal. Then, their preference of choice was re-tested in the touch screen chamber while they were cocaine challenged. All rats significantly changed their preference toward more risky choices when they were exposed to and challenged with cocaine, except those in the risk-averse/isolated housing group. These results indicate that the pre-existing trait toward risk and the housing condition interact to affect the quality of decision-making, and cocaine may help to aggravate this process.

Cocaine inhibits leptin-induced increase of the cocaine- and amphetamine-regulated transcript peptide in the nucleus accumbens in rats.

Two well-known appetite-regulatory peptides, leptin and cocaine- and amphetamine-regulated transcript (CART), are known to be involved in the brain rewarding pathway. However, it is not yet known how they interact in the nucleus accumbens, an important region mediating the rewarding effects of drugs of abuse. Using the immunoassay method, we found that a microinjection of leptin into the nucleus accumbens core induces an immediate and transient increase of the CART peptide in this site, whereas these effects were inhibited by cocaine. These results expand the role of accumbal leptin to the regulation of the CART peptide and further suggest that possible interaction of these appetite-regulating peptides may be involved in cocaine-mediated rewarding effects.

Amphetamine dephosphorylates ERM proteins in the nucleus accumbens core and lithium attenuates its effects.

The ezrin-radixin-moesin (ERM) proteins have been implicated not only in cell-shape determination but also in cellular signaling pathway. We have previously shown that cocaine decreases phosphorylation levels of these proteins in the nucleus accumbens (NAcc), an important brain area mediating addictive behaviors. Here we further revealed that the phosphorylation levels of ERM were decreased in the NAcc core, but not in the shell, by a single injection of amphetamine (AMPH) (2 mg/kg, i.p.). When lithium (100 mg/kg, i.p.) was co-administered with AMPH, the decreases of phosphorylation levels for ERM by AMPH were recovered back to basal levels in the NAcc core. Together, these results suggest that psychomotor stimulants like AMPH regulate phosphorylation levels of ERM in the NAcc core and lithium-involved signaling pathway has a regulatory role in the opposite direction in this site.

Carbohydrate intake interacts with SNP276G>T polymorphism in the adiponectin gene to affect fasting blood glucose, HbA1C, and HDL cholesterol in Korean patients with type 2 diabetes.

BACKGROUND/OBJECTIVE: The SNP276G>T polymorphism in the adiponectin gene has been reported to be associated with type 2 diabetes and impaired glucose tolerance. The objective of this study was to examine whether SNP276G>T polymorphism influences the blood glucose levels in relation to dietary carbohydrate intake. SUBJECTS/METHODS: In an ongoing, prospective study, 673 patients with type 2 diabetes (339 men and 334 women, aged 40-85 years) were recruited from one of two diabetes clinics in Seoul, Korea. The levels of carbohydrate intake were categorized as <55%, 55%-65%, and >65% of total energy intake. RESULTS: Significant gene-nutrient interactions between SNP276G>T polymorphism and the level of carbohydrate intake were found, which modulated plasma fasting blood glucose (p=0.0277), HbA1C (p=0.0407), and high-density lipoprotein (HDL) cholesterol (p=0.0134) concentrations. The G allele was associated with higher fasting blood glucose only in subjects consuming a low-carbohydrate diet (<55% of energy). However, when carbohydrate intake was intermediate (55%-65%), carriers of the T allele had greater fasting blood glucose and HbA1C concentrations. When carbohydrate intake was high (>65%), carriers of the T allele had greater HDL cholesterol concentrations. This interaction was significant even when carbohydrate intake was considered a continuous variable (p=0.0200 for fasting blood glucose, p=0.0408 for HbA1C, and p=0.0254 for HDL cholesterol), suggesting a strong dose-response relation. CONCLUSIONS: Our data show that the effect of the SNP276G>T polymorphism on plasma fasting blood glucose, HbA1C, and HDL cholesterol concentrations depends on dietary carbohydrate intake.

Microinjection of ghrelin in the nucleus accumbens core enhances locomotor activity induced by cocaine.

Ghrelin has a regulatory role not only in the rewarding aspect for feeding but also in drug addiction by interaction with mesolimbic dopaminergic pathways in the brain. Previously it has been shown that systemic ghrelin enhances cocaine-induced hyper-locomotor activity. However, it has not been determined yet what effects ghrelin may produce on cocaine-induced locomotor behavior when microinjected into the nucleus accumbens core, where cocaine actually produces its locomotor activating effects. In the present study, when rats were administered cocaine (15 mg/kg, i.p.) following a bilateral microinjection of ghrelin (0.1 or 0.5 µg/side) into the NAcc core, their locomotor activity was significantly enhanced, while these effects were inhibited by co-microinjection of ghrelin receptor antagonist (0.5 µg/side) into this site. When ghrelin alone microinjected, basal locomotor activity was unchanged. These results expand previous similar findings obtained by systemic ghrelin to more specific neuronal site, the nucleus accumbens core, and further suggest that it may work in this site to positively contribute to the expression of locomotor sensitization.

Inhibition of PKMζ in the nucleus accumbens core blocks the expression of locomotor sensitization induced by amphetamine.

Behavioral sensitization is a form of drug-induced, long-lasting, non-associative memory. Protein kinase M zeta (PKMζ) has been shown to play a role in maintaining associative long-term memory in various brain regions. In the present study, rats were pre-exposed to either saline or amphetamine (AMPH) (1 mg/kg, i.p.). After 1 week of drug-free withdrawal period, they were challenged with AMPH (1 mg/kg, i.p.) following a bilateral microinjection of either saline or zeta inhibitory peptide (ZIP) (5 nmole/side) into the NAcc core. In the AMPH pre-exposed group, the ZIP microinjection blocked the enhanced increase of locomotion by AMPH challenge. These results suggest that PKMζ in the NAcc core plays a role in the expression of AMPH-induced locomotor sensitization, and further extend the role of PKMζ in long-term memory to non-associative form of drug-related memory.

Development and evaluation of a food frequency questionnaire for Vietnamese female immigrants in Korea: the Korean Genome and Epidemiology Study (KoGES).

The objectives of this study were to develop a food-frequency questionnaire (FFQ) for Vietnamese female immigrants in Korea and to evaluate the validity of the FFQ. A total of 80 food items were selected in developing the FFQ according to consumption frequency, the contribution of energy and other nutrients, and the cooking methods based on one-day 24 hour recall (24HR) from 918 Vietnamese female immigrants between November 2006 and November 2007. The FFQ was validated by comparison with 24HR of 425 Vietnamese female immigrants between November 2008 and August 2009. The absolute nutrient intake calculated from the FFQ was higher than that estimated by 24HR for most nutrients. The correlation coefficients between 24HR and FFQ ranged from 0.10 (vitamin C) - 0.36 (energy) for crude intake, 0.05 (vitamin E) - 0.32 (calcium) for per 1000 kcal, and 0.08 (zinc) - 0.34 (calcium) for energy-adjusted, respectively. More than 70% of subjects were classified into the same or adjacent agreement groups for nutrients other than fiber, sodium, vitamin A, vitamin C, and vitamin E, while less than 10% of subjects were classified into complete disagreement groups. We conclude that the FFQ appears to be an acceptable tool for estimating nutrient intake and dietary patterns of Vietnamese female immigrants in Korea. Future studies to validate the FFQ using various biomarkers or other dietary assessment methods are needed.

Hypoadiponectinemia is strongly associated with metabolic syndrome in Korean type 2 diabetes patients.

OBJECTIVE: To evaluate the relationship between serum adiponectin level, dietary intake, and metabolic syndrome (MetS) in type 2 diabetes mellitus (DM) patients, and to identify factors associated with serum adiponectin level. METHODS: A cross-sectional study was performed using 789 type 2 DM patients (406 men and 383 women) 40-80 years old. Subjects were classified into 3 groups on the basis of serum adiponectin level. General characteristics and anthropometric, hematologic, and dietary data were obtained for each subject. RESULTS: The prevalence of hypoadiponectinemia (<4.0 µg/mL) was 57.4% in men and 32.4% in women. Serum adiponectin level was negatively correlated with body mass index (BMI), waist circumference, body fat percentage, and serum concentrations of insulin and triglyceride, and was positively correlated with high-density lipoprotein (HDL)-cholesterol level. Even though the direct association of nutrient intake with serum adiponectin concentration was not strong, various contributing factors for hypoadiponectinemia were strongly correlated with micronutrient intake, such as calcium, iron, and niacin. Both sexes in the group with the lowest adiponectin concentration had a higher prevalence of MetS and MetS components than corresponding sexes in the group with the highest adiponectin concentration. CONCLUSIONS: Our findings show that hypoadiponectinemia is strongly associated with MetS in type 2 DM patients. Dietary intake may be indirectly associated with adiponectin levels through factors such as BMI, waist circumference, insulin, homeostasis model assessment of insulin resistance (HOMA-IR), triglyceride, HDL-cholesterol, and blood pressure. Therefore, our results suggest that manipulation of the level of adiponectin may prevent MetS and reduce the risk of cardiovascular disease in type 2 DM patients.