Anushree N. Karkhanis

Chronic intermittent ethanol exposure reduces presynaptic dopamine neurotransmission in the mouse nucleus accumbens

BACKGROUND Increasing evidence suggests that chronic ethanol exposure decreases dopamine (DA) neurotransmission in the nucleus accumbens (NAc), contributing to a hypodopaminergic state during withdrawal. However, few studies have investigated adaptations in presynaptic DA terminals after chronic intermittent ethanol (CIE) exposure. In monkeys and rats, chronic ethanol exposure paradigms have been shown to increase DA uptake and D2 autoreceptor sensitivity. METHODS The current study examined the effects of ethanol on DA terminals in CIE exposed mice during two time-points after the cessation of CIE exposure. DA release and uptake were measured using fast scan cyclic voltammetry in NAc core slices from C57BL/6J mice, 0h and 72h following three weekly cycles (4 days of 16h ethanol vapor/8h room air/day+3 days withdrawal) of CIE vapor exposure. RESULTS Current results showed that DA release was reduced, uptake rates were increased, and inhibitory D2-type autoreceptor activity was augmented following CIE exposure in mice. CONCLUSIONS Overall, these CIE-induced adaptations in the accumbal DA system reduce DA signaling and therefore reveal several potential mechanisms contributing to a functional hypodopaminergic state during alcohol withdrawal.

Supersensitive Kappa Opioid Receptors Promotes Ethanol Withdrawal-Related Behaviors and Reduce Dopamine Signaling in the Nucleus Accumbens

Background: Chronic ethanol exposure reduces dopamine transmission in the nucleus accumbens, which may contribute to the negative affective symptoms associated with ethanol withdrawal. Kappa opioid receptors have been implicated in withdrawal-induced excessive drinking and anxiety-like behaviors and are known to inhibit dopamine release in the nucleus accumbens. The effects of chronic ethanol exposure on kappa opioid receptor-mediated changes in dopamine transmission at the level of the dopamine terminal and withdrawal-related behaviors were examined. Methods: Five weeks of chronic intermittent ethanol exposure in male C57BL/6 mice were used to examine the role of kappa opioid receptors in chronic ethanol-induced increases in ethanol intake and marble burying, a measure of anxiety/compulsive-like behavior. Drinking and marble burying were evaluated before and after chronic intermittent ethanol exposure, with and without kappa opioid receptor blockade by nor-binaltorphimine (10mg/kg i.p.). Functional alterations in kappa opioid receptors were assessed using fast scan cyclic voltammetry in brain slices containing the nucleus accumbens. Results: Chronic intermittent ethanol-exposed mice showed increased ethanol drinking and marble burying compared with controls, which was attenuated with kappa opioid receptor blockade. Chronic intermittent ethanol-induced increases in behavior were replicated with kappa opioid receptor activation in naïve mice. Fast scan cyclic voltammetry revealed that chronic intermittent ethanol reduced accumbal dopamine release and increased uptake rates, promoting a hypodopaminergic state of this region. Kappa opioid receptor activation with U50,488H concentration-dependently decreased dopamine release in both groups; however, this effect was greater in chronic intermittent ethanol-treated mice, indicating kappa opioid receptor supersensitivity in this group. Conclusions: These data suggest that the chronic intermittent ethanol-induced increase in ethanol intake and anxiety/compulsive-like behaviors may be driven by greater kappa opioid receptor sensitivity and a hypodopaminergic state of the nucleus accumbens.

Early-Life Social Isolation Stress Increases Kappa Opioid Receptor Responsiveness and Downregulates the Dopamine System.

Chronic early-life stress increases vulnerability to alcoholism and anxiety disorders during adulthood. Similarly, rats reared in social isolation (SI) during adolescence exhibit augmented ethanol intake and anxiety-like behaviors compared with group housed (GH) rats. Prior studies suggest that disruption of dopamine (DA) signaling contributes to SI-associated behaviors, although the mechanisms underlying these alterations are not fully understood. Kappa opioid receptors (KORs) have an important role in regulating mesolimbic DA signaling, and other kinds of stressors have been shown to augment KOR function. Therefore, we tested the hypothesis that SI-induced increases in KOR function contribute to the dysregulation of NAc DA and the escalation in ethanol intake associated with SI. Our ex vivo voltammetry experiments showed that the inhibitory effects of the kappa agonist U50,488 on DA release were significantly enhanced in the NAc core and shell of SI rats. Dynorphin levels in NAc tissue were observed to be lower in SI rats. Microdialysis in freely moving rats revealed that SI was also associated with reduced baseline DA levels, and pretreatment with the KOR antagonist nor-binaltorphimine (nor-BNI) increased DA levels selectively in SI subjects. Acute ethanol elevated DA in SI and GH rats and nor-BNI pretreatment augmented this effect in SI subjects, while having no effect on ethanol-stimulated DA release in GH rats. Together, these data suggest that KORs may have increased responsiveness following SI, which could lead to hypodopaminergia and contribute to an increased drive to consume ethanol. Indeed, SI rats exhibited greater ethanol intake and preference and KOR blockade selectively attenuated ethanol intake in SI rats. Collectively, the findings that nor-BNI reversed SI-mediated hypodopaminergic state and escalated ethanol intake suggest that KOR antagonists may represent a promising therapeutic strategy for the treatment of alcohol use disorders, particularly in cases linked to chronic early-life stress.

Adolescent social isolation as a model of heightened vulnerability to comorbid alcoholism and anxiety disorders

Individuals diagnosed with anxiety-related illnesses are at increased risk of developing alcoholism, exhibit a telescoped progression of this disease and fare worse in recovery, relative to alcoholics that do not suffer from a comorbid anxiety disorder. Similarly, preclinical evidence supports the notion that stress and anxiety represent major risk factors for the development of alcohol use disorder (AUD). Despite the importance of understanding the link between anxiety and alcoholism, much remains unknown about the neurobiological substrates underlying this relationship. One stumbling block has been the lack of animal models that reliably reproduce the spectrum of behaviors associated with increased vulnerability to these diseases. Here, we review the literature that has examined the behavioral and neurobiological outcomes of a simple rodent adolescent social isolation procedure and discuss its validity as a model of vulnerability to comorbid anxiety disorders and alcoholism. Recent studies have provided strong evidence that adolescent social isolation of male rats leads to the expression of a variety of behaviors linked with increased vulnerability to anxiety and/or AUD, including deficits in sensory gating and fear extinction, and increases in anxiety measures and ethanol drinking. Neurobiological studies are beginning to identify mesolimbic adaptations that may contribute to the behavioral phenotype engendered by this model. Some of these changes include increased excitability of ventral tegmental area dopamine neurons and pyramidal cells in the basolateral amygdala and significant alterations in baseline and stimulated catecholamine signaling. A growing body of evidence suggests that adolescent social isolation may represent a reliable rodent model of heightened vulnerability to anxiety disorders and alcoholism in male rats. These studies provide initial support for the face, construct, and predictive validity of this model and highlight its utility in identifying neurobiological adaptations associated with increased risk of developing these disorders.

Social isolation rearing increases dopamine uptake and psychostimulant potency in the striatum.

Social isolation rearing (SI) is a model of early life stress that results in neurobiological alterations leading to increased anxiety-like behaviors. These animals also exhibit an increased propensity to administer psychostimulants, such as cocaine; however, the mechanisms governing this increased addiction vulnerability remain to be elucidated. Long-term stressors have been shown to produce important alterations in nucleus accumbens core (NAc) function. The NAc regulates motivated and goal-directed behaviors, and individual differences in NAc function have been shown to be predictive of addiction vulnerability. Rats were reared in group (GH; 4/cage) or SI (1/cage) conditions from weaning (PD 28) into early adulthood (PD 77) and dopamine release was assessed using voltammetry in brain slices containing the NAc and dorsomedial striatum. SI rats exhibited enhanced dopamine release and uptake in both regions compared to GH rats. In regard to psychostimulant effects directly at the dopamine transporter (DAT), methylphenidate and amphetamine, but not cocaine, inhibited uptake more in SI than GH rats. The increased potencies were positively correlated with uptake rates, suggesting that increased potencies of amphetamine-like compounds are due to changes in DAT function. Cocaines effects on uptake were similar between rearing conditions, however, cocaine enhanced evoked dopamine release greater in SI than GH rats, suggesting that the enhanced cocaine reinforcement in SI animals involves a DAT independent mechanism. Together, the results provide the first evidence that greater psychostimulant effects in SI compared to GH rats are due to effects on dopamine terminals related to uptake dependent and independent mechanisms.

Chronic social isolation during adolescence augments catecholamine response to acute ethanol in the basolateral amygdala

Adolescent social isolation (SI) results in numerous behavioral alterations associated with increased risk of alcoholism. Notably, many of these changes involve the basolateral amygdala (BLA), including increased alcohol seeking. The BLA sends a strong glutamatergic projection to the nucleus accumbens and activation of this pathway potentiates reward‐seeking behavior. Dopamine (DA) and norepinephrine (NE) exert powerful excitatory and inhibitory effects on BLA activity and chronic stress can disrupt the excitation‐inhibition balance maintained by these catecholamines. Notably, the impact of SI on BLA DA and NE neurotransmission is unknown. Thus the aim of this study was to characterize SI‐mediated catecholamine alterations in the BLA. Male Long Evans rats were housed in groups of four (GH) or in SI for 6 weeks during adolescence. DA and NE transporter levels were then measured using Western blot hybridization and baseline and ethanol‐stimulated DA and NE levels were quantified using microdialysis. DA transporter levels were increased and baseline DA levels were decreased in SI compared to GH rats. SI also increased DA responses to an acute ethanol (2 g kg−1) challenge. While no group differences were noted in NE transporter or baseline NE levels, acute ethanol (2 g kg−1) only significantly increased NE levels in SI animals. Collectively, these SI‐dependent changes in BLA catecholamine signaling may lead to an increase in BLA excitability and a strengthening of the glutamatergic projection between the BLA and NAc. Such changes may promote the elevated ethanol drinking behavior observed in rats subjected to chronic adolescent stress. Synapse 69:385–395, 2015.

Distinct Effects of Nalmefene on Dopamine Uptake Rates and Kappa Opioid Receptor Activity in the Nucleus Accumbens Following Chronic Intermittent Ethanol Exposure

The development of pharmacotherapeutics that reduce relapse to alcohol drinking in patients with alcohol dependence is of considerable research interest. Preclinical data support a role for nucleus accumbens (NAc) κ opioid receptors (KOR) in chronic intermittent ethanol (CIE) exposure-induced increases in ethanol intake. Nalmefene, a high-affinity KOR partial agonist, reduces drinking in at-risk patients and relapse drinking in rodents, potentially due to its effects on NAc KORs. However, the effects of nalmefene on accumbal dopamine transmission and KOR function are poorly understood. We investigated the effects of nalmefene on dopamine transmission and KORs using fast scan cyclic voltammetry in NAc brain slices from male C57BL/6J mice following five weeks of CIE or air exposure. Nalmefene concentration-dependently reduced dopamine release similarly in air and CIE groups, suggesting that dynorphin tone may not be present in brain slices. Further, nalmefene attenuated dopamine uptake rates to a greater extent in brain slices from CIE-exposed mice, suggesting that dopamine transporter-KOR interactions may be fundamentally altered following CIE. Additionally, nalmefene reversed the dopamine-decreasing effects of a maximal concentration of a KOR agonist selectively in brain slices of CIE-exposed mice. It is possible that nalmefene may attenuate withdrawal-induced increases in ethanol consumption by modulation of dopamine transmission through KORs.

The individual and combined effects of phenmetrazine and mgluR2/3 agonist LY379268 on the motivation to self-administer cocaine

BACKGROUND The US Food and Drug Administration has not approved a treatment for cocaine addiction, possibly due in part to the fact that repeated cocaine use results in dysregulation of multiple neurotransmitter systems, including glutamate and dopamine, and an emergence of increased negative affective states and heightening motivation to take cocaine despite negative consequences. We used a combination therapy approach to assess whether modulation of both glutamate and dopamine transmission would reduce the motivation to self- administer cocaine compared to modulation of either system alone. METHODS The metabotropic glutamate 2/3 receptor agonist, LY379268, and the monoamine releaser, phenmetrazine, were used to assess their individual and combined ability to decrease the reinforcing efficacy of cocaine because they modulate glutamate and dopamine levels, respectively. Cocaine breakpoints and cocaine intake was assessed, using a progressive ratio schedule, at baseline in three groups based on dose of cocaine (0.19, 0.38, 0.75mg/kg/infusion), and following LY379268 (0.03 or 0.30mg/kg; i.p.), phenmetrazine (25mg/kg/day; osmotic minipump), and a combination of the two drugs. RESULTS LY379268 and phenmetrazine alone reduced breakpoints for all doses of cocaine. The combination of the two drugs showed a concerted effect in reducing breakpoints for all doses of cocaine, with the lowest dose of cocaine reduced by as much as 70%. CONCLUSIONS These data support combination therapy of dopamine and glutamate systems as an effective means to reduce the motivation to take cocaine since a combination of drugs can address neurobiological dysfunction in multiple neurotransmitter systems compared to therapies using single drugs.

Cross-Species Alterations in Synaptic Dopamine Regulation After Chronic Alcohol Exposure

Alcohol use disorders are a leading public health concern, engendering enormous costs in terms of both economic loss and human suffering. These disorders are characterized by compulsive and excessive alcohol use, as well as negative affect and alcohol craving during abstinence. Extensive research has implicated the dopamine system in both the acute pharmacological effects of alcohol and the symptomology of alcohol use disorders that develop after extended alcohol use. Preclinical research has shed light on many mechanisms by which chronic alcohol exposure dysregulates the dopamine system. However, many of the findings are inconsistent across experimental parameters such as alcohol exposure length, route of administration, and model organism. We propose that the dopaminergic alterations driving the core symptomology of alcohol use disorders are likely to be relatively stable across experimental settings. Recent work has been aimed at using multiple model organisms (mouse, rat, monkey) across various alcohol exposure procedures to search for commonalities. Here, we review recent advances in our understanding of the effects of chronic alcohol use on the dopamine system by highlighting findings that are consistent across experimental setting and species.