Symposium 27 Tuesday Sept. 25th 1.30 pm–3.00 pm; Room: Lecture Hall 3
Young Investigator Symposium: Chairpersons: Colombo G (Belgium), Kienast T (Germany)
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Presentation S27-1
Cellular mechanisms of ethanol and neuropeptide action in the amygdala: potential treatment for alcoholism
Roberto M (USA)
Aims. Alcohol dependence is a chronic relapsing disorder characterized by compulsive use of alcohol, loss of control over intake, and the presence of withdrawal syndrome, including both motivational and physical symptoms. Behavioral evidence indicates that the central nucleus of the amygdala (CeA) plays an important role in regulating voluntary ethanol intake and is involved in anxiety associated with ethanol dependence.
Our research is aimed to uncover the physiological mechanisms underlying ethanol effects on CeA neuronal communication and the involvement of neuropeptides such as corticotrophin-releasing factor (CRF) and nociceptin, as well as to reveal the neuroadaptations associated with ethanol-dependence.
Methods. We used the in vitro CeA slice preparation and in vivo brain microdialysis to study the interaction of ethanol and neuropeptides with the GABAergic system, in rats subjected to chronic ethanol treatment via ethanol inhalation.
Results. Using an electrophysiological approach, we found that acute ethanol increases inhibitory transmission in CeA neurons from both naïve and ethanol-dependent rats, in part by enhancing the vesicular release of Gamma-aminobutyric acid (GABA) in CeA. We confirmed these in vitro findings with in vivo microdialysis studies showing that both acute and chronic ethanol increase GABA release in the CeA. In addition, we found a critical mimicry between ethanol and CRF effects on GABAergic transmission that markedly adapts during the development of ethanol dependence. Nociceptin, however, decreased GABA release and opposed both ethanol and CRF effects. Interestingly, chronic ethanol enhanced the sensitivity of the GABAergic system in CeA to these two neuropeptides. Based on this evidence, we hypothesize that nociceptin functions as a ‘brake’ to limit the enhancement of inhibitory transmission in the neural circuitry involved in the reinforcing actions of ethanol.
Current studies are focused on understanding the cellular adaptations induced by chronic alcohol and how these changes contribute to the development of ethanol dependence and protracted abstinence. The discovery and characterization of such neuroadaptations will be useful toward developing new therapeutic agents to alleviate alcohol dependence. Therefore, we have also investigated the effects of gabapentin, a structural analogue of GABA that has anticonvulsant properties, on GABAergic transmission in CeA slices, and on ethanol intake and anxiety measures, using an animal model of ethanol dependence. Our data show that gabapentin increased the amplitudes of GABA responses in CeA neurons from non-dependent rats, but decreased their amplitudes in CeA of ethanol-dependent rats. Gabapentin reduced ethanol intake in dependent, but not in non-dependent rats and reversed a measure of delayed anxiety produced by ethanol. Because gabapentin opposes ethanol intake and ethanol actions in CeA neurons from ethanol-dependent rats, it may act therapeutically by controlling the inhibitory GABAergic system.
Conclusions. Thus, we found ethanol-dependence induced profound neuroadaptations in the CeA GABAergic system and in its sensitivity to CRF and nociceptin. Our findings also show that both nociceptin and gabapentin reverse cellular measures of ethanol dependence and suggest that these drugs represent potential medication for treatment of alcoholism.
Presentation S27-2
Hypersecretory haplotype of interleukin-1 gene cluster increases the risk of alcoholic liver cirrhosis
Petrasek J, Reznakova S, Hubacek JA, Stickel F, Sperl J, Trunecka P, Berg T, Spicak J, Jirsa M (Czech Republic)
Aims. Interleukin 1-β (IL-β) is an important pro-inflammatory cytokine. The objective of the study was to test the hypothesis that the IL-β promoter polymorphism –31C/T known to increase the expression of IL-1 and #946; and the penta-allelic 86-bp tandem repeat in intron 2 of IL-1 receptor antagonist (IL-1RN) known to increase the production of IL-β are associated with susceptibility to alcoholic liver cirrhosis (ALC).
Methods. Allele frequencies for both mentioned polymorphisms were determined in 198 heavy drinkers with clinically proven alcoholic cirrhosis and 246 non-abstaining age and sex matched healthy controls. The principle positive finding was further validated in an independent cohort of 173 patients with ALC.
Results. The haplotype [IL-1RN*2/*2; IL-1B –31T+] was associated with an increased risk of ALC in both the primary (odds ratio 4.9, 95% CI 1.4–17.8, P = 0.016) and the second confirmatory cohort (odds ratio 8.0, 95% CI 2.2–28.4, P = 0.001). The population atributable fraction that estimates the proportion of ALC in the study population that is attributable to the haplotype, was 4.9% (95% CI 1.3–8.6) in the pilot cohort and 7.5% (95% CI 3.1–12.0) in the confirmatory cohort.
Conclusions. Our observation is in line with the previous studies indicating that synergistic action of the IL-β -31T and IL-1RN*2 polymorphisms are responsible for IL-β high secretory phenotype. Such phenotype promotes inflammation and fibrosis and explains the molecular background of the increased risk of ALC in homozygotes for the IL-1RN*2 allele that are at least heterozygous for the IL-1 and #946; -31T allele.
Presentation S27-3
Beer but not ethanol stimulates amylase secretion in differentiated rat pancreatic acinar cell line AR4-2J and freshly isolated acinar cells
Gerloff A, Singer MV, Feick P (Germany)
Background. Ethanol is generally consumed in form of alcoholic beverages which contain numerous non-alcoholic constituents. Recent studies showed different pathophysiological effects of alcoholic beverages as compared to pure alcohol solutions. To differentiate the effects of beer and comparable ethanol solutions on protein secretion and signal transduction in the rat pancreatic acinar cell line AR4-2J.
Methods. AR4-2J cells were differentiated by dexamethasone treatment for 72 h and dispersed pancreatic acini were prepared from CD-1 mice using collagenase type V. After incubation of cells with beer (1–10%) or appropriate ethanol concentrations for up to 60 minutes, protein secretion using amylase activity assay and calcium mobilization were measured.
Results. Incubation of AR4-2J cells with beer caused a dose-dependent stimulation of basal amylase secretion and had an additive effect on amylase release maximally induced by CCK. In contrast, ethanol showed no effect on amylase release. LDH measurement after long-term treatment (24 and 48 h) of AR4-2J cells showed a cytoprotective effect of beer as compared to control, suggesting that the increase of amylase secretion was not due to cell membrane damage. Using selective antagonists we showed that beer-induced amylase release was mediated by PLC and the activation of IP3-Receptor whereas the inhibition of PKC, adenylate cyclase and PKA had no significant effect. Investigations using the fluorescent Ca2+ indicator Fura-2/AM as well as the calcium chelator BAPTA/AM revealed that beer induced an increase of cytosolic free calcium concentration. Beer showed also a stimulatory effect on protein secretion of primary acinar cells.
Conclusions. Our data demonstrate that beer dose-dependently increases amylase release which may be mediated by activation of PLC and subsequent calcium release from internal stores. Since ethanol is not responsible for this effect our findings clearly indicate that non-alcoholic constituents of alcoholic beverages have to be considered in the study of ethanol-induced metabolic, pathological and functional changes.
Presentation S27-4
Decision-making deficits in addiction
Kahnt T, Beck A, Park S-Y, Kienast T, Heinz A, Wrase J (Germany)
Successful decision-making behavior can be defined as the ability to learn which actions maximize reward, and flexibly adjust decision strategies to changing environmental conditions. During the last decade it has been proposed that decision-making deficits are one of the hallmarks of addiction. A lot of studies consistently demonstrate decreased performance of populations with substance abuse disorder relative to control groups on decision-making paradigms such as the Iowa Gambling Task (IGT). These subjects perform similar to patients with lesions of the ventromedial prefrontal cortex (VMPFC) and orbitofrontal cortex (OFC), regions which are essential for successful performance in decision-making tasks. These regions are involved in processing of reward value, inhibitory decision-making processes and assessment of the future consequences of the individual's own action. Thus, in line with neurobiological addiction research, it has been proposed that functional deficits in prefrontal areas may constitute a characteristic of addiction, leading to impaired decision-making behavior. Consistent with this hypothesis, neuroimaging studies report hypoactivty of prefrontal and orbitofrontal areas during decision-making in detoxified addicts relative to controls. Here we report preliminary data from an experiment using a probabilistic reversal learning task and functional magnetic resonance imaging (fMRI) in detoxified alcoholic patients. This task is designed to study both, the learning of a decision strategy by operant conditioning and the flexible adjustment of decision strategies to changing environmental conditions.
These findings might explain the well known addiction process of continued drug use despite harmful consequences.