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P.6.b.004 Effect of amino acid derivatives and lipoic acid on lipid peroxidation and antioxidant systems indices in the rat brain during alcohol withdrawal
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P.6.b Addiction – Alcohol (basic)
P.6.b.003 Dopamine-related gene polymorphisms and alcohol withdrawal symptoms in patients with alcohol dependence B. Kee1 ° , Y.S. Lee1 , K.J. Min1 , K. Paik2 , D. Park1 . 1 Chung-Ang University, Psychiatry, Seoul, Republic of Korea; 2 Dankook University, Psychiatry, Cheonan, Republic of Korea Introducton: Genetic determination of individual alcohol withdrawal symptom can be helpful to find out genetic factors of alcohol dependence and to predict forthcoming alcohol withdrawal symptoms according to his or her own genes. Dopaminergic system at VTA (Ventral Tegmental Area) projects to cerebral cortex and limbic system, and especially NA (Nucleus Accumbens), where is known to be related with sensation and reward. We investigated the relationship between the polymorphism of the genes associated with dopamine and the alcohol withdrawal symptoms in the alcohol dependent patients. Methods: Subjects were 108 hospitalized male alcoholic patients whose ages were between 19 and 65 years, and age and sex matched 76 healthy controls were selected to compare the gene distrubution. In order to evaluate the severity of the alcohol withdrawal symptoms, the Clinical Institute Withdrawal Assessment for Alcohol (CIWA-Ar) was checked 48 hours after the last alcohol consumption. We analyzed the polymorphisms of DRD2 Taq1, dopamine transporter (DAT1), and catechol-O-methyltransferase (COMT). We analyzed allele frequencies, genotypes, heterozygotes, homozygotes of candidate genes such as DRD2 and DAT polymorphism, and catecholO-methyltransferase (COMT) polymorphism, using Monte-Carlo test. Genotypic distribution was compared to the expected results from Hardy-Weinberg equilibrium equation. The relationship between the scores of CIWA-Ar subscales and the genetic polymorphism in alcohol withdrawal patients were analyzed. Results: The ages, duration of education and genotypes for the alcohol dependent patients did not differ from those of the healthy control group. The genetic distribution between the two groups were similar to each other. None of the genotypes counts deviated significantly from those expected Hardy-Weinberg equilibrium. DRD2 Taq I: The auditory hallucination subscale score of the CIWA-Ar scale in homozygotes was significantly higher than that in heterozygotes, with its odd ratio at 1.34. The total score of CIWA-Ar scale in heterozygote was significantly higher than that in homozygote. DAT1: In the subjects without DAT-9 gene allele, the subscale scores of sweating and anxiety were significantly higher than those in the subject with DAT-9 gene allele. The total score of CIWA-Ar scale in the subject without DAT-9 gene allele was significantly higher than that in the subject with DAT-9 gene allele. COMT: The total scores of CIWA-Ar scale in the heterozygotes were significantly higher than those in homozygote. Conclusions: The genetic polymorphisms of DRD2, DAT1 and COMT are associated with various alcohol withdrawal symptoms, which are developed by the neurotrasmitters which were modulated by specific genes. Furthermore, these genetic polymorphisms are considered to have close association with withdrawal symptoms, which may be a important factor in the future treatment of alcohol withdrawal symptoms. Since multiple genes involve in the development of the alcohol withdrawal symptoms, individual symptom and genetic factors must be considered in the pharmacological treatment. Future large-scale investigations on the vulnerable genes, associated with the alcohol withdrawal symptoms, maybe necessary to confirm any associations between them.
References [1] Finckh U., Rommelspacher H., Kuhn S., Dufeu P., Otto G., Heinz A., 1997, Influence of the dopamine D2 receptor (DRD2) genotype on neuroadaptive effects of alcohol and the clinical outcome of alcoholism, Pharmacogenetics, 7, 271–281. [2] Gorwood P., Limosin F., Batel P., Hamon M., Ades J., Boni C., 2003, The A9 allele of the dopamine transporter gene is associated with delirium tremens and alcohol-withdrawal seizure, Biol Psychiatry, 53, 1, 85−92. [3] Kohnke M.D., Wiatr G., Kolb W., Kohnke A.M., Schick S., Lutz U., Vonthein R., Gaertner I., 2003, Plasma homovanillic acid: A significant association with alcoholism isindependent of a functional polymorphism of the human catechol – O – methyltransferase gene, Neuropsychopharmacology, 28, 5, 1004–1010.
P.6.b.004 Effect of amino acid derivatives and lipoic acid on lipid peroxidation and antioxidant systems indices in the rat brain during alcohol withdrawal P.S. Pronko ° , T.I. Khomich, L.R. Bardina, V.I. Satanovskaya, O.A. Borodich. Institute of Biochemistry NASB, Laboratory of Alcohols and Aldehydes Metabolism, Grodno, Belarus In detoxification of alcoholic patients, not only manifestations of pathologic craving for alcohol and alcohol withdrawal become the therapeutic targets, but also alcohol-damaged organs (brain, liver, heart and others). Many studies have demonstrated that alcohol enhances lipid peroxidation as well as modification of proteins in the brain. Both necrotic and apoptotic cell death can be triggered by oxidative stress. Earlier, scientists at our institute revealed the capacity of glutamine to reduce ethanol consumption and to decrease the severity of alcohol withdrawal syndrome signs, to accelerate alcohol elimination from the body and to diminish ethanol hepatotoxicity in rats. Ethanolamine decreases voluntary ethanol consumption by animals, manifests anticonvulsive and antistress activities in alcohol withdrawal and possesses antioxidant properties (Ostrovsky Yu.M., Ostrovsky S.Yu., 1995). In experiments in vitro using hippocampal cell culture, lipoic acid prevented ethanol-induced neurotoxicity and protein intracellular oxidation (Pirlich et al., 2002). The purpose of the study was to use a rat ethanol withdrawal model to study the effect of glutamine, N-acetylcysteine, ethanolamine and lipoic acid on the metabolic impairments in rat brain. The ethanol withdrawal syndrome was simulated in male albino rats by administering ethanol twice a day at a dose of 4−6 g/kg, i.g. for 5 days (Tezikov E.B. et al., 1991). The control animals received equal volumes of water in a similar way. Either lipoic acid (40 mg/kg, i.g.), or glutamine (340 mg/kg, i.g.) or ethanolamine (100 mg/kg, i.g.), or N-acetylcysteine (100 mg/kg, i.g.) were administered twice (after 3 h following the last ethanol administration and 2 h before decapitation). After 18 h following the ethanol withdrawal and 2 h following the last administration of the substances, the rats were killed by decapitation and parameters of lipid peroxidation, antioxidant systems and aldehyde metabolism were determined in the brain. After 5-day alcohol intake and 18 h following the cessation of its administration, brain tissue showed considerable elevation of thiobarbituric acid-reactive substances and activated catalase activity. The administration of glutamine, lipoic acid and N-acetylcysteine significantly decreased lipid peroxidation and increased reduced glutathione concentration, as well as glutathione reductase and glutathione peroxidase activities in brain tissue. The activity of catalase was normalized after the lipoic acid and N-acetylcysteine administration. There has been revealed lipoic
P.6.b Addiction – Alcohol (basic) acid, glutamine and N-acetylcysteine ability of eliminating the manifestations of ethanol-induced oxidative stress and stimulating the activities of the brain antioxidant defense systems in alcohol withdrawal. The activity of low KM brain aldehyde dehydrogenase diminished during ethanol withdrawal. N-acetylcycteine elevated the activity of aldehyde dehydrogenase, whereas ethanolamine decreased enzyme activity. The experimental data obtained confirm the manifestation of oxidative stress during alcohol withdrawal in brain tissue and suggest that the mechanism of the glutamine, lipoic acid and N-acetylcysteine antioxidant effects in the brain is probably related to activation of the nonenzymatic and enzymatic antioxidant defense systems. Our results substantiate the advisability of applying these substances as agents for metabolic therapy in patients with alcohol withdrawal. References [1] Y.M. Ostrovsky and S.Yu Ostrovsky, 1995, Amino acids in the pathogenesis, diagnosis and treatment of alcoholism, Science and Technology, Minsk. [2] M. Pirlich, K. Kick, G. Sanding, H. Lochs, T. Grune, 2002, Alpha-lipoic acid prevents ethanol-induced protein oxidation in mouse hippocampal HT22 cells, Neurosci. Lett., 328, 93−96. [3] E.B. Tezikov, V.P. Nuzhny, A.Ch. Abdrashitov, V.P. Listvina, 1991, Influence of the tolerance to ethanol, severity of ethanol withdrawal and age of rats on alcohol postintoxication heard damage, Vopr. Narcology, 1, 7−9.
P.6.b.005 Effects of olanzapine on ethanol withdrawal syndrome in rats N. Unsalan1 , H. Kayir2 , E. Saglam3 , I.T. Uzbay2 ° . 1 Maltepe University Faculty of Medicine, Department of Psychiatry, Istanbul, Turkey; 2 Gulhane Military Medical Academy, Medical Pharmacology Dept., Ankara, Turkey; 3 Maltepe University Faculty of Medicine, Department of Pharmacology, Istanbul, Turkey Introduction: Dopaminergic system, especially dopamine D2 receptors, has a specific importance in addiction for its impact to the reward mechanisms. Thus, drugs affecting dopaminergic system may be useful to control ethanol dependence. Olanzapine, an atypical antipsychotic drug, has antagonistic effects at a wide range of receptors, but has a higher affinity for D2 and 5-HT2A. Although it has been suggested that olanzapine reduces craving for alcohol (Hutchiason et al., 2003), there is no information about the effects of olanzapine on ethanol withdrawal syndrome (EWS). Thus, the present study was designed to investigate the effects of olanzapine on EWS in rats. Methods: Adult male Wistar rats (228±4.93 g) were subjects (n = 8 for each group). Ethanol (7.2%, v/v) was given to rats by a liquid diet for 21 days as previously described (Uzbay et al., 1995). Control rats were pair fed an isocaloric liquid diet containing sucrose as a caloric substitute to ethanol. The last day, ethanol was withdrawn from the diet and at the 2nd, 4th and 6th hrs of EWS, rats were evaluated for 5 min for the withdrawal signs, included locomotor hyperactivity, agitation, stereotyped behaviours and wet dog shakes. After 6 h of withdrawal testing, rats were exposed to an audiogenic stimulus (100 dB) for 60 s in a separate and sound-proof place in the laboratory. The incidence and latency of the audiogenic seizures were recorded. Locomotor activity was recorded (Opto Varimex Minor, Columbus, USA) as a total of horizontal, vertical and ambulatory activities. Other parameters were evaluated by an observer blind to the test groups. Olanzapine (0.5−2 mg/kg) or saline were injected to rats intraperitoneally
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30 min prior to 2nd and 6th hrs tests. Data were evaluated by analysis of variance (one-way ANOVA) followed by Dunnett’s test or Kruskal-Wallis test followed by Mann Whitney-U, and Chisquare tests. The level of significance was set at p < 0.05 levels. Results: Olanzapine (1 and 2 mg/kg), at the 2nd h experiment, produced some inhibitory effects on locomotor activity in control rats [F(3, 20) = 17.865; p < 0.0001]. Thus, we did not evaluate its effects on 2nd h of EWS. Olanzapine, only at 2 mg/kg dose, produced some significant inhibitory effects on stereotyped behaviours [F(3, 28) = 3.259; p = 0.036] and wet dog shakes [F(3, 28) = 3.010; p = 0.047] at the 6th h of EWS. Although olanzapine did not produce any significant change on the incidence of audiogenic seizures (p = 0.677, Chi-square test) in ethanol-dependent rats, it elicited some audiogenic seizures with an incidence, did not reach statistically significant levels (p = 0.427, Chi-square test), in naive controls. Conclusion: Our results suggest that acute olanzapine treatment has limited beneficial effects on stereotyped behaviours and wet dog shakes during EWS in rats. Thus, olanzapine does not seem to be a safe and effective choice in the treatment of EWS. References [1] Hutchison KE, Wooden A, Swift RM, Smolen A, McGeary J, Adler L, Paris L, 2003, Olanzapine reduces craving for alcohol: A DRD4 VNTR polymorphism by pharmacotherapy interaction, Neuropsychopharmacology, 28, 1882–1888. [2] Uzbay IT, Kayaalp SO, 1995, A modified liquid diet of chronic ethanol administration: Validation by ethanol withdrawal syndrome in rats, Pharmacol Res, 31, 37−42.
P.6.b.006 The role of epigenetic HERP dysregulation in patients with alcohol dependence S. Bleich ° , B. Lenz, D. B¨onsch, H. Frieling. University of Erlangen-Nuremberg, Department of Psychiatry and Psychotherapy, Erlangen, Germany Background: Elevated plasma homocysteine concentrations can influence genomic and gene specific DNA methylation in peripheral blood cells. The aim of this study was to investigate whether in patients with alcohol dependence, who show chronically elevated homocysteine levels, DNA methylation pattern within the HERP (homocysteine-induced ER protein) promoter region and expression of HERP-mRNA is altered. Methods: The HERP mRNA expression level was measured by quantitative PCR in the blood of 66 male alcoholics and 55 nondrinking healthy controls. Epigenetic genomic DNA methylation status and HERP promoter methylation was measured with a nonradioactive elongation-assay. Results: We observed a significant increase (7.6%) of the HERP promoter DNA methylation in patients with alcohol dependence (t-Test, t = −2.45, p < 0.02) when compared with healthy controls (80.4%, SD 14.5) which was significantly associated with their elevated homocysteine levels (multiple linear regression, p < 0.007). Furthermore, we found a significant lower HERP mRNA expression in patients with alcohol dependence (t-Test, −7.61 DCT; SD 1.87, p < 0.001) when compared with healthy controls (−6.04 DCT; SD 2.41). The lowered HERP mRNA expression in alcoholic patients was best explained by the hypermethylation of the regulatory HERP gene promoter (regression analysis, p = 0.004). Conclusions: To our knowledge, this is the first study evaluating HERP mRNA expression and its specific gene promoter methylation in alcoholics. Since hypermethylation of DNA is
P.6.b Addiction – Alcohol (basic)
P.6.b.003 Dopamine-related gene polymorphisms and alcohol withdrawal symptoms in patients with alcohol dependence B. Kee1 ° , Y.S. Lee1 , K.J. Min1 , K. Paik2 , D. Park1 . 1 Chung-Ang University, Psychiatry, Seoul, Republic of Korea; 2 Dankook University, Psychiatry, Cheonan, Republic of Korea Introducton: Genetic determination of individual alcohol withdrawal symptom can be helpful to find out genetic factors of alcohol dependence and to predict forthcoming alcohol withdrawal symptoms according to his or her own genes. Dopaminergic system at VTA (Ventral Tegmental Area) projects to cerebral cortex and limbic system, and especially NA (Nucleus Accumbens), where is known to be related with sensation and reward. We investigated the relationship between the polymorphism of the genes associated with dopamine and the alcohol withdrawal symptoms in the alcohol dependent patients. Methods: Subjects were 108 hospitalized male alcoholic patients whose ages were between 19 and 65 years, and age and sex matched 76 healthy controls were selected to compare the gene distrubution. In order to evaluate the severity of the alcohol withdrawal symptoms, the Clinical Institute Withdrawal Assessment for Alcohol (CIWA-Ar) was checked 48 hours after the last alcohol consumption. We analyzed the polymorphisms of DRD2 Taq1, dopamine transporter (DAT1), and catechol-O-methyltransferase (COMT). We analyzed allele frequencies, genotypes, heterozygotes, homozygotes of candidate genes such as DRD2 and DAT polymorphism, and catecholO-methyltransferase (COMT) polymorphism, using Monte-Carlo test. Genotypic distribution was compared to the expected results from Hardy-Weinberg equilibrium equation. The relationship between the scores of CIWA-Ar subscales and the genetic polymorphism in alcohol withdrawal patients were analyzed. Results: The ages, duration of education and genotypes for the alcohol dependent patients did not differ from those of the healthy control group. The genetic distribution between the two groups were similar to each other. None of the genotypes counts deviated significantly from those expected Hardy-Weinberg equilibrium. DRD2 Taq I: The auditory hallucination subscale score of the CIWA-Ar scale in homozygotes was significantly higher than that in heterozygotes, with its odd ratio at 1.34. The total score of CIWA-Ar scale in heterozygote was significantly higher than that in homozygote. DAT1: In the subjects without DAT-9 gene allele, the subscale scores of sweating and anxiety were significantly higher than those in the subject with DAT-9 gene allele. The total score of CIWA-Ar scale in the subject without DAT-9 gene allele was significantly higher than that in the subject with DAT-9 gene allele. COMT: The total scores of CIWA-Ar scale in the heterozygotes were significantly higher than those in homozygote. Conclusions: The genetic polymorphisms of DRD2, DAT1 and COMT are associated with various alcohol withdrawal symptoms, which are developed by the neurotrasmitters which were modulated by specific genes. Furthermore, these genetic polymorphisms are considered to have close association with withdrawal symptoms, which may be a important factor in the future treatment of alcohol withdrawal symptoms. Since multiple genes involve in the development of the alcohol withdrawal symptoms, individual symptom and genetic factors must be considered in the pharmacological treatment. Future large-scale investigations on the vulnerable genes, associated with the alcohol withdrawal symptoms, maybe necessary to confirm any associations between them.
References [1] Finckh U., Rommelspacher H., Kuhn S., Dufeu P., Otto G., Heinz A., 1997, Influence of the dopamine D2 receptor (DRD2) genotype on neuroadaptive effects of alcohol and the clinical outcome of alcoholism, Pharmacogenetics, 7, 271–281. [2] Gorwood P., Limosin F., Batel P., Hamon M., Ades J., Boni C., 2003, The A9 allele of the dopamine transporter gene is associated with delirium tremens and alcohol-withdrawal seizure, Biol Psychiatry, 53, 1, 85−92. [3] Kohnke M.D., Wiatr G., Kolb W., Kohnke A.M., Schick S., Lutz U., Vonthein R., Gaertner I., 2003, Plasma homovanillic acid: A significant association with alcoholism isindependent of a functional polymorphism of the human catechol – O – methyltransferase gene, Neuropsychopharmacology, 28, 5, 1004–1010.
P.6.b.004 Effect of amino acid derivatives and lipoic acid on lipid peroxidation and antioxidant systems indices in the rat brain during alcohol withdrawal P.S. Pronko ° , T.I. Khomich, L.R. Bardina, V.I. Satanovskaya, O.A. Borodich. Institute of Biochemistry NASB, Laboratory of Alcohols and Aldehydes Metabolism, Grodno, Belarus In detoxification of alcoholic patients, not only manifestations of pathologic craving for alcohol and alcohol withdrawal become the therapeutic targets, but also alcohol-damaged organs (brain, liver, heart and others). Many studies have demonstrated that alcohol enhances lipid peroxidation as well as modification of proteins in the brain. Both necrotic and apoptotic cell death can be triggered by oxidative stress. Earlier, scientists at our institute revealed the capacity of glutamine to reduce ethanol consumption and to decrease the severity of alcohol withdrawal syndrome signs, to accelerate alcohol elimination from the body and to diminish ethanol hepatotoxicity in rats. Ethanolamine decreases voluntary ethanol consumption by animals, manifests anticonvulsive and antistress activities in alcohol withdrawal and possesses antioxidant properties (Ostrovsky Yu.M., Ostrovsky S.Yu., 1995). In experiments in vitro using hippocampal cell culture, lipoic acid prevented ethanol-induced neurotoxicity and protein intracellular oxidation (Pirlich et al., 2002). The purpose of the study was to use a rat ethanol withdrawal model to study the effect of glutamine, N-acetylcysteine, ethanolamine and lipoic acid on the metabolic impairments in rat brain. The ethanol withdrawal syndrome was simulated in male albino rats by administering ethanol twice a day at a dose of 4−6 g/kg, i.g. for 5 days (Tezikov E.B. et al., 1991). The control animals received equal volumes of water in a similar way. Either lipoic acid (40 mg/kg, i.g.), or glutamine (340 mg/kg, i.g.) or ethanolamine (100 mg/kg, i.g.), or N-acetylcysteine (100 mg/kg, i.g.) were administered twice (after 3 h following the last ethanol administration and 2 h before decapitation). After 18 h following the ethanol withdrawal and 2 h following the last administration of the substances, the rats were killed by decapitation and parameters of lipid peroxidation, antioxidant systems and aldehyde metabolism were determined in the brain. After 5-day alcohol intake and 18 h following the cessation of its administration, brain tissue showed considerable elevation of thiobarbituric acid-reactive substances and activated catalase activity. The administration of glutamine, lipoic acid and N-acetylcysteine significantly decreased lipid peroxidation and increased reduced glutathione concentration, as well as glutathione reductase and glutathione peroxidase activities in brain tissue. The activity of catalase was normalized after the lipoic acid and N-acetylcysteine administration. There has been revealed lipoic
P.6.b Addiction – Alcohol (basic) acid, glutamine and N-acetylcysteine ability of eliminating the manifestations of ethanol-induced oxidative stress and stimulating the activities of the brain antioxidant defense systems in alcohol withdrawal. The activity of low KM brain aldehyde dehydrogenase diminished during ethanol withdrawal. N-acetylcycteine elevated the activity of aldehyde dehydrogenase, whereas ethanolamine decreased enzyme activity. The experimental data obtained confirm the manifestation of oxidative stress during alcohol withdrawal in brain tissue and suggest that the mechanism of the glutamine, lipoic acid and N-acetylcysteine antioxidant effects in the brain is probably related to activation of the nonenzymatic and enzymatic antioxidant defense systems. Our results substantiate the advisability of applying these substances as agents for metabolic therapy in patients with alcohol withdrawal. References [1] Y.M. Ostrovsky and S.Yu Ostrovsky, 1995, Amino acids in the pathogenesis, diagnosis and treatment of alcoholism, Science and Technology, Minsk. [2] M. Pirlich, K. Kick, G. Sanding, H. Lochs, T. Grune, 2002, Alpha-lipoic acid prevents ethanol-induced protein oxidation in mouse hippocampal HT22 cells, Neurosci. Lett., 328, 93−96. [3] E.B. Tezikov, V.P. Nuzhny, A.Ch. Abdrashitov, V.P. Listvina, 1991, Influence of the tolerance to ethanol, severity of ethanol withdrawal and age of rats on alcohol postintoxication heard damage, Vopr. Narcology, 1, 7−9.
P.6.b.005 Effects of olanzapine on ethanol withdrawal syndrome in rats N. Unsalan1 , H. Kayir2 , E. Saglam3 , I.T. Uzbay2 ° . 1 Maltepe University Faculty of Medicine, Department of Psychiatry, Istanbul, Turkey; 2 Gulhane Military Medical Academy, Medical Pharmacology Dept., Ankara, Turkey; 3 Maltepe University Faculty of Medicine, Department of Pharmacology, Istanbul, Turkey Introduction: Dopaminergic system, especially dopamine D2 receptors, has a specific importance in addiction for its impact to the reward mechanisms. Thus, drugs affecting dopaminergic system may be useful to control ethanol dependence. Olanzapine, an atypical antipsychotic drug, has antagonistic effects at a wide range of receptors, but has a higher affinity for D2 and 5-HT2A. Although it has been suggested that olanzapine reduces craving for alcohol (Hutchiason et al., 2003), there is no information about the effects of olanzapine on ethanol withdrawal syndrome (EWS). Thus, the present study was designed to investigate the effects of olanzapine on EWS in rats. Methods: Adult male Wistar rats (228±4.93 g) were subjects (n = 8 for each group). Ethanol (7.2%, v/v) was given to rats by a liquid diet for 21 days as previously described (Uzbay et al., 1995). Control rats were pair fed an isocaloric liquid diet containing sucrose as a caloric substitute to ethanol. The last day, ethanol was withdrawn from the diet and at the 2nd, 4th and 6th hrs of EWS, rats were evaluated for 5 min for the withdrawal signs, included locomotor hyperactivity, agitation, stereotyped behaviours and wet dog shakes. After 6 h of withdrawal testing, rats were exposed to an audiogenic stimulus (100 dB) for 60 s in a separate and sound-proof place in the laboratory. The incidence and latency of the audiogenic seizures were recorded. Locomotor activity was recorded (Opto Varimex Minor, Columbus, USA) as a total of horizontal, vertical and ambulatory activities. Other parameters were evaluated by an observer blind to the test groups. Olanzapine (0.5−2 mg/kg) or saline were injected to rats intraperitoneally
S499
30 min prior to 2nd and 6th hrs tests. Data were evaluated by analysis of variance (one-way ANOVA) followed by Dunnett’s test or Kruskal-Wallis test followed by Mann Whitney-U, and Chisquare tests. The level of significance was set at p < 0.05 levels. Results: Olanzapine (1 and 2 mg/kg), at the 2nd h experiment, produced some inhibitory effects on locomotor activity in control rats [F(3, 20) = 17.865; p < 0.0001]. Thus, we did not evaluate its effects on 2nd h of EWS. Olanzapine, only at 2 mg/kg dose, produced some significant inhibitory effects on stereotyped behaviours [F(3, 28) = 3.259; p = 0.036] and wet dog shakes [F(3, 28) = 3.010; p = 0.047] at the 6th h of EWS. Although olanzapine did not produce any significant change on the incidence of audiogenic seizures (p = 0.677, Chi-square test) in ethanol-dependent rats, it elicited some audiogenic seizures with an incidence, did not reach statistically significant levels (p = 0.427, Chi-square test), in naive controls. Conclusion: Our results suggest that acute olanzapine treatment has limited beneficial effects on stereotyped behaviours and wet dog shakes during EWS in rats. Thus, olanzapine does not seem to be a safe and effective choice in the treatment of EWS. References [1] Hutchison KE, Wooden A, Swift RM, Smolen A, McGeary J, Adler L, Paris L, 2003, Olanzapine reduces craving for alcohol: A DRD4 VNTR polymorphism by pharmacotherapy interaction, Neuropsychopharmacology, 28, 1882–1888. [2] Uzbay IT, Kayaalp SO, 1995, A modified liquid diet of chronic ethanol administration: Validation by ethanol withdrawal syndrome in rats, Pharmacol Res, 31, 37−42.
P.6.b.006 The role of epigenetic HERP dysregulation in patients with alcohol dependence S. Bleich ° , B. Lenz, D. B¨onsch, H. Frieling. University of Erlangen-Nuremberg, Department of Psychiatry and Psychotherapy, Erlangen, Germany Background: Elevated plasma homocysteine concentrations can influence genomic and gene specific DNA methylation in peripheral blood cells. The aim of this study was to investigate whether in patients with alcohol dependence, who show chronically elevated homocysteine levels, DNA methylation pattern within the HERP (homocysteine-induced ER protein) promoter region and expression of HERP-mRNA is altered. Methods: The HERP mRNA expression level was measured by quantitative PCR in the blood of 66 male alcoholics and 55 nondrinking healthy controls. Epigenetic genomic DNA methylation status and HERP promoter methylation was measured with a nonradioactive elongation-assay. Results: We observed a significant increase (7.6%) of the HERP promoter DNA methylation in patients with alcohol dependence (t-Test, t = −2.45, p < 0.02) when compared with healthy controls (80.4%, SD 14.5) which was significantly associated with their elevated homocysteine levels (multiple linear regression, p < 0.007). Furthermore, we found a significant lower HERP mRNA expression in patients with alcohol dependence (t-Test, −7.61 DCT; SD 1.87, p < 0.001) when compared with healthy controls (−6.04 DCT; SD 2.41). The lowered HERP mRNA expression in alcoholic patients was best explained by the hypermethylation of the regulatory HERP gene promoter (regression analysis, p = 0.004). Conclusions: To our knowledge, this is the first study evaluating HERP mRNA expression and its specific gene promoter methylation in alcoholics. Since hypermethylation of DNA is