1. the reasons for that, if there are no receptors to ethanol and most of ethanol
effects are conditioned by its metabolites? Relying on our own histochemical
studies on the comparative distribution of the main ethanol-metabolizing
enzymes in brain structures and literature data, we believe that structure-
specific action of alcohol in the brain depends on different ability of its
structures to oxidize ethanol and its first, highly active metabolite acet-
aldehyde (AA). It results in local accumulation or deficiency of ethanol
metabolites (AA, salsolinol, acetate) mediating the selective disturbances in
the brain structures and their involvement in the behavioral effects of alcohol
and alcoholism pathogenesis. In those mechanisms, the ethanol-oxidizing
enzymes catalase and, to the less extent, cytochrome Ð450 2Å1 ensure the
local formation of AA, which mediates the positive reinforcing, motor dis-
order as well as local neurotoxic effects of alcohol. The AA oxidizing
enzyme aldehyde dehydrogenase protects the brain structures from the toxic
effect of the excess of AA, but in specific brain structures it may initiate the
local inborn or acquired deficiency of AA and, at the behavioral level, the
addiction to ethanol as metabolic precursor of AA. Our theory explains
numerous discrete, often contradictory, data concerning the alcohol action in
the brain, indicates the central oxidative mechanisms of alcoholism patho-
genesis and possible ways of its metabolic treatment.
O3.4
INVOLVEMENT OF CHROMATIN REMODELLING IN THE
DEVELOPMENT OF ETHANOL-INDUCED BEHAVIORAL
SENSITIZATION
R. Legastelois, M. Naassila and B. Botia
Université de Picardie Jules Verne, Faculté de Pharmacie, INSERM
ERI24-GRAP, Amiens, France
Alcoholism is a major public health priority and a devastating illness world-
wide despite the current pharmacotherapies. Studies have shown that epige-
netic mechanisms such as chromatin remodelling are deeply involved in the
induction and the persistence of addictive behaviors. We focused on the
involvement of histone acetylation in the model of ethanol-induced behav-
ioral sensitization. Behavioral sensitization is defined by a progressive and
long-lasting enhancement of locomotor response to a drug after repeated
injections reflecting the sensitization of the dopaminergic pathway of the
reward circuitry. We used sodium butyrate (NaB), a histone deacetylase
inhibitor (HDACi), on behavioral sensitization induced by low (1 g/kg) or
high (2 g/kg) doses of ethanol. In our protocol, DBA/2J mice received daily
ethanol injections during 10 days (induction phase: days 1–10) followed by
locomotor activity tests. Mice were then let undisturbed in their home cages
for 1 week and were finally challenged with an ethanol injection at Day 17
(expression phase). Very interestingly, NaB inhibited the induction and the
expression of sensitization when ethanol was administered at the low dose
but not at the high dose. Furthermore, daily injections of NaB from Day 11
to 16 reversed sensitization of mice previously sensitized with the low dose
of ethanol. Moreover, several brain structures have been isolated and PCR
arrays have been performed to identify the modifications of gene expression
associated with sensitization development or with its inhibition by NaB. The
studied genes belong to various families like neurotransmitters, chromatin
remodelling enzymes or transduction pathways. Our data suggest that the
neuroadaptations occurring during the induction of ethanol sensitization are
different depending of the ethanol dose. Our findings also suggest that the
induction and expression of ethanol-induced sensitization with low dose of
ethanol may involve chromatin remodelling mechanisms. Finally, the interest
of studying HDACi in alcohol addiction is increased by our results showing
the reversion of sensitization by NaB. (This study was supported by the
Regional Council of Picardie (RCP), INSERM, MiLDT/Inca/INSERM
project A09119ES. R.L. and B.B. are respectively supported by doctoral and
post-doctoral fellowship from RCP.)
O3.5
INTERPRETATION OF THE CARBOHYDRATE-DEFICIENT
TRANSFERRIN (CDT) ASSAY OF CIRRHOTIC PATIENTS
P. Gonzalo1,2, M. Pecquet1, C. Bon1, S. Gonzalo3 and J. Souquet1,2
1
Hôpital de la Croix-Rousse, Lyon, France, 2
Université Lyon I, Lyon,
France and 3
Laboratoire Biomnis, Lyon, France
Background. The percentage of CDT in the serum is a biological marker to
estimate chronic alcohol abuse. This assay is both difficult and critical for
cirrhotic patients, especially those awaiting liver transplantations. In this
presentation, we propose to display the performances of the CDT-Capillarys
assay (Sebia) to discriminate abstainers from abusers among cirrhotic
patients and to expose recommendations to improve its reliability in this
patient group.
Methods. A total of 110 patients with known hepatic status of cirrhosis had
their CDT measured by Capillarys 2 and confronted to their daily alcohol
intake. CDT assays by the Bio-Rad %CDT by HPLC test or the N-Latex
CDT assay (Siemens) were also performed as alternative methods.
Results. Many electrophoretic profiles displayed by the Capillarys2 are
extensively processed by the Phoresis software in case of cirrhosis. This was
not observed with control sera. In about one-third of the cases, the unpro-
cessed profiles displayed major base-lane drifts and peaks with less resol-
ution between disialo- and trisialo-transferrin fractions. In order to decide
when the processed electrophoretic profiles could reliably be used, we
defined a qualitative criterion. This criterion consisted in applying an indi-
cator of the resolution between the disialo- and trisialo-transferrin peaks on
the electrophorectic profile. Thus, detecting abusers with cirrhosis using the
CDT-Capillarys assay is possible when the indicator is in the normal range.
However, only 54% of the profiles from cirrhotic patients fulfilled this cri-
terion, and no alternative CDT assay (either the Bio-Rad %CDT by HPLC
test or the N-Latex CDT assay by immunoprecipitation, Siemens) demon-
strated satisfying performance for excluded samples.
Conclusions. An attentive analysis of Capillarys CDT electrophoretic profiles
is required for CDT validation of cirrhotic patients. The reliability of this par-
ameter being a major point for biologists, we provide here a new criterion to
select reliable cirrhotic CDT profiles. In the future, decreasing the number of
uninterpretable samples in case of cirrhosis will require an improvement in all
CDT assays, which might be possible in capillary zone electrophoresis by
increasing the resolution of the transferrin isoform profiles.
O3.6
FRONTAL LOBE FUNCTION IN ALCOHOLICS CLASSIFIED
ACCORDING TO LESCH’S TYPOLOGY
E. M. Nakamura-Palacios and M. P. Zago-Gomes
Federal University of Espírito Santo, Vitória, Brazil
This study examined frontal lobe cognitive function and mental state among
patients with different types of alcohol dependence according to Lesch’s
typology. The Frontal Assessment Battery (FAB) and the Mini-Mental
Status Examination (MMSE) were given to 170 patients with alcoholism
from a Brazilian outpatient service classified by Lesch’s typology and to 40
non-alcoholic controls matched for age, gender, socio-demographic charac-
teristics and education. Of the alcoholic sample, 21.2% were classified as
Type I, 29.4% as Type II, 28.8% as Type III and 20.6% as Type IV.
Alcoholics showed significantly lower overall scores on the MMSE and the
FAB, when compared with non-alcoholic subjects. Type IV alcoholics had
lower MMSE and FAB overall scores, when compared with non-alcoholic
controls and also with all other types of alcoholic subjects. However, Type
II and III subjects with alcoholism also had lower overall FAB scores, but
not overall MMSE scores, when compared with non-alcoholic controls. The
FAB subsets of motor programming, sensitivity to interference and inhibi-
tory control, were significantly reduced in Types II, III and IV alcoholics,
when compared with non-alcoholic subjects, but only motor programming
remained impaired in Type IV alcoholics with preserved mental function.
Executive dysfunctions in alcohol dependence seem to vary depending upon
the type of alcoholism. Therefore, the determination of clinical type of
alcohol dependence by applying Lesch’s typology, along with brief mental
state and frontal function examinations, is of clinical relevance in the exam-
ination of alcoholics and provides significant clues for more directed forms
of alcohol dependence treatment.
FREE ORAL COMMUNICATIONS 4: ALCOHOL
DEPENDENCE: TREATMENT APPROACHES
doi:10.1093/alcalc/agr096
O4.1
EFFECTIVENESS OF STIMULANT MEDICATION IN FETAL
ALCOHOL SPECTRUM DISORDERS
M. A. Infante, C. C. Humber, S. N. Mattson and E. P. Riley
San Diego State University, San Diego, CA, USA
Fetal alcohol spectrum disorders (FASDs) are often characterized by symp-
toms of inattention and hyperactivity. Many children with FASDs receive a
diagnosis of attention-deficit/hyperactivity disorder (ADHD). However, it is
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2. unclear whether treatment with standard stimulant medication is effective in
FASDs. This study evaluated response to stimulant medication in a group of
children with heavy prenatal alcohol exposure and ADHD (ALC-ADHD)
and nonexposed children with ADHD (Con-ADHD). Children, aged 7–15,
were assessed with the QuotientTM
ADHD System, which provides an objec-
tive measure of three core symptoms of ADHD (motion, attention and shifts
in attention state). Subjects were evaluated twice, with and without stimulant
medication. A significant group × stimulant interaction was found for
measures of accuracy (P = 0.06), commission errors (P = 0.02), head move-
ment (P = 0.03), head displacement (P < 0.01) and magnitude of head move-
ment (P = 0.02). Subjects in the Con-ADHD group evidenced greater
improvement with stimulant medication than those in the ALC-ADHD
group. Importantly, in contrast, children in the ALC-ADHD group per-
formed worse on several measures when tested on stimulant medication than
when tested without a stimulant. These findings suggest that stimulant treat-
ment for attention deficits in children with FASD is questionable. They also
suggest that prenatal alcohol exposure should be considered as a contributory
factor in the etiology of ADHD in individuals who do not show a positive
response to stimulant medication. Future research should evaluate which
aspects of ADHD are affected by stimulants in individuals with FASD and
whether other treatments, including nonstimulant medication, may be more
effective. (The research was supported by NIAAA grants R01 AA010417
T32 AA013525 to EPR and AA019605 to SNM.)
O4.2
NEUROCOGNITIVE FACTORS MODERATE THE
EFFECTIVENESS OF GROUP MOTIVATIONAL ENHANCEMENT
THERAPY AMONG HIGH-RISK ADOLESCENTS
A. D. Bryan1, S. Feldstein Ewing2, A. Brock3, R. Magnan4 and
K. Hutchison3,1
1
University of Colorado, Boulder, CO, USA, 2
Mind Research Network,
Albuqerque, NM, USA, 3
Mind Research Network, Albuquerque, NM, USA
and 4
University of New Mexico, Albuquerque, NM, USA
Adolescents involved in the juvenile justice system experience high levels of
alcohol use disorders and related risk behavior, including alcohol-related
sexual risk. Interventions to date are somewhat effective, but do not work
well for all youth. A better understanding of neurocognitive factors under-
lying risk behavior may facilitate the development of better interventions
and the targeting and tailoring of those that are currently available. This
ongoing study explores whether neurocognitive factors might moderate the
effectiveness of an intervention to decrease alcohol-related sexual risk behav-
ior. Adolescents in the juvenile-justice system (age 14–18; 55% male) were
randomly assigned to a group-level MET-based intervention to decrease
alcohol-related risky sexual behavior (n = 94) or an information-based
control intervention (n = 95). Over 80% of these youth have engaged in
sexual intercourse with an average age at first intercourse of 13.09 and an
average of 6.6 sexual partners (lifetime), yet only 15.2% had used condoms
100% of the time they had had sex. Almost 20% drank alcohol once a week
or more, and over 40% consumed four or more drinks per drinking occasion.
Only 34% of participants said they ‘never’ had sex under the influence of
alcohol. Clearly, these young people in engage in high levels of sexual risk,
alcohol use and the co-occurrence of these behaviors. In a task assessing
response inhibition (Go/NoGo), we find robust main effects consistent with
prior work with both adolescents and adults. We also show significant corre-
lations between activation during the task (correct rejects versus hits) and
both alcohol use and risky sexual behavior. Importantly, brain activation
during the task significantly moderates the effectiveness of the intervention
on both sexual risk (regions: IFG, DLPFC, SMA) and alcohol use (region:
IFG). Understanding why adolescents with different levels of activation
during a response inhibition task respond more positively to the MET inter-
vention is an important first step in understanding why these interventions
are not universally effective, and lay the founding for discovering easily
identifiable markers of variability in brain activation (e.g. genetic factors)
that may help to tailor and target psychosocial intervention.
O4.3
SEROTONERGIC, DOPAMINERGIC AND NORADRENERGIC
FUNCTIONS IN ALCOHOL-DEPENDENT INDIVIDUALS
K. J. Berglund1, C. Fahlke1, U. Berggren2, H. Zetterberg2, K. Blennow2,
J. Engel3 and J. Balldin2
1
Department of Psychology, Göteborg, Sweden, 2
Department of Psychiatry
and Neurochemistry, Göteborg, Sweden and 3
Department of Pharmacology,
Göteborg, Sweden
Background. Alcohol-dependence (AD) has been associated with a reduced
function of serotonin (5-HT), dopamine (DA) as well as noradrenaline (NA)
activities in several neuroendocrine studies. To our knowledge, there are,
however, no studies investigating all three systems with the use of neuro-
endocrine methods in one and the same individual.
Methods. In this study, 42 AD individuals and 28 controls participated
in the neuroendocrine tests. Central 5-HT neurotransmission was assessed
by the prolactin (PRL) response to citalopram (CIT), the postsynaptic
DRD2 function by the growth hormone (GH) response to apomorphine
(APO) and the postsynaptic α2-adrenoceptor function by GH response to
clonidine (CLON). The difference between baseline hormonal levels and
the highest level after challenge drug administration (delta-values) was
calculated and comparisons in these values were made between AD indi-
viduals and controls. Based on a median-split procedure for delta values
in the controls, individuals were also defined as ‘high’ and ‘low’ respon-
ders in each system and an individual monoaminergic profile was con-
structed when combining these responses. The frequency of different
combinations of these profiles was compared between AD individuals
and controls.
Results. There was a significant difference in mean delta PRL values
(PRL response to CIT) between AD individuals and controls (P < 0.05).
Consequently, the mean delta PRL value was reduced by 45% in AD
individuals. There were no significant differences in APO-GH and
CLON-GH concentrations in mean delta GH values between the groups.
Regarding combinations of monoamonergic profiles, AD individuals had
a combined low response of 5-HT and DA (P < 0.04); 5-HT and NA (P
< 0.01) as well as a low response in all three systems (5-HT, DA and
NA; P < 0.01).
Conclusion. The monoaminergic dysfunction was mainly restricted to
an impairment of the serotonergic system, suggesting that this system
is the most vulnerable to long term and excessive alcohol consumption.
Moreover, impaired monoaminergic profiles including low responses in
two or three systems were more frequently observed in AD individuals.
Since, this is the first study that investigated all three monoaminergic
systems in one and the same AD individual, further studies are
needed.
O4.4
A PRACTICAL SCALE OF CLINICAL IMPRESSION AS BASIS
FOR TREATMENT DECISIONS AND LONG-TERM FOLLOW-UPS
OF ADDICTED PATIENTS
A. Ulmer
Gemeinschaftspraxis, Stuttgart, Germany
Objective. Patients with addiction diseases often suffer more severely from
their disease than patients with other severe chronic diseases (e.g. HIV,
hepatitis, diabetes). The potency of addiction diseases to destroy life, the
most important social relationships and a central aspect: the honour of the
person, threatens them especially. Many patients do not find their way to
regular treatment and many remain unhappy despite treatment. We have to
strengthen our reflections: What are we doing? How good is the status of
our patients?
Methods. For >17 years, we have been using a very easy scale on clinical
impression for a prospective documentation of the particular general develop-
ment, the actual medication, its dosage and special events like detoxification
or arrests in the life of our addicted patients. We call it clinical impression,
according to our clinical view. First, we made four steps from ‘no improve-
ment’ over ‘better, but not good’ and ‘good, but not yet completely’ to
‘good’. A fifth step is for patients who remain stable after a successful termin-
ation of treatment. The frequent addition of + and − led to an effective division
of each step into three steps, finally resulting in a 15-step scale.
Results. The scale and subsequently developed charts became a precious
help for our daily treatment decisions. The quality of decisions made
improved substantially. Secondly, the diagrams gradually extended and
changed our view of addiction diseases, like statistical analyses or comple-
menting them, thus leading to a further improvement of the therapy.
Conclusions. An easy scale on clinical impressions, guided by the question:
‘what is good?’ and subsequent diagrams influence and change our view on
addiction diseases. Strongly showing us the chronic character of this disease
and the limited effect of most active measures against it, they force us to
ask: How can our patients live as well as possible with the disease, if it is
impossible to surmount it? What is good? Orientation is a perfect treatment
of other chronic diseases, leading to a stable, satisfied and unthreatened life
and working capability like in a healthy life. This, finally, is a new para-
digma.
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3. Practical scale of clinical impression about the development of patients in
treatment because of alcohol dependence
The following aspects are includet in the categories ‘good’ to ‘not good’:
Sureness of the assessment
Clearness of an attitude
Panic and anxiety attacks
Mood
Excitability/balance
Craving
Reliability and readiness to change
Appellation of the problem
Social stability
Laboratory results
Frequency of irritations and any relapses
Heaviness of irritations and any relapses
Complications (e.g. pancreatitis, suizide, criminality, insurances)
Medication compliance
Cooperation with the medical setting
Cooperation with the further therapeutic setting
V Good, without
further therapy
15 Very stable condition, no actual danger
from addiction or accompanying
problems can be recognized any longer.
14 Sustained stability, lasting over a period of
at least months. Good general result, no
further treatment required.
13 Good general results, even without further
treatment, outcome does not yet appear to
be absolutely sure.
IV Stable good, no
use of drugs.
Use of alcohol,
if any, is
absolutely
unproblematic
12 Similar to 11, still more sure, near to a
regular completion of the treatment.
11 Sustained stable condition, no recognizable
endangerment from alcohol or
accompanying problems.
10 Similar to 11, but stability not yet
absolutely sure.
III Good, but not yet
completely so
9 Not yet really stable and persistently good,
but approaching that condition.
8 Good, but not yet stable, e.g. not yet
sustained, or still emotionally
compromised.
7 Similar to 8, but still very unstable or
compromised.
II Situation better,
but not yet
good
6 Overall results not yet a good, but nearly so.
5 Similar to 4, but general results are not very
bad, or improvements noted over a short
period of time.
4 In some respects, considerable
improvements, but overall results not
good (e.g. dysphoria, negative events
indicating some conflictual behaviour,
alcohol).
I No improvement 3 No general improvement, but actually not
the deepest relapse, use of alcohol is
rather lighter.
2 Similar to 1, but slightly milder.
1 actually again completely relapsed, failing
control
O4.5
ACAMPROSATE FOR IMPROVING CONTROLLED DRINKING:
IMPACT OF TREATMENT DURATION ON THERAPY SUCCESS
P. Lehert1 and B. Mason2
1
University of Louvain, Faculty of Economics (FUCAM), Mons, Belgium
and 2
C-Scripps Institute, San Diego, CA, USA
Acamprosate is commonly used to reduce consumption of alcoholic patients.
Meta-analyses provided evidence of efficacy of this drug, with relapse relative
risk compared with placebo estimated to be 0.65 and 0.8. However, different
treatment durations were used in the trials, varying from 3 months until 1 year,
thus the pooled results were based on the assumption of a constant effect of
treatment in time. A recent meta-analysis (Mann et al., 2004), noticed better
efficacy for longer studies, however, these results suffer from the assumption of
homogeneity of treatment effect across studies. Our objective was to re-assess
the varying effect of acamprosate with treatment duration, in conducting an
Individual Patient Data IPD meta-analysis based on all studies with a duration
of at least 1 year. Our main endpoint was time to first severe relapse (TFR,
severe relapse defined as more than 5 drinks for men and 3 drinks for women)
and was analyzed on an intent to treat basis. We conducted a specific IPD test
based on mixed multilevel proportional time Hazard regression in stratifying on
study interaction, by using a time-dependent covariate to test time-treatment
first-order interaction and adjusting on baseline covariates. Constant treatment
and a linear increase of treatment effect in time were compared.
Results. Seven studies with a duration of at least 1 year were found
(n = 2194). The linearly increasing treatment effect on TFR was more determi-
nant than the constant effect. For the three periods, Day 90, 180 and 360, we
found hazard ratios (HRs) = 0.56, 95%CI [0.46 to 0.69] (P < 0.0001), 0.68
[0.57, 0.80], P < 0.0001 and 0.80 [0.71, 0.89], P = 0.004, respectively.
Conclusion. The acamprosate effect of acamprosate remains small after
3 months duration, although the effect becomes clinically significant at D180
and still improves until 1 year of treatment. Based on an IPD estimate of
patients followed for at least 1 year, our analysis confirms an expected linear
effect of acamprosate in time in reducing severe relapses, and provides
evidence that this treatment must be provided for at least 6 months, ideally
1 year.
REFERENCE
Mann K, Lehert P, Morgan MY (2004) The efficacy of acamprosate in the
maintenance of abstinence in alcohol-dependent individuals: results of a
meta-analysis. Alcohol Clin Exp Res 28:51–63.
O4.6
ALCOHOLISM EFFECTS ON DIFFERENT COMPONENTS
OF THEORY OF MIND
H. Beaunieux1, C. Cauvin2, C. Lannuzel2, C. Duval1, A. Le Berre1,
F. Vabret2, A. Pitel1, P. Allain3, B. Desgranges1 and F. Eustache1
1
Inserm- EPHE- Université de Caen Basse-Normandie, Caen Cedex, France,
2
CHU Caen Côte de Nacre, Caen Cedex, France and 3
Université d’Angers,
Angers, France
Introduction. Alcoholism is associated with a wide range of cognitive and
affective disorders, which have been explained in part by a specific vulner-
ability of the frontal lobes to the neurotoxic effects of alcohol. The ability to
infer representations or mental states to others has been referred to as
‘Theory of Mind’ (ToM). ToM is thought to be subtended, in part, by the
frontal lobes, but so far only a few studies have addressed the issue of ToM
deficits in alcoholism. Thus, the aim of the present study was to investigate
ToM and its links with other cognitive processes, such as information pro-
cessing speed, executive functions and episodic memory.
Method. A comprehensive neuropsychological test battery was administered
to patients with chronic alcoholism (AL) and age-matched healthy controls
(HC). The test battery included measurements of processing speed, executive
functions, episodic memory, subjective and objective assessment ToM tasks.
The subjective assessment of ToM abilities was performed via the ToM
scale, an original self-rating questionnaire. The objective tasks were designed
to probe both components (affective and cognitive) of ToM: The Eyes test,
the False Belief Task and the Faux Pas Recognition test. The two groups
were matched with respect to intellectual abilities, age and performance on
ToM tasks control conditions.
Results. Group comparisons revealed a significant difference between AL
and HC on objective ToM tasks but not on subjective ToM task. Group com-
parisons revealed no significant impairment of processing speed, executive
functions and episodic memory in AL. Discussion. To our knowledge, this is
the first study to have examined the effects of alcoholism on measures of both
ToM components using two types of assessment. AL, spared from other cog-
nitive deficits, were less efficient than HC to appreciate the mental states of
others but were unaware of their ToM deficits and believed to be as effective
as HC. These ToM deficits, which may contribute to interpersonal difficulties,
raise a fundamental question: are these deficits harmful consequences of
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4. excessive alcohol consumption or a premorbid risk factor for addiction? (This
study was supported by the grant No. RPE10001EEA from MILDT.)
FREE ORAL COMMUNICATIONS 5: ALCOHOL-
RELATED LIVER DISEASES: BIOLOGICAL MARKERS
doi:10.1093/alcalc/agr097
O5.1
THE CYTOCHROME P450 2E1 INHIBITOR CHLORMETHIAZOLE
INHIBITS HEPATIC ETHANOL-MEDIATED CARCINOGENESIS
INDUCED BY DIETHYLNITROSAMINE
Q. Ye1, F. Lian1, P. R. G. Chavez1, J. Chung1, W. Ling2, H. K. Seitz3 and
X. D. Wang4
1
Nutrition & Cancer Biology Lab, Human Nutrition Res. Center on Aging,
Tufts University, Boston, MA, USA, 2
Nutrition & Cancer Biology Lab,
Human Nutrition Res. Centre on Aging, Tufts University, Boston, MA,
USA, 3
Universitätsklinikum Heidelberg, Medizinische Klinik, Krankenhaus
Salem, Heidelberg, Germany and 4
Nutrition & Cancer Biology Lab.,
Human Nutrition Res. Center on Aging, Tufts University, Boston,
MA, USA
Background. Alcohol is a carcinogen for the liver. Chronic alcohol con-
sumption increases Cytochrome P450 2E1 (CYP2E1) in liver which is
associated with a generation of highly carcinogenic DNA lesions. The
purpose of the present study was to investigate whether the CYP2E1 inhibi-
tor chlormethiazole (CMZ) has an effect on hepatic carcinogenesis induced
by diethylnitrosamine (DEN).
Methods. Rats received Lieber–DeCarli ethanol containing control diets
with and without CMZ and a single intraperitoneal injection of a low dose
of DEN (20 mg per kg body weight). The formation of hepatic preneoplastic
foci was assessed by immunostaining of placental glutathione-S-transferase
(p-GST). In addition, hepatic proliferation was assessed by immunohisto-
chemistry for proliferating cellular nuclear antigen, Ki-67 and cyclin D1. In
addition, hepatic expression of TNF-α and activation of NF-κB and
CYP2E1 were assessed by either real-time PCR or western blotting analysis.
Results. Ethanol feeding resulted in a significant induction of hepatic
CYP2E1 increased nuclear accumulation of NF-κB protein and TNF-α
expression which was associated with increased hepatocyte proliferation
markers and cyclin D1 protein expression as well as increased
p-GST-positive altered hepatic foci in rat liver tissues. Chlormethiazole treat-
ment significantly inhibited ethanol-induced CYP2E1 expression, TNF-α
mRNA expression, NF-κB activation, hepatocyte proliferation and altered
hepatic foci formation after a 1-month period.
Conclusion. These data indicate that the inhibition of ethanol-induced
CYP2e1 can block hepatic preneoplastic lesion by decreasing TNF-α
expression and NF-κB activation in DEN-treated rats.
O5.2
BETAINE FEEDING PREVENTS THE BLOOD ALCOHOL CYCLE
IN RATS FED ALCOHOL CONTINUOUSLY FOR 1 MONTH USING
THE INTRAGASTRIC TUBE FEEDING MODEL
S. W. French1, X. Li2, J. Li1, F. Bardag-Gorce1, J. Oliva1 and B. A. French1
1
Harbor UCLA Medical Center, Torrance, CA, USA and 2
UCLA, Los
Angeles, CA, USA
Background. Blood alcohol levels (BAL) cycle up and down over a 7- to
8-day period when ethanol is fed continuously for 1 month in the intragas-
tric tube feeding rat model (ITFRM) of alcoholic liver disease. The cycling
phenomenon is due to an alternating increase and decrease in the metabolic
rate. Recently, we found that S-adenosylmethionine (SAMe) fed with
alcohol prevented the BAL cycle.
Method. Using the ITFRM, we fed betaine (2 g/kg/day) with ethanol for 1
month and recorded the daily 24 h urine ethanol level (UAL) to measure the
BAL cycle. UAL is equivalent to BAL because of the constant ethanol infu-
sion. Liver histology, steatosis and BAL were measured terminally after
1-month treatment. Microarray analysis was done on the RNA liver extracts
to determine the effects of betaine and alcohol on changes in gene
expression.
Results. Betaine fed with ethanol completely prevented the BAL cycle as
did SAMe. Betaine also significantly reduced the BAL compared with
ethanol fed without betaine. This was also observed when SAMe was fed
with ethanol. The mechanism involved in both cases is that SAMe is
required for the conversion of norepinephrine to epinephrine by PNMT.
Betaine feeding increases SAMe levels. Epinephrine induction of an
increased metabolic rate is 5–10-fold greater than norepinephrine, which
explains why SAMe and betaine prevented the cycle. Microarray analysis
showed that betaine feeding prevented the upregulation of a large number of
genes, including TLR2/4, IL-1b, Jax3, Sirt3, Fas, Ifnγ-1, Tgfgr2, Tnfrsf21,
Lbp and Stat 3, which could explain how betaine prevented fatty liver.
Conclusion. Betaine and SAMe feeding lower the BAL and prevent the
BAL cycle by increasing the metabolic rate. This increases the rate of
ethanol elimination. This reduces the blood alcohol level in ITFRM. (This
study was supported by NIH/NIAAA grant 8116.)
O5.3
BONE LOSS IN ALCOHOL ABUSE IS ASSOCIATED
WITH OSTEOCYTE APOPTOSIS, BONE MARROW
AND MICRO-VESSELS FAT INCORPORATION
D. B. Maurel1, C. Jaffré1, N. L. Fazzalari2, R. Uzbekov3, N. Boisseau4,
G. Y. Rochefort1, S. Pallu1 and C. Benhamou1
1
INSERM Unit U658, Orleans, France, 2
Bone and Joint Research
Laboratory, Adelaide, Australia, 3
Departement des Microscopies, Tours,
France and 4
Laboratoire de Biologie des Activités Physiques et Sportives,
Clermont-Ferrand, France
Excessive alcohol consumption has deleterious effects on many organs
including bone tissue. Alcohol abuse decreases bone mineral density, due to
osteoblast and osteoclast activity changes. It has been shown that alcohol
induces osteocyte apoptosis and marrow fat excess, but their relationship is
not well understood. The aim of this in vivo study was to assess the relation-
ship between the increase of adiposity in the marrow and micro-vessels and
osteocyte apoptosis in the case of alcohol-induced bone loss. A total of 12
eight-week-old male Wistar rats were treated with 35% v/v ethanol in the
drinking water during 17 weeks (A) and compared with 12 controls (C). At
the end of the study, we measured bone mineral density by absorptiometry.
Osteocyte morphology was assessed with transmission electron microscopy
on ultra-thin tibia sections. Osteocyte apoptosis was assessed through
caspase-3 and toluidine blue staining. Fat bone marrow and cortical bone
micro-vessels content were evaluated on semi-thin sections stained with
toluidine blue. Chronic alcohol consumption induced bone loss leading to
osteoporosis. After alcohol treatment, the number of apoptotic osteocyte was
increased, as shown by the greater caspase-3 staining and the higher empty
osteocyte lacunae in A vs. C. Lipid droplets accumulation were observed
within the osteocytes, the bone marrow and the cortical bone micro-vessels
in A vs. C. An inverse correlation was noted between BMD and osteocyte
apoptosis (r = −0.72, P = 0.002) and strong significant correlations were
showed between osteocyte apoptotic number and lipidic droplet accumu-
lation in osteocyte (r = 0.95, P < 0.001), bone marrow (r = 0.65, P < 0.02)
and bone micro-vessels (r = 0.83, P < 0.005). The bone loss observed follow-
ing the alcohol treatment was linked to high osteocyte apoptosis and lipid
accumulation in bone, which was correlated to increased bone marrow and
blood micro-vessels fat content.
O5.4
ALCOHOL INTOXICATIONS DURING ADOLESCENCE
INCREASE MOTIVATION FOR ALCOHOL IN ADULT RATS
M. Naassila1, V. Warnault2, R. Legastelois1, C. Vilpoux1, B. Botia1,
H. Houchi1, O. Pierrefiche1, J. Jeanblanc1, M. Kervern3 and S. Alaux1
1
Université de Picardie Jules Verne, Amiens, France, 2
Gallo Research
Center, Emeryville, CA, USA and 3
Centre de Psychiatrie & Neurosciences,
Paris, France
Background. Recent studies suggested that individuals who start drinking at
an early age are more likely to develop alcohol dependence. We analyzed in
Sprague–Dawley rats the consequences of intermittent ethanol (EtOH)
intoxications (IEI) during adolescence on later motivation to consume
EtOH.
Methods. Every 2 days, rats received an EtOH injection (3.0 g/kg) for 2 con-
secutive days and across 14 days. The adolescent period window (adolescence
vs peri-adolescence/puberty), frequency of EtOH exposure (every 2 days vs
every day) and specificity towards EtOH rewarding effects (ethanol or
amphetamine-induced place preference) were analyzed. Susceptibility to
relapse, sensitivity to motivational properties of EtOH and basal level of
anxiety-like behaviour were tested. An array of neurotransmitter-specific genes
was created to assess persistent gene expression alterations in adult brain.
Results. We found that IEI specifically during adolescence and including
2-day periods without EtOH (‘withdrawal’) increased later preference and
i32 ESBRA 2011 ABSTRACTS
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