Effect of Ethanol Deprivation and Re-Exposure on the Ethanol Drinking Behaviour of the High-Alcohol-Drinker (UChB) Rats

doi:10.4236/jbbs.2011.11001

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Journal of Behavioral and Brain Science, 2011, 1, 1-5

doi:10.4236/jbbs.2011.11001 Published Online February 2011 (http://www.SciRP.org/journal/jbbs)

Effect of Ethanol Deprivation and Re-Exposure on the

Ethanol Drinking Behaviour of the High-Alcohol-Drinker

(UChB) Rats

Lutske Tampier, Maria Elena Quintanilla

Molecular and Clinical Pharmacology Program, Institute of Biomedical Sciences, Faculty of Medicine,

University of Chile, Santiago, Chile

E-mail: [email protected]

Received January 4, 2011; revised February 20, 2011; accepted February 22 , 20 11

Abstract

Alcohol addiction constitutes a major health problem in the general population, it is a complex pathology

characterized by the development of tolerance, physical dependence and compulsive ethanol-seeking behav-

iour that often manifests as a chronic relapsing syndrome. 0ne of the major concerns in the treatment of al-

cohol-dependent patients is the prevention of relapse during periods of abstinence. The alcohol deprivation

effect (ADE) is defined as a temporary increase in the voluntary intake of ethanol when it is reinstated after a

period of alcohol deprivation and has been used as an animal model of relapse-like drinking. ADE can be

used to evaluate the efficacy of possible pharmacological agents to prevent relapse drinking. The current

study was undertaken to examine whether the high-alcohol-drinker UChB rats would display an ADE. Rats

were given either continuous or periodic concurrent access to 10%, 20% (vol/vol) of ethanol across depriva-

tion cycles. UChB rats consuming ethanol voluntarily for two months, exhibit a robust ADE after a single

deprivation period of two weeks. The increased alcohol intake during the early days of re-exposure, follow-

ing a withdrawal phase, is attributed to a shift in preference towards the higher concentration of ethanol that

might reflect an increase in craving for alcohol. Since an ADE is also observed in UChB rats, make this line

of rats selectively bred for their high voluntary ethanol consumption, a useful model for study the efficacy of

pharmacological agents for the treatment of relapse of alcohol consumption.

Keywords: Alcohol Intake, Alcohol Deprivation, Alcohol Drinking Rats, UChB Rats

1. Introduction

Addiction can be best defined as the loss of control over

drug use. I t is caused by the action of a drug of abuse and

generally requires repeated drug exposure. One of the

drugs more abused is alcohol. Alcohol addiction (Alco-

holism) constitutes a major health problem in th e general

population, it is a complex pathology characterized by

the development of tolerance, physical dependence and

compulsive ethanol-seeking behaviour that often mani-

fests as a chronic relapsing syndrome. As a matter of fact,

one of the major concerns in the treatment of alco-

hol-depe ndent patients is the pre vention of rela pse during

periods of abstinence.

According to Spanagel and Hölter (1999) [1], alcohol

drinking over a long period of time can be separated into

three stages. First, the initiation or acquisition of alcohol

drinking, followed by a second stage of controlled alco-

hol-drinking behaviour and eventually a third stage

where uncontrolled alcohol-drinking behaviour occurs,

leading to the development of dependence, which has

been characterized as an progressive dysregulation of

brain reward and stress circuits [2,3]. Although these

various stages of alcohol drinking occur in humans, it is

not clear whether those transitions in alcohol drinking

from one stage to another can also be observed in all

animals with a high preference an d a high daily intake of

alcohol Various approaches have been used to evaluate

the occurrence of addictive behaviour in laboratory ani-

mals; most of them based on changes in the amount of

alcohol intake following a period of abstinence and a

shift of preference towards higher concentration of etha-

nol solutions [1]. The alcohol deprivation effect (ADE)

has been proposed as a model of “loss of control” and

2 L. TAMPIER ET AL.

“relapse-like” drinking [4,5] and that can be used to

evaluate the efficacy of possible pharmacological agents

to prevent relapse drinking. ADE is defined as a tempo-

rary increase in the ethanol preference and in the volun-

tary intake of ethanol solutions over baseline drinking

conditions, when alcohol is reinstated after a period of

abstinence [6]. The negative consequences of withdrawal,

trigger further drinking and has been hypothesized to be

an animal model of alcohol craving [5]. An ADE and the

modification of ethanol intake by exposure to repeated

cycles of alcohol availability and deprivation has been

demonstrated in the selectively bred alcohol-preferring

(P) and high-alcohol-drinking (HAD) line of rats [4,7,8]

but not in Sardinian alcohol preferring (sP) rats [9].

However the effects of repeated deprivations or access to

different ethanol concentrations have not been tested in

the high-alcohol-drinking (UChB) rats, selectively bred

in this laboratory for high volun tary ethano l consumption

using the two-bottle choice (10% v/v ethanol and water)

paradigm [10,11]. The UChB rats slowly increased their

intake of ethanol over 20 days in the free choice condi-

tion to a steady state point reaching 7-9 g ethanol per kg

body weight per day. The gradual increase in the ethanol

consumption displayed by the UChB rats across the time

appeared to be the results of neuroadaptative changes

subsequent to chronic intake. The objective of this study

is to examine whether UChB rats would display an ADE

and a shift in ethanol concentration preference from

lower concentrations (10%) to higher concentration

(20%) of ethanol across deprivation cycles.

2. Animals and Methods

2.1. Animals

The animals were female UChB rats from the 82nd gen-

eration (bred at Schoo l of Medicine, University of Chile).

At three months old, rats were housed individually in

cages where they received concurrent free access to 10%

(v/v) , 20% (v/v) ethanol and water and solid food ad

libitum for 8 weeks Rats were maintained on a 12-hr

reversed light/dark cycle (lights off at 19.00 hr). During

the duration of testing, animals were weighted weekly.

All procedures used in this study were in compliance

with the Chilean Ethical animals rules.

2.2. Experimental Procedure

After the period of 8 weeks under the concurrent free

choice condition between 10%, 20% ethanol and water

paradigm, nine animals were deprived from ethanol for

15 consecutives days (alcohol deprivation period). After

the first deprivation period, rats received a continuous

free-choice access to both ethanol solutions (10%, 20%)

and water for an additional two-week period and ethanol

and water consumption recorded every 24 hours (1st

ADE). After these 2 weeks of ethanol re-exposure, rats

were again deprived of ethanol for 2 additional weeks,

followed by 2 weeks of ethanol exposure (2nd ADE).

This cycle was again repeated (3rd ADE). Results are

expressed as grams of total ethanol intake related to kg-

body weight per day, and as grams of ethanol of each

ethanol solution (10% or 20%) consumed per kg- body

weight per day. Total water intake (water present in

ethanol solution plus water from the water tube) was

expressed as ml/kg/day). Base data were the average of 7

days immediately before deprivation. An additional

group of nine rats was never deprived of ethanol and

served as non deprived controls and their consumption

were evaluated at 8 weeks and at 18 weeks of continuous

ethanol consumption.

2.3. Statistical Analysis

Data is expressed as the mean SEM of each condition.

To determine whether consumption of total ethanol or for

a particular ethanol solution was increased after ethanol

re-exposure, individual repeated measures ANOVA were

performed separately on each of the deprivation groups

and compared to the base data of the non deprived group.

All post hoc comparisons were performed by using the

Neuman-Keuls test. P < 0.05 was considered statistically

significant.

3. Results

Effect of Deprivation on Post-Daily Ethanol In-

take by UChB Rats

After the first deprivation period (1st ADE), UChB rats

displayed a significant increase in the total amount of

ethanol consumed during ethanol re-exposure (Figure 1).

Studies over a period of 24 hours a day showed a sig-

nificant increase in ethanol intake (F(6,69) = 8.55; p <

001). A post hoc analysis revealed that the mean daily

ethanol intake of the rats was significantly higher during

the initial 2 days after re-exposure, compared to base

ethanol intake. Also after the second deprivation period

(2nd ADE) and third deprivation period (3rd ADE) rats

significantly increased their total ethanol consumption

after ethanol re-exposure (F6,69) = 9.74; p < 0.001 and

(F(6,69) = 7.21; p< 0.001 respectively (Figure 1). The

post hoc analysis revealed a significant increase of etha-

nol intake following the first 2 days of re-exposure (Fig-

ure 1). No significant statistical difference in the total

ethanol intake, analyzed by the one way ANOVA

(F(3,165) = 0.3 was observed between 1st, 2nd or 3rd

ADE (Figure 1). Each time ethanol intake returns to the

L. TAMPIER ET AL.

baseline around 7 .5 g/kg/day. Total water intake (su m of

water from ethanol solution plus water from the water

tube) about 100 to 120 ml/kg body weight/day, remains

stable during the entire experimental period.

The patterns of the two ethanol solutions, consumed dur-

ing 24 hours of consumption are depicted in Figure 2.

The intake of 20% ethanol increased after the ethanol

re-exposure (F(6, 69) = 3.62; p < 0.005). The post hoc

analysis revealed that the amount of 20% ethanol solu-

tion increased during the initial 2 days of ethanol re-ex-

posure. Whereas, the amount of the 10% ethanol solution

did not show a significant difference (F6,69) = 2.8; n.s.

The preference for the 10% ethanol solution remains

stable. The increase in ethanol consumption after the 2nd

ADE and 3rd ADE was also due to a shift in the prefer-

ence for the more concentrated 20% ethanol solution

(F6,69) = 6.94; p < 0.001 and (F6,69) = 4.64; p <

0.001) respectively. The post hoc analysis revealed that

the intake of the 20% ethanol solution is significantly

higher during the first two days of ethanol re-exposure

(Figure 2). Rats show a preference for the more concen-

trated ethanol solution. Overall there was a significant

increase in ethanol intake and an effect of ethanol con-

centrations.

The total ethanol consumption and the relative intakes

of the 10 and 20% ethanol solutions and water, in the

non deprived UChB rats, under a 24-hr free-choice con-

ditions appears in Table 1 . Each value is the mean of th e

last seven consecutive days of the period of 8-weeks of

*Significantly higher than base data.

Figure 1. The mean (SEM) total amount of ethanol con-

sumed (g/kg/day) by UChB rats under 24-hr free-choice

condition, deprived of ethanol for 2 weeks and then sub-

jected to one to three cycles of 2 weeks of ethanol drinking

and 2 weeks of deprivation (1st, 2nd and 3rd ADE).

*Significantly higher than the base data of the 20% ethanol

solution.

Figure 2. The mean (SEM) of ethanol 10% and 20% con-

sumed (g/kg/day) by UChB rats under 24-hr free-choice

condition, deprived of ethanol for 2 weeks and then sub-

jected to one to three cycles of 2 weeks of ethanol drinking

and 2 weeks of deprivation (1st, 2nd and 3rd ADE)

Table 1. Ethanol, water and total water intake in non de-

prived UChB controls rats at two periods of continuous

access to a 10%, 20% v/v ethanol solution and water.

After 8 weeks 18 weeks

Ethanol intake g/kg/day g/kg/day

Total ethanol 7.35  0.44 7.80  0.5

10% ethanol 6.36  0.42 6.00  0.31

20% ethanol 0.99  0.19 1.36  0.50

Water intake ml/kg/day ml/kg/day

Water 25  5 20  8

Total water intake118.1  6.7 116.6  7.8

ethanol free choice consumption and the last seven con-

secutive days after a period of 18 weeks of continuous

ethanol consumption.

The non deprived group that was allowed to have 24-hr

continuous access to ethanol during all the 18 weeks of

the experiments never approached intakes as seen after

4 L. TAMPIER ET AL.

each deprivation. Also, the base ethanol intake (g/kg/

day), the average of the 7 days immediately before dep-

rivation did not change as a result of the deprivation pe-

riods. Data analyzed by one-way ANOVA indicate no

significant difference in the total ethanol intake (F2,29)

= 0.02; ns; 10% ethanol intake (F2,29)=0.27; ns and

20% ethanol intake (F2,29) = 0.19; ns between base

data.

4. Discussion

UChB rats consuming ethanol voluntarily for two

months, exhibit a robust ADE after a single deprivation

period of two weeks. The increased alcohol intake during

the early days of re-exposure, following a withdrawal

phase, is attributed to a shift in preference towards the

higher concentration of ethanol that might reflect an in-

crease in craving for alcohol, which returns to baseline

after a period of two day s .

Examining the effects of the second or third depriva-

tion on ethanol intake, during the initial 24-hr period

after the reinstatement of ethanol, the second and third

deprivation cycle did not enhance the amount of ethanol

intake above that was attained with the first deprivation.

This could be the result of a ceiling effect (i.e. rats are

drinking as much as possible under the experimental

conditions). Repeated ADE evoked a drinking pattern in

UChB rats very similar to that ob serv ed in P rats [8]. The

increase in the amount of ethanol consumed was not the

result of an indiscriminating polydipsia generated by the

period of abstinence. The total fluid intake by rats (sum

of water from ethanol solution plus water from the water

tube) was not altered during all the period of the study.

The effects observed after a single deprivation or re-

peated deprivations were not observed in the non de-

prived control group, suggesting that chronic exposure to

ethanol alone is not sufficient to produce such a marked

increase in ethanol intake or a shift in preference for

higher concentrations of ethanol.

Uncontrolled alcohol drinking behavior can be as-

sessed after prolonged drug self-administration in some

individuals, and similar to those humans, some rats se-

lectively bred for their high voluntary ethanol consump-

tion can also develop addiction-like behaviours, sug-

gesting that specific genetic factors are involved in the

transition between high ethanol intake and development

of an alcohol deprivation effect [12,13]. Therefore, the

results suggest that neurobiological alterations may be

occurring during time and these alterations result in a

marked increased of ethanol consumption and to the de-

velopment of uncontrolled alcohol-drinking behavior.

Another approach to evaluate the occurrence and the

degree of addiction in laboratory animals are based on

changes in alcohol intake by making drug seeking con-

sistently resistant to interfering factors like concurrent

choice of preferred flavours such as saccharin or sucrose

[14]. This condition can also been ob served in UChB rats.

The placement of a third bottle containing a highly pal-

atable 0.2% saccharin solution, decreased the ethanol

intake in UChB rats that were exposed for a period of 7

days to a 10% ethanol and water under free choice con-

dition, but not in UChB rats exposed to the 10% ethanol

solution for a period of 60 days [15]. Moreover, it is

known that addicts lost the ability to stop drinking and

continue seeking drugs in spite of adverse environmental

conditions. As a matter of fact, it has been previously

observed that under chronic ethanol intake UChB rats

leads to marked tolerance to the aversive effects of disul-

firam and cyanamide on ethanol intake despite the pres-

ence of consistently high levels of blood acetaldehyde

[16]. These results are in accord also to the notion that

UChB rats develop uncontrolled alcohol-drinking be-

haviour, because of its resistance to the aversive effects

produced by an increase in blood acetaldehyde levels.

There are several independent lines of evidence for

genetic contributions to vulnerability to alcoholism [17].

Genetics is just beginning to illuminate the variations

among individuals in their response to alcohol and one

question is what are the changes in the brain that underlie

the transition from contro lled to uncontro lled alcohol use.

According to Nestler [18,19], repeated exposure to a

drug of abuse alters the amounts, and even the types of

genes expressed in specific brain regions resulting in

alterations in molecular and cellular mechanisms that

underlie long-term associative memories in several fore-

brain circuits. Whereas some alcoholics can cease drink-

ing, others cannot. Despite multiple episodes of treat-

ment, and despite risk of significant life problems, re-

lapses to alcohol drinking is very frequent. As a matter of

fact, the alcohol deprivation effect (ADE) has been pro-

posed as an animal model of “loss of control” and “re-

lapse-like” drinking [4,5]. Since an alcohol deprivation

effect (ADE) is also observed in UChB rats make this

line of rats selectively bred for their high voluntary

ethanol consumption, a useful model for study the effi-

cacy of pharmacological agents for the treatment of re-

lapse of ethanol consumption.

5. Conclusions

Relapse to alcohol use after prolonged withdrawal peri-

ods is the major problem in the treatment of alcohol de-

pendence in humans. However, there are relatively few

studies concentrated on elucidation of the neurochemical

events underlying relapse to alcohol. The ADE is a use-

ful model for studying alcohol relapse in both rats and

L. TAMPIER ET AL.

mice. Genetics, stress, and environmental cues are all

important factors that influence relapse. Long-term al-

terations in neuronal activity within the glutamatergic

and dopaminergic/serotoninergic systems, appears to

underlie alcohol relapse.

6. Acknowledgements

We thank Mr. Juan Santibáñez for the expert care of the

animals.

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