Alcohol, Vol. 19, No. 1, pp. 15–22, 1999© 1999 Elsevier Science Inc.

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15

Sex Differences in Alcohol Drinking Patterns During Forced and Voluntary Consumption

in Rats

JORGE JUÁREZ AND ELIANA BARRIOS DE TOMASI

Institute of Neurosciences, University of Guadalajara, Guadalajara, Mexico

Received 2 December 1998; Accepted 10 December 1998

JUÁREZ, J. AND E. BARRIOS DE TOMASI.

Sex differences in alcohol drinking patterns during forced and voluntaryconsumption in rats.

ALCOHOL

19

(1) 15–22, 1999.—Wistar rats were studied during forced and voluntary alcohol consump-tion, and continuous or periodic access to ethanol (6%) v/v with different availability of fluids. Absolute volume of alcoholconsumption was not different between sexes in any condition; however, females consumed significantly more alcohol thanmales on a g/kg basis in all conditions. These differences were significantly more extensive during continuous free-choice toalcohol and water than during forced alcohol consumption. Females showed greater alcohol preference than males only dur-ing continuous free-choice to alcohol and water. During periodic free-choice to alcohol and water condition, alcohol con-sumption was distributed during more hours throughout the day in females than males. During periodic free-choice to alco-hol and to an isocaloric sweetened solution (ISS), intakes of ISS were very high compared to regular intakes of daily water;nevertheless, alcohol consumption was maintained to similar levels observed in continuous free-choice to alcohol and waterand represented almost 50% of regular daily consumes of water in males and females. Free-choice for alcohol and ISS modi-fied the usual pattern of alcohol consumption during the daily light-dark cycle in males and females and reduced the time de-voted to drinking alcohol compared to other conditions, in which similar intakes were observed. Results show that the extentof the higher alcohol consumption in females than males and the changes in patterns of alcohol intake were dependent on thenature of the ingestion schedule. © 1999 Elsevier Science Inc. All rights reserved.

Sex differences Patterns of alcohol Voluntary consumption Forced consumption Ethanol

GENDER has been extensively studied as a factor that influ-ences alcohol consumption: It has been reported that menconsume alcohol more frequently, and larger amounts of al-cohol, than women (2,4,5,11,19,22); however, experimentalstudies describe higher consumption of alcohol in femalesthan in males (1,3,12,15,16,20). There are sex differences re-lated to pharmacokinetics of alcohol that may influence dif-ferentially drinking patterns of alcohol. First, metabolic rateof alcohol seems to be higher in females than males (18,21,23,25). Second, rates of first-pass alcohol metabolism, relatedto gastric alcohol dehydrogenase, are lower in women than inmen (9,13,14). Third, it is well known that females have lessbody water than males; this is important because ethanol dis-tributes from the blood into the tissues and fluids in propor-tion to their relative contents of water; therefore, volume of

distribution of ethanol in the body is equivalent to the totalbody water (13). Last, there are data suggesting that neuro-esteroid activity may play a role in producing differential alco-hol effects in males and females (14).

Studies on sex differences in alcohol consumption gener-ally analyze mean volume of ethanol intake; however, it hasbeen described that although this measure is a useful sum-mary for illustrating gender differences, it is important to con-sider several components of ethanol intake such as, frequencyof drinking, quantity of drinks consumed per occasion, andethanol content per drink, before interpreting the sex differ-ences (6). This consideration is intimately related to patternsof alcohol consumption, which can change from the time fromthe first exposure to alcohol to the establishment of a likingfor alcohol. It is well known that differences between occa-

Requests for reprints should be addressed to Dr. Jorge Juárez G., Instituto de Neurociencias, Universidad de Guadalajara, Calle Rayo 2611,Col. Jardines del Bosque, Guadalajara, Jal. C.P. 44520, México. Tel/Fax:

1

52-3-647-77-76; E-mail: jjuarez@udgserv.cencar.udg.mx

16 JUÁREZ AND BARRIOS DE TOMASI

sional drinkers and alcoholics are not only based on theamount of consumed alcohol but also in the distribution ofdrinking behavior along the time. Therefore, sex differencescan be expressed not only by the amount of alcohol consump-tion but also by patterns of alcohol intake. Change in one ofthese two measures does not necessarily presuppose a changein the other. The support of this assumption is one of the ob-jectives in the present study. An additional factor to consider,in relation to sex differences, is the volitive characteristic ofalcohol consumption (i.e., it has been described that forced, incontrast with voluntary ethanol consumption, can significantlyaffect the alcohol intake in rats) (1). Therefore sex differencesin patterns and amount of alcohol consumption could changedepending on forced versus free-choice access to alcohol,which to the best of our knowledge, have not been fully stud-ied.

The purpose of the present study was to analyze sex differ-ences in patterns of alcohol consumption and in the amount ofalcohol intake during forced and voluntary conditions of ex-posure to alcohol.

METHOD

Wistar rats, 8 males and 8 females, were used. Before be-ginning the study, subjects had not had any experience in theintake of any other liquid besides to tap water. Subjects weremaintained on a 12-12-h light-dark cycle, lights on at 8:00

A

.

M

., and food pellets were available ad libitum during allconditions of the study. After weaning, rats of the same sexwere placed in collective cages of four rats each. One weekbefore the study was started, subjects were placed in individ-ual cages.

Starting at 90 days of age, rats were individually exposed todifferent drinking conditions, which will be described in thefollowing sections. With the purpose of studying the patternsof alcohol consumption during the day and its possible modifi-cation along the different drinking conditions, the subjectswere periodically tested in a lickometer system (MED Associ-ates, Inc.), which accurately records the rate of licking of sip-per tubes attached to bottles containing liquid. Temperature,feeding, and light-dark cycle were maintained identical in allconditions. Total consumption of liquids was measured ineach condition, both during availability of a sole liquid andduring free-choice to two fluids. Body weight was recordedperiodically.

Lickometer System Test

On the lickometer test days, subjects were individuallyplaced in cages with bottles attached to the lickometer systemduring 24 h, starting off at 10:00

A

.

M

. each day. Dimensions ofthese cages were similar to the home cages. Frequency of licksto drinking spouts were accumulated and recorded each hourto study the patterns of alcohol consumption during the day.With these data, time distribution of different intervals of ac-cess frequency/hour to drinking spouts was studied in malesand females. Considering the changes in activity dependingon the daily light-dark cycle, the hours of each day were di-vided and analyzed in two phases: light phase from 7:00

A

.

M

.to 7:00

P

.

M

., and dark phase from 7:01

P

.

M

. to 6:59

A

.

M

. Theanalyzed intervals of licking frequency/hour were the follow-ings: 0–20, 21–150, 151–300, 301–450, 451–600, 601–750, andmore than 750 accesses/h. We have observed that very few re-corded accesses to sipper tubes occur by accidental contact orexploratory behavior; therefore, the interval of 0–20 licks/hwas considered as no access to drinking spouts.

Continuous Access to Alcohol (Forced Consumption)

At 90 days of age males and females were continuously ex-posed during 30 days to a solution containing 6% ethanol(99.8%, Merck) v/v, water and 2 g of sucrose/100 ml (ETOH6%). This solution was the only available liquid in their homecage, and it was measured daily at the same hour. Subjectswere tested on the lickometer one day per week under thesame continuous exposure to ETOH 6%. Therefore, eachsubject was tested four days on the lickometer during this firstcondition.

Continuous Free-Choice to Alcohol and Water

After 30 days of continuous access to alcohol, one bottlecontaining tap water was added to the cage, therefore, free-choice to ETOH 6% and water was permitted continuouslyduring eight days. Each subject was tested two days on thelickometer, at beginning (day 2 or 3) and at final (day 7 or 8)of this period under the same free-choice availability of liq-uids. Consumption of both liquids was measured daily at thesame hour.

Periodic Free-Choice to Alcohol and Water

Immediately after the last day of the previous condition,subjects were periodically deprived of alcohol solution: threedays of alcohol deprivation for one day of free-choice toETOH 6% and tap water. In the days of alcohol deprivation,tap water was available ad libitum. This condition lasted twoweeks; therefore, rats were exposed to both liquids two daysper week. On two of the four days of free choice to alcohol andwater, each subject was tested on the lickometer (one day perweek), and the patterns of alcohol along the day were studied.Liquid consumption was measured daily at the same hour.

Periodic Free-Choice to Alcohol andIsocaloric-Sweetened Solution

This condition was equal to the previous one in all re-spects, except that after three days of alcohol deprivation,free-choice to ETOH 6% and to an isocaloric sweetened solu-tion (ISS) (sucrose and water) was available. In the days ofethanol deprivation, tap water was available ad libitum. Thiscondition lasted two weeks; therefore, free-choice to alcoholand ISS was offered in four days. On two of these four days,each subject was tested on the lickometer (one day per week)and the patterns of alcohol along the day were studied. Liquidconsumption was measured daily at the same time.

Statistical Analyses

In the condition where only alcohol was available, ANOVAsex

3

week was used for the amount of consumption in mland an ANOVA sex

3

phase for the frequency of accesses todrinking spouts. When two liquids were available simulta-neously, ANOVA sex

3

liquid

3

day was used for theamount of consumption in ml and an ANOVA sex

3

liquid

3

phase for the frequency of accesses to drinking spouts. In thislast case, the average of accesses in the days that subjectswere tested in the lickometer system was considered. Distribu-tion in time of the different intervals of licking frequency/hourto drinking spouts was analyzed by an ANOVA sex

3

interval

3

phase. Tukey Student’s

t

-test was used to compare pairs. A

p

value of

,

0.05 was considered statistically significant.

SEX DIFFERENCES IN ALCOHOL DRINKING PATTERNS 17

RESULTS

Continuous Access to Alcohol (Forced Consumption)

The daily volume of alcohol consumption was averaged foreach of the four weeks that lasted this condition. There werenot significant differences in the absolute volume of alcoholconsumption, neither between males an females nor betweenweeks. However, analyses of grams of alcohol/kg of bodyweight showed higher mean daily consumption of alcohol infemales (8.7 g/kg) than in males (5.5 g/kg), (

t

(14)

5

4.63,

p

5

0.0004) (Fig. 1A, 1B).During the four weeks of continuous access to alcohol,

subjects were tested four days on the lickometer system, oncea week. In these four days, access frequency to drinkingspouts was higher in the dark phase than in the light phase re-gardless of sex [

F

(1, 26)

5

6.93,

p

5

0.013]. Analysis of the dis-tribution of alcohol drinking behavior along the day for eachof these 4 days (Table 1) showed significant differences be-tween intervals of licking frequency/h (rate). On these days,

the interval that represented absence of access to alcohol con-sumption (0–20), was more frequent along of the 24 dailyhours than other intervals, (Table 1): Day 1 [

F

(6, 72)

5

52.55,

p

,

0.001], day 2 [

F

(6, 72)

5

62.06,

p

,

0.00001], day 3 [

F

(6,72)

5

43.93,

p

,

0.00001] and day 4 [

F

(6, 72)

5

42.03,

p

,

0.001]. The mean percentage of daily time devoted to drink-ing alcohol, regardless of sex, was 49.3, 48.3, 48.0 ,and 47.4 forweeks 1,2,3, and 4, respectively. The intervals that indicateddrinking behavior (21–150 access/h and higher) did not showdifferences between them; however, the more usual rates ofdrinking behavior observed in this period were 21–150, 151–300, and 301–450 access/h, with a mean time of 9.94%,11.15%, and 10.2%, respectively (Table 1). Interactions be-tween interval and phase were significant in all weeks: week1, [

F

(6, 72)

5

16.23,

p

,

0.001]; week 2, [

F

(6, 72)

5

15.03,

p

,

0.00001]; week 3, [

F

(6, 72)

5

14.39,

p

,

0.00001] and week 4,[

F

(6, 72)

5

17.41,

p

,

0.001]. These interactions showed thatabsence of alcohol intake occurred more time in the lightphase than in the darkness. Although drinking behavior oc-curred more frequently in the darkness than in the light phase,differences between phase were not significant (Table 1).

Free-Choice to Alcohol and Water (Continuous Exposure)

Analysis for the eight continuous days of free-choice to al-cohol and water showed that subjects consumed higher vol-ume of water than of alcohol regardless of sex and days[

F

(1, 11)

5

23.07,

p

,

0.001]; however, interaction sex

3

liq-uids

3

days [

F

(7, 77)

5

4.49,

p

,

0.001] showed that althougha tendency to consume more water than alcohol was observedin females, there were not significant differences between theconsumption of both liquids in no one day (Fig. 2A). In con-trast, males, showed significant higher consumption of waterthan of alcohol in all days of this condition with the exceptionof the two initial days (Fig. 2B). There were no sex differencesin the total volume of alcohol consumed in this period; how-ever, alcohol consumption g/kg of body weight, per day, wassignificantly higher in females (5.07 g/kg) than in males (1.93g/kg), [

F

(1, 11)

5

28.72,

p

5

0.0004], (Fig. 1A, 1B). Duringthis period of eight days, each subject was tested two days onthe lickometer, but the data of the initial day in females waslost for technical problems. Thus, ANOVA sex

3

liquid

3

phase for the accesses to the drinking spouts, was performedonly for the final day. This analysis showed higher frequencyof accesses in females than in males regardless of liquids andphase [

F

(1, 13)

5

5.11,

p

5

0.04] and higher frequency of ac-cess to the drinking spouts in the dark phase than in the lightphase regardless of sex and available liquid [

F

(1, 13)

5

45.29,

p

,

0.001]. Distribution of alcohol drinking behavior, duringthis day, showed that absence of alcohol intake, representedby interval of 0–20 licks/h, occupied significantly more time(54.5%) than any interval of alcohol consumption [

F

(6, 84)

5

73.55,

p

,

0.00001] (Table 2). At the same time, the rate of20–151 access/h occupied significantly more time (18.15%)than higher rates of drinking behavior. Finally, absence of al-cohol intake occurred more time in the light than in the darkphase [

F

(6, 84)

5

19.61,

p

,

0.00001] (Table 2).

Periodic Exposure to Alcohol and Continuous Accessto Water

During this condition that lasted two weeks, subjects wereexposed to continuous access to water and periodic access toalcohol two times a week. Three days of alcohol deprivationelapsed between each exposure of free-choice to water andalcohol. ANOVA sex

3

liquid

3

days, for the four days of ac-

FIG. 1. Mean 6 SEM of alcohol intake in ml/day (A), alcohol intakein g/kg of body weight (B), and alcohol intake in % h/day (C). Bars showdifferences between males and females during forced consumption,continuous free-choice to alcohol and water (CFAW), periodic free-choice to alcohol and water (PFAW), and periodic free-choice toalcohol and to an isocaloric sweetened solution (PFAISS). Significantsex differences (*).

18 JUÁREZ AND BARRIOS DE TOMASI

cess to both liquids, showed that there were neither sex norliquid significant differences. However, the day’s main effectshowed that the volume of liquid consumed was higher onday 4 than in the first and the third day, and the consumptionon the second day was higher than the first day (

F

(3, 42)

5

9.31,

p

,

0.001). In this period, as during the previous condi-tions, there were no sex differences in the total volume of al-cohol consumed; however, mean daily alcohol intake g/kg ofbody weight was significantly higher in females (6.04 g/kg)

than in males (3.22 g/kg), (

F

(1, 14)

5

16.58,

p

5

0.001) (Fig.1A, 1B).

Of the four days of free-choice to alcohol and water, sub-jects were tested two days on the lickometer. Accesses to eachliquid in these two days were averaged and the ANOVA sex

3

liquid

3

phase showed higher frequency of access to liquidsduring dark phase than in the light phase [

F

(1, 14)

5

26.41,

p

5

0.0003]. There were no significant differences in the accessfrequency, neither in terms of sex nor of available liquid.Analysis of alcohol intake distribution, along the days of teston the lickometer (Table 3), showed significant differencesbetween access frequency intervals [

F(6, 84) 5 156.62, p ,0.00001]: Subjects drank 40.3% of total time, and the rate of21–150 access/h occupied significantly more time (16.6%)than higher rates. At the same time, rate of 151–300 occupiedsignificantly more time (10.1%) than rates over 450 accesses/h(Table 3). In this condition of periodic exposure to alcoholwith continuous access to water, females distributed their al-

TABLE 1CONTINUOUS EXPOSURE TO ALCOHOL AS ONLY AVAILABLE LIQUID (FORCED CONSUMPTION)

Weeks

1 2 3 4

Licks/hour L D T L D T L D T L D T T

0–20(no access) 33.1* 17.5 50.65† 34.1* 17.6 51.7† 34.9* 17.05 51.9† 35* 17.55 52.5† 51.71

21–150 4.51 5.62 10.13 3.51 5.67 9.18 3.54 5.01 8.55 4.74 7.18 11.92 9.94151–300 4.16 6.75 10.92 4.86 6.7 11.56 4.85 6.41 10.99 3.81 7.35 11.16 11.15301–450 3.29 7.18 10.47 2.67 7.42 10.1 3.02 9.53 12.55 2.77 4.86 7.63 10.18451–600 1.38 5.1 6.48 3.56 5.38 8.94 1.73 5.11 6.84 1.47 5.3 6.77 7.25601–750 1.73 3.89 5.62 0.32 2.74 3.06 0.71 3.02 3.73 1.33 4.33 5.66 4.51.750 1.38 3.71 5.09 0.91 4.31 5.22 1.21 3.97 5.18 0.81 3.08 3.84 4.83

Distribution in time (mean % hours/day) of different rates of alcohol drinking in the light (L) and dark (D) phases and themean % total (T) of hours/day

*L . D; Tukey test p , 0.01.†0–20 . higher rates; Tukey test p , 0.01.

FIG. 2. Mean 6 SEM of ethanol consumption (ml) duringcontinuous free-choice to alcohol and water in females (A) and males(B). Significant differences between alcohol and water (*), p , 0.05(Tukey test).

TABLE 2CONTINUOUS EXPOSURE TO FREE-CHOICE

TO ALCOHOL AND WATER

Licks/hour L (%) D (%) T (%)

0–20(no access) 36.16* 18.33 54.49†

21–150 8.83 9.32 18.15‡151–300 2.08 7.01 9.09301–450 0.07 5.45 5.52451–600 1.3 4.67 5.97601–750 0.28 2.34 2.62.750 0.54 2.86 3.4

Distribution in time (mean % hours/day) of dif-ferent rates of alcohol drinking in the light (L) anddark (D) phases and the mean % total (T) ofhours/day.

*L . D; Tukey test p , 0.01.†0–20 . higher rates; Tukey test p , 0.01.‡21–150 . higher rates; Tukey test p , 0.01.

SEX DIFFERENCES IN ALCOHOL DRINKING PATTERNS 19

cohol drinking behavior along more hours a day than males[F(6, 84) 5 5.24, p 5 0.00026] (Figure 1C). As in the otherconditions, subjects drank more during the dark phase than inthe light phase [F(6, 84) 5 17.15, p , 0.00001]. On the otherhand, number of hours with accesses to drinking spouts wasnot different between males and females during light phase, butfemales distributed their drinking behavior along more hoursduring the dark phase than did males [F(6, 84) 5 2.53, p 5 0.026].

Total Daily Liquid Consumption

In this condition of periodic free-choice to alcohol and water,subjects consumed a mean of 57.3 ml of liquid in the days of onlyaccess to tap water, and there were no sex differences. In days offree-choice to alcohol and water, a mean of 74.79 ml of total fluid(alcohol solution plus water) was consumed, and there were nosex differences; however, significant differences were observedbetween total fluid consumed in days of only water and that ofdays of access to both liquids [F(1, 26) 5 31.41, p 5 0.00005].

Periodic Free-Choice of Alcohol andIsocaloric-Sweetened Solution

During this condition that lasted two weeks, free-choice toalcohol and to an isocaloric sweetened solution was available

two days a week. The rest of the days tap water was availablead libitum. Three days with only water elapsed between eachexposure to alcohol and to ISS. ANOVA sex 3 solution 3days for the four days of simultaneous exposure to alcoholand sweetened solution showed the following significant dif-ferences: First, females showed higher liquid consumptionthan males regardless of solution and days [F(1,14) 5 23.5, p ,0.001]. Second, subjects showed higher consumption of sweet-ened solution than alcohol regardless of sex and days [F(1,14) 5182.45, p , 0.001]. Third, sweetened solution consumptionshowed a steady increase from day 1 to day 4 regardless ofsex, while alcohol consumption did not show significant varia-tions between days [F(3,42) 5 10.34, p , 0.001]. Last, femalesconsumed higher quantity of sweetened solution than males,but sex differences were not significant in the alcohol con-sumption [F(1,14) 5 24.54, p , 0.001]. As in the other condi-tions, there were no sex differences in the total volume of al-cohol intake; however, on a g/kg basis, the mean daily ofalcohol consumption was significantly higher in females (3.73g/kg) than in males (2.35 g/kg), [F(1,14) 5 18.26, p 5 0.001],(Fig. 1A, 1B).

In two of the four days that subjects were exposed to si-multaneous access to alcohol and to ISS, patterns of alcoholconsumption on the lickometer were studied. Analysis for ac-cess frequency to drinking spouts showed very similar resultsthan those previously described for the amount of fluids con-sumption in this same condition; therefore, only the more rel-evant data are described. Interaction sex 3 solution 3 phase[F(1,14) 5 16.45, p 5 0.0014] indicated that only the accessesto sweetened solution in females occurred more frequently inthe dark than in the light phase (Fig. 3). Therefore, the usualpattern of higher drinking behavior during dark phase waseliminated in the alcohol consumption in both sexes and inthe ISS intake in males.

Analysis of the distribution of alcohol drinking behaviorthroughout the day (Table 4) showed significant differencesbetween intervals [F(6, 84) 5 267.35, p , 0.00001]: Absenceof alcohol intake represented by interval of 0–20 licks/h oc-curred in 68.9% of the daily time (Table 4). Of the 31.1% ofthe time that subjects spent in drinking behavior, the intervalof 21–150 licks/h occupied significantly more time (18.3%)than higher rates (Table 4). Number of hours that subjectsspent in drinking behavior was higher in the dark than in thelight phase [F(6,84) 5 4.18, p 5 0.0012], but specific rates ofaccesses/h that indicated drinking behavior (21–150 and over)did not show differences between phases (Table 4).

Total Daily Liquid Consumption

Means of total daily liquid consumption, in this period,were the following: In the days of only access to tap water, fe-males consumed 51.7 ml, and males consumed 56.7 ml; thissex difference was not significant. However, sex differenceswere significant [F(1,28) 5 24.14, p 5 0.00012] in the days offree-choice to alcohol and sweetened solution: Females con-sumed a mean of 151.56 ml of total fluid (alcohol solutionplus ISS), and males consumed a mean of 98.72 ml. Differ-ences in the total fluid consumption between days of only wa-ter and days of free-choice to alcohol and to ISS also were sig-nificant [F(1,28) 5 144.62, p , 0.00001].

DISCUSSION

There were no sex differences in the absolute volume ofalcohol consumption in any condition, but when alcohol in-take was calculated in g/kg of body weight, females consumed

TABLE 3PERIODIC EXPOSURE TO FREE-CHOICE

TO ALCOHOL AND WATER

Licks/hour L (%) D (%) T (%)

0–20(no access) F 34.1 6 1.9 17.16 6 3.1 51.26*

M 38.2 6 1.8 29.9 6 2.1 68.1T 36.15‡ 23.53 59.68§

21–150 F 7.54 6 1.4 9.87 6 1.9 17.41M 5.45 6 1.5 10.41 6 1.9 15.86T 6.49 10.14 16.63¶

151–300 F 3.12 6 1.04 9.37 6 2.04 12.49M 3.38 6 1.00 4.41 6 1.3 7.79T 3.25 6.89 10.13#

301–450 F 1.82 6 0.6 6.75 62.6 8.57M 1.01 6 0.4 3.64 6 1.1 4.68T 1.43 5.19 6.62

451–600 F 2.34 6 0.9 3.12 6 1.04 5.46M 0.52 6 0.3 1.82 6 0.6 2.34T 2.86 2.47 3.9

601–750 F 0.52 6 0.3 1.82 6 0.6 2.34M 0.52 6 0.3 0.78 6 0.4 1.3T 0.52 1.3 1.82

.750 F 0.52 6 0.3 2.86 6 1.8 3.38M 0.52 6 0.3 0.52 6 0.3 1.04T 0.52 1.69 2.21

Distribution in time (mean % hours/day) of different rates of al-cohol drinking for males (M) and females (F) in the light (L) anddark (D) phases and the mean % total (T) of hours/day.

*F , M in the interval 0–20; Tukey test p , 0.01.†DM . DF in the interval 0–20; Tukey test p , 0.01.‡L . D in the interval 0–20; Tukey test p , 0.01.§0–20 . higher rates; Tukey test p , 0.01.¶21–150 . higher rates; Tukey test p, 0.01.#151–300 . rates above 450 licks/hour; Tukey test p, 0.01.

20 JUÁREZ AND BARRIOS DE TOMASI

significantly more alcohol than males in all conditions. Thesedifferences were significantly more extensive during continu-ous free-choice to alcohol and water than during continuousexposure to alcohol as sole available fluid, which indicateddifferences between voluntary and forced conditions of alco-hol consumption. This difference could be explained in thefollowing terms: In forced consumption, subjects had to sat-isfy their daily basic liquid requirements by means of the onlyavailable alcohol solution. These liquid requirements seem tobe similar in males and females at this age because there areno sex differences in daily water consumption. Therefore, sexdifferences in alcohol intake, during forced consumption,seem to depend on the normal sex differences in body weightand not on differences in the volume of liquid consumed. Onthe other hand, in free-choice condition where alcohol was si-multaneously available with water, the daily liquid require-ments could be satisfied by nonalcoholic fluid. Therefore, ifalcohol intake occurred voluntarily and sex differences weremore extensive than in forced consumption, an additionalsupport is given to the existence of sex differences in alcoholconsumption. Differences in alcohol intake between forcedand free-choice consumption it has been reported in relationto different strains (1), but, to the best of our knowledge, noevidence has been compiled in relation to sex differences insubjects tested in different conditions. Higher alcohol con-sumption in females than males has been supported in otherstudies (1,15,16,20). It is possible that these sex differencescould be related to sex differences in the pharmacokinetics ofalcohol (e.g., it has been described that alcohol metabolic rateis higher in females than in males) (18,21,23,25). Therefore, ifalcohol elimination is faster in females, they would need drinkmore amount to maintain a given blood alcohol level in time.Sex hormones after puberty, mainly testosterone, seem playan important role in these sex differences; that is to say, cas-tration in males produce an increase in the ethanol metabolicrate, but ovariectomy does not produce any significant changein females. When these males and females, previously gonad-ectomized, are treated with testosterone, a decrease in the al-cohol metabolic rate is observed (21).

The highest consumption, in voluntary alcohol intake, wasfound during periodic free-choice to alcohol and water, which

suggests that alcohol deprivation for a few days is sufficient toincrease alcohol intake in males and females. It has been de-scribed that number and duration of accesses per ethanoldrinking bout are increased when the number of access peri-ods per day is decreased in male rats (8), thus patterns of al-cohol intake can be modified when opportunity to obtain al-cohol is limited. It is possible that high levels of alcoholconsumption showed by subjects in the periodic free-choiceto alcohol and water could have been influenced, at least par-tially, by the previous five weeks of experience drinking alco-hol; however, it has been reported lower levels of alcohol con-sumption in subjects that previously had been exposedcontinuously to alcohol during 30 days than subjects that hadbeen exposed to alcohol only once (15).

During free-choice to alcohol and sweetened solution,males and females consumed a great amount of ISS. Thispreference for ISS was higher in females than in males andcan be explained by the well known preference for sweetenedsolutions in females compared to males (24,26). It is notewor-thy that nevertheless this great preference for sweet, femalesdrank higher levels of alcohol (3.73 g/kg of b.w.) than didmales in any condition of voluntary consumption. Males alsoshowed a clear preference for ISS compared with water; nev-ertheless, they drank higher amounts of alcohol, in this condi-tion, than the observed during continuous free-choice to alco-hol and water. Although ISS intake was very high comparedto normal daily water consumption, alcohol solution intake,in the same condition, meant almost the 50% of regular dailyvolume of water consumption. Therefore, a liking for alcoholcan not be explained by either a need to satisfy liquid require-ments or a need to taste properties of alcohol solution. Thissuggests that seeking alcohol probably was directly related toits pharmacological effects.

Ethanol preference is defined as a ratio calculated fromthe amount of ethanol consumed, divided by the amount oftotal fluid consumed in a test of free-choice intake (15). Obvi-ously, changes in the total fluid consumption can modify thisratio although the volume of ethanol remain constant; there-fore, it is possible to find differences in ethanol preference,but not in real ethanol consumption. This condition has beendescribed in alcohol-preferring (P) rats (17) and in a newhigh-ethanol-preferring (HEP) strain of rats (20), in which

FIG. 3. Mean 6 SEM of the rate of licking of sipper tubes attachedto bottles containing alcohol (Alc) or an isocaloric sweetenedsolution (ISS) in females and males during the light and the darkphase. Significant differences between phases (*), p , 0.05 (Tukeytest).

TABLE 4PERIODIC EXPOSURE TO FREE-CHOICE

TO ALCOHOL AND TO ISS

Licks/hour L (%) D (%) T (%)

0–20(no access) 37.46* 31.47 68.93†

21–150 8.30 9.99 18.29‡151–300 1.95 4.53 6.48301–450 1.30 2.08 3.38451–600 0.05 0.91 0.96601–750 0.39 0.52 0.91.750 0.52 0.39 0.91

Distribution in time (mean % hours/day) of dif-ferent rates of alcohol drinking in the light (L) anddark (D) phases and the mean % total (T) ofhours/day.

*L . D; Tukey test p , 0.01.†0–20 . higher rates; Tukey test p , 0.01.‡21–150 . higher rates; Tukey test p , 0.01.

SEX DIFFERENCES IN ALCOHOL DRINKING PATTERNS 21

the proportional consumption of ethanol declines because ofthe high consumption of very palatable solutions, but rats donot modify substantially their alcohol intake. In the presentstudy, a similar condition was observed during the period offree-choice to alcohol and sweetened solution: Ethanol pref-erence was higher in males than in females, although the vol-ume of alcohol consumed was very similar in both genders. Inthis case, sex differences in alcohol preference were due tothe higher intake of sweetened solution in females than inmales. In contrast, during continuous exposure to free-choiceto alcohol and water, females showed higher ethanol prefer-ence than males. In this case, the amount of fluid consump-tion/day was similar in both genders, so these sex differenceswere due to a real ethanol preference. These sex differencesapparently depended on a continuous exposure to alcohol, asthe same availability of liquids under a schedule of periodicexposure did not produce these effects.

Study of drinking behavior distribution during the day wasuseful to analyze sex differences in patterns of alcohol intakebeyond the measures of amount of alcohol consumption.Drinking behavior was significantly more frequent during thedark phase than in the light one in most conditions studiedand in both sexes, which is not surprising because the greateractivity in rodents occur during the dark phase. During free-choice to alcohol and sweetened solution, this higher fre-quency of consumption, observed during the dark phase, didnot occur either, for alcohol accesses in males and females norfor accesses to ISS in males. These data suggest that tasteproperties of available solutions can modify not only theamount of alcohol consumed but also the cyclic usual patternsof alcohol consumption in males and females.

Analyses of distribution of alcohol drinking behavior dur-ing the day showed that during forced condition of alcohol in-take, subjects had access to drinking spouts only in 48.6% h/day and the more usual rates of drinking were comprised be-tween 21 to 450 access/h. During continuous free-choice to al-cohol and water, subjects had access to alcohol drinkingspouts in 46% h/day, but the more frequent rate observed wasof 21–150 access/h. Therefore, the only difference betweenthese two conditions was the rate of licks/hour. This indicatedthat subjects consumed different amount of alcohol in forcedand free-choice conditions, but this consumption was distrib-uted in a similar way along the day in males and females. Dur-ing periodic free-choice to alcohol and water, patterns of alco-hol were different in each sex: Males and females drank asimilar volume of alcohol solution, but females distributed

their alcohol intake in more hours along the day (48.7%) thandid males (31.8%). These data support the notion that pat-terns of alcohol can be modified independently of changes inthe volume consumed and suggest that alcohol deprivationfor a few days can modify differentially the patterns of alco-hol consumption in each sex.

It has been reported that P rats show a greater number ofethanol bouts per day rather than larger bout size comparedwith nonselected rats (7). This difference is similar to the ob-served between females and males during the periodic free-choice to alcohol and water condition in the present study.Files suggests that part of the genetic selection for ethanolpreference in P rats could be an altered regulation of ethanolintake pattern (7); however, the causal relation with a similarpattern in the females of the present study remain to be inves-tigated. It has been reported that frequency, size, and dura-tion of alcohol bouts integrally produce the pattern of alcoholintake in each individual, and it is possible to influence thelevel of intoxication achieved (10). Similarly, the characteris-tics of alcohol drinking behavior, common in subjects of eachsex, seem to shape the sex differences in the patterns of alco-hol consumption.

It is possible that sequence of the different conditions ofalcohol exposure could have influenced in the observedchanges in alcohol consumption along the present study, andit would be necessary to analyze this possibility in future ex-perimental designs. However, the sequence of conditions wasidentical for males and females and sex differences were evi-dent. Females drank more and showed higher ethanol prefer-ence than males during free access than during forced condi-tions. Changes in the alcohol exposure schedule from continuousto periodic access produced a broader distribution of alcoholconsumption during the day in females than males, and tasteproperties of available fluids affected the usual pattern of al-cohol consumption in the daily light-dark cycle in both gen-ders. Therefore, the extent of sex differences in alcohol con-sumption and the characteristics of the patterns of alcoholintake in each sex were dependent on the nature of the alco-hol ingestion schedule. Additionally, present data supportthat the amount of alcohol consumed and the patterns of al-cohol intake can be modified each independently of the other.

ACKNOWLEDGEMENTS

This study was supported by CONACyT, project N8 4178P-H9607.

REFERENCES

1. Adams, N.; Shihabi, Z. K.; Blizard, D. A.: Ethanol preference inthe Harrington derivation of the Maudsley reactive and nonreac-tive strains. Alcohol Clin. Exp. Res. 15:170–174; 1991.

2. Alvarez, F.J.; del Río, M. C.: Gender differences in patterns of alcoholconsumption in Spain. Alcohol Clin. Exp. Res. 18:1342–1347; 1994.

3. Blanchard, B. A.; Steindorf, S.; Wang, S.; Glick, S. D.: Sex differ-ences in ethanol-induced dopamine release in nucleus accumbensand in ethanol consumption in rats. Alcohol Clin. Exp. Res.17:968–973; 1993.

4. Crawford, S.; Ryder, D. A.: A study of sex differences in cogni-tive impairment in alcoholics using traditional and computer-based tests. Drug Alcohol Depend. 18:369–375; 1986.

5. Dawson, D. A.; Archer, L.: Gender effects in alcohol consump-tion: effects of measurement. Br. J. Addict. 87:119–123; 1992.

6. Dawson, D. A.: Patterns of alcohol consumption: beverageeffects on gender differences. Addiction 88:133–138; 1993.

7. Files, F. J.; Andrews C. M.; Samson, H. H.; Lumeng, L.; Li, T. K.:

Alcohol self-administration in a nonrestricted access situation withalcohol preferring (P) rats. Alcohol Clin. Exp. Res. 16:751–756; 1992.

8. Files, F. J.; Lewis, R. S.; Samson, H. H.: Effects of continuous ver-sus limited access to ethanol on ethanol self-administration.Alcohol 11:523–531; 1994.

9. Frezza, M.; Di Padova, C.; Pozzato, G.; Terpin, M.; Baraona, E.;Lieber, C. S.: High blood ethanol levels in women. The role ofdecreased gastric alcohol dehydrogenase activity and first passmetabolism. N. Engl. J. Med. 322:95–99; 1990.

10. Gill, K.; Amit, Z.; Smith, B. R.: Alcohol as a food: a commentaryon Richter. Physiol. Behav. 60:1485–1490; 1996.

11. Israwlowitz, R. E.; Hong, O. T.: Sex differences in drinking pat-terns and behaviors among college students in Singapore. Psy-chol. Rep. 64:364–366; 1989.

12. Juárez, J.; Guzmán-Flores, C.; Ervin, F. R.; Palmour, R. M.: Vol-untary alcohol consumption in Vervet monkeys: individual, sex,and age differences. Pharmacol. Biochem. Behav. 46:985–988; 1993.

22 JUÁREZ AND BARRIOS DE TOMASI

13. Kalant, H.: Pharmacokinetics of ethanol: absorption, distribution,and elimination. In: Kissin, B.; Begleiter, H., eds. The pharmacol-ogy of alcohol and alcohol dependence. New York: Oxford Uni-versity Press; 1996:15–58.

14. Lancaster, F. E.: Gender differences in the brain: implications for thestudy of human alcoholism. Alcohol. Clin. Exp. Res. 18:740–746; 1994.

15. Lancaster, F. E.; Brown, T. D.; Coker, K. L.; Elliott J. A.; Wren,S. B.: Sex differences in alcohol preference and drinking patternsemerge during the early postpubertal period in Sprague-Dawleyrats. Alcohol. Clin. Exp. Res. 20:1043–1049;1996.

16. Lancaster, F. E.; Spiegel, K. S.: Sex differences in pattern ofdrinking. Alcohol 9:415–420; 1992.

17. Lankford, M. F.; Roscoe, A. K.; Pennington, S. N.; Myers, R. D.:Drinking of high concentrations of ethanol versus palatable fluidsin alcohol-preferring (P) rats: valid animal model of alcoholism..Alcohol 8:293–299; 1991.

18. Mishra, L.; Sharma, S.; Potter, J. J.; Mezey, E.: More rapid elimi-nation of alcohol in women as compared to their male siblings.Alcoholism 13:752–754; 1989.

19. Mooney, D. K.; Fromme, K.; Kivlahan, D. R.; Marlatt, G. A.:Correlates of alcohol consumption: sex, age, and expectanciesrelate differentially to quantity and frequency. Addict. Behav.12:235–240; 1987.

20. Myers, R. D.; Robinson, D. E.; West, M. W.; Biggs, T. A. G.;McMillen, B. A.: Genetics of alcoholism: rapid development of anew high ethanol preferring (HEP) strain of female and malerats. Alcohol 16:343–358; 1998.

21. Rachamin, G.; MacDonald, A.; Wahid, S.; Clapp, J. J.; Khanna, J. M.;Israel, Y.: Modulation of alcohol dehydrogenase and ethanol metab-olism by sex hormones in the spontaneously hypertensive rat. Effectof chronic ethanol administration. Biochem. J. 186:483–490; 1980.

22. Robbins, C.: Sex differences in psychosocial consequences ofalcohol and drug abuse. J. Health Soc. Behav. 30:117–130; 1989.

23. Sutker, P. B.; Goist, K. C., Jr.; Allain, A. N.; Bugg, F.: Acute alco-hol intoxication: sex comparisons on pharmacokinetic and moodmeasures. Alcoholism 11:507–521; 1987.

24. Valenstein, E. S.; Kakolewski, J. W.; Cox, V. C.: Sex differencesin taste preference for glucose and saccharin solutions. Science156:942–943; 1967.

25. Van Thiel, D. H. and Gavaler, J. S.: Ethanol metabolism andhepatotoxicity. Does sex make a difference? Recent Dev. Alco-hol 6:291–304; 1988.

26. Wade, G. N.; Zucker, I.: Taste preferences of female rats: modifi-cation by neonatal hormones, food deprivation, and prior experi-ence. Physiol. Behav. 4:935–943; 1969.