Ethanol Preference and Behavioral

8/11/2019 Ethanol Preference and Behavioral

1/9

Journal

of Comparative an d Physiological Psychology

1973 ,

Vol.

82, No. 3,

466-474

ETH A N O L PREFERENCE AND BEHAVIORAL

TO LERA N CE IN

MICE:

BIOCHEMICAL

A N D

NEUROPHYSIOLOGICAL MECHANISMS

1

CARL W.SCHNEIDER,

2

Indiana University

of

Pennsylvania

SALLY K. E VA NS, MA YNA R D B .

CHENOWETH,

AN DFLOYD L. BEM AN

Dow

Chemical Com pany, Midland, Michigan

Determinations were made ofethanol preference and behavioral tolerance in

inbred strains of mice. High- and low-preferencestrains were compared on

neural tolerance to ethanol and metabolic capacity.

High

preference fo r

ethanol w as accompanied by higher behavioral and neural tolerance than

that found

in

low-preference mice. Differences

in

metabolism

of

ethanol

be-

tween high- and low-preferring mice were small. However, low-preference

animals did not

metabolize acetaldehyde

as

rapidly

as

high-p reference ani-

mals. Differences in preferenc e for propylene glycol were in the same direc-

tion

and as

extreme

as

those

for

ethanol. Both substances

are CNS

depres-

sants; but

unlike alcohol, propylene glycol

is not

metabolized

to a

toxic

metaboli te

that

might induce a conditioned aversion. This finding in ad-

dition

to the difference

observed

in

neural tolerance suggests that neural

sensitivity may play a part in the acceptance or rejection of ethanol and

propylene glycol.

Differences

in the ethanol preferenceof

inbred strains

of

mice have been found

re -

peatedly

since the initial demonstrations by

McClearn and Rodgers (1959, 1961). In a

two-choice

situation wherethe animalsm ay

obtain water or 10%ethanolthe miceof the

C57BL strain will obtain as much as 90%

of their

fluid

from

the

ethanol bottle, while

the mice of the DBA/2 strain almost to-

tally avoid drinking ethanol (Rodgers,

1967). The pronounced difference between

these

tw o

strains

has

prompted

a

number

of

investigators

to

attempt

to

gain

a

better

un-

derstanding

of

underlying mechanisms, pri-

marily through th e study of biochemical

phenomena.

The most obv ious starting place in this

regard

is the initial step in the metabolism

of ethanol, i.e.,

its

oxidation

to

acetalde-

hyde by alcohol dehydrogenase (ADH),

presumably

th e

rate limiting step

in the

metabolic process (Jacobsen, 1952; New-

Data

for

this investigation were collected

while Carl W. Schneider was an employee of the

Dow

Chemical Company.

2

Requests

fo r

reprints should

be

sent

to

Carl

W . Schneider, Dep artmen t of Psychology, In diana

University of Pennsylvania, Indiana, Pennsylvania

15701.

man, 1947; W esterfeld, 19 55). In vitro

studies

(Bennett

&

Hebert , 19 60; M cClearn,

Bennett , Hebert, Kakihana, &Schlesinger,

1964; Rodgers , McClearn, Bennett ,

& He-

bert, 1963) have shown

the

C57BL/Crgl

(high ethanol preferring mice)

to

have

greater liver

ADH

activity than

th e

low-

preferringDBA/2 Crgl.

In

another

in

vitro

investigation, Sheppard, Albersheim , and

McClearn

(1968) found

the

high-preferr ing

C57BL/6J

mice

to

have

ADH

activity

about

3 0% higher

than

the low-preferring

DBA/2]

strain.

This positive relationship between liver

alcohol

dehydrogenase

activity

and ethanol

preference has

been tested further

by

deter-

minations of the

rate

of metabolism. In

general, these findings indicate that

differ-

ences

in

metabolicrate between high-

and

low-preference

animals tend

to be

quite

small or equivocal an d seem to be insuffi-

cient

to account for the large beha vioral

differences (Bennett

&

Hebert , 1960; Rod-

gers, 1967; Schlesinger, Bennett, & Hebert,

1967; Sheppard, Albersheim, & M cClearn,

1970; W ilson, 1967 ).

The next step in the metabolism of alco-

hol involves

the

oxidation

of

acetaldehyde

466


2/9

E T HAN OL

PREFERENCE

IN

MICE

467

to acetylcoenzyme A

either

directly or via

an intermediate

step involving acetic acid

(Forsander & Raiha, 196 0). A cetaldehyde

ism etabolized

at a

very

rapid

rate

by

alde-

hyde dehydrogenase (Hald & Larsen, 1949;

Lubin & Westerfeld,

1945).

It is an ex-

tremely toxic

substance

and accumulation

during

the

metabolic process could lead

to

deleterious effects (Jacobsen, 1952) that

might

be followed by

conditioned avoidance

of

alcohol.

Schlesinger,Kakihana, and

Ben-

nett

(1966) made determinations

of

blood

acetaldehyde levels 1 and 2 hr. after injec-

tion

ofC57BL/Crgl and DBA/2 Crgl

mice

with ethanol.

They

found

higher levels of

acetaldehyde

in thelatterstrain

af ter

1hr.,

but the two strains were equivalent 2 hr.

after

injection. Sheppard et al. (1968) have

demonstrated thatthe

C57BL/6J

mice have

higher

aldehyde dehydrogenase

activity in

the liver than the DBA/2J mice. In addi-

tion,

gas

chromatographic analysis indi-

cates

that

C57BL/6J mice metabolize

acet-

aldehyde at a more rapid

rate

than the

DBA/2J

animals (Sheppard

etal., 1970) .

In two

extensive reviews (Lester, 196 6;

Mendelson, 1968)

it was

concluded

that ad-

equate evidence unequivocally tying prefer-

ence for ethanol to some aspect of meta-

bolic

capacity

was

lacking. However,

the

recent

findings

of

Sheppard

et al. (1968,

1 9 7 0 )

suggest

that

metabolic capacity

may

play

a

part

in

determining strain differences

in

ethanol preference.

Another areathat

has

received lessatten-

tion

is

that

of the relative

tolerance

for

ethanol between

th e

high-

and

low-prefer -

r ing strains. In one investigation

(Kaki-

hana, Brown,

McClearn, & Tabershaw,

1966) ,

mice of the C57BL/Crgl and

Balb/

cCrgl ( low-p reference s train) s trains were

injected

intraperitoneally with an anes-

thetic

dose

of

ethanol.

The animals

were

placed

in a trough and the amount of

time

until they righted themselves

was

deter-

mined as

sleeping

time.

Brain alcohol

levels determined

at 40,

100,

and 140

min.

by

the gas

chromatograph

method after

injection

were

the

same

for

both strains,

indicating no difference in the metabolic

rate. Of particular interest is the finding

that t h f i

high- prefer r in g C57BL animals

awoke at a

t ime

when

their bra in-a lcohol

level w as significantly above that of the

Balb animals

at

their time

of

waking.Rate

of

absorption of alcoholw as determined in -

directly

by

measur ing

th e

amount

of

time

before th e

mouse

fell

from

th e

underside

of

a

wire mesh where

it was

placed immedi-

ately after injection, and it was found to be

th e

same

in

both groups.

The

results

of

this

investigation demonstrated a greater cen-

tral nerv ous system (CN S) sensitivityto al-

cohol in the

low-preference mouse

strain

than

in the high-preference

strain.

In light of these

findings

one may specu-

late

that

neura l

as

well

as

metabol ic factors

may

play

apartin

both tolerance

and

pre f -

erence

fo r

ethanol.What

th e

relationship

is

among th e possible mechanisms underlying

the

pronounced behavioral

differences be-

tween high-

and low-e thanol -pre ferr ing

mice

is not at all

clear.

T he

series

of

experiments

described in this paper was designed for the

purpose

of

gaining

a

fur ther unders tanding

of the

possible relationship

of

behavioral

and neural tolerance

to

ethanol , ethanol

preference,

and

metabol ic capacity.

E X P E R I M E N T

1

In the first experiment, determinations

were

made

of the preference for and

behav-

ioral tolerance

to

ethanol

in

threestrains

of

mice.

T he behavioral tolerance testing in -

volved measures of the effects of alcohol on

nest-building behavior. Nest building

is a

behavior displayed by all mice and has

proven to be very

useful

in del ineating drug

effects (Schneider

&

Chenoweth, 1970,

1971).

Method

Subjects.

Three

groups

of 20

male

mice

from

three strains (C57BL/6J, Swiss-Webster,

DBA/ 2 j )

were

used in measures of ethanol preference an d

subsequent tests

of the effects of the drug on

nest-

building behavior .

The

C57BL/6J

an d

DBA/2J

mice were

obtained from

Jackson Lab oratory,Ba r

Harbor, Maine, and the Swiss-Webstermice

from

Spartan Research Animals, Inc.,

Haslett,

Michi-

g a n . All animalswere 60-65 days old

upon

arrival

an d

70-75 days

old at the

beginning

of the

experi-

ment .

Housing and testing. The animals were housed

individually in clear plastic boxes (28 X 28cm.)

equipped with

wire

floors and special

covers

con-


3/9

468

SCHNEIDER,

EVANS , CHENOWETH, AND B E M A N

taining a cotton dispenser, two fluid delivery

holes,

and a food

hopper. During preference deter-

minations each animal was presented with two

15-ml.

centrifuge tubes graduated

in

,1-ml. incre-

ments.

One tube

contained distilled

Hs>0 and the

other

distilled H

2

0

plus

95%

ethanol

in a 10%

(V/V) solution. During nest-building tests, cotton

could be

obtained from

a 7 X 10 cm.

cylinder

attached to the

covers.

The

cotton

was

tightly

packed against

th e

cover with

a

ki logram weight

that

fit

snugly

into the

cylinder.

The

cover area

supporting the cotton contained

12-mm.

holes

through w hich a mouse could, with some

effort,

pull

cotton into

th e

cage. Amount

of

cotton used

fo r

nest building was determined by weighing unused

cottonon aMettlerbalance with

10-mg.

divisions.

Procedure.

The entire experiment was run in

a

windowless room with tem pera ture, hum idity,

and a 7A.M.-5P.M.lightperiod heldconstant.

The

experimental design

was as

follows:

(a) 3

days

of adaptation to plastic boxes,

(b )

10 days

of prefere nce testing, (c) 3 days of rest,

(d)

10

days

of

nest building

preethanol, (e) 10

days

of

nest building with forced consumption

of 10%

ethanol, (/) 10 days of nest building postethanol.

(Five

animals

in

each group received

no

alcohol

during

the

10-day forced-consu mp tion period

and

thus served

as

environmental controls.)

During preference testing

the

animals

had a

choice

between

the 10%

ethanol solution

or

dis-

tilled water contained

in the

15-ml. graduated

tubes. Measures of amount consumed were made

every

24 hr.

beginning

at 10A .M . , an d

each

day the

position of the bottles was switched in order to

control fo r position effects. The preference index

for each animal

w as

calculated

by

dividing

th e

volume of ethanol solution consumed by the

volume ofethanol solution plus water consumed.

During the nest-building tests, cotton in excess

of th e amount required fo r nest building was

weighed and placed in each cylinder. Twenty-four

hours later the cotton remaining in the cylinder

was

weighed

to

determine

the

amount used

to

build

the

nest.

After

the

weighing,

the old

nest

w as

removed

and a

more than adequate supply

of

cotton was preweighed and returned to the cyl-

inder for the next

24-hr,

period. The procedurew as

th e

same every

day for the

entire 30-day period .

Results

Figure

1 illustrates the

preference

index

for

10%

ethanol

and the

effect

of the

solu-

tion on the nest building of the same sub-

jects during exposure

to

alcohol

as the

sole

source of fluid. In accordance

with previous

findings, the C57BL/6J

strain

has the

highest

preference for

ethanol while

th e

DBA/2] animalsavoid it. The Swiss-Web-

stermice are low

preference,

but do notuni-

formly avoid

alcohol

like the DBA mice.

The weight of the nest is an indirect meas-

ure of the amount of work the animals

would

do

over

a 24-hr,

period.

The

high-

preferring C57BL animals showed abso-

lutely

no

change

in

nest w eight dur ing

forced-ethanol consumption, while th e low-

preference Swiss-Webster and DBA strains

showed a significant reduction in nest size

duringthat period. The DB A strain showed

th e greatest effect with a reductionin nest

size of 27%. Of particular interest are the

results ob tained duringth epostexposurepe-

riod. The DBA mice, most affected during

exposure, increased nest weight over the

preexposure

mean

by 8%.

This

was in

marked contrast

to the C57

animals'statis-

ticallysignificant decrease in nest size after

withdrawal of ethanol. This

effect

on nest

building

may serveas a model fo rlow-level

withdrawalsymptoms.

E X P E R I M E N T 2

The

results

of the

previous experiment

clearly

demonstrate a positive relationship

between preference fo r ethanol and behav-

ioral

tolerance.

The

work

of

Kakihana

et

al. (1966) suggeststhat this difference

in

tolerance

between high-

and

low-preference

strains

may be due to

greater

CN S

sensitiv-

ity in the low-preference (in their case,

Balb) strain. This possibility

was

explored

further

by

examining

the CN S

susce ptibility

to ethanolbyq uantitative m easurementof a

centrallymediatedreflex activity. Thismay

be

accomplished

in the mouse by use of the

j aw- je rk

reflex

described originally by

Sher-

rington

(1917).The afferent pathwayofthis

reflex has

been delineated

by

Harrison

and

Cor

bin

(1941)

and

shown

to be the

mesen-

cephalic root of the fifth cranial nerve.

Method

Apparatus.

Jaw jerk s were sensed by a min iature

strain gauge cemented to awatch spring.The out-

put

from

the

strain gauge

wasreceivedby a

Beck-

man

Model

RB

Dynograph system where

it was

amplified

and recorded on a constant-speed chart

recorder.

Stimulation was achieved with a flat-ended

bipolar

stainless-steel electrod e. The electrod e was

insulated with Epoxylite 6001-M cement, and the

tw o

poles at the t ip w ere .5 mm . apart. Electrical

s t imulat ion

was

provided

by a

Grass S88 st imu-

lator.


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Ethanol Preference and Behavioral