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1992. Vol. 16. pp. Q69-976

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REDUCING THE TIME NEEDED TO CONDUCT CONDITIONED PLACE PREFERENCE TESTING DANIEL J. CALCAGNETTI and MARTIN D. SCHECHTER Department of Pharmacology Northeastern Ohio Universities College of Medicine Rootstown, OH 44272-0095 U.S.A.

(Final form, January 1992) Abstract Calcagnetti, Daniel J. and Martin D. Schechter: Reducing the time needed to conduct conditioned place preference testing. Prog. Neuro-psychopharmacol. 81 Biol. Psychiat. 1992, 16(6): 969-976. The objective of the present experiment was to demonstrate whether four days of twice-a-day conditioned place conditioning produces a preference that is equivalent to Two doses of the that produced by using eight days of once-a-day tmining. amphetamine-like stimulant drug cathinone (0.2 and 1.6 mg/kg) were selected to demonstrate the effectiveness of twice-a-day (morning-vehicle; afternoon-drug pairings) conditioning. The 0.2 mg/kg dose of cathinone failed to affect the expression of place preference, whereas the 1.6 mg/kg dose produced a significant (p < 0.002) shift in CPP from baseline when compared to previous measurements. These results demonstrate that twice-a-day pairings over four days effectively shorten the total duration of training without changing the development of place preference produced by once-a-day over eight conditioning day schedule. Kevwords: Cathinone, Abbreviations:

Conditioned

Conditioned

place preference, locomotor activity

place preference (CPP)

Introduction

when rats are confined on multiple administration drug-associated

occasions to a cue-specific environment/place

after

of a rewarding drug, they will chose to spend a significantly greater time in that place in contrast to a place that had not been conditioned

procedure is known as “conditioned assess the rewarding,

with the drug. This

place preference” (CPP) and it is an effective method to

as well as the aversive, effects of centrally or peripherally 969

administered

D. J. Calcagnettland M. D. Schechter

970

drugs of abuse [for reviews see Hoffman

1989; Swerdlow et al., 19891.

Although CPP has

been extensively used to access the rewarding effects of drugs, a deterrent to its more widespread use resides in the fact that the procedure requires an extended period of time to condition the drug-environment. In a report (Bozarth, 1987) of parametric experiments conducted to determine the number of conditioning trials required for each of the two conditions, i.e., drug and its vehicle, the critical minimum of conditioning trials was three. Thus, trials in excess of six did not increase the rewarding effects of the drug, as assessed by CPP. With six trials optimum and with each conditioning of the experimentation investigators

trial performed once-a-day, this phase

would require a total of six days. In order to speed this process, a few

have administered

the vehicle in the morning and conditioned

and followed this with the afternoon

administration

opposite side. In fact, in one case, conditioning

the rats in one side

of the drug and conditioning

on the

with the vehicle actually followed conditioning

with the drug (Acquas, et al., 1990), a practice that might compromise the results if the drug under testing had an extended half-life.

The purpose psychostimulant

of the present investigation

cathinone failed to produce CPP at 0.2 mg/kg in one group of rats, whereas

a dose of 1.6 mg/kg conditioned significantly

greater

amount

a place preference in another group wherein they spent a

of time in a previously

experiment involved administration period.

was to expand recent evidence in which the

environment.

to use these two conditions

on the same day.

produces an effect upon preference that is not significantly

from the once-a-day conditioning,

This

of either vehicle or cathinone once-a-day over an eight day

The present study intended

twice-a-day conditioning

non-preferred

If the

different

then the time (in days) needed to conduct CPP experiments

may be compressed in a shorter interval without decrement to the strength of conditioning. Cathinone was selected to test the shortened conditioning

procedure since it is a stimulant with

a known duration of action (Schechter, 1989) and for which the dose-response well characterized

(Meehan and Schechter, in review).

Additionally,

curve has been

doses of cathinone above

1 mg/kg have been reported to significantly increase locomotor behavior (Kalix, 1980) and this will provide a separate indicator effective. cathinone

as to the doses of cathinone

In this light, locomotor

activity was measured

doses of 0.2 and 1.6 mg/kg.

tested being physiologically

with and without

injection

of

971

Shortening cathinone place conditioning MATERIALS AND METHODS

Male rats of Sprague-Dawley from Z&-Miller

Laboratories

descent, weighing

175195

g upon arrival, were purchased

Inc. (Allison Park, PA). All subjects were individually

housed

in stainless steel hanging cages equipped with ad libitum access to food (Purina 5008) and water.

They were maintained

in a colony room with a constant temperature

and humidity on

a 12:12 hr light:dark cycle (dark onset at 18:00 hr). Place preference conditioning/testing

was

conducted in a room separate from the colony room. All subjects were handled in compliance with the “Guide for the Care and Use of Laboratory Animals,” Dept. of Health, Education and Welfare Publication, Dnm

1985.

and Injection

(-)~t~none

hy~o~o~de

vehicle control injection. ~~~ton~y

(NIDA) was dissolved in sterile water that also served as the All doses tested are expressed as the salt and were administered

(IP) in an injection voice

Conditioned

Place Preference Annaratus

Place conditioning/testing

of 1 ml/kg of body weight.

and Procedure

was conducted

in one of four modular

(Lafayette Inst. Co., IN). The three-chambered

test component

stainless steel apparatus consisted of a center

section through which the subjects were allowed access into two end sections,

A constraint

wall served to restrict a subject’s egress from the right or left side of the apparatus conditioning.

units

during

The right and left end sections (20.5 x 30.5 x 20 cm), originally identical, were

altered in three sensory modalities to provide the following discriminable of the chamber was illuminated

cues: the “dark” side

by a 6W, 30V red light bulb and had a solid black Plexiglas

floor; the *light” side was coated

with a 6W, 3OV white light bulb, with a bar grid floor

and pine shavings in the drop pan. Location of the subject while in the chamber was detected by weight pivot sensors connected to a computer that automatic~y

recorded the time (in set)

spent in each side of the apparatus. Two groups of subjects (n= 10 each) underwent testing, drug and vehicle conditioning days of habituation appears. baseline

to the conditioning

three treatment phases: habitation/baseline

and place preference resting.

Subjects were given two

room and 15 mm of free access in the place preference

On the third day, 15 min of free access served to establish a pa-conditio~ng of place preference

per subject.

The side in which the rats spent less time was

972

D. J.

Calcagnetti and M. D. Schechter

considered its less-preferred side for the remainder of the study. Place conditioning

was then

initiated and conducted in two daily 30 min sessions.

The twice-a-day confinement

conditioning

phase consisted of four days of pairing with vehicle and

for 30 rain in the preferred

side of the apparatus

followed by a second pairing session after administration afternoon

15 min.

(10:00 h)

of cathinone and conllnement,

(14:OO h) in the less-preferred side. Twenty-four

conditioning,

in the morning

in the

hr following the 4th (last) day of

each subject was allowed free access to explore all sections of the chamber for

Place preference was, thus, redetermined

Snontaneous

in the non-drugged

state.

Locomotor Activity

Activity was measured by the interruption

of one of four photosensor

light sources placed

in the wall of a 45.5 X 35.5 X 20.5 cm Plexiglas cage. The sensors were oriented 5.5 cm above the floor and 9.5 cm apart along the wall of the longer side. constituted

Each photocell interruption

one activity count. A computer recorded activity counts every 5 rain of the 30 min

testing duration. administration administered

Activity was measured

in the light phase and began immediately

of vehicle on the day of non-drug CPP testing.

Two days later, subjects were

their assigned dose of cathinone and a second activity session was measured.

allowed for a comparison

after

This

of activity with and without drug.

Statistical Analvsis

The critical measurement less-preferred t-test.

side of the CPP apparatus.

Additionally,

measurements

was the actual time (in set) that the subjects

independent

between

These measurements

spent in the

were compared by a paired

i-tests and paired t-tests were employed to compare activity

and among the dose groups, respectively.

The level of statistical

significance was set at p < 0.05.

Results

Table 1 shows the mean and standard deviation (S.D.) of the set spent in the less-preferred side of the apparatus in a drug-free test after conditioning

trials with either 0.2 or 1.6 mg/kg

of cathinone.

significantly

Rats conditioned

with 1.6 mg/kg cathinone

increased the amount

973

Shortening cathinone place conditlontng of time they spent in the less-preferred t=4.5;

p < 0.002).

cathinone

side of the apparatus

In contrast, the group of rats conditioned

failed to show a reliable change from baseline

cathinone pairings with the less-preferred

(df-9;

with the 0.2 mg/kg dose of

(df=9;

t=0.87;

p=O.41).

Thus,

side produced a place preference at the 1.6 mg/kg

dose, but not with the 0.2 mg/kg dose using the twice-a-day present findings, using this compressed conditioning recent report in which cathinone

compared to baseline

conditioning

conditioning

procedure.

The

procedure, yielded the same results as a

was conducted

over eight days, once-a-day,

pairings of either cathinone or vehicle (Meehan and Schechter, in review), and are included in Table 1 for comparative

purposes.

Table 1 Mean (kS.D.) Time (in set) Spent in the Non-Preferred Side during Baseline and after Four Twice-a-Day (A) or Four Once-a-Day (B) Pairing with Either 0.2 or 1.6 mg/kg Cathinone (i.p.).

A

0.2 mz/kg

TWICE-A-DAY

1.6 mg/kg

BASELINE

MEAN S.D.

234.8 (57.3)

207.5 (113.7)

PREFERENCE TEST

MEAN SD.

210.0 (134.0)

385.8” (82.1)

-10.6

85.9

% INCREASE B.

ONCE-A-DAY

1.6 mz/kg

0.2 mdkg

BASELINE

MEAN S.D.

209.1 126.0

190.6 (118.6)

PREFERENCE TEST

MEAN S.D.

330.5 (172.3)

371.6 (165.0)

58.1

95.0

O/6CHANGE

a Significant difference from baseline, paired i-test; p < 0.05.

Comparison by independent

t-tests of activity counts after vehicle administration

between

the 0.2 and 1.6 mg/kg dose groups failed to reveal reliable differences between groups (df=18; t=O.45; p=O.6).

Paired i-test comparison of activity counts after administration

(1.6 mg/kg) revealed a significant t=5.64;

of cathinone

increase (82.7%) in activity compared to vehicle (df=9;

p c O.OOl), whereas there was no reliable difference in activity between vehicle and

D. J. Calcagnetti and M. D. Schechter

974

drug administration

(df=9; t=0.13;

p=O.22) using the 0.2 mg/kg dose.

mean [and standard deviation (SD.)] of photocell interruptions IP administration

of vehicle and cathinone.

Table 2 shows the

over a 30 rain period after the

These results indicate that the 1.6 mg/kg dose of

cathinone produced a significant shift to the less-preferred

side of the CPP apparatus and also

signif?cantly increased locomotor activity.

Table 2 Mean (S.D.) Photocell Interruptions per 30 min Period with Vehicle or Cathinone 1.6 mg/kg, IP) as a Measure of Activity.

0.2

1.6

MEAN S.D.

279.7 (124.4)

256.2 (110.2)

MEAN S.D.

300.5 (100.0)

468.2” (153.2)

% INCREASE IN ACTMTY

7.4

82.7

(0.2 or

VEHICLE

CATHINONE

“Significant difference from baseline, paired t-test; p < 0.05.

A twice-a-day

conditioning

procedure,

with morning

pairings, resulted in equivalent preference measurements which vehicle or cathinone conditioning

vehicle and afternoon

cathinone

compared to identical experiments in

sessions were carried out once-a-day over eight days.

These findings support the possibility that the interval between vehicle and drug conditioning sessions of 3 hr (on the same day) is as effective in producing a CPP as an interval of 24 hr.

The 1.6 mg/kg, but not the 0.2 mg/kg, dose of cathinone significantly increased locomotor activity. cathinone

This finding is important

because it confirms that a physiologically

is also capable of producing

effective dose of

a CPP. Moreover, a lower dose of cathinone

mg/kg), that does not increase activity, fails to alter the conditioning also in agreement with the findings that cathinone significantly

(0.2

of place. These data are

increases basal activity (Kalix,

1980).

In conclusion,

it is hoped that the evidence herein regarding

the use of this compressed

975

Shortening cathinone place ~~i~o~g

conditioning Shortened duration

procedure conditioning

of action.

may encourage

greater use of this technique

but with one caveat.

for CPP testing should be employed only for drugs with a known

Using a shortened

conditioning

procedure with a drug of unknown,

or

possibly long half-life, may result in the drug effect remaining active in the next days’ morning conditioning

session with vehicle and, hence, confounding

The present

findings

are particularly

the non-drug conditioning

timely and relevant

employing shortened (l-4 days) conditioning et al., X991) but have yet to demonstrate

since several

session.

laboratories

are

procedures (Corrigall and Linseman, 1988; Pam

what, if any, impact shortened conditioning

have on the expression of place preference compared to longer conditioning

sessions

intervals.

The

present research is the only report to-date that has assessed the impact of a 4 day twice-a-day conditioning

procedure with the once-a-day 8 day conditioning

employed in many laboratories

procedure that is commonly

(Brockwell et al., 1991; Morency and Beninger, 1986; Spyraki

et al., 1987).

Conclusion

In conchrsion, these results demonstrate (afternoon)

that twice-a-day pairings, of (mo~g)

vehicle and

drug, over a four day period can effectively shorten the number of days needed to

tram rats in a conditioned

place preference task.

Acknowledgements

We thank The National Institute of Drug Abuse for the gift of I-> cathinone and for funding grant No. 3591 to M.D.S. Funding for D.J.C. was provided by the State of Ohio Academic Challenge Program.

ACQUAS, E., CABBONI, E., GABAU, L. AND DI CHIABA, G. (1990). Blockade of acquisition of drug-conditioned place aversion by 5HT, antagonist. Psychopharmacology m 459-463. BKOCKWELL, N.T., EIEELBOOM, R AND BENINGER RJ. (1991). Cake-educed taste con~tio~: suction of dose-dependent preference and aversion, Biochem. Behav. s 513-517.

place and Pha~acol.

D. J. Catcagnettl and M. D. Schechter

976

BOZARTH, MA. (1987). Conditioned place preference: a parametric analysis using systemic heroin injections. In: Methods of Assessing the Reinforcing Properties of Abused Drugs, M.A. Bozarth (Ed), pp 241-273 (Springer-Verlag:New York). CORRIGALL, W.A. AND LINSEMAN, M.A. (1988). Conditioned place preference produced by intra-hippocampal morphine. Pharmacol. Biochem. Behav. a 787-789. HOFFMAN, D.C. (1989). of drug reinforcement.

The use of place conditioning Brain Res.Bull. a 373-387.

KALIK, P. (1980). H~~o~~ of the ~phet~e leaves. Brit. J. Pharmacol. &, 11-13.

in studying the neuropharmacology

type induced by a constituent

of khat

MEEHAN, S.M. AND SCHECHTER, M.D. (m review). Effect of dopamine release inhibition upon conditioned place preference produced by the psychostimulant cathinone. Psychopharmacology Q(&00-00 MORENCY, M.A. AND BENINGER, RJ. (1986). place conditioning. Brain Res. B 33-41.

Dopaminergic

substrates of cocaine-induced

PANI, L., KUZMIN, A., ~TELLO~~ MC., GESSA, G.L. AND FRATTA, W. (1991). The calcium antagonist PN 200-110 inhibits the reinforcing properties of cocaine. Brain Res Bull. 26.445447. SCHECHTER, M.D. (1989). Temporal parameters of cathinone compared to amphetamine cocaine. Pharmacol. Biochem. Behav. & 289-292. SPYRAKI, C., NOMIKOS, G.G. AND VARONOS, D.D. (1987). Intravenous place preference: attenuation by haloperidol. Brain Res. Bull. 24, 57-62.

and

cocaine-induced

SWERDLOW, N.R., GILBERT, D. AND KOOB, G.F. (1989). Conditioned drug effects on spatial preference. In: Neuromethods Series l- Psychoph~acolo~, Vol. 13, A.A. Boulton, G.B. Baker, and A.J. Greenshaw (Eds), pp 399-446 (Humana Press, Clifton, NJ).

Inquiries and reprint requests should be addressed to: Daniel J. Calcagnetti, Ph.D. Northeastern Ohio Universities College of Medicine Department of Pharmacology P.O. Box 95 Rootstown, OH 44272-9989 U.S.A.