Abstract. Place preferences induced by the indirect dopa- mine (DA) receptor agonists amphetamine (AMP) and methylphenidate (MPD) were investigated using ...
Psychopharmacology
Psychopharmacology (1986) 90:247-252
© Springer-Verlag 1986
The effects of haloperidol on amphetamineand methylphenidate-induced conditioned place preferences and locomotor activity S. Mithani 1, M.T. Martin-Iverson 1, A.G. Phillips 2, and H.C. Fibiger 1 Division of Neurological Sciences, Department of Psychiatry, and 2 Department of Psychology, University of British Columbia, Vancouver, B.C., Canada Abstract. Place preferences induced by the indirect dopamine (DA) receptor agonists amphetamine (AMP) and methylphenidate (MPD) were investigated using an unbiased compartment procedure. In this procedure, prior to drug conditioning, rats did not exhibit preferences for either of the two compartments in a shuttle box. Both stimulants produced place preferences. Repeated testing of the MPD conditioned animals revealed an extinction-like decrease in preferences, suggesting that place preferences produced by MPD result from conditioning of MPD's reinforcing properties to environmental cues. During conditioning, the DA receptor antagonist haloperidol was administered prior to drug (S + ) treatments, or prior to both drug and vehicle ( S - ) treatments. Haloperidol pretreatment blocked place preferences induced by AMP but not by MPD. In contrast, haloperidol blocked locomotor activity stimulated by either AMP or MPD. These results suggest that the reinforcing properties of MPD and AMP may be mediated by different mechanisms, while the locomotor stimulant effects of the two drugs have common neural substrates. Key words: d-Amphetamine - Methylphenidate - Haloperidol Conditioned place preference - Dopamine Reward
The conditioned place preference (CPP) paradigm has been used to demonstrate reinforcing effects of psychostimulants such as amphetamine (AMP) (Sherman et al. 1980), cocaine (Spyraki et al. 1982 a), methylphenidate (MPD), and nomifensine (Martin-Iverson et al. 1985), as well as opioids such as heroin (Spyraki et al. 1983; Bozarth and Wise 1981), morphine (Schwartz and Marchok 1974), and the endogenous opioids (Katz and Gormezano 1979; Stapleton et al. 1979). The finding that both AMP and cocaine have agonistic effects at dopamine (DA) synapses (Moore et al. 1977), and that DA receptor antagonists can attenuate AMP (Spyraki et al. 1982b), heroin (Bozarth and Wise 1981), and morphine (Schwartz and Marchok 1974) CPP suggests a dopaminergic involvement in the effect of these drugs on place preferences. Furthermore, microinjections of morphine and an enkephalinamide into the ventral tegmental area, an area rich in dopaminergic cell bodies, have also been shown to produce place preferences (Phillips and LePlane 1980 1982). There is, however, conflicting evidence regarding the involvement of dopaminergic systems in the reinforcing properties of other psychostimulants. For instance, haloperOffprint requests to." H.C. Fibiger
idol, at doses that block stimulant-induced locomotor activity (Martin-Iverson et al. 1985) and attenuate AMP CPP (Spyraki et al. 1982b), does not affect CPP produced either with cocaine (Spyraki et al. 1982a), MPD or nomifensine (Martin-Iverson et al. 1985). Similarly, preferences for a place associated with cocaine is unaffected by the destruction of accumbens DA terminals (Spyraki et al. 1982b). The CPP procedures of previous investigations of psychostimulants used boxes in which rats exhibited preferences for one compartment, and the less preferred compartment was associated with drug administration (Martin-Iverson et al. 1985; Spyraki et al. 1982a, b). It is therefore possible that MPD-like psychostimulants induce place preferences by reducing aversiveness of the less preferred compartments. This could occur by facilitating habituation to neophobia of the less preferred compartment, or by drug actions on brain systems underlying aversive reactions, rather than from associations of compartment cues with the reinforcing properties of the drugs. That MPD facilitation of habituation may underlie the changes in place preferences reported earlier is supported by the increase in preference for the less preferred compartment that is often shown by control groups after the conditioning trials (Martin-Iverson et al. 1985). This problem can be overcome by using unbiased compartments. If rats do not exhibit initial preferences for either compartment, then it is unlikely that drug-induced changes result from a reduction in the aversiveness of one compartment. Thus, the present experiments tested the influence of AMP and MPD on place preferences in "unbiased" boxes. It was also reasoned that changes in compartment preference related to increased habituation would be long lasting. Repeated testing would increase the drug effect, thus reducing aversiveness more strongly. On the other hand, place preferences resulting from conditioning of "pleasurable" drug effects should undergo extinction during repeated tests without the primary drug reward. Therefore, the effect of repeated testing of animals conditioned with MPD was examined to determine if the rats' place preferences would exhibit extinction-like decreases. In order to clarify the involvement of dopaminergic systems in the rewarding effects of psychostimulants, the ability of haloperidol (a DA receptor antagonist) to alter preferences induced with AMP and MPD in "unbiased" compartments was investigated. In addition, the influence of haloperidol on locomotor activity induced by MPD and AMP was examined by following a protocol similar to that used in place preference conditioning. This was undertaken to assess whether or not differences in the susceptibility
248 of place preference with AMP and MPD to blockade with haloperidol is associated with differences in the ability of haloperidol to block the motor stimulant effects of these drugs.
Methods
Animals Male Wistar rats (Woodlyn) weighing 270-300 g at the beginning of the experiment were housed five per cage and maintained on a light-dark cycle (light: 08.00-20.00 hours) with food and water continuously available. Testing and training were performed between 09.00 and 15.00 hours. Since the rats suffered from diarrhoea upon arrival, they were treated with chloramphenicol (Rogar-Maine Powder, Rogar/STB) in the drinking water (5 g/300 ml) for the 1st week, and were then handled for 1 week.
Apparatus Place preference. Each animal was conditioned and tested in one of four shuttle boxes (80 x 25 x 36 cm) divided into two compartments ( 3 4 x 2 5 c m ) joined by a tunnel (8 x 8 x 6 cm) which could be closed by guillotine doors. Each of the two compartments was distinctive in the color of the walls and in the type of floor, having brown walls or black and white striped walls (1 cm) and a grid floor or a mesh floor (1.2 cm square). The wall and floor cues were counterbalanced. Thus, rats of each group were assigned a drug-conditioned compartment consisting of either l) brown wall+grid floor, 2) brown wall+mesh floor, 3) striped wall + grid floor, or 4) striped wall + mesh floor. The shuttle boxes were mounted on a fulcrum, permitting the time spent in each compartment and the number of crossings from one compartment to the other to be recorded by electromechanical equipment.
Locomotor activity. Five circular (61 cm diameter) activity cages (BRS/LVE) transected by six infra-red beams were used to measure locomotor activity. Each photocell beam interruption and the time of its occurrence were recorded with a NOVA IV (Data General) minicomputer equipped with M A N X (GC Controls) software and interface.
Behavioral procedure Place preference. The behavioral procedure consisted of three phases: pretest (Phase 1), conditioning (Phase 2) and test (Phase 3). During Phase 1, rats were placed in one of the compartments (hereafter called the start-box) following a counterbalanced order. They were given free access to both compartments and the time spent in the non-start box was recorded over a 15-rain period for 3 consecutive days. Phase 2 consisted of 8 days. On days 1, 3, 5 and 7 (drug days, S +), rats were injected (IP) with the drug and confined to the non-start box for 30 rain. On alternate days (control days, S - ) , rats were confined to the start-box for 30 min following vehicle injections. In Phase 3, each rat was placed in the start-box and given free access to both compartments for 15 min: the time spent in each compartment, and the number of crossings between compartments, were recorded.
Locomotor activity. The protocol used was similar to place preference conditioning. It consisted of three pretests (Phase 1), where locomotor activity was recorded over 15 min for 3 consecutive days. This was followed by Phase 2, consisting of 8 days, whereby on days 1, 3, 5 and 7, drugs were administered and locomotor activity recorded over a period of 30 rain. On days 2, 4, 6 and 8, animals were injected with vehicle and placed singly, in holding cages, in the locomotor activity room, for 30 min. Phase 2 was followed by Phase 3 (test day), when animals were not injected and locomotor activity was recorded over a period of 15 rain. Experiment 1 In order to confirm the "unbiased" nature of the shuttle box, animals were conditioned to both compartments of the testing apparatus following vehicle (distilled water). On days 1, 3, 5 and 7, rats were injected with vehicle 70 rain and 10 rain before placement in the non-start compartment; on alternate days, animals were placed in the start compartment following vehicle injection at similar times. On the test day (Phase 3), animals did not receive any injections and were given free access to both compartments. The amount of time spent in each compartment, and the number of crossings between compartments, were recorded over 15 rain.
Experiment 2 Animals were randomly assigned to one of six groups (n = 10 per group, except Group 5 where n = 20) receiving the following treatments on drug days and control days respectively. Group 1 : vehicle + AMP and vehicle + vehicle; Group 2 : haloperidol + AMP and vehicle + vehicle; Group 3 : haloperidol + AMP and haloperidol + vehicle; Group 4 : vehicle + MPD and vehicle + vehicle; Group 5 : haloperidol + MPD and vehicle + vehicle; Group 6 : haloperidol + MPD and haloperidol + vehicle. On drug days haloperidol (0.2mg/kg) was injected 60 rain before AMP (1.5 mg/kg) or MPD (5.0 mg/kg). Animals were placed in the non-start compartment 10 rain after stimulant injection. The doses of AMP, MPD and haloperidol were chosen to be consistent with those used in previous place preference studies (Spyraki et al. 1982b; Martin-Iverson et al. 1985). Injections were given at similar times on control days, and animals were placed in the start compartment. On the test day (Phase 3), animals were given free access to both compartments, and the time spent in each compartment, as well as the number of crossings from one compartment to the other, were recorded over 15 min. Groups 3 and 6 were included in order to determine whether place preferences result from some absolute level of DA receptor activation, or from relative differences in DA transmission between S + and S - days. Therefore, if for instance, the attenuation of AMP-induced place preference by haloperidol is due to the latter, then less attenuation of AMP-induced place preference would be observed when haloperidol is given on both S + and S - days.
Experiment 3 The effect of repeated testing on the MPD place preferences was examined to ascertain whether place preferences un-
249 dergo extinction. Animals were injected with MPD (5.0 rag/ kg) or vehicle on drug days, 10 min before placement in the non-start box for 30 rain. On control days, animals were injected with vehicle 10 rain before being confined to the start compartment for 30 rain. In Phase 3, the time spent in the non-start compartment was recorded over a period of 15 min for 3 consecutive days.
600
500 ~- 400.
3o0 200
Experiment 4
100
Locomotor activity. Naive animals were randomly assigned to one of four groups (n = 12 per group), receiving the following treatment on drug days. Group 1: vehicle+AMP (1.5 mg/kg); Group 2: vehicle+MPD (5.0 mg/kg); Group 3 : halopefidol (0.2 mg/kg) + AMP; Group 4: haloperidol + MPD. Haloperidol or vehicle was injected 60 rain before AMP or MPD, and animals were placed in the locomotor boxes 10 min after stimulant injection. On control days, all animals were injected with vehicle.
Drugs used d-Amphetamine sulfate (AMP) was purchased from Smith Kline and French Pharmaceuticals, haloperidol from McNeil Pharmaceuticals, and methylphenidate HCI (MPD) from CIBA Pharmaceuticals. All drugs were dissolved in distilled water.
Statistical analysis Experiment 1 was analysed with a correlated t-test, as were the number of crossings in all CPP experiments. Experiment 2 was analysed with ANOVA with two between factors, the first consisting of two levels (stimulants), and the second composed of three levels (haloperidol pretreatment) and with one within factor, consisting of two levels (mean of three pretests x test). Planned comparisons were conducted. Analysis of variance with one between factor of two levels and one within factor of four levels was applied to the results of Experiment 3. As the validity of this analysis is dependent upon compound symmetry, an additional multivariate test (Rao's R, distributed as exact F) was utilized for the interaction term of the two factors when there were more than two levels within a repeated measures factor. Planned comparisons were made with univariate tests. In experiment 4, locomotor activity data on S + days was analysed by ANOVA with four factors; two betweensubject factors (stimulant treatment and haloperidol pretreatment), and two within subject factors (days and blocks of 5 rain). Tests for linear trends over days were carried out for selected groups. Individual post-hoc comparisons were made using Student's t-test. Locomotor activity on pretest days and test day was analysed with ANOVA with a similar design. Results
Experiment 1. Figure 1 depicts the mean time spent in the non-start (conditioned) compartment before (on all 3 pretest days), and after pairing both compartments of the testing apparatus with vehicle. No significant alteration in time spent on the non-start side was observed (P>0.05, n = 10)
Prel
Pre2
Pre3
Test
Fig. 1. Mean (+ SE) time spent in the non-start compartment before (pretests, open bars) and after (test, striped bar) pairing with vehicle
800 -
ul
600
a ,~ 400
o
~ 200-
I
~1~~ ~ / ~ Treatment schedule, S +/S- days
Fig. 2. Time (s) spent in the non-start (drug paired) side of a shuttle box before (pretest, open bars) and after (test, striped bars) pairing of a distinctive side of the box with 1.5 mg/kg AMP (A), or 5.0 rag/ kg MPD (M), pretreated with VEH (V), or 0.2 mg/kg haloperidoI (H) on drug (S +) days, and the other side with V or H on control (S - ) days. Rats were pretreated with haloperidol or vehicle 60 rain before stimulant injection. *** P < 0.001 Test significantly different from pretest after conditioning. During pretests, there was no significant preference for either compartment. The number of crossings also showed little change (P > 0.1, n = 10, Table 1).
Experiment 2. The ability of both AMP and MPD to produce place preferences, and the effects of prior treatment with haloperidol, given only in conjunction with stimulants, or also on the control days, are illustrated in Fig. 2. Analysis of variance revealed a significant interaction between the type of stimulant given and the haloperidol treatment on the change in time spent in the non-start compartment from pretest to test [3-way interaction, F(2,64)= 6.591, P < 0.05]. Rats pretreated with vehicle prior to AMP injections on conditioning days, and vehicle prior to vehicle injections on control days, exhibited a significant increase in the time spent in the AMP-paired compartment on the post-conditioning test relative to the mean of the three pretests [F(1,64) = 24.6, P < 0.0001]. No such increase was observed in animals pretreated with haloperidol prior to AMP injections, regardless of whether or not they also were given haloperidol injections on control days [pretreatment only prior to AMP: F(1,64)=1.18, P > 0 . 1 or on both AMP and control days : F(1,64) = 2.25, P > 0.1]. In contrast, animals conditioned with MPD exhibited significant increases in the time spent in the MPD-paired
250 Table 1. Mean (4-_SE) number of crossings between compartments before (pretests) and after (test) conditioning
28002400 -
Treatment
Mean of 3 pretests
Test day
c 2000 E
o
Veh + Veh/Veh + Veh* Veh + AMP/Veh + Veh HAL + AMP/Veh + Veh HAL + AMP/HAL + Veh Veh + MPD/Veh + Veh HAL + MPD/Veh + Veh HAL + MPD/HAL + Veh
9.9±1.2 8.7±2.1 9.0±5.0 50.3±1.1 8.8±5.3 10.6±1.9 7.8±1.7
7.8±1.0 9.5±1.9 7.9±0.9 8.6±1.3 9.3±1.4 52.4±2.2 9.2±1.5
"~ 600y///~
+
Day 2
Day 3
Day 4
v//z
I
haloperidol, at the present dose, attenuates place preferences induced by AMP, but not by MPD. There was no change in the number of crossings for any of the groups, relative to pretest (Table 1).
200100
600 "6 '~ 400
Fig. 4. Locomotor activity counts during 30 rain on all 4 days of drug conditioning (S+) following AMP (1.5 mg/kg, KS3) MPD (5 mg/kg, W/A), haloperidol (0.2 mg/kg)+ AMP ( ~ ) , or haloperidol + MPD (NN). Rats were pretreated with haloperidoI or vehicle 60 rain before stimulant injection. *** P < 0.001 Haloperidol pretreatment with AMP and MPD significantly different from stimulant alone. * P
