10
Behavioural Brain Research 116 (2000) 187 – 196 Research report Involvement of nitric oxide in phencyclidine-induced place aversion and preference in mice Yoshiaki Miyamoto a , Yukihiro Noda a , Yumiko Komori b , Hishayoshi Sugihara b , Hiroshi Furukawa c , Toshitaka Nabeshima a, * a Department of Neuropsychopharmacology and Hospital Pharmacy, Nagoya Uni6ersity Graduate School of Medicine, 65 Tsuruma -cho, Showa -ku, Nagoya, Japan b Department of Microbiology, Faculty of Pharmaceutical Sciences, Meijo Uni6ersity, 150 Yagotoyama, Tenpaku -ku, Nagoya, Japan c Department of Medicinal Chemistry, Faculty of Pharmaceutical Sciences, Meijo Uni6ersity, 150 Yagotoyama, Tenpaku -ku, Nagoya, Japan Received 3 April 2000; received in revised form 19 June 2000; accepted 19 June 2000 Abstract The present study investigated the involvement of nitric oxide (NO) in phencyclidine (PCP)-induced place aversion and preference in the place conditioning paradigm. PCP-induced place aversion in naive mice was dose-dependently attenuated by administration of N G -nitro-L-arginine methyl ester (L-NAME), a NO synthase (NOS) inhibitor, during the conditioning. The NOS activity and dopamine (DA) turnover in the hippocampus in mice showing PCP-induced place aversion were decreased, such changes being restored by administration of L-NAME during the conditioning. On the other hand, PCP-induced place preference in mice pretreated with PCP for 28 days was not attenuated by administration of L-NAME during the conditioning. Although NOS activity was not changed, the DA turnover in the cerebral cortex was increased in mice showing PCP-induced place preference. In mice pretreated with L-NAME and PCP for 28 days before the place conditioning paradigm, PCP neither induced place preference, nor changed the NOS activity or DA turnover. These results suggest that NO is involved in the acquisition of PCP-induced aversive effects, and in the development of PCP-induced preferred effects. Further, the functional change of the DAergic neuronal system mediated by NO in the hippocampus and cerebral cortex may be necessary for the expression of aversive effects and development of preferred effects, respectively, induced by PCP. © 2000 Elsevier Science B.V. All rights reserved. Keywords: Nitric oxide; Phencyclidine; Place aversion; Place preference; Dopaminergic neuronal system; Mouse www.elsevier.com/locate/bbr 1. Introduction Nitric oxide (NO), discovered as a neurotransmitter in the central nervous system [11], is an important messenger molecule in a number of organ systems [33,59]. NO is a gaseous compound formed by NO synthase (NOS) in a Ca 2 + /calmodulin-dependent reac- tion between L-arginine and oxygen [46]. NOS activa- tion has been considered a biochemical event resulting from N -methyl-D-aspartate (NMDA) receptor channel activation [5,28,57,58]. Namely, glutamate-induced opening of the NMDA receptor channel results in Ca 2 + influx, which in turn activates calmodulin, and then calmodulin activation causes a stimulation of NOS, which converts from L-arginine and oxygen to NO and L-citrulline [36,57]. The non-competitive NMDA receptor antagonist phencyclidine [1-(1-phenyl- cyclohexyl)piperidine; PCP] binds to PCP binding sites of NMDA receptor channel complexes [19,21,22,30,56], and blocks NMDA receptor channel activation in the glutamatergic neuronal system. Thus, PCP has an influ- ence on NOS activation in the central nervous system [44]. However, PCP affects not only NMDA receptor channels in the glutamatergic neuronal system, but also several neuronal systems including the dopaminergic (DAergic) and the serotonergic (5-HTergic) neuronal systems [10,14,38,51,52,54]. * Corresponding author. Tel.: +81-52-7442674; fax: +81-52- 7442979. E-mail address: [email protected] (T. Nabeshima). 0166-4328/00/$ - see front matter © 2000 Elsevier Science B.V. All rights reserved. PII:S0166-4328(00)00274-6

Involvement of nitric oxide in phencyclidine-induced place aversion and preference in mice

Embed Size (px)

Citation preview

Behavioural Brain Research 116 (2000) 187–196

Research report

Involvement of nitric oxide in phencyclidine-induced place aversionand preference in mice

Yoshiaki Miyamoto a, Yukihiro Noda a, Yumiko Komori b, Hishayoshi Sugihara b,Hiroshi Furukawa c, Toshitaka Nabeshima a,*

a Department of Neuropsychopharmacology and Hospital Pharmacy, Nagoya Uni6ersity Graduate School of Medicine, 65 Tsuruma-cho,Showa-ku, Nagoya, Japan

b Department of Microbiology, Faculty of Pharmaceutical Sciences, Meijo Uni6ersity, 150 Yagotoyama, Tenpaku-ku, Nagoya, Japanc Department of Medicinal Chemistry, Faculty of Pharmaceutical Sciences, Meijo Uni6ersity, 150 Yagotoyama, Tenpaku-ku, Nagoya, Japan

Received 3 April 2000; received in revised form 19 June 2000; accepted 19 June 2000

Abstract

The present study investigated the involvement of nitric oxide (NO) in phencyclidine (PCP)-induced place aversion andpreference in the place conditioning paradigm. PCP-induced place aversion in naive mice was dose-dependently attenuated byadministration of NG-nitro-L-arginine methyl ester (L-NAME), a NO synthase (NOS) inhibitor, during the conditioning. TheNOS activity and dopamine (DA) turnover in the hippocampus in mice showing PCP-induced place aversion were decreased, suchchanges being restored by administration of L-NAME during the conditioning. On the other hand, PCP-induced place preferencein mice pretreated with PCP for 28 days was not attenuated by administration of L-NAME during the conditioning. AlthoughNOS activity was not changed, the DA turnover in the cerebral cortex was increased in mice showing PCP-induced placepreference. In mice pretreated with L-NAME and PCP for 28 days before the place conditioning paradigm, PCP neither inducedplace preference, nor changed the NOS activity or DA turnover. These results suggest that NO is involved in the acquisition ofPCP-induced aversive effects, and in the development of PCP-induced preferred effects. Further, the functional change of theDAergic neuronal system mediated by NO in the hippocampus and cerebral cortex may be necessary for the expression of aversiveeffects and development of preferred effects, respectively, induced by PCP. © 2000 Elsevier Science B.V. All rights reserved.

Keywords: Nitric oxide; Phencyclidine; Place aversion; Place preference; Dopaminergic neuronal system; Mouse

www.elsevier.com/locate/bbr

1. Introduction

Nitric oxide (NO), discovered as a neurotransmitterin the central nervous system [11], is an importantmessenger molecule in a number of organ systems[33,59]. NO is a gaseous compound formed by NOsynthase (NOS) in a Ca2+/calmodulin-dependent reac-tion between L-arginine and oxygen [46]. NOS activa-tion has been considered a biochemical event resultingfrom N-methyl-D-aspartate (NMDA) receptor channelactivation [5,28,57,58]. Namely, glutamate-induced

opening of the NMDA receptor channel results inCa2+ influx, which in turn activates calmodulin, andthen calmodulin activation causes a stimulation ofNOS, which converts from L-arginine and oxygen toNO and L-citrulline [36,57]. The non-competitiveNMDA receptor antagonist phencyclidine [1-(1-phenyl-cyclohexyl)piperidine; PCP] binds to PCP binding sitesof NMDA receptor channel complexes [19,21,22,30,56],and blocks NMDA receptor channel activation in theglutamatergic neuronal system. Thus, PCP has an influ-ence on NOS activation in the central nervous system[44]. However, PCP affects not only NMDA receptorchannels in the glutamatergic neuronal system, but alsoseveral neuronal systems including the dopaminergic(DAergic) and the serotonergic (5-HTergic) neuronalsystems [10,14,38,51,52,54].

* Corresponding author. Tel.: +81-52-7442674; fax: +81-52-7442979.

E-mail address: [email protected] (T. Nabeshima).

0166-4328/00/$ - see front matter © 2000 Elsevier Science B.V. All rights reserved.PII: S 0 1 6 6 -4328 (00 )00274 -6

Y. Miyamoto et al. / Beha6ioural Brain Research 116 (2000) 187–196188

PCP was originally developed as an anesthetic agentin surgery, but fell into disuse due to its psychoto-mimetic effects. It has since become a common drug ofabuse in the USA, since it produces both psychologicaland physical dependence in humans [47]. In the placeconditioning paradigm, which is widely used for deter-mining the aversive and preferred effects of drugs inanimals [49], PCP produces place aversion in naiverodents [3,18,26], whereas it produces place preferencein rodents pretreated with PCP repeatedly [25,39,42].These phenomena observed in animals are similar tothose in humans; although a single use of PCP pro-duces aversive effects, long-term use has rewardingeffects in humans [16]. We have previously found thatPCP-induced place aversion in rats is attributed tointeraction with the 5-HTergic neuronal system [26,39],and that PCP-induced place preference in mice is de-pendent on the DAergic neuronal system, particularly,the D1 receptors [42]. However, the mechanisms respon-sible for PCP-induced place aversion and preference arenot yet clear.

Recent studies have indicated that NO is involved inthe behavioral effects of PCP [20,43,44,55]. For exam-ple, NOS inhibitor prevents the PCP-induced impair-ment of prepulse inhibition and hyperlocomotion [20],and enhances the PCP-induced ataxia, but not sniffingand head movement [43]. NO plays an important rolein the development, but not in the maintenance, oftolerance and sensitization to the effects of PCP [44].Further, other recent studies have indicated that NO isinvolved in the preferred effect of cocaine [17,23], apsychostimulant like PCP, in the place conditioningparadigm. Thus, there is a possibility that NO is in-volved in PCP-induced place aversion and preference inthe place conditioning paradigm.

Accordingly, we attempted to investigate the involve-ment of NO in PCP-induced place aversion and prefer-ence in the place conditioning paradigm. The roles ofNO were investigated in two different phases, in the

‘acquisition phase’ during the conditioning to PCP andin the ‘development phase’ during the repeated PCPpretreatment for 28 days before the conditioning toPCP in the place conditioning paradigm (Table 1).Further, we measured the NOS activity, and DA and5-HT turnovers in various brain regions in mice show-ing PCP-induced place aversion and preference as the‘expression phase’ of aversive and preferred effects,respectively, of PCP (Table 1). NO has been function-ally linked to DAergic and 5-HTergic neurotransmis-sions in the brain, which are implicated in the aversiveand preferred effects of drugs.

2. Materials and methods

2.1. Animals

Male mice of the ddY strain (Japan SLC, Shizuoka,Japan), weighing 25–28 g at the beginning of theexperiments, were used. The animals were housed inplastic cages and were kept in a regulated environment(2491°C, 5095% humidity), with a 12/12 h light-darkcycle (lights on at 08:00 h). Food (CE2, Clea Japan,Tokyo, Japan) and tap water were available ad libitum.

All experiments were performed in accordance withthe Guidelines for Animal Experiments of the NagoyaUniversity School of Medicine. The procedures involv-ing animals and their care were conducted in confor-mity with the international guidelines ‘Principles oflaboratory animal care’ (NIH publication No. 85-23,revised 1985).

2.2. Drugs

Phencyclidine HCl [(1-(1-phenylcyclohexyl)piperidinehydrochloride; PCP] was synthesized by the authorsaccording to a report by Maddox et al. [35] and waschecked for purity. NG-nitro-L-arginine methyl ester

Table 1Experimental schedule

Acquisition phase Expression phaseDevelopment phasePlace conditioningparadigm

Post-conditioning testConditioning (mg/kg)Pretreatment for 28 days(mg/kg/day)

Measurement of NOS activity and of monoamine– PCP (8) and/orPCP-inducedplace aversion L-NAME (0.5–50) metabolism

D-NAME (50)PCP-induced PCP (10) PCP (8) and/or Measurement of NOS activity and of monoamine

L-NAME (5 or 50) metabolismplace preferenceD-NAME

PCP (10) and/or PCP (8) Measurement of NOS activity and of monoamineL-NAME (50) metabolismD-NAME (50)

Y. Miyamoto et al. / Beha6ioural Brain Research 116 (2000) 187–196 189

(L-NAME) and NG-nitro-D-arginine methyl ester (D-NAME) were purchased from Sigma Chemical (St.Louis, MO, USA). Other agents were obtained fromstandard commercial sources. PCP, L-NAME and D-NAME were dissolved in saline (0.9% NaCl).

2.3. Place conditioning paradigm

2.3.1. Experimental apparatusThe apparatus used for the place conditioning

paradigm consisted of two compartments: a black Plex-iglass box and a transparent Plexiglass box (both 15×15×15 cm high) with a metal grid floor. To enable themice to distinguish easily the transparent box from theblack one, the floor of the transparent and black boxeswere covered with white plastic mesh and with frostingPlexiglass, respectively. Each box could be divided by asliding door (10×15 cm high).

2.3.2. Experimental procedurePre-conditioning test: The place conditioning

paradigm was performed according to the method ofNoda et al. [42]. In the pre-conditioning test, the slidingdoor was opened and the mouse was allowed to movefreely between both boxes for 15 min once a day for 3days. On day 3, we measured the number of secondsthat the mouse spent in the black and transparent boxesby using a Scanet SV-10 LD (Toyo Sangyo, Toyama,Japan). The box in which the mouse spent the mosttime was referred to as the ‘preferred side’, and theother box the ‘non-preferred side’.

Conditioning: conditioning was performed during sixsuccessive days. Mice were given drugs or vehicle inthe apparatus with the sliding door closed. On days4, 6 and 8, a mouse was given PCP and placed in itspreferred side when investigating the aversive effectsof PCP or its non-preferred side when investigatingthe preferred effects of it for 20 min. On days 5, 7and 9, the mouse was given saline and placed oppo-site to the drug-conditioning side for 20 min.Post-conditioning test: On day 10 (the next day ofconditioning), a post-conditioning test was per-formed without drug treatment. In the post-condi-tioning test, the sliding door was opened, and wemeasured the number of seconds that the mousespent in the black and transparent boxes for 15 minusing the Scanet SV-10 LD.

2.3.3. Drug administrationPCP (8 mg/kg s.c.) was injected immediately before

the conditioning according to our previous report [42].L-NAME (0.5, 5 or 50 mg/kg i.p.) and D-NAME (50mg/kg i.p.) were administered 60 min before everytreatment with PCP. These drugs were administered forthree alternating days in the 6 day-conditioning period,and corresponding vehicles were administered for the

other 3 days (acquisition phase of PCP-induced placeaversion and preference in Table 1).

In the experiment of repeated administration of PCP,mice were administered PCP (10 mg/kg/day s.c.) for 28days in the home cage as in a previous report [42]. Inthe experiment of repeated administration of L-NAMEor D-NAME and PCP, mice were administered L-NAME (5 and 50 mg/kg/day i.p.) or D-NAME (50mg/kg/day i.p.) 60 min before every treatment withPCP (10 mg/kg/day s.c.) for 28 days in the home cage(development phase of PCP-induced place preference inTable 1).

2.3.4. Data analysisPlace conditioning behaviors were expressed by

Post−Pre, which was calculated as: ((post value)−(pre value)), where post and pre values were the differ-ence in the number of seconds spent in thedrug-conditioning and the saline-conditioning sites dur-ing the post-conditioning and pre-conditioning tests,respectively.

2.4. NOS acti6ity, and DA and 5-HT turno6er assays

Immediately after the post-conditioning test (expres-sion phase of PCP-induced place aversion and prefer-ence in Table 1), on the second day of the finaladministration of PCP and/or L-NAME in the condi-tioning, drug-treated mice were sacrificed by decapita-tion, and divided into two groups to determine theNOS activity and monoamine (DA and 5-HT)turnover. Brains were rapidly removed and the cerebralcortex, hippocampus and striatum were dissected outon an ice-cold plate according to the method ofGlowinski and Iversen [12]. Each tissue sample wasquickly frozen and stored in a deep freezer at −80°Cuntil assayed.

2.4.1. Measurement of NOS acti6ityUsing tissue samples, brain NOS activity was deter-

mined as described previously [5], with a minor modifi-cation [29]. Each frozen tissue sample was weighed,then homogenized with an ultrasonic processor (475 W,Model XL2020, Heat Systems, New York City, USA)in 5 vol. (w/v) of 50 mM Tris-HCl buffer (pH 7.4)containing 0.1 mM EDTA, 0.1 mM EGTA, 1 mMpepstatin, 2 mM leupeptin, 1 mM phenylmethylsulpho-nyl fluoride and 0.5 mM dithiothreitol. The ho-mogenate was centrifuged at 20 000×g for 45 min at4°C and the supernatant was used in the assay. NOSactivity was measured by monitoring the conversion of[3H]-arginine to [3H]-citrulline. Briefly, the supernatantwas incubated for 12 min at 37°C in a final volume of100 ml NADPH containing 50 ml L-arginine, 2 mMCaCl2, 0.3 mg calmodulin, 10 mM tetrahydrobiopterinand 200 000 d.p.m. of L-[3H]-arginine. The assays were

Y. Miyamoto et al. / Beha6ioural Brain Research 116 (2000) 187–196190

Fig. 1. Effect of L-NAME and D-NAME on the acquisition of place aversion induced by PCP in mice. L-NAME (0.5, 5 or 50 mg/kg i.p.) orD-NAME (50 mg/kg i.p.) was administered 60 min before PCP (8 mg/kg s.c.) treatment during the conditioning in the place conditioningparadigm. Each column represents the mean9SEM. Numbers in parentheses are the number of animals used. **PB0.01 versus (saline+saline)-treated group. �PB0.05, ��PB0.01 versus (saline+PCP)-treated group.

terminated by the addition of 2 ml of ice-cold acetatebuffer (pH 5.5) containing 1 mM citrulline, 2 mMEDTA and 0.2 mM EGTA. The sample was applied to1-ml columns of Dowex AG50-X8 (Na+ form) and theeluate was collected. The column was then furthereluted with 2 ml of water. [3H]-Citrulline in the com-bined eluate was quantified with a liquid scintillationspectrometer. Protein content was determined accord-ing to the method of Lowry et al. [34], with bovineserum albumin used as standard.

2.4.2. Measurement of DA and 5-HT turno6ersUsing tissue samples, the contents of DA, 5-HT and

their metabolites were determined by using a highpressure liquid chromatography (HPLC) system withan electrochemical detector (Eicom, Kyoto, Japan) asdescribed [41]. Each frozen tissue sample was weighed,then homogenized with an ultrasonic processor in 350ml of 0.2 M perchloric acid containing isoproterenol asinternal standard. The homogenate was placed in icefor 30 min and then centrifuged at 20 000×g for 15min at 4°C. The supernatant was mixed with 1 Msodium acetate to adjust the pH to 3.0 and then in-jected into a liquid chromatography system equippedwith a reversed-phase ODS-column [4.6×150 mm, Ei-compak MA-5 ODS (diameter of stationary phasegrains; 5 mm), Eicom] and an electrochemical detector(Model ECD-100, Eicom). The column temperaturewas maintained at 25°C and the detector potential wasset at +750 mV. The mobile phase was 0.1 M citricacid and 0.1 M sodium acetate, pH 3.9, containing 14%methanol, 160 mg/l sodium-L-octanesulfonate and 5mg/l EDTA: the flow rate was 1 ml/min. The turnoversof DA and 5-HT were calculated from the contents ofDA, 5-HT and their metabolites determined by HPLC.

2.5. Statistical analysis

All data were expressed as the mean9SEM Statisti-cal differences among values for individual groups weredetermined by one-way analysis of variance (ANOVA),followed by the Student-Newmann-Keuls multiple com-parisons test when F ratios were significant (PB0.05).

3. Results

3.1. Effect of L-NAME and D-NAME on theacquisition of PCP-induced place a6ersion

As shown in Fig. 1, PCP (8 mg/kg s.c.) significantlyinduced place aversion in naive mice consistent withour previous report. The acquisition of PCP-inducedplace aversion was dose-dependently and significantlyattenuated by coadministration of L-NAME (0.5–50mg/kg i.p.) with PCP during the conditioning. How-ever, the coadministration of D-NAME (50 mg/kg i.p.)with PCP did not modify place aversion induced byPCP. L-NAME (50 mg/kg i.p.) itself did not produceeither place aversion or place preference.

3.2. NOS acti6ity in the brain of mice showingPCP-induced place a6ersion

Immediately after the post-conditioning test, NOSactivity in the brain was measured (Fig. 2). The mostNOS activity in mice treated with saline was found inthe hippocampus, followed by the cerebral cortex andstriatum. The NOS activity in the hippocampus of micetreated with L-NAME alone was not different fromthat of control mice in the post-conditioning test. How-

Y. Miyamoto et al. / Beha6ioural Brain Research 116 (2000) 187–196 191

ever, the NOS activity in the hippocampus was signifi-cantly lower in mice showing PCP-induced place aver-sion than in mice treated with saline. This change ofNOS activity in the hippocampus was restored by coad-

ministration of L-NAME (50 mg/kg) with PCP duringthe conditioning. NOS activity in the cerebral cortexand striatum did not significantly change in any drug-treated mice.

3.3. DA and 5-HT turno6ers in the hippocampus ofmice showing PCP-induced place a6ersion

DA and 5-HT turnovers in the hippocampus immedi-ately after the post-conditioning test are shown in Fig.3. The ratios of 3,4-dihydroxyphenylacetic acid (DO-PAC) and homovanillic acid (HVA) to DA (DOPAC/DA and HVA/DA) in mice showing PCP-induced placeaversion, were significantly decreased to 63.8 and71.5%, respectively, compared with those in micetreated with saline. This decrease in the ratios of DO-PAC/DA and HVA/DA was reversed by coadministra-tion of L-NAME (50 mg/kg) with PCP during theconditioning. L-NAME alone did not change the ratiosof DOPAC/DA and HVA/DA. The ratio of 5-hydrox-yindoleacetic acid (5-HIAA) to 5-HT (5-HIAA/5-HT)was unaffected in all drug-treated mice.

DA and 5-HT turnovers in the cerebral cortex andstriatum in drug-treated mice did not change signifi-cantly compared with those in mice treated with saline(data not shown).

3.4. Effect of L-NAME and D-NAME on theacquisition and on the de6elopment of PCP-inducedplace preference

As shown in Fig. 4, PCP (8 mg/kg s.c.) significantlyinduced place preference in mice pretreated with PCP(10 mg/kg/day s.c.) for 28 days. When PCP was admin-istered in combination with L-NAME (5 and 50 mg/kgi.p.) or D-NAME (50 mg/kg i.p.) during the condition-ing, the aquisition of PCP-induced place preference wasnot affected (Fig. 4A). However, in mice pretreatedwith L-NAME (50 mg/kg/day i.p.), but not with D-NAME (50 mg/kg/day i.p.), and PCP for 28 days,PCP-induced place preference was not developed (Fig.4B). In mice pretreated with L-NAME alone for 28days, PCP induced neither place aversion nor placepreference (Fig. 4B).

3.5. NOS acti6ity in the brain of mice showingPCP-induced place preference

Immediately after the post-conditioning test, NOSactivity in the brain was measured (Fig. 5). The NOSactivity in the hippocampus, cerebral cortex and stria-tum of mice treated with drugs during the conditioning(Fig. 5A) and during the pretreatment for 28 daysbefore the place conditioning paradigm (Fig. 5B), didnot significantly change compared with those in micetreated with saline.

Fig. 2. Nitric oxide synthase activity in the brain of mice showingPCP-induced place aversion. Mice were treated with saline, PCP (8mg/kg s.c.), L-NAME (50 mg/kg i.p.)+PCP (8 mg/kg s.c.) orL-NAME (50 mg/kg i.p.) during the conditioning in the place condi-tioning paradigm. Immediately after the post-conditioning test on thesecond day of the final administration of PCP and/or L-NAME, eachmouse was sacrificed, and then NOS activity was determined. Eachcolumn represents the mean9SEM from eight samples. *PB0.05versus (saline+saline)-treated group. �PB0.05 versus (saline+PCP)-treated group.

Fig. 3. Dopamine and serotonin turnovers in the hippocampus ofmice showing PCP-induced place aversion. Mice were treated withsaline, PCP (8 mg/kg s.c.), L-NAME (50 mg/kg i.p.)+PCP (8 mg/kgs.c.) or L-NAME (50 mg/kg i.p.) during the conditioning in the placeconditioning paradigm. Immediately after the post-conditioning teston the second day of the final administration of PCP and/or L-NAME, each mouse was sacrificed, and then the monoamineturnover was determined. Each column represents the mean9SEMfrom seven samples. **PB0.01, ***PB0.001 versus (saline+saline)-treated group. �PB0.05, ��PB0.01 versus (saline+PCP)-treatedgroup.

Y. Miyamoto et al. / Beha6ioural Brain Research 116 (2000) 187–196192

Fig. 4. Effect of L-NAME and D-NAME on the acquisition (A) andon the development (B) of place preference induced by PCP in mice.(A) On the day after the final injection of PCP (10 mg/kg/day s.c.) for28 days, the place conditioning paradigm was performed as describedin Section 2. L-NAME (5 or 50 mg/kg i.p.) or D-NAME (50 mg/kgi.p.) was administered 60 min before PCP (8 mg/kg s.c.) treatmentduring the conditioning in the place conditioning paradigm. Eachcolumn represents the mean9SEM. Numbers in parentheses are thenumber of animals used. **PB0.01 versus (saline/saline+saline)-treated group. (B) On the day after the final injection of PCP (10mg/kg/day s.c.), L-NAME (50 mg/kg/day i.p.)+PCP (10 mg/kg/days.c.), L-NAME (50 mg/kg/day i.p.) or D-NAME (50 mg/kg/day i.p.)for 28 days, the place conditioning paradigm was performed asdescribed in the Section 2. PCP (8 mg/kg s.c.) was administeredduring the conditioning in the place conditioning paradigm. Eachcolumn represents the mean9SEM. Numbers in parentheses are thenumber of animals used. *PB0.05 versus (saline+saline/saline)-treated group. �PB0.05 versus (saline+PCP/PCP)-treated group.

Fig. 6. The ratios of DOPAC/DA and HVA/DA inmice showing PCP-induced place preference, whichwere treated with PCP and/or L-NAME (50 mg/kg)during the conditioning, were significantly increasedcompared with those in mice treated with saline (Fig.6A). Although the ratios of DOPAC/DA and HVA/DA in mice pretreated with PCP for 28 days before theplace conditioning paradigm were significantly in-creased, such increases in the cerebral cortex were

Fig. 5. Nitric oxide synthase activity in the brain of mice showingPCP-induced place preference. (A) Mice were pretreated with PCP(10 mg/kg/day s.c.) for 28 days, and then treated with saline, PCP (8mg/kg s.c.) or L-NAME (50 mg/kg i.p.)+PCP (8 mg/kg s.c.) duringthe conditioning in the place conditioning paradigm. Immediatelyafter the post-conditioning test on the second day of the finaladministration of PCP and/or L-NAME, each mouse was sacrificed,and then NOS activity was determined. Each column represents themean9SEM from eight samples. (B) Mice were pretreated withsaline, PCP (10 mg/kg/day s.c.), L-NAME (50 mg/kg/day i.p.)+PCP(10 mg/kg/day s.c.) or L-NAME (50 mg/kg/day i.p.) for 28 days, andthen treated with PCP (8 mg/kg s.c.) during the conditioning in theplace conditioning paradigm. Immediately after the post-conditioningtest on the second day of the final administration of PCP, each mousewas sacrificed, and then NOS activity was determined. Each columnrepresents the mean9SEM from eight samples.

3.6. DA and 5-HT turno6ers in the cerebral cortex ofmice showing PCP-induced place preference

DA and 5-HT turnovers in the cerebral cortex imme-diately after the post-conditioning test are shown in

Y. Miyamoto et al. / Beha6ioural Brain Research 116 (2000) 187–196 193

Fig. 6. Dopamine and serotonin turnovers in the cerebral cortex ofmice showing PCP-induced place preference. (A) Mice were pre-treated with PCP (10 mg/kg/day s.c.) for 28 days, and then treatedwith saline, PCP (8 mg/kg s.c.) or L-NAME (50 mg/kg i.p.)+PCP (8mg/kg s.c.) during the conditioning in the place conditioningparadigm. Immediately after the post-conditioning test on the secondday of the final administration of PCP and/or L-NAME, each mousewas sacrificed, and then NOS activity was determined. Each columnrepresents the mean9SEM from seven samples. *PB0.05 versus(saline/saline+saline)-treated group. (B) Mice were pretreated withsaline, PCP (10 mg/kg/day s.c.), L-NAME (50 mg/kg/day i.p.)+PCP(10 mg/kg/day s.c.) or L-NAME (50 mg/kg/day i.p.) for 28 days, andthen treated with PCP (8 mg/kg s.c.) during the conditioning in theplace conditioning paradigm. Immediately after the post-conditioningtest on the second day of the final administration of PCP, each mousewas sacrificed, and then the monoamine turnover was determined.Each column represents the mean9SEM from seven samples. *PB0.05 versus corresponding (saline+saline/saline)-treated group. �PB0.05 versus (saline+PCP/PCP)-treated group.

conditioning and during the pretreatment for 28 daysbefore the place conditioning paradigm (Fig. 6A andB).

DA and 5-HT turnovers in the striatum andhippocampus in drug-treated mice did not change sig-nificantly compared with those in mice treated withsaline (data not shown).

4. Discussion

Several studies have reported that PCP producesplace aversion in naive rodents [3,18,26] and placepreference in rodents pretreated with PCP repeatedly[25,39,42] in the place conditioning paradigm. Thesephenomena are consistent with those in humans; al-though a single use of PCP produces aversive effects,long-term use has rewarding effects in humans [16],indicating that PCP has aversive and preferred effects.Acute and subchronic treatment with PCP has beendemonstrated to produce several functional alterationsin neuronal systems such as changes in DA turnover,5-HT turnover [37], NOS activity [44] and the densitiesof NMDA, alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA), benzodiazepine and mus-carinic receptors [9]. Thus, there is a possibility thatsuch functional alterations are involved in PCP-inducedaversive and/or preferred effects. In the place condition-ing paradigm, NO is involved in the acquisition ofdrug-induced place preference [17,23,27]. For example,the coadministration of NOS inhibitor 7-nitroindazoleand L-N-nitroarginine during the conditioning, attenu-ates place preference induced by cocaine [17] and mor-phine [27], respectively, in rodents. Mice deficient forthe neuronal NOS gene are resistant to cocaine-inducedplace preference [17]. In the present study, to clarify theroles of NO in PCP-induced place aversion and prefer-ence, we investigated the effects of a NOS inhibitor inthe ‘acquisition phase’ during the conditioning to PCPand in the ‘development phase’ during repeated PCPpretreatment for 28 days before the conditioning toPCP in the place conditioning paradigm (Table 1).Further, we measured the NOS activity and mono-amine turnover in the brain in mice showing PCP-in-duced place aversion and preference as the ‘expressionphase’ of aversive and preferred effects, respectively, ofPCP (Table 1).

In the experiment to investigate the involvement ofNO in the acquisition of PCP-induced place aversion,we confirmed previous reports that PCP produces placeaversion in naive mice [3,18,26], and found that theacquisition of PCP-induced place aversion was blockedby coadministration of L-NAME, a NOS inhibitor,with PCP, whereas it was not blocked by D-NAME, aless active enantiomer of L-NAME. These findingssuggest that NO is involved in the acquisition of aver-

restored by co-pretreatment with L-NAME and PCPfor 28 days (Fig. 6B). The ratios of DOPAC/DA andHVA/DA in mice pretreated with L-NAME alone for28 days did not change (Fig. 6B). The ratio of 5-HIAA/5-HT was unaffected in all drug-treated mice during the

Y. Miyamoto et al. / Beha6ioural Brain Research 116 (2000) 187–196194

sive effects of PCP. Several lines of evidence haveindicated that the 5-HTergic neuronal system plays animportant role in the aversive effects of drugs in theplace conditioning paradigm [1,2,6,26,39,40,48]. For in-stance, the 5-HT2 receptor antagonist, ritanserin [39]and the 5-HT3 receptor antagonists, ICS 205-903 andMDL 72222 [2], attenuate the acquisition of PCP-in-duced place aversion. Further, the 5-HT2 receptor ago-nists, 2,5-dimethoxy-4-iodoamphetamine and1-(3-chlorophenyl) piperazine, induce place aversion inmice (unpublished data). Thus, the aversive effects ofPCP may be acquired by action of the 5-HTergicneuronal system mediated via the NO system. Thishypothesis may be supported by reports that NO re-lease-inducing substances increase the extracellular con-centration of 5-HT in the rat brain [13,32]. Since themechanisms of interaction between the NO system and5-HTergic neuronal system are unclear, further studiesare necessary to clarify the biochemical mechanisms inthe acquisition of PCP-induced place aversion.

Biochemical changes were observed in mice showingPCP-induced place aversion. In the hippocampus ofmice treated with PCP in the conditioning, a significantdecrease of NOS activity was observed. In addition, theanalysis of the neurochemical effects revealed a reduc-tion of DA turnover, but not 5-HT turnover in thesame area. No such biochemical changes of NOS activ-ity and DA turnover in the hippocampus were recog-nized in mice treated with L-NAME and PCP, whichdid not show PCP-induced place aversion. These resultssuggest that the biochemical changes in the hippocam-pus such as decrease of NOS activity and DA turnoverare necessary for the expression of the aversive effectsof PCP. The 5-HT turnover in the hippocampus, cere-bral cortex and striatum in all drug-treated mice wasunchanged compared with in mice treated with saline,suggesting that 5-HT turnover is not involved in theexpression of the aversive effects of PCP. Thus, theabove findings indicate that different neuronal mecha-nisms mediate the acquisition and expression of PCP-induced place aversion. Namely, although NO isinvolved in both the acquisition and the expression, theformer may be mediated by the 5-HTergic neuronalsystem, the latter by the DAergic neuronal system.

In the next experiments, we investigated the involve-ment of NO in PCP-induced place preference. Thepresent results were consistent with our previous re-ports that PCP produces preferred effects in mice pre-treated with PCP for 28 days [25,39,42]. ThePCP-induced place preference was not blocked bycoadministration of L-NAME with PCP during theconditioning in mice pretreated with PCP alone,whereas PCP did not induce place preference in micepretreated with L-NAME and PCP for 28 days beforethe place conditioning paradigm. Thus, some functionalalterations occurred to the neuronal system via the NO

system during the treatment with PCP for 28 daysbefore the place conditioning paradigm. These findingsdemonstrate that NO is involved in the development,but not the acquisition, of the preferred effects of PCP.

In the biochemical analysis on the expression of thepreferred effects of PCP, NOS activity in the brain inmice showing PCP-induced place preference was notchanged by any drug treatment either during the condi-tioning or before the place conditioning paradigm.These results indicate that NO may not be involved inthe expression of the preferred effects of PCP. The DAturnover in the cerebral cortex was increased in miceshowing PCP-induced place preference, which weretreated with not only PCP, but also L-NAME and PCPduring the conditioning (Fig. 4A and 6A). In addition,only mice showing PCP-induced place preference hadan increase of DA turnover (Fig. 4B and 6B). However,such a change may not be reflected by the biochemicalchanges in the expression of the preferred effects ofPCP, because the increase in the DA turnover in micepretreated with PCP was consistent with a report thatrepeated PCP pretreatment produces an increase of DAturnover in the rat brain [37]. The DAergic neuronalsystem has been demonstrated important to the pre-ferred and abuse properties of drugs in the place condi-tioning paradigm [31]. Our previous study indicatedsimilar findings; that PCP-induced place preference inmice pretreated with PCP for 28 days is mediated viathe DAergic neuronal system [42]. Further, severalstudies have described that there is an interaction be-tween the NO system and DAergic neuronal system[4,7,15,24,45,50,60]. For example, endogenous NO maybe involved in the regulation of DA turnover [45,60],and may influence the activity of DA transporters [24].In the present study, in mice pretreated with PCP andL-NAME for 28 days, we could find neither PCP-in-duced place preference, nor an increase in the DAturnover. Thus, the functional up-regulation of theDAergic neuronal system induced by repeated PCPpretreatment may be mediated via the NO system.Further, along with the behavioral experiments on thedevelopment of PCP-induced place preference, the bio-chemical experiments suggest that NO is involved in thedevelopment of preferred effects of PCP. Ellison andco-workers have reported that long-term treatment withPCP induced a decrease in the extent of AMPA bindingin the rat brain [9]. NO release is associated withaltered sensitivity of AMPA receptors [8]. Thus, theinteraction between the AMPA receptor and NO sys-tem following the repeated PCP treatment may beessential for the development of PCP-induced placepreference. In fact, AMPA receptor has been demon-strated to be involved in morphine and amphetamine-induced place preference in animals [53]. However, thispoint must be considered with caution, as the neu-ropharmacology of PCP and association with the NOsystem remain to be clarified.

Y. Miyamoto et al. / Beha6ioural Brain Research 116 (2000) 187–196 195

In summary, the present study demonstrated thatNO is involved in the acquisition of PCP-induced placeaversion, and in the development, but not acquisition,of PCP-induced place preference. Further, the decreaseof DA turnover due to change of NOS activity in thehippocampus may be necessary for the expression ofPCP-induced aversive effects, and an increase of DAturnover mediated by the NOS system in the cerebralcortex may be essential for the development of PCP-in-duced preferred effects. Thus, the NO plays a role innot only the behavioral effects of PCP, but also thepsychostimulant effects of PCP.

Acknowledgements

This research was partly supported by a Grant-in Aidfor Drug Abuse Research (92-2) from the Ministry ofHealth and Welfare of Japan, by a Grant-in-Aid forCOE Research and Scientific Research (c10044260)from the Ministry of Education, Science, Sports andCulture of Japan, and from INSERM-JSPS Joint Re-search Project, and a Sasakawa Scientific ResearchGrant (9-220).

References

[1] Acquas E, Carboni E, Leone P, Di Chiara G. SCH 23390 blocksdrug-conditioned place-preference and place-aversion: anhedonia(lack of reward) or apathy (lack of motivation) after dopamine-receptor blockade? Psychopharmacology 1989;99:151–5.

[2] Acquas E, Carboni E, Leone P, Di Chiara G. Blockade ofacquisition of drug-conditioned place aversion by 5-HT3 antago-nists. Psychopharmacology 1990;100:459–63.

[3] Barr GA, Paredes W, Bridger WH. Place conditioning withmorphine and phencyclidine; dose dependent effects. Life Sci1985;36:363–8.

[4] Bowyer JF, Clausing P, Gough B, Slikker W, Jr., Holson RR.Nitric oxide regulation of methamphetamine-induced dopaminerelease in caudate/putamen. Brain Res 1995;699:62–70.

[5] Bredt DS, Snyder SH. Nitric oxide mediates glutamate-linkedenhancement of cGMP levels in the cerebellum. Proc Natl AcadSci USA 1989;86:9030–3.

[6] Carboni E, Acquas E, Leone P, Di Chiara G. 5-HT3 antagonistsblock morphine- and nicotine- but not amphetamine-inducedreward. Psychopharmacology 1989;97:175–8.

[7] Desvignes C, Bert L, Vinet L, Denoroy L, Renaud B, Lambas-Senas L. Evidence that the neuronal nitric oxide synthase in-hibitor 7-nitroindazole inhibits monoamine oxidase in the rat: invivo effects on extracellular striatal dopamine and 3,4-dihydrox-yphenylacetic acid. Neurosci Lett 1999;246:5–8.

[8] Dev KK, Morris BJ. Modulation of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) binding sites by nitricoxide. J Neurochem 1994;63:946–52.

[9] Ellison G, Keys A, Noguchi K. Long-term changes in brainfollowing continuous phencyclidine administration: an autora-diographic study using flunitrazepam, ketanserin, mazindol,quinuclidinyl benzilate, piperidyl-3,4-3H(N)-TCP, and AMPAreceptor ligands. Pharmacol Toxicol 1999;84:9–17.

[10] Garey RE, Weisberg LA, Heath RG. PCP(Phencyclidine): anupdate. J Psyched Drugs 1979;9:280–5.

[11] Garthwaite J, Charle SL, Chess-Williams R. Endothelium-derived relaxing factor release on activation of NMDA receptorssuggests role as intracellular messenger in the brain. Nature1988;336:385–8.

[12] Glowinski J, Iversen LL. Regional studies of catecholamines inthe rat brain. I. the disposition of [3H]norepinephrine, [3H]dopamine and [3H]dopa in various regions of the brain. JNeurochem 1966;13:655–69.

[13] Guevara-Guzman R, Emson PC, Kendrick KM. Modulation ofin vivo striatal transmitter release by nitric oxide and cyclicGMP. J Neurochem 1994;62:807–10.

[14] Imperato A, Honora T, Jenssen LH. Dopamine release in thenucleus caudatus and in the nucleus accumbens is under gluta-matergic control through non-NMDA receptors: a study infreely-moving rats. Brain Res 1990;530:223–8.

[15] Inoue H, Arai I, Shibata S, Watanabe S. NG-nitro-L-argininemethyl ester attenuates the maintenance and expression ofmethamphetamine-induced behavioral sensitization and enhance-ment of striatal dopamine release. J Pharmacol Exp Ther1996;277:1424–30.

[16] Issacs SO, Martin P, Washington AJ. Phencyclidine (PCP)abuse. Oral Surg Oral Med Oral Pathol 1986;61:126–9.

[17] Itzhak Y, Martin JL, Black MD, Huang PL. The role ofneuronal nitric oxide synthase in cocaine-induced conditionedplace preference. NeuroReport 1998;9:2485–8.

[18] Iwamoto ET. Place-conditioning properties of mu, kappa andsigma opioid agonists. Alcohol Drug Res 1986;6:327–39.

[19] Javitt DC, Zukin SR. Recent advances in the phencyclidinemodel of schizophrenia. Am J Psych 1991;148:1301–8.

[20] Johansson C, Jackson DM, Svensson L. Nitric oxide synthaseinhibition blocks phencyclidine-induced behavioral effects onprepulse inhibition and locomotor activity in the rat. Psy-chopharmacology 1997;131:167–73.

[21] Jones SM, Snell LD, Johnson KM. Inhibition by phencyclidineof excitatory amino acid-stimulated release of neurotransmitterin the nucleus accumbens. Neuropharmacology 1987;26:173–9.

[22] Jones SM, Snell LD, Johnson KM. Phencyclidine selectivityinhibits N-methyl-D-aspartate-induced hippocampal[3H]norepinephrine release. J Pharmacol Exp Ther1987;240:492–7.

[23] Kim HS, Park WK. Nitric oxide mediation of cocaine-induceddopaminergic behaviors: ambulation-accelerating activity, re-verse tolerance and conditioned place preference in mice. JPharmacol Exp Ther 1995;275:551–7.

[24] Kiss JP, Hennings EC, Zsilla G, Vizi ES. A possible role of nitricoxide in the regulation of dopamine transporter function in thestriatum. Neurochem Int 1999;34:345–50.

[25] Kitaichi K, Noda Y, Hasegawa T, Furukawa H, Nabeshima T.Acute phencyclidine induces aversion, but repeated phency-clidine induces preference in the place conditioning test in rats.Eur J Pharmacol 1996;318:7–9.

[26] Kitaichi K, Noda Y, Miyamoto Y, Numaguchi A, Osawa H,Hasegawa T, Furukawa H, Nabeshima T. Involvement of theserotonergic neuronal system in phencyclidine-induced placeaversion in rats. Behav Brain Res 1999;103:105–11.

[27] Kivastik T, Rutkauskaite J, Zharkovsky A. Nitric oxide synthe-sis inhibition attenuates morphine-induced place preference.Pharmacol Biochem Behav 1996;53:1013–5.

[28] Knowles RG, Palacios M, Palmer RMJ, Moncada S. Formationof nitric oxide from L-arginine in the central nervous system: atransduction mechanism for stimulation of the soluble guanylatecyclase. Proc Natl Acad Sci USA 1989;86:5159–62.

[29] Komori Y, Chiang KT, Fukuto JM. The effects of nonionicdetergents on the activity and/or stability of rat brain nitric oxidesynthase. Arch Biochem Biophys 1993;307:311–5.

Y. Miyamoto et al. / Beha6ioural Brain Research 116 (2000) 187–196196

[30] Largent BL, Gundlach AL, Snyder SH. Pharmacological andautoradiographic discrimination of sigma and phencyclidine re-ceptor binding sites in brain with (+ )-[3H]-SKF 10,047, (+ )-[3H]-3-[3-hydroxyphenyl]-N-(1-propyl) piperidine and [3H]-1-[1-(2-thienyl) cyclohexyl] piperidine. J Pharmacol Exp Ther 1986;238:739–48.

[31] Leone P, Di Chiara G. Blockade D-1 receptors by SCH-23390antagonizes morphine and amphetamine-induced place preferenceconditioning. Eur J Pharmacol 1987;134:251–4.

[32] Lorrain DS, Hull EM. Nitric oxide increase dopamine andserotonin release in the medial preoptic area. NeuroReport1993;5:87–9.

[33] Lowenstein CJ, Dinerman JL, Snyder SH. Nitric oxide: a physi-ologic messenger. Ann Intern Med 1994;120:227–37.

[34] Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. Proteinmeasurement with the folin phenol regent. J Biol Chem1951;193:265–75.

[35] Maddox VH, Godefri EF, Parcell RF. The synthesis of phency-clidine and other 1-arylcyclohexylamines. J Med Chem 1965;8:230.

[36] Marletta MA. Approaches toward selective inhibition of nitricoxide synthase. J Med Chem 1994;37:1899–907.

[37] Nabeshima T, Fukaya H, Yamaguchi K, Ishikawa K, FurukawaH, Kameyama T. Development of tolerance and supersensitivityto phencyclidine in rats after repeated administration of phency-clidine. Eur J Pharmacol 1987;135:23–33.

[38] Nabeshima T, Fukaya H, Yamaguchi K, Ishikawa K, FurukawaH, Kameyama T. Potentiation in phencyclidine-induced sero-tonin-mediated behaviors after intracerebroventricular adminis-tration of 5,7-dihydroxytryptamine in rats. J Pharmacol Exp Ther1987;223:669–74.

[39] Nabeshima T, Kitaichi K, Noda Y. Functional changes inneuronal systems induced by phencyclidine administration. AnnNY Acad Sci 1996;801:29–38.

[40] Neisewander JL, McDougall SA, Bowling SL, Bardo MT. Condi-tioned taste aversion and place preference with buspirone andgepirone. Psychopharmacology 1990;100:485–90.

[41] Noda Y, Mamiya T, Furukawa H, Nabeshima T. Effects ofantidepressants on phencyclidine-induced enhancement of immo-bility in a forced swimming test in mice. Eur J Pharmacol1997;324:135–40.

[42] Noda Y, Miyamoto Y, Mamiya T, Kamei H, Furukawa H,Nabeshima T. Involvement of dopaminergic system in phency-clidine-induced place preference in mice pretreated with phency-clidine repeatedly. J Pharmacol Exp Ther 1998;286:44–51.

[43] Noda Y, Yamada K, Furukawa H, Nabeshima T. Involvementof nitric oxide in phencyclidine-induced hyperlocomotion in mice.Eur J Pharmacol 1995;286:291–7.

[44] Noda Y, Yamada K, Komori Y, Sugihara H, Furukawa H,Nabeshima T. Role of nitric oxide in the development of toleranceand sensitization to behavioural effects of phencyclidine in mice.Br J Pharmacol 1996;117:1579–85.

[45] Noda Y, Yamada K, Nabeshima T. Role of nitric oxide in theeffect of aging on spatial memory in rats. Behav Brain Res1997;83:153–8.

[46] Palmer RM, Ashton DS, Moncada S. Vascular endothelial cellssynthesize nitric oxide from L-arginine. Nature 1988;333:664–6.

[47] Petersen RC, Stillman RC. Phencyclidine abuse: an appraisal. In:National Institute of Drug Abuse Research Monograph, Vol. 21.Washington DC: Department of Health, Education and Welfare,1978. p. 1–17.

[48] Rocha B, Di Scala G, Rigo M, Hoyer D, Sander G. Effect of5,7-dihydroxytryptamine lesion on mianserin-induced conditionedplace aversion and 5-hydroxytryptamine1C receptors in the tatbrain. Neuroscience 1993;56:687–93.

[49] Schechter MD, Calcagnetti DJ. Trends in place preference condi-tioning with a cross-indexed bibliography; 1957–1991. NeurosciBiobehav Rev 1993;17:21–41.

[50] Shibata M, Araki N, Ohta K, Hamada J, Shimazu K, FukuuchiY. Nitric oxide regulates NMDA-induced dopamine release in ratstriatum. NeuroReport 1996;7:605–8.

[51] Smith RC, Meltzer HY, Arora RC, Davis JM. Effects of phency-clidine on [3H]catecholamine and [3H]serotonin uptake in synap-tosomal preparations from rat brain. Biochem Pharmacol1977;126:1435–9.

[52] Steinpreis RE, Salamone JD. The role of nucleus accumbensdopamine in the neurochemical and behavioral effects of phency-clidine: a microdialysis and behavioral study. Brain Res1993;612:263–70.

[53] Tzschentke TM, Schmidt WJ. Interactions of MK-801 and GYKI52466 with morphine and amphetamine in place preferenceconditioning and behavioural sensitization. Behav Brain Res1997;84:99–107.

[54] Vickroy TW, Johnson KM. Similar dopamine-releasing effects ofphencyclidine and nonamphetamine stimulants in striatal slices. JPharmacol Exp Ther 1982;223:669–74.

[55] Wiley JL. Nitric oxide synthase inhibitors attenuate phencyclidine-induced disruption of prepulse inhibition. Neuropsychopharma-cology 1998;19:86–94.

[56] Wroblewski JT, Nicoletti F, Fada E, Costa E. Phencyclidine is anegative allosteric modulator of signal transduction at two sub-classes of excitatory amino acid receptors. Proc Natl Acad SciUSA 1988;84:5068–72.

[57] Yamada K, Komori Y, Tanaka T, Senzaki K, Nikai T, SugiharaH, Kameyama T, Nabeshima T. Brain dysfunction associated withan induction of nitric oxide synthase following an intracerebralinjection of lipopolysaccharide in rats. Neuroscience 1999;88:281–94.

[58] Yamada K, Nabeshima T. Simultaneous measurement of nitriteand nitrate levels as indices of nitric oxide release in the cerebellumof conscious rats. J Neurochem 1997;68:1234–43.

[59] Yamada K, Nabeshima T. Nitric oxide and cyclic GMP signallingpathway in learning and memory processes. Curr Top Pharmacol1998;4:77–86.

[60] Yamada K, Noda Y, Nakayama S, Komori Y, Sugihara H,Hasegawa T, Nabeshima T. Role of nitric oxide in learning andmemory and in monoamine metabolism in the rat brain. Br JPharmacol 1995;115:852–8.

.