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The effects of phencyclidine on amphetamine stereotypy in rats
European Journal of Pharmacology, 48 (1978) 445--450 © Elsevier/North-Holland Biomedical Press
445
T H E E F F E C T S OF PHENCYCLIDINE ON AMPHETAMINE S T E R E O T Y P Y IN RATS ROBERT L. BALSTER and LARRY D. CHAIT Pharmacology Department, Medical College of Virginia, Virginia Commonwealth University, Richmond, Virginia 23298, U.S.A.
Received 13 September 1977, revised MS received 28 December 1977, accepted 13 January 1978
R.L. BALSTER and L.D. CHAIT, The effects of phencyclidine on amphetamine stereotypy in rats, European J. Pharmacol. 48 (1978) 445--450. In two separate experiments a 9 point rating scale was used to assess the effects of various doses of phencyclidine on the behavioral stereotypy produced by d-amphetamine in rats. A dose of phencyclidine (2.5 mg/kg) which had no effect when given alone, enhanced the behavioral effects of 1 and 3 mg/kg of d-amphetamine. Higher doses (5 and 10 mg/kg) of phencyclidine produced some stereotypy when given alone but they also produced ataxia which confounded the rating of their other behavioral effects. These higher doses did not enhance the effects of d-amphetamine. This study provides further evidence that phencyclidine may have dopaminergic activity similar to amphetamine. Phencyclidine
Amphetamine
Stereotypy
1. I n t r o d u c t i o n Phencyclidine possesses an unusual spect r u m o f central nervous system effects in animals and man (Domino, 1964; Balster and Chait, 1976). Originally developed as an anesthetic, phencyclidine produces a characteristic dissociative state in man (Greifenstein et al., 1958). Often, however, hallucinatory emergence reactions f r om phencyclidine anesthesia occurred ( J ohns t one et al., 1959). Unusual psychological effects have also been seen in studies o f lower doses of phencyclidine in man. These effects have been t e r m e d p s y c h o t o m i m e t i c and the drug has recently been subject to considerable abuse because o f these hallucinogenic properties (Lindgren et al., 1969; Burns and Lerner, 1976). At t h e present time t he neuropharmacology of phencyclidine is unclear. There are, however, t w o lines o f evidence to suggest t h a t increased dopaminergic activity may be responsible f o r some of t he behavioral effects o f this drug. Phencyclidine has been shown
Drug interactions
Behavior
to be a p o t e n t competitive inhibitor of dopamine reupt ake in rat striatum (Garey and Heath, 1976; Smith et al., 1975). It has been shown to produce ipsilateral r o t a t i o n in rats with unilateral lesions in the substantia nigra (Kanner et al., 1975). This is generally taken as evidence of an indirect-acting dopam i ne agonist (Ungerstedt, 1971). In addition, these effects on r o t a t i o n behavior can be reversed by p r e t r e a t m e n t with a-methyl-p-tyrosine, haloperidol and pimozide (Kanner et al., 1975; Finnegan et al., 1976). Phencyclidine also produces a s t e r e o t y p e d side to side head m o v e m e n t (Kanner et al., 1975; Finnegan et al., 1976) similar in some respects to the behavioral effects of high doses of amphetamines. Since the effects of phencyclidine on dopamine reuptake and its p r o d u c t i o n of ipsilateral turning are similar to those of amphetamine, phencyclidine might be expected to enhance some o f the behavioral effects of amphetamine. We have studied the effects of phencyclidine on d-amphetamine p r o d u c e d
446 stereotypy using a standarized, quantitative scale in rats.
EXPERIMENT I
1.2. Materials and methods The subjects were 16 male Sprague--Dawley (ASR; Madison, Wisconsin) albino rats weighing 220--270 g at the beginning of experimentation. They were randomly divided into 4 groups of 4 and were individually housed and had ad libitum access to food and water except during the experimental sessions. One week prior to the first drug dosing the animals were placed in the experimental cages and allowed to adapt to the observation room for 4 h. The experimental cages were standard, clear plastic, laboratory animal housing, 47 X 27 X 20 cm, open on the top with a wire mesh cover. A layer of bedding lined the b o t t o m of each cage. Eight of these cages were placed side by side in a row in a room which could be sealed off from outside disturbances. On drug test days, the animals were brought into the experimental room 15 min prior to testing. Following adaptation to the experimental environment the rating procedure was begun. Stereotypy was measured using the scale described b y Ellinwood and Balster (1974). Each animal was rated for 20 sec at 5 min pre-injection and then at 5, 15, 30, 45, 60, 75 and 90 min post-injection. For each observation period each subject was assigned a rating of 1--9 based upon which of the following behaviors predominated during that period: (1) asleep, (2) inactive, (3) inplace activities, (4) normal activity, (5) hyperactive, (6) slow patterned, (7) fast patterned, (8) restricted or (9) reactive. A more complete definition of each of these behaviors can be found in Ellinwood and Balster (1974). Two observers who were blind as to the dosing conditions were used for each time period. Each group of 4 rats was assigned to one of the four following doses of d-amphetamine:
R.L. BALSTER, L.D. CHAIT 0, 3, 6 or 12 mg/kg. Each group was injected with that dose of d-amphetamine and simultaneously with 0, 2.5, 5 or 10 mg/kg phencyclidine. The order of testing phencyclidine doses was different for each subject in a group and at least 4 days intervened between each test. All injections were i.p. and doses are calculated on the sulfate salt of d-amphetamine and the hydrochloride salt of phencyclidine. Both drugs were dissolved in normal saline. The 0 dose refers to saline vehicle alone.
1.3. Results
Interrater reliability was calculated using Pearson's R for each rating period. The R value ranged from 0.74 to 0.90. The lowest agreement occurred at 5 min post-injection when behavior was undergoing considerable transition. These reliability values correspond well with those reported by Ellinwood and Balster (1974) using the same scale. For purposes of data analysis, the rating for each observation was expressed as the average of the two raters. The mean pre-injection rating ranged from 2.8 to 3.6. No systematic differences were seen between any of the four groups or over any of the four test days; consequently, the pre-injection data have been excluded from the figures and statistical analysis. Fig. 1 presents the mean rating at each time period for all of the dose combinations of damphetamine and phencyclidine. The upper left shows the results of phencyclidine + vehicle. The lowest dose of phencyclidine (2.5 mg/kg) did not produce any stereotypy and could not be distinguished from vehicle control. Phencyclidine at 5 and 10 mg/kg produced a dose-related elevation in the scale, with the effects being greatest in the early post-injection period and decreasing b y 75 min. Although these doses produced clear repetitious behavior (very similar to that seen with d-amphetamine) which resulted in high scores on the stereotypy scale, there was a
PHENCYCLIDINE--AMPHETAMINE INTERACTIONS
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Fig. 1. The effects of phencyclidine on d-amphetamine stereotypy in the rat. Each point represents the mean rating given 4 subjects by 2 raters at each of various time periods after the i.p. administration of both drugs. A dose of 0 mg/kg designates a vehicle injection. Ordinate: mean rating; abscissa: time (min). A, 0 mg/kg d-amphetamine + phencyclidine; B, 3 mg/kg d-amphetamine + phencyclidine; C, 6 mg/kg d-amphetamine + phencyclidine; D, 12 mg/kg d-amphetamine + phencyclidine, o o, 0 mg/kg phencyclidine; a . . . . . . c], 2.5 mg/kg phencyclidine;/1. . . . . -/1, 5 mg/kg phencyclidine; o . . . . . o, 10 mg/kg phencyclidine.
definite behavioral difference between these effects of phencyclidine and the steretyped behavior produced by d-amphetamine. Doses of 5 m g / k g and above of phencyclidine produce marked p s y c h o m o t o r ataxia. At 5 mg/kg these effects are typically over by 15--30 min whereas at 10 mg/kg they usually last 60--75 min. This gross ataxia makes the scoring of stereotypy difficult. Therefore, the use of this scale to study the behavioral effects of phencyclidine given alone is clearly not appropriate.
Amphetamine alone (open circles on each of the remaining three panels) shows a doserelated production of stereotypy which asymptotes at 6 mg/kg. Since 6 and 12 mg/kg d-amphetamine already produce almost maximal scores on this scale it is not possible to show enhancement. There is some suggestion in the lower panels that some of the doses of phencyclidine enhanced the effects of 6 and 12 mg/kg d-amphetamine at the beginning and at the end of the observation period. The effect of 2.5 mg/kg phencyclidine
448 is clearer when combined with a submaximally effective dose of d-amphetamine (3 mg/kg) as seen in the upper right panel. From 45 min post-injection on, there is a clear enhancement of the effect of d-amphetamine. Interestingly, this enhancement by phencyclidine only occurs reliably at the lowest dose, a dose which is inactive w h e n given alone. We are not certain why 5 or 10 mg/kg phencyclidine did not enhance d-amphetamine, although we might conjecture that the m o t o r ataxia produced by these doses prevented the expression of stereotyped behavior. The complete experiment was subjected to a three-way analysis of variance (d-amphetamine X phencyclidine X time period) with repeated measures on the last two factors (Winer, 1962). This analysis indicated amphetamine dose (P ~< 0.01), phencyclidine dose (P ~ 0.05) and time (P ~< 0.01) to be significant as well as all possible interaction effects (P ~< 0.05).
R.L. BALSTER, L.D. CHAIT environment as described previously. The same two raters were used and the testing protocol and rating scale were the same as in Experiment I. Each group of rats was assigned to one dose of d-amphetamine (either 1 or 3 mg/kg). Each group was tested with that dose of d-amphetamine in combination with both saline vehicle and 2.5 mg/kg phencyclidine. The order of testing vehicle and phencyclidine was counter9-
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EXPERIMENT II In Experiment I, the only evidence t h a t phencyclidine can enhance the behavioral effects of d-amphetamine was found at one dose combination. In large part, this was due to the already maximal effects produced by the two highest doses of d-amphetamine. In the second experiment we repeated the interaction of a behaviorally inactive dose of phencyclidine (2.5 mg/kg) with the low dose (3 mg/kg) of d-amphetamine in eight additional animals. We also studied the combination of this dose of phencyclidine with a lower dose of d-amphetamine (1 mg/kg).
II.2. Materials and methods
The experimental design for this experiment was very similar to Experiment I. For t h i s study we used 16 naive male Sprague--Dawley rats divided into 2 groups of 8. They were housed and adapted to the experimental
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Fig. 2. The effects of phencyclidine on d-amphetamine stereotypy in the rat. Each point represents the mean rating-+ S.E.M. given 8 subjects by 2 rats at each of various time periods after the i.p. administration o f both drugs. A dose of 0 mg/kg designates a vehicle injection. Ordinate: mean rating; abscissa: time (min). Upper: 1 m g / k g d-amphetamine + phencyclidine. Lower: 3 mg/kg d-amphetamine + phencyclidine, o o, 0 mg/kg phencyclidine ; [] . . . . . . o, 2.5 mg/kg phencyclidine.
PHENCYCLIDIN E--AMPHETAMINE INTERACTIONS
balanced within each group. Four days intervened between the two tests.
II.3. Results
As in Experiment I, the rating for each observation was expressed as the average of the two raters and standard errors were calculated for each group for these 8 values. The mean pre-injection rating ranged from 2.7 to 3.2. The results of the pre-injection rating were omitted from subsequent data analysis. Fig. 2 presents the mean rating + S.E.M. at each time period for each of the dose combinations of d-amphetamine and phencyclidine. The open circles in b o t h panels represent the effects of d-amphetamine alone. 1 mg/kg d-amphetamine did not produce stereotyped behavior. However, in combination with 2.5 mg/kg phencyclidine an enhancement which averaged a b o u t 2 rating scale divisions occurred. The dose of 3 mg/kg damphetamine alone had a greater effect in this group of animals than in Experiment I; however, it t o o was enhanced b y 2.5 mg/kg phencyclidine at every observation period except 60 min. Three-way analysis of variance on these data indicated that the effects of amphetamine dose (P~< 0.01), phencyclidine dose (P~< 0.05) and time (P ~< 0.01) were significant; however, none of the interactions was significant.
4. General discussion A rating scale which was developed to assess the continuum of behavioral effects following high dose amphetamine administration in rats was used to study the interaction between phencyclidine and d-amphetamine. A dose of phencyclidine which b y itself could not be distinguished from vehicle on this scale enhanced the effects of low doses (1--3 mg/kg) of d-amphetamine. Higher doses of phencyclidine by themselves produced m o t o r ataxia
449
and some stereotyped behavior. They did not result in an enhancement of d-amphetamine effects. It is possible that phencyclidine alters the pharmacokinetics of d-amphetamine in some way to increase the effective level o f damphetamine. In the rat, oxidative hydroxylation in the liver is the major m o d e of metabolism for b o t h phencyclidine (Wong and Biemann, 1976) and amphetamine (Axelrod, 1954; Dring et ah, 1970}. There have been no studies on the pharmacokinetic interaction of these two c o m p o u n d s b u t it is possible that phencyclidine could block the inactivation of d-amphetamine resulting in the behavioral enhancement we observed. The explanation for our findings which is most consistent with previous research on phencyclidine neuropharmacology concerns the role of phencyclidine as a dopaminergic agonist. It produces ipsilateral turning in substantia nigra lesioned rats (Kanner et al., 1975; Finnegan et al., 1976) and blocks the reuptake of dopamine in rat striatum (Garey and Heath, 1976; Smith et ah, 1975). Since these effects are the same as those produced b y high doses of amphetamines it is tempting to conclude that the enhancement is due to their c o m m o n dopaminergic activity. Increased dopaminergic activity is particularly interesting in light of some other behavioral and pharmacological properties of phencyclidine. A number of investigators have argued that phencyclidine is more truly psychotomimetic than other hallucinogenic drugs since its subjective effects in humans more closely resemble certain symptoms of schizophrenia (Luby et al., 1959; Cohen et ah, 1962). In view of the current interest in the role of increased dopamine activity in schizophrenia (Snyder et al., 1974} and the use of amphetamine intoxication as a behavioral and neuropharmacological animal model for schizophrenia (Randrup and Munkvad, 1967; Ellinwood, 1967), the neuropharmacological and behavioral effects of phencyclidine warrant further attention in this context. The turning behavior in substantia nigra lesioned
450
rats produced by phencyclidine is antagonized by the dopamine receptor blocker haloperidol (Kanner et al., 1975) and there is some clinical evidence that haloperidol reduces the incidence of traumatic emergence reactions seen in phencyctidine anesthesia (Helrich and Atwood, 1964). There has also been a report on the use of chlorpromazine to treat acute phencyclidine intoxication (Luisada and Brown, 1976). On the other hand, we (Balster and Chait, 1976) failed to find evidence that chlorpromazine antagonizes the effects of phencyclidine on operant behavior of rhesus monkeys, and other clinical reports do not advise the use of phenothiazines in acute phencyclidine intoxication (Fauman et al., 1975; Burns and Lerner, 1976). Clearly, a more thorough investigation of the neuropharmacological basis for the behavioral effects of phencyclidine will help in the development of a rationale for the treatment of acute phencyclidine emergencies.
Acknowledgements The research was supported by Grant DA-01442 from the National Institute of Drug Abuse. L.D. Chait is a predoctoral fellow supported by training grant DA-07027.
References Axelrod, J., 1954, Studies on sympathomimetic amines. I. The biotransformation and physiological disposition of d-amphetamine, d-p-hydroxyamphetamine and d-methamphetamine, J. Pharmacol. Exptl. Therap. 110, 315. Balster, R.L. and L.D. Chait, 1976, The behavioral pharmacology of phencyclidine, Clin. Toxicol. 9, 513. Burns, R.S. and S.E. Lerner, 1976, Perspective: acute phencyclidine intoxication, Clin. Toxicol. 9,477. Cohen, B.D., G. Rosenbaum, E.D. Luby and J.S. Gottlieb, 1962, Comparison o f phencyclidine (Sernyl) with other drugs, Arch. Gem Psychiat. 6, 79. Domino, E.F., 1964, Neurobiology of phencyclidine (Sernyl), a drug with an unusual spectrum of pharmacological activity, Intern. Rev. Neurobiol. 6, 303. Dring, L.G., R.L. Smith and R.T. Williams, 1970, The metabolic fate of amphetamine in man and other species, Biochem. J. 1 1 6 , 4 2 5 .
R.L. BALSTER, L.D. CHAIT Ellinwood, E.H., Jr., 1967, Amphetamine psychosis: I. Description of the individuals and process, J. Nerv. Mental Dis. 44, 273. Ellinwood, E.H., Jr. and R.L. Balster, 1974, Rating the behavioral effects of amphetamine, European J. Pharmacol. 28, 35. Fauman, F., F. Baker, L.W. Coppleson, P. Rosen and M.B. Segal, 1975, Psychosis induced by phencyclidine, J. Amer. Col. Emergency Physicians 4, 223. Finnegan, K.T., M.I. Kanner and H.Y. Meltzer, 1976, Phencyclidine-induced rotational behavior in rats with nigrostriatal lesions and its modulation by dopaminergic and cholinergic agents, Pharmacol. Biochem. Behav. 5 , 6 5 1 . Garey, R.E. and R.G. Heath, 1976, The effects of phencyclidine on the uptake of 3H-catecholamines by rat striatal and hypothalamic synaptosomes, Life Sci. 18, 1105. Greifenstein, F.E., J. Yoshitake, M. DeVault and J.E. Gajewski, 1958, A study of 1-aryl cyclohexylamine for anesthesia, Anesth. Analg. 37,283. Helrich, M. and J.M. Atwood, 1964, Modification of Sernyl anesthesia with haloperidol, Anesth. Analg. 43,471. Johnstone, M., V. Evans and S. Baigel, 1959, Sernyl (CI-395) in clinical anesthesia, Brit. J. Anesth. 31, 433. Kanner, M., K. Finnegan and H.Y. Meltzer, 1975, Dopaminergic effects of phencyclidine in rats with nigrostriatal lesions, Psychopharmacol. Commun. 1,393. Lindgren, J.E., C.G. Hammer, R. Hessling and B. Holmstedt, 1969, The chemical identity of "hog" -- a new hallucinogen, Amer. J. Pharm. 141, 86. Luby, E.D., B.D. Cohen, G. Rosenbaum, J.S. Gottlieb and R. Kelley, 1959, A study of a new schizophrenimimetic drug, Sernyl, A.M.A. Arch. Neurol. Psychiat. 8 1 , 3 6 3 . Luisada, P.V. and B.I. Brown, 1976, Clinical management of the phencyclidine psychosis, Clin. Toxicol. 9,539. Randrup, A. and I. Munkvad, 1967, Stereotypic activities produced by amphetamine in several species and man, Psychopharmacologia (Berlin) 11,300. Smith, R.C., H.Y. Meltzer, H. Dekirmenjian and J.M. Davis, 1975, Effects of phencyclidine on biogenic amines in rat brain, Neurosci. Abstracts 1, No. 468. Snyder, S.H., S.P. Banerjee, H.I. Yamamura and D. Greenberg, 1974, Drugs, neurotransmitters and schizophrenia, Science 184, 1243. Ungerstedt, U., 1971, Striatal dopamine release after amphetamine or nerve degeneration revealed by rotational behavior, Acta Physiol. Scand. Suppl. 367, 49. Winer, B.J., 1962, Statistical Principles in Experimental Design (McGraw Hill, N.Y.). Wong, L.K. and K. Biemann, 1976, Metabolites of phencyclidine, Clin. Toxicol. 9, 583.
445
T H E E F F E C T S OF PHENCYCLIDINE ON AMPHETAMINE S T E R E O T Y P Y IN RATS ROBERT L. BALSTER and LARRY D. CHAIT Pharmacology Department, Medical College of Virginia, Virginia Commonwealth University, Richmond, Virginia 23298, U.S.A.
Received 13 September 1977, revised MS received 28 December 1977, accepted 13 January 1978
R.L. BALSTER and L.D. CHAIT, The effects of phencyclidine on amphetamine stereotypy in rats, European J. Pharmacol. 48 (1978) 445--450. In two separate experiments a 9 point rating scale was used to assess the effects of various doses of phencyclidine on the behavioral stereotypy produced by d-amphetamine in rats. A dose of phencyclidine (2.5 mg/kg) which had no effect when given alone, enhanced the behavioral effects of 1 and 3 mg/kg of d-amphetamine. Higher doses (5 and 10 mg/kg) of phencyclidine produced some stereotypy when given alone but they also produced ataxia which confounded the rating of their other behavioral effects. These higher doses did not enhance the effects of d-amphetamine. This study provides further evidence that phencyclidine may have dopaminergic activity similar to amphetamine. Phencyclidine
Amphetamine
Stereotypy
1. I n t r o d u c t i o n Phencyclidine possesses an unusual spect r u m o f central nervous system effects in animals and man (Domino, 1964; Balster and Chait, 1976). Originally developed as an anesthetic, phencyclidine produces a characteristic dissociative state in man (Greifenstein et al., 1958). Often, however, hallucinatory emergence reactions f r om phencyclidine anesthesia occurred ( J ohns t one et al., 1959). Unusual psychological effects have also been seen in studies o f lower doses of phencyclidine in man. These effects have been t e r m e d p s y c h o t o m i m e t i c and the drug has recently been subject to considerable abuse because o f these hallucinogenic properties (Lindgren et al., 1969; Burns and Lerner, 1976). At t h e present time t he neuropharmacology of phencyclidine is unclear. There are, however, t w o lines o f evidence to suggest t h a t increased dopaminergic activity may be responsible f o r some of t he behavioral effects o f this drug. Phencyclidine has been shown
Drug interactions
Behavior
to be a p o t e n t competitive inhibitor of dopamine reupt ake in rat striatum (Garey and Heath, 1976; Smith et al., 1975). It has been shown to produce ipsilateral r o t a t i o n in rats with unilateral lesions in the substantia nigra (Kanner et al., 1975). This is generally taken as evidence of an indirect-acting dopam i ne agonist (Ungerstedt, 1971). In addition, these effects on r o t a t i o n behavior can be reversed by p r e t r e a t m e n t with a-methyl-p-tyrosine, haloperidol and pimozide (Kanner et al., 1975; Finnegan et al., 1976). Phencyclidine also produces a s t e r e o t y p e d side to side head m o v e m e n t (Kanner et al., 1975; Finnegan et al., 1976) similar in some respects to the behavioral effects of high doses of amphetamines. Since the effects of phencyclidine on dopamine reuptake and its p r o d u c t i o n of ipsilateral turning are similar to those of amphetamine, phencyclidine might be expected to enhance some o f the behavioral effects of amphetamine. We have studied the effects of phencyclidine on d-amphetamine p r o d u c e d
446 stereotypy using a standarized, quantitative scale in rats.
EXPERIMENT I
1.2. Materials and methods The subjects were 16 male Sprague--Dawley (ASR; Madison, Wisconsin) albino rats weighing 220--270 g at the beginning of experimentation. They were randomly divided into 4 groups of 4 and were individually housed and had ad libitum access to food and water except during the experimental sessions. One week prior to the first drug dosing the animals were placed in the experimental cages and allowed to adapt to the observation room for 4 h. The experimental cages were standard, clear plastic, laboratory animal housing, 47 X 27 X 20 cm, open on the top with a wire mesh cover. A layer of bedding lined the b o t t o m of each cage. Eight of these cages were placed side by side in a row in a room which could be sealed off from outside disturbances. On drug test days, the animals were brought into the experimental room 15 min prior to testing. Following adaptation to the experimental environment the rating procedure was begun. Stereotypy was measured using the scale described b y Ellinwood and Balster (1974). Each animal was rated for 20 sec at 5 min pre-injection and then at 5, 15, 30, 45, 60, 75 and 90 min post-injection. For each observation period each subject was assigned a rating of 1--9 based upon which of the following behaviors predominated during that period: (1) asleep, (2) inactive, (3) inplace activities, (4) normal activity, (5) hyperactive, (6) slow patterned, (7) fast patterned, (8) restricted or (9) reactive. A more complete definition of each of these behaviors can be found in Ellinwood and Balster (1974). Two observers who were blind as to the dosing conditions were used for each time period. Each group of 4 rats was assigned to one of the four following doses of d-amphetamine:
R.L. BALSTER, L.D. CHAIT 0, 3, 6 or 12 mg/kg. Each group was injected with that dose of d-amphetamine and simultaneously with 0, 2.5, 5 or 10 mg/kg phencyclidine. The order of testing phencyclidine doses was different for each subject in a group and at least 4 days intervened between each test. All injections were i.p. and doses are calculated on the sulfate salt of d-amphetamine and the hydrochloride salt of phencyclidine. Both drugs were dissolved in normal saline. The 0 dose refers to saline vehicle alone.
1.3. Results
Interrater reliability was calculated using Pearson's R for each rating period. The R value ranged from 0.74 to 0.90. The lowest agreement occurred at 5 min post-injection when behavior was undergoing considerable transition. These reliability values correspond well with those reported by Ellinwood and Balster (1974) using the same scale. For purposes of data analysis, the rating for each observation was expressed as the average of the two raters. The mean pre-injection rating ranged from 2.8 to 3.6. No systematic differences were seen between any of the four groups or over any of the four test days; consequently, the pre-injection data have been excluded from the figures and statistical analysis. Fig. 1 presents the mean rating at each time period for all of the dose combinations of damphetamine and phencyclidine. The upper left shows the results of phencyclidine + vehicle. The lowest dose of phencyclidine (2.5 mg/kg) did not produce any stereotypy and could not be distinguished from vehicle control. Phencyclidine at 5 and 10 mg/kg produced a dose-related elevation in the scale, with the effects being greatest in the early post-injection period and decreasing b y 75 min. Although these doses produced clear repetitious behavior (very similar to that seen with d-amphetamine) which resulted in high scores on the stereotypy scale, there was a
PHENCYCLIDINE--AMPHETAMINE INTERACTIONS
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Fig. 1. The effects of phencyclidine on d-amphetamine stereotypy in the rat. Each point represents the mean rating given 4 subjects by 2 raters at each of various time periods after the i.p. administration of both drugs. A dose of 0 mg/kg designates a vehicle injection. Ordinate: mean rating; abscissa: time (min). A, 0 mg/kg d-amphetamine + phencyclidine; B, 3 mg/kg d-amphetamine + phencyclidine; C, 6 mg/kg d-amphetamine + phencyclidine; D, 12 mg/kg d-amphetamine + phencyclidine, o o, 0 mg/kg phencyclidine; a . . . . . . c], 2.5 mg/kg phencyclidine;/1. . . . . -/1, 5 mg/kg phencyclidine; o . . . . . o, 10 mg/kg phencyclidine.
definite behavioral difference between these effects of phencyclidine and the steretyped behavior produced by d-amphetamine. Doses of 5 m g / k g and above of phencyclidine produce marked p s y c h o m o t o r ataxia. At 5 mg/kg these effects are typically over by 15--30 min whereas at 10 mg/kg they usually last 60--75 min. This gross ataxia makes the scoring of stereotypy difficult. Therefore, the use of this scale to study the behavioral effects of phencyclidine given alone is clearly not appropriate.
Amphetamine alone (open circles on each of the remaining three panels) shows a doserelated production of stereotypy which asymptotes at 6 mg/kg. Since 6 and 12 mg/kg d-amphetamine already produce almost maximal scores on this scale it is not possible to show enhancement. There is some suggestion in the lower panels that some of the doses of phencyclidine enhanced the effects of 6 and 12 mg/kg d-amphetamine at the beginning and at the end of the observation period. The effect of 2.5 mg/kg phencyclidine
448 is clearer when combined with a submaximally effective dose of d-amphetamine (3 mg/kg) as seen in the upper right panel. From 45 min post-injection on, there is a clear enhancement of the effect of d-amphetamine. Interestingly, this enhancement by phencyclidine only occurs reliably at the lowest dose, a dose which is inactive w h e n given alone. We are not certain why 5 or 10 mg/kg phencyclidine did not enhance d-amphetamine, although we might conjecture that the m o t o r ataxia produced by these doses prevented the expression of stereotyped behavior. The complete experiment was subjected to a three-way analysis of variance (d-amphetamine X phencyclidine X time period) with repeated measures on the last two factors (Winer, 1962). This analysis indicated amphetamine dose (P ~< 0.01), phencyclidine dose (P ~ 0.05) and time (P ~< 0.01) to be significant as well as all possible interaction effects (P ~< 0.05).
R.L. BALSTER, L.D. CHAIT environment as described previously. The same two raters were used and the testing protocol and rating scale were the same as in Experiment I. Each group of rats was assigned to one dose of d-amphetamine (either 1 or 3 mg/kg). Each group was tested with that dose of d-amphetamine in combination with both saline vehicle and 2.5 mg/kg phencyclidine. The order of testing vehicle and phencyclidine was counter9-
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22
115
EXPERIMENT II In Experiment I, the only evidence t h a t phencyclidine can enhance the behavioral effects of d-amphetamine was found at one dose combination. In large part, this was due to the already maximal effects produced by the two highest doses of d-amphetamine. In the second experiment we repeated the interaction of a behaviorally inactive dose of phencyclidine (2.5 mg/kg) with the low dose (3 mg/kg) of d-amphetamine in eight additional animals. We also studied the combination of this dose of phencyclidine with a lower dose of d-amphetamine (1 mg/kg).
II.2. Materials and methods
The experimental design for this experiment was very similar to Experiment I. For t h i s study we used 16 naive male Sprague--Dawley rats divided into 2 groups of 8. They were housed and adapted to the experimental
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Fig. 2. The effects of phencyclidine on d-amphetamine stereotypy in the rat. Each point represents the mean rating-+ S.E.M. given 8 subjects by 2 rats at each of various time periods after the i.p. administration o f both drugs. A dose of 0 mg/kg designates a vehicle injection. Ordinate: mean rating; abscissa: time (min). Upper: 1 m g / k g d-amphetamine + phencyclidine. Lower: 3 mg/kg d-amphetamine + phencyclidine, o o, 0 mg/kg phencyclidine ; [] . . . . . . o, 2.5 mg/kg phencyclidine.
PHENCYCLIDIN E--AMPHETAMINE INTERACTIONS
balanced within each group. Four days intervened between the two tests.
II.3. Results
As in Experiment I, the rating for each observation was expressed as the average of the two raters and standard errors were calculated for each group for these 8 values. The mean pre-injection rating ranged from 2.7 to 3.2. The results of the pre-injection rating were omitted from subsequent data analysis. Fig. 2 presents the mean rating + S.E.M. at each time period for each of the dose combinations of d-amphetamine and phencyclidine. The open circles in b o t h panels represent the effects of d-amphetamine alone. 1 mg/kg d-amphetamine did not produce stereotyped behavior. However, in combination with 2.5 mg/kg phencyclidine an enhancement which averaged a b o u t 2 rating scale divisions occurred. The dose of 3 mg/kg damphetamine alone had a greater effect in this group of animals than in Experiment I; however, it t o o was enhanced b y 2.5 mg/kg phencyclidine at every observation period except 60 min. Three-way analysis of variance on these data indicated that the effects of amphetamine dose (P~< 0.01), phencyclidine dose (P~< 0.05) and time (P ~< 0.01) were significant; however, none of the interactions was significant.
4. General discussion A rating scale which was developed to assess the continuum of behavioral effects following high dose amphetamine administration in rats was used to study the interaction between phencyclidine and d-amphetamine. A dose of phencyclidine which b y itself could not be distinguished from vehicle on this scale enhanced the effects of low doses (1--3 mg/kg) of d-amphetamine. Higher doses of phencyclidine by themselves produced m o t o r ataxia
449
and some stereotyped behavior. They did not result in an enhancement of d-amphetamine effects. It is possible that phencyclidine alters the pharmacokinetics of d-amphetamine in some way to increase the effective level o f damphetamine. In the rat, oxidative hydroxylation in the liver is the major m o d e of metabolism for b o t h phencyclidine (Wong and Biemann, 1976) and amphetamine (Axelrod, 1954; Dring et ah, 1970}. There have been no studies on the pharmacokinetic interaction of these two c o m p o u n d s b u t it is possible that phencyclidine could block the inactivation of d-amphetamine resulting in the behavioral enhancement we observed. The explanation for our findings which is most consistent with previous research on phencyclidine neuropharmacology concerns the role of phencyclidine as a dopaminergic agonist. It produces ipsilateral turning in substantia nigra lesioned rats (Kanner et al., 1975; Finnegan et al., 1976) and blocks the reuptake of dopamine in rat striatum (Garey and Heath, 1976; Smith et ah, 1975). Since these effects are the same as those produced b y high doses of amphetamines it is tempting to conclude that the enhancement is due to their c o m m o n dopaminergic activity. Increased dopaminergic activity is particularly interesting in light of some other behavioral and pharmacological properties of phencyclidine. A number of investigators have argued that phencyclidine is more truly psychotomimetic than other hallucinogenic drugs since its subjective effects in humans more closely resemble certain symptoms of schizophrenia (Luby et al., 1959; Cohen et ah, 1962). In view of the current interest in the role of increased dopamine activity in schizophrenia (Snyder et al., 1974} and the use of amphetamine intoxication as a behavioral and neuropharmacological animal model for schizophrenia (Randrup and Munkvad, 1967; Ellinwood, 1967), the neuropharmacological and behavioral effects of phencyclidine warrant further attention in this context. The turning behavior in substantia nigra lesioned
450
rats produced by phencyclidine is antagonized by the dopamine receptor blocker haloperidol (Kanner et al., 1975) and there is some clinical evidence that haloperidol reduces the incidence of traumatic emergence reactions seen in phencyctidine anesthesia (Helrich and Atwood, 1964). There has also been a report on the use of chlorpromazine to treat acute phencyclidine intoxication (Luisada and Brown, 1976). On the other hand, we (Balster and Chait, 1976) failed to find evidence that chlorpromazine antagonizes the effects of phencyclidine on operant behavior of rhesus monkeys, and other clinical reports do not advise the use of phenothiazines in acute phencyclidine intoxication (Fauman et al., 1975; Burns and Lerner, 1976). Clearly, a more thorough investigation of the neuropharmacological basis for the behavioral effects of phencyclidine will help in the development of a rationale for the treatment of acute phencyclidine emergencies.
Acknowledgements The research was supported by Grant DA-01442 from the National Institute of Drug Abuse. L.D. Chait is a predoctoral fellow supported by training grant DA-07027.
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