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Long-term motor stimulant effects of (+)-4-propyl-9-hydroxynapthoxazine (PHNO), a dopamine D-2 receptor agonist: interactions with a dopamine D-1 receptor antagonist and agonist
European Journal of Pharmacology. 149 (1988) 25- 31
25
Elsevier F.IP 50238
Long-term motor stimulant effects of ( +)-4-propyl-9-hydroxynaphthoxazine (PHNO), a dopamine D-2 receptor agonist: interactions with a dopamine D-1 receptor antagonist and agonist Mathew T. Martin-Iverson *, Susan D. Iversen and Stephen M. Stahl Merck Sharpe & Dohme. Neurosciem'e Research ('entre. Terlings Park. Eastwick Road. llarh~w, E~'sex ('M20 2QR. U.K.
Received 26 October 1987, revised MS received 26 January 1988, accepted 9 February 1988
Rats were given continuous infusions of ( + )-4-propyl-9-hydroxynaphthoxazine (PHNO, 5 ttg/h), a dopamine D-2 receptor agonist, using subcutaneous implants of ALZET osmotic minipumps. It was observed that tolerance occurred to the motor stimulant effects of PHNO during the light cycle of each day, but not during the dark cycle. Rather, thc motor stimulant actions of PHNO were gradually augmented during successive nights. Daytime tolerance to the stimulant actions of PI-|NO was reversed by a mild environmental stress or by administration of the D-I receptor agonist, SKF 38393 (6 m g / k g i.p.). Co-administration of the dopamine D-I receptor antagonist, (SCH 23390, 20 p,g/h s.c. by ALZET osmotic minipumps), initally blocked the motor stimulant actions of PHNO and also attenuated the reversal of tolerance to PHNO produced by stress, without blocking the actions of stress on activity in vehicle-infused animals. These results indicate that tolerance to the behavioural effects of PHNO may result from a loss of activation of D-I receptors by endogenous DA. Dopamine receptor subtypes: Locomotor acitivity; PHNO (( + )-4-propyl-9-hydroxynaphthoxazine); (Tolerance, Sensitization)
I. Introduction ( + )-4-Propyl-9-hydroxynaphthoxazine (PHNO) is an extremely p o t e n t d o p a m i n e ( D A ) agonist with a m a r k e d selectivity for the D-2 r e c e p t o r s u b t y p e ( M a r t i n et al., 1984). In a previous s t u d y ( M a r t i n - l v e r s o n et al., in press), it was o b s e r v e d that chronic c o n t i n u o u s a d m i n i s t r a t i o n of P H N O to rats by Alzet o s m o t i c m i n i p u m p s resulted in a r a p i d d e v e l o p m e n t of tolerance to the m o t o r s t i m u l a n t effects of P H N O d u r i n g the light p e r i o d
* To whom all correspondence should be addressed at present address: Neurc.chemical Research Unit, Department of Psychiatry, 1E7.44 Walter Mackenzie lleahh Sciences Centre, University of Alberta, Edmonton, Alberta, Canada T6G 2B7.
of the light-dark cycle. However, the m o t o r s t i m u l a n t actions of P H N O were g r a d u a l l y augm e n t e d d u r i n g the d a r k p e r i o d s of successive 24 h periods, on the s a m e d a y s that tolerance occurred d u r i n g the light periods. It was further o b s e r v e d that stress could t e m p o r a r i l y reverse "daytime' tolerance. To explain these o b s e r v a t i o n s , a hypothesis was p r o p o s e d that is b a s e d on the recent suggestion that there is a functional interaction between different s u b t y p e s of D A receptors. Specifically, some b e h a v i o u r a l effects of D A agonists selective for the D-2 r e c e p t o r s u b t y p e c a n n o t be elicited without the c o n c o m i t a n t a c t i v a t i o n of the D-I receptor s u b t y p e ( M o l l o y et al., 1986). In a n o r m a l animal, the acute a d m i n i s t r a t i o n of a selective D-2 receptor agonist can elicit m a n y of its b e h a v i o u r a l effects b e c a u s e of c o n c o m i t a n t activation of D-1
0014-2999/88/$03.50 :': 1988 Elsevier Science Publishers B.V. (Biomedical I)ivision)
26 receptors, presumably by endogenous release of DA. That endogenous DA may activate D-I receptors, allowing the expression of motor stimulation by the D-2 agonists, is supported by the findings that extensive DA depletions block the motor stimulation of bromocriptine, a D-2 receptorselective agonist (Jackson and Jenkins, 1985: Jackson and Hashizume, 1986). Agents that promote DA release (Jackson and Jenkins, 1985) or a D-1 receptor-selective agonist (Jackson and Hashizume, 1986) can re-instate the motor stimulant actions of bromocriptine blocked by DA depletion. Furthermore, these behavioural interactions are paralleled by similar interactions between agonists selective for different DA receptor subtypes, in relation to effects on the firing rates of basal ganglia neurons (Waiters et al., 1987). It has been observed that DA release is high at night and low during the day (O'Neill and Fillenz, 1985), and that mild stress can induce DA release (Antelman et al., 1975). These data, in conjunction with the evidence that DA autoreceptors are particularly effective under conditions of low basal release of DA (Mereu et al., 1986) prompted the speculation that continuous administration of a [)-2 receptor agonist leads eventually to a decrease in endogenous DA release during the day, but not during the night. Therefore, behavioural tolerance to continuous infusions of P H N O occurs only during the day. as a consequence of loss of activation of D-1 receptors by endogenous I)A. The observation that a mild stress (which may induce DA release) temporarily reversed tolerance was taken as support for this interpretation. In brief, it was proposed that tolerance to the motor stimulant effects of P H N O was a result of loss of activation of D-I receptors by endogenous DA. This hypothesis of the mechanism of tolerance to PHNO, a D-2 agonist, makes several testable predictions. Firstly, a D-1 antagonist, such as SCH 23390 should block PHNO-induced motor stimulation. Secondly, SCH 23390 should block the stress-induced reversal of tolerance to PHNO. Thirdly, a D-I agonist, like SKF 38393, should reverse tolerance to PHNO. The following experiments were undertaken to test these predictions.
2. Materials and methods
2. I. Experimental animals and al~paratus Male Sprague-Dawley rats ( n = 6 for each group) weighing 30(-)-350 g were housed in cages equipped with infrared photocell apparatus which allowed continuous monitoring of locomotor activity over periods of weeks. The cage dimensions were 24 (W) × 16 (H) × 38 (l,) cm, with two photocells placed 3 cm from the floor, and spaced 13 cm apart, equidistant from both end walls. The rats lived in these cages (under a light-dark cycle in which the light period was from 9:00 to 21:00 h) for at least 10 days prior to any drug treatments. Drug treatments were continued for 11-15 days, depending upon the experiment, with continuous recording of photobeam interruptions in hourly blocks. On Mondays, Wednesdays and Fridays the rats were given fresh water and food, and the litter underneath the cages was changed. This procedure was accompanied with a fair amount of noise, lasting close to 20 min, and constitutes the 'stress' variable of the present experiments. Average activity counts were calculated over blocks of 12 h (nocturnal activity), 10 h (diurnal activity) and 2 h (time during and 100 min following the care of the animals (stress), or the equivalent periods on days without stress. 2.2. ('ompound~ P H N O H('I (Merck Sharpe & Dohme) was dissolved in distilled water and A L Z E T osmotic minipumps (No. 2002) were filled with the solution to provide a release rate of 5.0 p,g/h (weight expressed as free base), which gave a dose of approximately 15 #g kg 1 h - i . SCH 23390 (RBI) was dissolved in distilled water and given by ALZ E T osmotic minipumps at the rate of 20 # g / h (about 60 p,g kg i h - i ) . SKF 38393 (RB1) was dissolved in distilled water for a dose of 6.0 m g / k g (Lp.). 2.3. Continuous administration of P l t N O and S ( ' I I 233~)0 All animals received two osmotic minipumps implanted s.c. between the shoulders while the rats
27
were under ether anaesthesia. Different groups of rats were given two pumps containing only vehicle (VEH + VEH), one p u m p containing vehicle and the other P H N O (VEH + PHNO), one p u m p loaded with SCH 23390 and the other with vehicle (SCI-I + VEH), or one pump containing P H N O and the other loaded with SCH 23390 (SCH + PHNO). The n of each group was 6. Interruptions of photobeams were recorded in hourly blocks throughout drug treatments. 2.4. Administration of S K F 38393
ACTIVITY
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3. Results
3.1. Lffe¢'ts of continuous administration of PItNO, S C t l 23390 and both with s.c. osmotic minipumps on locomotor actit'ity. Figure 1 displays the effects of continuous administration of vehicle, PHNO, SCH 23390 and both drugs on locomotor activity, presented as the average photobeam interruptions from the 10 h of the light cycle (exclusive of the 2 h during and after environmental disturbance (stress) or the equivalent times on days without stress). Analysis of variance revealed a significant SCH 23390 x P H N O x Day interaction (F(10,200) = 8.20, P < 0.001). As previously observed (Martin-Iverson et al., in press), continuous administration of this dose of P H N O resulted in tolerance to the motor
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Four groups of rats (n = 6 for each group) were used in this experiment. Two groups were implanted with osmotic minipumps containing vehicle (sterile water), and two groups were given osmotic pumps containing P H N O (5 # g / h ) . Seven days after osmotic p u m p implants, one vehicle-infused group and one PHNO-infused group were given injections of SKF 38393 (6.0 m g / k g i.p.) at 12:00 h on each of 4 consecutive days. The remaining two groups (one with vehicle infusions and one with P H N O infusions) were given vehicle injections (sterile water, 1 m l / k g i.p.) at the same times. The photobeam interruptions occurring during the 4 h after injections were collapsed across days to obtain an average for each animal.
COUNTS
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Fig. l. Mean daytime locomotor activity counts of rats given continuous infusions of vehicle and vehicle ( V E H + V E H ) , vehicle and SCH 23390 (20 p.g/h, VFH + SCH), PHNO (5.0 p.g/h) and vehicle (PHNO + VEIl), or PHNO and SCH 23390 ( P H N O + S C H ) . Note that tolerance to the motor stimulant effects of PHNO, a selective D-2 receptor agonist, occurs rapidly over the first 3 days. The D-I receptor antagonist, SCH 23390, completely blocked the initial stimulant action of PHNO, but partial 'escape' from this blockade was evident on days 7, 9 and 11.
stimulant effects developing rapidly (within 3-4 days) during the light periods. The D-1 antagonist, SCH 23390, appeared to reducc acitivity when given with vehicle, but this effect was not significant. However, when given with PHNO, it significantly blocked the increase in activity on the first 2 days (P < 0.01 on both days, Tukey's HSD test). Interestingly, on days 7. 9 and 11 (days without stress) activity levels in the group receiving both P H N O and SCH 23390 began to exhibit increases in activity (significantly different from vehicle: P < 0 . 0 1 (day 9) and P < 0 . 0 5 (day 11), Tukcy's HSD). Figure 2 shows the effects of drug treatments on nocturnal activity, expressed as the average photobeam counts over the 12 11 dark periods. Analysis of variance indicated a significant SCH 23390 × P H N O × Night interaction (F(11,220) = 4.53, P < 0.001). The locomotor activity induced by P H N O gradually increased over successive nights (fig. 2). This effect was significantly different from that of the VEH + VEH group on every night (P < 0.01), and the effect of P H N O on the last night was significantly higher than the motor
28
A C ' I V I T Y COUNTS
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P < 0.01), reflecting that the tolerance to PHNO activity-inducing effects observed on days without stress was reversed by stress, and that this stressinduced reversal of behaviour tolerance to PHNO was blocked by SCH 23390. SCH 23390 did not significantly attenuate stress-induced activity m vehicle-infused rats. 3.3. Effects of administration of S K F 38393 to rats exhibiting tolerance m the motor stimulant actions of PHNO.
12
SUCCESSIVE NIGHTS
Fig. 2. Mean nocturnal locomotor activity counts of rats given c o n t i n u o u s infusions of vehicle and vehicle ( V E H + V E t t ) , vehicle and SCH 23390 (20 p , g / h , V E H + S C H ) , P H N O (5 # g / h ) and vehicle ( P H N O + VEH). or P H N O and SCH 23390 (PHNO+SCH). U n l i k e daytime activity, ncx:turnal activity was gradually augmented over successive nights in the groups receiving P H N O , a D-2 receptor agonist. The D-1 receptor antagonist, SCH 23390, blocked the initial motor stimulant effects of P H N O . but this action was no longer apparent by the sixth night.
Figure 4 displays the results of the administration of the D-1 receptor agonist. SKF 38393, on locomotor behaviour of rats exhibiting tolerance to the locomotor stimulant actions of PHNO. or of rats treated with vehicle. There were no significant effects of PHNO in rats given injections of vehicle in addition to the PHNO infusions. Neither wcre effects of SKF 38393 on activity apparent in rats given vehicle infusions. On the other hand.
All VTI" CO~NIS/4OUR
activity induced on the first night (P < 0.01). The group given SCH 23390 + VEH again exhibited an apparent decrease in activity on the first few nights, which was not statistically significant. SCH 23390 significantly blocked the PHNO effect on locomotor activity on the first 5 nights. However, on subsequent nights, the activity in animals treated with SCH 23390 and PHNO increased. until the level was the same (or higher) than in the group receiving VEH + PHNO.
3000.
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3.2. Effects of S C H 23390 on stress-induced ret:ersal of tolerance to the stimulant actions of PHNO. The actions of the D-I receptor antagonist, SCH 23390 on locomotor activity for the 2 h after stress, and on the equivalent times of days without stress, in rats treated continuously with PHNO or vehicle, are depicted in fig. 3. Only the averages for the last 6 days of treatment were included to examine the effects during tolerance to the motor stimulant effects of PHNO. Analysis of variance revealed that there was a significant stress × PHNO × SCH 23390 interaction (F(I,20) = 9.31.
~:,,"H * V FH v'[H, PHNO TR'_-AT M.=NT
S( ~* PHNO
Fig. 3. Mean hourly d a y t i m e l o c o m o t o r activity over a 2 h period of mild environmental stress (cross-hatched bars) or at the equivalent times on days without stress (open bars) in rats given continuous infusions of vehicle ( V E H + VEt 1), S ( ' H 23390 120 ,~g/h. S C H , - V E H ) , P t I N O (5 I,t g / h , VEI-! + I)HNO), or both SCH 23390 anti P H N O ( S C H + P I t N ( ) ) . Only data for the last 6 days of an 11 day treatment regimen wcrc included; days on which tolerance to the motor stimulant effects of P H N O are apparent. Note that stress reverses tolerance to the stimulant action of P H N O , a D-2 receptor agonist. The I)-1 receptor antagonist attenuates stress-induced reversal of tolerance to P H N O ' s effects, but does not block stress-induced activity in vehicle-infused rats. Error bars reflect S.E.s of the difference scores.
29 ACTIVITY COUNTS/HOUR 1200-
T
1000" BOO 600
oolmW oNI
VEH*VEH
VEH÷SKF VEH*PHNO PHNO~'SKF TREATMENT
Fig. 4. Mean hourly locomotor activity counts ( + S.E.M.) over a 4 h period in rats treated with continuous infusions of P H N O (5 p.g/h) on days 8-11 of treatment, a time during which tolerance to the stimulant effects of PHNO, a D-2 receptor agonist, is observed. Injections of a D-1 receptor agonist, SKF 38393 (6.0 m g / k g i.p., SKF) reverses tolerance to P H N O ' s actions, without influencing activity in the vehicle (VEH)-infused group. See text for statistical details.
there was a significant interaction between P H N O and SKF 38393 (F(1,20) = 7.19, P < 0.02), indicating that SKF 38393 reversed the tolerance to the motor stimulant actions of PHNO.
4. Di~ussion
The present results replicate and confirm the previous findings (Martin-Iverson et al., in press) that continuous administration of the DA agonist, P H N O (selective for the D-2 receptor subtype) leads to the rapid development of tolerance to the motor stimulant actions of P H N O during the light periods of the day, while sensitization of the same behavioural effects occurs during the dark periods. The previous report that stress temporarily reverses tolerance was also replicated. It was hypothesized (Martin-Iverson et al., in press) that day time tolerance may have been the result of a progressive inhibition of endogenous DA release, possibly via action on presynaptic DA autoreceptors. While PHNO, a direct D-2 agonist, does not itself induce DA release (Martin et al., 1985), there is evidence that many of the behavioural
(Jackson and Jenkins, 1985; Jackson and Hashizume, 1986) and some neurophysiological (Waiters et al., 1987) effects of selective D-2 agonists require concomitant activation of D-1 receptors, either by endogenous DA, or exogenous DA agonists, such as SKF 38393. A progressive loss of endogenous DA release would therefore attenuate the behavioural actions of P H N O by decreasing activation of D-I receptors. Presumably, stress reverses tolerance by increasing the release of DA (Antelman et al., 1975). Tolerance does not occur at night, possibly because of the large increase in DA release during the night (O'Neill and Fillenz, 1986) with the relative ineffectiveness of presynaptic inhibition of DA release under conditions of high basal DA release (Mereu et al., 1986). The experiments of the present study were designed to test the validity of some of the assumptions underlying this hypothesis of tolerance to the motor stimulant effects of PHNO. Specifically, the assumption that the motor stimulant actions of P H N O require concomitant activation of D-1 receptors was examined by co-administration of the D-I receptor antagonist, SCH 23390. As predicted, it was observed that SCH 23390 blocked the locomotor activity induced by PHNO, both during the night, and during the day, before the development of tolerance to the actions of PHNO. Thus, it can be concluded that the motor stimulation induced by the D-2 receptor-selective agonist, PHNO, depends upon concomitant activation of D-1 receptors, presumably by DA, the endogenous ligand. It is of interest that the blockade of the locomotor effects of P H N O during the night by continuous administration of SCH 23390 was not maintained over successive nights. Indeed, on the last few nights of treatment, locomotor activity in the group of rats receiving SCH 23390 + P H N O was actually higher than that of the group receiving VEH + P H N O (see fig. 2). The possibility that this results from the development of supersensitivity of DA D-1 receptors is supported by previous observations of such actions of SCH 23390 (Creese and Chen, 1985). It is also of interest that chronic treatment with the D-1 antagonist results in an augmentation of the behavioural effects of a D-2 agonist, in the absence of influences on D-2 recep-
3O tots (Hess et al., 1986). Finally, the finding that chronic treatment with a non-selective DA indirect agonist attenuates D-2 receptor supersensitivity, but a u g m e n t s D-1 receptor supersensitivity in DA-depleted a n i m a l s (Parenti et al., 1986) or in patients with P a r k i n s o n ' s disease ( R i n n e et al.. 1985) suggests the appealing hypothesis that sensitization to the behavioural actions of D A agonists observed d u r i n g the night with chronic c o n t i n u o u s a d m i n i s t r a t i o n , or d u r i n g the day with chronic i n t e r m i t t e n t a d m i n i s t r a t i o n ( M a r t i n - I v e r s o n el al.. in press) is the result of D-1 receptor supersensitivity in the presence of either e n d o g e n o u s or exogenous ligand. The time course of the a u g m e n tation of n o c t u r n a l locomotor activity in rats treated with VEH + P H N O was very similar to that of recovery from SCH 23390-induced blockade of P H N O - i n d u c e d n o c t u r n a l activity, supporting this view. It was previously suggested that the reversal of tolerance to the behavioural effects of P H N O induced by stress is mediated by the action of e n d o g e n o u s DA on D-1 receptors, with stress ind u c i n g the release of DA ( M a r t i n - l v e r s o n et al., in press). The present observation that the D-1 receptor antagonist bk~cked the stress-induced reversal of tolerance supports this suggestion. It is of interest that while SCH 23390 blocked the stress reversal of P H N O tolerance, it had no effect on stress-induced activity itself, in vehicle-infused rats. This indicates that the s t i m u l a t i o n of m o t o r activity by stress does not d e p e n d u p o n a DA I)-I receptor substrate. Finally, if tolerance to the motor s t i m u l a n t effects of P H N O develops as a c o n s e q u e n c e of loss of activation of D-1 receptors by e n d o g e n o u s DA. then it follows that an exogenous D-I receptor agonist would reverse tolerance. This was observed in the present experiments, using S K F 38393, an agonist selective for the D-I s u b t y p e of D A receptors. The present experiments establish that (11 an agonist selective for the D-2 subtype of DA receptor requires c o n c o m i t a n t activation of the D-I receptor subtype for expression of its behavioural actions; (2) stress-reversal of tolerance to the behavioural effects of a D-2 agonist is mediated by actions on the D-1 receptor, p r e s u m a b l y by en-
dogenous DA: (3) tolerance to the behavioural effects of a D-2 agonist can be reversed by a I)-I agonist. These results offer strong support to the hypothesis that tolerance to some of the behavioural actions of a D-2 agonist develops as a consequence of a rapidly progressive loss of activation of D-1 receptors by e n d o g e n o u s DA. They also suggest the possibility that behavoural sensitization may result from release of DA, u n d e r certain circumstances such as after stress or d u r i n g the night, in the presence of D-I receptors made supersensitive by daytime, non-stressed periods of i n h i b i t i o n of e n d o g e n o u s DA release.
Acknowledgements This research was ~,upported by Merck Sharpe & Dohme. M.T.M.-I. v,a,,, supported by a NATO Science Fellowship. We appreciate the excellent typing skills of H Stelte.
References Antelman. S.M., 17. Szechtman, P. Chin and A.I). Fisher, 1975. Tail pinch-induced eating, gnawing and licking behavior in rats: dependence on the nigrostriatal dopamine system, Brain Res. 99, 319. Creese, I. and A. ('hen, 1985, Selective D-1 dopamine receptor increase following chronic treatment with St'It 23390, |:,uropean J. Pharmacol. 109, 127. Hess, I'.J., I,.H Albers, II. l,e and 1. ('recsc, 1986. Effects of chronic S('H 23390 treatment on the bi(~.'hemical and behavioral properties of t) I and I'), dopamine receptors: potentiated behavioral responses to a D: dopaminc agonist after selectivc D 1 dopamine receptor upregulation. J. Pharmacol. Exp. l'her. 238, 846. Jackson, D.M. antt M. Hashizun]e, 1986, Bromocriptine induces marked locomotor stimulation in dopamine-depleted nficc when D-I dopamine receptors are stimulated with SKF 38393, Psychopharmacology 90, 147. Jackson, I).M. and ().F. Jenkins, 1985, Ilypothesis: bromtvriptine lacks intrinsic dopamine receptor stimulating pr¢>pcrties. ,I. Neural Transm. 62, 219. Martin, GE., M. Williams. I).J. Pettibone, G.(,;. Yarbrough. B.V. ('lineschmidt and .IH. Jones, 1984, Phammcologic profile of a novel potent direct-acting dopamine agonist, ( ) )-4-propyl-9-hydroxynaphthoxazine [(+)-PttN()]. ,1. Pharmacol. Exp. Ther. 230, 869. Martin-lverson, M.T.. S.M. Stahl and S.D. Iversen. Chronic administration of a selective dopamine 1)-2 agonist: factors determining behavioura[ tolerance and sensitiTation, P,~'.'chopharmacology (in press).
31 Mereu, G., F. Miuntoni, P. Calabresi, F. Romani, V. Boi and G.L. Gessa, 1986, Responsiveness to 'autoreceptor' doses of apomorphine is inversely correlated with the firing rate of dopaminergic A9 neurons: action of baclofen, Neurosci. Left. 65, 161. Molloy, A.G., K.M. O'Boyle, M.T. Pugh and J.L. Waddington, 1986. Locomotor behaviors in response to new selective D-1 and D-2 dopamine receptor agonists, and the influence of selective antagonists, Pharmacol. Biochem Behav. 25. 249. O'Neill, R.D. and M. Fillenz, 1985, Simultaneous monitoring of dopamine release in rat frontal cortex, nucleus accumhens and striatum: effect of drugs, circadian changes and correlations with motor activity, Neuroscience 16, 49.
Parenti, M., C. Flauto, E. Parati, A. Vescovi and A. Groppetti, 1986, Differential effect of repeated treatment with I,DOPA on dopamine-D1 or -D2 receptors, Neuropharmacology 25, 331. Rinne, J.O., J.K. Rinne, K. Laakso, P. Lonnberg and U . K Rinne. 1985, Dopamine D-1 receptors in the Parkinsonian brain, Brain Res. 359, 306. Waiters, J.R., D.A. Bergstrom, J.H. Carlson, T.N. Chase and A.R. Braun, 1987. D 1 dopamine receptor activation required for postsynaptic expression of D 2 agonist effects, Science 236. 719.
25
Elsevier F.IP 50238
Long-term motor stimulant effects of ( +)-4-propyl-9-hydroxynaphthoxazine (PHNO), a dopamine D-2 receptor agonist: interactions with a dopamine D-1 receptor antagonist and agonist Mathew T. Martin-Iverson *, Susan D. Iversen and Stephen M. Stahl Merck Sharpe & Dohme. Neurosciem'e Research ('entre. Terlings Park. Eastwick Road. llarh~w, E~'sex ('M20 2QR. U.K.
Received 26 October 1987, revised MS received 26 January 1988, accepted 9 February 1988
Rats were given continuous infusions of ( + )-4-propyl-9-hydroxynaphthoxazine (PHNO, 5 ttg/h), a dopamine D-2 receptor agonist, using subcutaneous implants of ALZET osmotic minipumps. It was observed that tolerance occurred to the motor stimulant effects of PHNO during the light cycle of each day, but not during the dark cycle. Rather, thc motor stimulant actions of PHNO were gradually augmented during successive nights. Daytime tolerance to the stimulant actions of PI-|NO was reversed by a mild environmental stress or by administration of the D-I receptor agonist, SKF 38393 (6 m g / k g i.p.). Co-administration of the dopamine D-I receptor antagonist, (SCH 23390, 20 p,g/h s.c. by ALZET osmotic minipumps), initally blocked the motor stimulant actions of PHNO and also attenuated the reversal of tolerance to PHNO produced by stress, without blocking the actions of stress on activity in vehicle-infused animals. These results indicate that tolerance to the behavioural effects of PHNO may result from a loss of activation of D-I receptors by endogenous DA. Dopamine receptor subtypes: Locomotor acitivity; PHNO (( + )-4-propyl-9-hydroxynaphthoxazine); (Tolerance, Sensitization)
I. Introduction ( + )-4-Propyl-9-hydroxynaphthoxazine (PHNO) is an extremely p o t e n t d o p a m i n e ( D A ) agonist with a m a r k e d selectivity for the D-2 r e c e p t o r s u b t y p e ( M a r t i n et al., 1984). In a previous s t u d y ( M a r t i n - l v e r s o n et al., in press), it was o b s e r v e d that chronic c o n t i n u o u s a d m i n i s t r a t i o n of P H N O to rats by Alzet o s m o t i c m i n i p u m p s resulted in a r a p i d d e v e l o p m e n t of tolerance to the m o t o r s t i m u l a n t effects of P H N O d u r i n g the light p e r i o d
* To whom all correspondence should be addressed at present address: Neurc.chemical Research Unit, Department of Psychiatry, 1E7.44 Walter Mackenzie lleahh Sciences Centre, University of Alberta, Edmonton, Alberta, Canada T6G 2B7.
of the light-dark cycle. However, the m o t o r s t i m u l a n t actions of P H N O were g r a d u a l l y augm e n t e d d u r i n g the d a r k p e r i o d s of successive 24 h periods, on the s a m e d a y s that tolerance occurred d u r i n g the light periods. It was further o b s e r v e d that stress could t e m p o r a r i l y reverse "daytime' tolerance. To explain these o b s e r v a t i o n s , a hypothesis was p r o p o s e d that is b a s e d on the recent suggestion that there is a functional interaction between different s u b t y p e s of D A receptors. Specifically, some b e h a v i o u r a l effects of D A agonists selective for the D-2 r e c e p t o r s u b t y p e c a n n o t be elicited without the c o n c o m i t a n t a c t i v a t i o n of the D-I receptor s u b t y p e ( M o l l o y et al., 1986). In a n o r m a l animal, the acute a d m i n i s t r a t i o n of a selective D-2 receptor agonist can elicit m a n y of its b e h a v i o u r a l effects b e c a u s e of c o n c o m i t a n t activation of D-1
0014-2999/88/$03.50 :': 1988 Elsevier Science Publishers B.V. (Biomedical I)ivision)
26 receptors, presumably by endogenous release of DA. That endogenous DA may activate D-I receptors, allowing the expression of motor stimulation by the D-2 agonists, is supported by the findings that extensive DA depletions block the motor stimulation of bromocriptine, a D-2 receptorselective agonist (Jackson and Jenkins, 1985: Jackson and Hashizume, 1986). Agents that promote DA release (Jackson and Jenkins, 1985) or a D-1 receptor-selective agonist (Jackson and Hashizume, 1986) can re-instate the motor stimulant actions of bromocriptine blocked by DA depletion. Furthermore, these behavioural interactions are paralleled by similar interactions between agonists selective for different DA receptor subtypes, in relation to effects on the firing rates of basal ganglia neurons (Waiters et al., 1987). It has been observed that DA release is high at night and low during the day (O'Neill and Fillenz, 1985), and that mild stress can induce DA release (Antelman et al., 1975). These data, in conjunction with the evidence that DA autoreceptors are particularly effective under conditions of low basal release of DA (Mereu et al., 1986) prompted the speculation that continuous administration of a [)-2 receptor agonist leads eventually to a decrease in endogenous DA release during the day, but not during the night. Therefore, behavioural tolerance to continuous infusions of P H N O occurs only during the day. as a consequence of loss of activation of D-1 receptors by endogenous I)A. The observation that a mild stress (which may induce DA release) temporarily reversed tolerance was taken as support for this interpretation. In brief, it was proposed that tolerance to the motor stimulant effects of P H N O was a result of loss of activation of D-I receptors by endogenous DA. This hypothesis of the mechanism of tolerance to PHNO, a D-2 agonist, makes several testable predictions. Firstly, a D-1 antagonist, such as SCH 23390 should block PHNO-induced motor stimulation. Secondly, SCH 23390 should block the stress-induced reversal of tolerance to PHNO. Thirdly, a D-I agonist, like SKF 38393, should reverse tolerance to PHNO. The following experiments were undertaken to test these predictions.
2. Materials and methods
2. I. Experimental animals and al~paratus Male Sprague-Dawley rats ( n = 6 for each group) weighing 30(-)-350 g were housed in cages equipped with infrared photocell apparatus which allowed continuous monitoring of locomotor activity over periods of weeks. The cage dimensions were 24 (W) × 16 (H) × 38 (l,) cm, with two photocells placed 3 cm from the floor, and spaced 13 cm apart, equidistant from both end walls. The rats lived in these cages (under a light-dark cycle in which the light period was from 9:00 to 21:00 h) for at least 10 days prior to any drug treatments. Drug treatments were continued for 11-15 days, depending upon the experiment, with continuous recording of photobeam interruptions in hourly blocks. On Mondays, Wednesdays and Fridays the rats were given fresh water and food, and the litter underneath the cages was changed. This procedure was accompanied with a fair amount of noise, lasting close to 20 min, and constitutes the 'stress' variable of the present experiments. Average activity counts were calculated over blocks of 12 h (nocturnal activity), 10 h (diurnal activity) and 2 h (time during and 100 min following the care of the animals (stress), or the equivalent periods on days without stress. 2.2. ('ompound~ P H N O H('I (Merck Sharpe & Dohme) was dissolved in distilled water and A L Z E T osmotic minipumps (No. 2002) were filled with the solution to provide a release rate of 5.0 p,g/h (weight expressed as free base), which gave a dose of approximately 15 #g kg 1 h - i . SCH 23390 (RBI) was dissolved in distilled water and given by ALZ E T osmotic minipumps at the rate of 20 # g / h (about 60 p,g kg i h - i ) . SKF 38393 (RB1) was dissolved in distilled water for a dose of 6.0 m g / k g (Lp.). 2.3. Continuous administration of P l t N O and S ( ' I I 233~)0 All animals received two osmotic minipumps implanted s.c. between the shoulders while the rats
27
were under ether anaesthesia. Different groups of rats were given two pumps containing only vehicle (VEH + VEH), one p u m p containing vehicle and the other P H N O (VEH + PHNO), one p u m p loaded with SCH 23390 and the other with vehicle (SCI-I + VEH), or one pump containing P H N O and the other loaded with SCH 23390 (SCH + PHNO). The n of each group was 6. Interruptions of photobeams were recorded in hourly blocks throughout drug treatments. 2.4. Administration of S K F 38393
ACTIVITY
I0'0001 80004_ f
3. Results
3.1. Lffe¢'ts of continuous administration of PItNO, S C t l 23390 and both with s.c. osmotic minipumps on locomotor actit'ity. Figure 1 displays the effects of continuous administration of vehicle, PHNO, SCH 23390 and both drugs on locomotor activity, presented as the average photobeam interruptions from the 10 h of the light cycle (exclusive of the 2 h during and after environmental disturbance (stress) or the equivalent times on days without stress). Analysis of variance revealed a significant SCH 23390 x P H N O x Day interaction (F(10,200) = 8.20, P < 0.001). As previously observed (Martin-Iverson et al., in press), continuous administration of this dose of P H N O resulted in tolerance to the motor
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Four groups of rats (n = 6 for each group) were used in this experiment. Two groups were implanted with osmotic minipumps containing vehicle (sterile water), and two groups were given osmotic pumps containing P H N O (5 # g / h ) . Seven days after osmotic p u m p implants, one vehicle-infused group and one PHNO-infused group were given injections of SKF 38393 (6.0 m g / k g i.p.) at 12:00 h on each of 4 consecutive days. The remaining two groups (one with vehicle infusions and one with P H N O infusions) were given vehicle injections (sterile water, 1 m l / k g i.p.) at the same times. The photobeam interruptions occurring during the 4 h after injections were collapsed across days to obtain an average for each animal.
COUNTS
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Fig. l. Mean daytime locomotor activity counts of rats given continuous infusions of vehicle and vehicle ( V E H + V E H ) , vehicle and SCH 23390 (20 p.g/h, VFH + SCH), PHNO (5.0 p.g/h) and vehicle (PHNO + VEIl), or PHNO and SCH 23390 ( P H N O + S C H ) . Note that tolerance to the motor stimulant effects of PHNO, a selective D-2 receptor agonist, occurs rapidly over the first 3 days. The D-I receptor antagonist, SCH 23390, completely blocked the initial stimulant action of PHNO, but partial 'escape' from this blockade was evident on days 7, 9 and 11.
stimulant effects developing rapidly (within 3-4 days) during the light periods. The D-1 antagonist, SCH 23390, appeared to reducc acitivity when given with vehicle, but this effect was not significant. However, when given with PHNO, it significantly blocked the increase in activity on the first 2 days (P < 0.01 on both days, Tukey's HSD test). Interestingly, on days 7. 9 and 11 (days without stress) activity levels in the group receiving both P H N O and SCH 23390 began to exhibit increases in activity (significantly different from vehicle: P < 0 . 0 1 (day 9) and P < 0 . 0 5 (day 11), Tukcy's HSD). Figure 2 shows the effects of drug treatments on nocturnal activity, expressed as the average photobeam counts over the 12 11 dark periods. Analysis of variance indicated a significant SCH 23390 × P H N O × Night interaction (F(11,220) = 4.53, P < 0.001). The locomotor activity induced by P H N O gradually increased over successive nights (fig. 2). This effect was significantly different from that of the VEH + VEH group on every night (P < 0.01), and the effect of P H N O on the last night was significantly higher than the motor
28
A C ' I V I T Y COUNTS
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P < 0.01), reflecting that the tolerance to PHNO activity-inducing effects observed on days without stress was reversed by stress, and that this stressinduced reversal of behaviour tolerance to PHNO was blocked by SCH 23390. SCH 23390 did not significantly attenuate stress-induced activity m vehicle-infused rats. 3.3. Effects of administration of S K F 38393 to rats exhibiting tolerance m the motor stimulant actions of PHNO.
12
SUCCESSIVE NIGHTS
Fig. 2. Mean nocturnal locomotor activity counts of rats given c o n t i n u o u s infusions of vehicle and vehicle ( V E H + V E t t ) , vehicle and SCH 23390 (20 p , g / h , V E H + S C H ) , P H N O (5 # g / h ) and vehicle ( P H N O + VEH). or P H N O and SCH 23390 (PHNO+SCH). U n l i k e daytime activity, ncx:turnal activity was gradually augmented over successive nights in the groups receiving P H N O , a D-2 receptor agonist. The D-1 receptor antagonist, SCH 23390, blocked the initial motor stimulant effects of P H N O . but this action was no longer apparent by the sixth night.
Figure 4 displays the results of the administration of the D-1 receptor agonist. SKF 38393, on locomotor behaviour of rats exhibiting tolerance to the locomotor stimulant actions of PHNO. or of rats treated with vehicle. There were no significant effects of PHNO in rats given injections of vehicle in addition to the PHNO infusions. Neither wcre effects of SKF 38393 on activity apparent in rats given vehicle infusions. On the other hand.
All VTI" CO~NIS/4OUR
activity induced on the first night (P < 0.01). The group given SCH 23390 + VEH again exhibited an apparent decrease in activity on the first few nights, which was not statistically significant. SCH 23390 significantly blocked the PHNO effect on locomotor activity on the first 5 nights. However, on subsequent nights, the activity in animals treated with SCH 23390 and PHNO increased. until the level was the same (or higher) than in the group receiving VEH + PHNO.
3000.
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3.2. Effects of S C H 23390 on stress-induced ret:ersal of tolerance to the stimulant actions of PHNO. The actions of the D-I receptor antagonist, SCH 23390 on locomotor activity for the 2 h after stress, and on the equivalent times of days without stress, in rats treated continuously with PHNO or vehicle, are depicted in fig. 3. Only the averages for the last 6 days of treatment were included to examine the effects during tolerance to the motor stimulant effects of PHNO. Analysis of variance revealed that there was a significant stress × PHNO × SCH 23390 interaction (F(I,20) = 9.31.
~:,,"H * V FH v'[H, PHNO TR'_-AT M.=NT
S( ~* PHNO
Fig. 3. Mean hourly d a y t i m e l o c o m o t o r activity over a 2 h period of mild environmental stress (cross-hatched bars) or at the equivalent times on days without stress (open bars) in rats given continuous infusions of vehicle ( V E H + VEt 1), S ( ' H 23390 120 ,~g/h. S C H , - V E H ) , P t I N O (5 I,t g / h , VEI-! + I)HNO), or both SCH 23390 anti P H N O ( S C H + P I t N ( ) ) . Only data for the last 6 days of an 11 day treatment regimen wcrc included; days on which tolerance to the motor stimulant effects of P H N O are apparent. Note that stress reverses tolerance to the stimulant action of P H N O , a D-2 receptor agonist. The I)-1 receptor antagonist attenuates stress-induced reversal of tolerance to P H N O ' s effects, but does not block stress-induced activity in vehicle-infused rats. Error bars reflect S.E.s of the difference scores.
29 ACTIVITY COUNTS/HOUR 1200-
T
1000" BOO 600
oolmW oNI
VEH*VEH
VEH÷SKF VEH*PHNO PHNO~'SKF TREATMENT
Fig. 4. Mean hourly locomotor activity counts ( + S.E.M.) over a 4 h period in rats treated with continuous infusions of P H N O (5 p.g/h) on days 8-11 of treatment, a time during which tolerance to the stimulant effects of PHNO, a D-2 receptor agonist, is observed. Injections of a D-1 receptor agonist, SKF 38393 (6.0 m g / k g i.p., SKF) reverses tolerance to P H N O ' s actions, without influencing activity in the vehicle (VEH)-infused group. See text for statistical details.
there was a significant interaction between P H N O and SKF 38393 (F(1,20) = 7.19, P < 0.02), indicating that SKF 38393 reversed the tolerance to the motor stimulant actions of PHNO.
4. Di~ussion
The present results replicate and confirm the previous findings (Martin-Iverson et al., in press) that continuous administration of the DA agonist, P H N O (selective for the D-2 receptor subtype) leads to the rapid development of tolerance to the motor stimulant actions of P H N O during the light periods of the day, while sensitization of the same behavioural effects occurs during the dark periods. The previous report that stress temporarily reverses tolerance was also replicated. It was hypothesized (Martin-Iverson et al., in press) that day time tolerance may have been the result of a progressive inhibition of endogenous DA release, possibly via action on presynaptic DA autoreceptors. While PHNO, a direct D-2 agonist, does not itself induce DA release (Martin et al., 1985), there is evidence that many of the behavioural
(Jackson and Jenkins, 1985; Jackson and Hashizume, 1986) and some neurophysiological (Waiters et al., 1987) effects of selective D-2 agonists require concomitant activation of D-1 receptors, either by endogenous DA, or exogenous DA agonists, such as SKF 38393. A progressive loss of endogenous DA release would therefore attenuate the behavioural actions of P H N O by decreasing activation of D-I receptors. Presumably, stress reverses tolerance by increasing the release of DA (Antelman et al., 1975). Tolerance does not occur at night, possibly because of the large increase in DA release during the night (O'Neill and Fillenz, 1986) with the relative ineffectiveness of presynaptic inhibition of DA release under conditions of high basal DA release (Mereu et al., 1986). The experiments of the present study were designed to test the validity of some of the assumptions underlying this hypothesis of tolerance to the motor stimulant effects of PHNO. Specifically, the assumption that the motor stimulant actions of P H N O require concomitant activation of D-1 receptors was examined by co-administration of the D-I receptor antagonist, SCH 23390. As predicted, it was observed that SCH 23390 blocked the locomotor activity induced by PHNO, both during the night, and during the day, before the development of tolerance to the actions of PHNO. Thus, it can be concluded that the motor stimulation induced by the D-2 receptor-selective agonist, PHNO, depends upon concomitant activation of D-1 receptors, presumably by DA, the endogenous ligand. It is of interest that the blockade of the locomotor effects of P H N O during the night by continuous administration of SCH 23390 was not maintained over successive nights. Indeed, on the last few nights of treatment, locomotor activity in the group of rats receiving SCH 23390 + P H N O was actually higher than that of the group receiving VEH + P H N O (see fig. 2). The possibility that this results from the development of supersensitivity of DA D-1 receptors is supported by previous observations of such actions of SCH 23390 (Creese and Chen, 1985). It is also of interest that chronic treatment with the D-1 antagonist results in an augmentation of the behavioural effects of a D-2 agonist, in the absence of influences on D-2 recep-
3O tots (Hess et al., 1986). Finally, the finding that chronic treatment with a non-selective DA indirect agonist attenuates D-2 receptor supersensitivity, but a u g m e n t s D-1 receptor supersensitivity in DA-depleted a n i m a l s (Parenti et al., 1986) or in patients with P a r k i n s o n ' s disease ( R i n n e et al.. 1985) suggests the appealing hypothesis that sensitization to the behavioural actions of D A agonists observed d u r i n g the night with chronic c o n t i n u o u s a d m i n i s t r a t i o n , or d u r i n g the day with chronic i n t e r m i t t e n t a d m i n i s t r a t i o n ( M a r t i n - I v e r s o n el al.. in press) is the result of D-1 receptor supersensitivity in the presence of either e n d o g e n o u s or exogenous ligand. The time course of the a u g m e n tation of n o c t u r n a l locomotor activity in rats treated with VEH + P H N O was very similar to that of recovery from SCH 23390-induced blockade of P H N O - i n d u c e d n o c t u r n a l activity, supporting this view. It was previously suggested that the reversal of tolerance to the behavioural effects of P H N O induced by stress is mediated by the action of e n d o g e n o u s DA on D-1 receptors, with stress ind u c i n g the release of DA ( M a r t i n - l v e r s o n et al., in press). The present observation that the D-1 receptor antagonist bk~cked the stress-induced reversal of tolerance supports this suggestion. It is of interest that while SCH 23390 blocked the stress reversal of P H N O tolerance, it had no effect on stress-induced activity itself, in vehicle-infused rats. This indicates that the s t i m u l a t i o n of m o t o r activity by stress does not d e p e n d u p o n a DA I)-I receptor substrate. Finally, if tolerance to the motor s t i m u l a n t effects of P H N O develops as a c o n s e q u e n c e of loss of activation of D-1 receptors by e n d o g e n o u s DA. then it follows that an exogenous D-I receptor agonist would reverse tolerance. This was observed in the present experiments, using S K F 38393, an agonist selective for the D-I s u b t y p e of D A receptors. The present experiments establish that (11 an agonist selective for the D-2 subtype of DA receptor requires c o n c o m i t a n t activation of the D-I receptor subtype for expression of its behavioural actions; (2) stress-reversal of tolerance to the behavioural effects of a D-2 agonist is mediated by actions on the D-1 receptor, p r e s u m a b l y by en-
dogenous DA: (3) tolerance to the behavioural effects of a D-2 agonist can be reversed by a I)-I agonist. These results offer strong support to the hypothesis that tolerance to some of the behavioural actions of a D-2 agonist develops as a consequence of a rapidly progressive loss of activation of D-1 receptors by e n d o g e n o u s DA. They also suggest the possibility that behavoural sensitization may result from release of DA, u n d e r certain circumstances such as after stress or d u r i n g the night, in the presence of D-I receptors made supersensitive by daytime, non-stressed periods of i n h i b i t i o n of e n d o g e n o u s DA release.
Acknowledgements This research was ~,upported by Merck Sharpe & Dohme. M.T.M.-I. v,a,,, supported by a NATO Science Fellowship. We appreciate the excellent typing skills of H Stelte.
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Parenti, M., C. Flauto, E. Parati, A. Vescovi and A. Groppetti, 1986, Differential effect of repeated treatment with I,DOPA on dopamine-D1 or -D2 receptors, Neuropharmacology 25, 331. Rinne, J.O., J.K. Rinne, K. Laakso, P. Lonnberg and U . K Rinne. 1985, Dopamine D-1 receptors in the Parkinsonian brain, Brain Res. 359, 306. Waiters, J.R., D.A. Bergstrom, J.H. Carlson, T.N. Chase and A.R. Braun, 1987. D 1 dopamine receptor activation required for postsynaptic expression of D 2 agonist effects, Science 236. 719.