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Amphetamine withdrawal: A behavioral evaluation
Life Sciences, Vol. 38, pp. 1617-1623 Printed in the U.S.A.
Pergamon Press
AMPHETAMINE WITHDRAWAL: A BEHAVIORAL EVALUATION Larry Kokkinidis, Robert M. Zacharko and Hymie Anisman Department of Psychology, University of Saskatchewan Saskatoon, Saskatchewan S7N OWO and Department of Psychology, Carleton University, Ottawa, Ontario KIS 5B6 Canada (Received in final form February 18, 1986) Summary The effects of withdrawal from long-term amphetamine treatment on intracranial self-stimulation, forced swiminduced immobility, shuttle escape performance, acoustic startle and locomotor activity were evaluated. Mice implanted with stimulating electrodes in the lateral hypothalamus demonstrated stable and reliable rates of self-stimulation responding. After exposure to a chronic schedule of amphetamine treatment response rates were severely depressed. In addition to modifying intracranial self-stimulation responding, amphetamine withdrawal increased the duration of immobility in a forced-swim situation. Although chronic amphetamine exposure induced pronounced behavioral changes in the intracranial self-stimulation and forced swim tasks, drug withdrawal had little effect on shuttle escape performance, acoustic startle and locomotor activity. Based on these findings it was suggested that the development of postamphetamine depression in the self-stimulation and forced swim paradigms was not related to variations in motoric or arousal mechanisms resulting from amphetamine withdrawal, but rather involved drug-induced changes in motivational processes. Several behavioral paradigms currently employed as animal models of clinical depression have proven to be valuable tools in evaluating the potenti~ therapeutic efficacy of antidepressant agents. One behavioral model of depression that has received considerable attention is the development of behavioral deficits in animals following exposure to inescapable shock (1). Specifically, animals exposed to inescapable shock with little or no control of their response to the stressor, developed performance deficits during subsequent escape testing (2). In a somewhat related behavioral situation animals forced to swim with no chance of escape often become immobile and assume a floating posture (3). Tricyclic antidepressant treatment has proven effective in mitigating the stress-induced deficits in escape responding (4), as well as the development of immobility in the forced-swim paradigm (3). Recent work involving the effects of long-term amphetamine administration on intracranial self-stimulation (ICSS) has shown that withdrawal from a chronic drug schedule of amphetamine treatment provoked a pronounced and sustained depression of rates of responding for electrical brain stimulation (5, 6, 7, 8). The available data suggests that the post-amphetamine depression 0024-3205/86 $3.00 + .00 Copyright (c) 1986 Pergamon Press Ltd.
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of ICSS is not related to a general lethargy induced by drug withdrawal that may exert a non-specific influence on ICSS performance, but rather involves an attenuation of the reinforcing effects of electrical brain stimulation. In accordance with this view it was shown that rats exhibited depressed rates of responding and an elevation of current threshold for several weeks after cessation of amphetamine treatment (8). Using a rate-independent measure of ICSS, Cassens and co-workers (9) found increased reinforcement thresholds after animals were exposed to chronic amphetamine treatment, and these authors suggested as have others, that the post-amphetamine depression of ICSS may have clinical implications when it is considered that anhedonia is characteristic of both amphetamine-induced and naturally occurring depression in humans (8, 9). Consistent with this position we found that chronic administration of the tricyclic antidepressants imipramine and amitriptyline, attenuated the postamphetamine depression of ICSS (lO). Although the ICSS paradigm, the shuttle escape task and the forced swim test have all been offered as potential models of depression and useful predictors of antidepressant drugs (for review see ll), these tasks most likely reflect different features of performance. Indeed, shuttle escape performance and the forced swim procedure appear to be sensitive to manipulations which affect the motoric capacity of the animals (2, 12). In contrast, the ICSS paradigm provides direct evaluation of changes in central reward processes associated with pharmacological treatment, although contribution of performance variables cannot be totally discounted (13). Since it is well established that withdrawal from amphetamine results in a pronounced depression of ICSS, it was of interest to determine whether a schedule of amphetamine administration that impaired ICSS responding would also modify shuttle escape performance and immobility in a forced-swim test. There is reason to believe that this might well be the case given the data showing that stressors and amphetamine are largely interchangeable with respect to their effects on behavior (14, 15). In addition to ICSS, shuttle escape, and swim-induced immobility, the effects of amphetamine withdrawal on locomotor activity and acoustic startle were assessed to determine directly whether amphetamine withdrawal modified arousal processes. Material and Methods Subjects Eighty male CD-I mice, 80 days of age obtained from the Canadian Breeding Laboratories (Quebec, Canada) served as subjects. Mice were housed individually in standard polypropylene cages and allowed free access to food and water. Behavioral testing was carried out during the light portion of the light/dark cycle. Apparatus The ICSS apparatus consisted of a black Plexiglas box 0.6 cm thick and 15.0 cm in width, length and height. A hole 1.5 cm in diameter was located in the centre of the Plexiglas floor of the apparatus. A photobeam unit was situated directly beneath the hold (0.5 cm) such that interruption of the photobeam by a head-poke resulted in electrical brain stimulation (0.3 sec in duration). Brain stimulation (300 uA base to peak) was delivered from an Ortec Dual Channel Stimulator and consisted of monophasic square wave pulses (lO0 Hz) with a pulse duration of O.l msec. Current was determined by monitoring on an oscilloscope the drop in voltage output across a lO K resistor placed in series with the stimulating electrode. LeHigh Valley timers and counters controlled the duration of the brain stimulation and recorded response
Vol. 38, No. 17, 1986
Post-amphetamine Depression
1619
rates. The shuttle boxes used for escape testing consisted of black Plexiglas and measured 26.4 x 9.0 x 15.5 cm. The two compartments of each of the six boxes were separated by a horizontally moveable stainless steel gate. The grid floor of each compartment consisted of stainless steel rods (0.32 cm) spaced l.O cm apart. Footshock (150 uA, AC) was delivered through a 3000 V source. Shuttle boxes were housed in sound attenuated chambers and were controlled by a microcomputer system. Acoustic startle and locomotor activity was recorded in two styrofoam (2.0 cm thick) circular chambers 28.0 cm in diameter and 21.0 cm high. The styrofoam floor of each chamber was covered with a plastic sheath and was positioned over an 8~W speaker (28.0 cm in diameter). Voltages produced by movements on the floor were fed to a Commodore PET Series 2001 computer. The analogue signal from the speaker was amplified and digitized by an 8 bit A/D converter. The digitized output was printed out on a Data Terminal Mart printer. In the case of acoustic startle, only activity during the tone presentation was measured. The 2700 Hz tone (700 msec in duration, 5-msec rise-fall time) was generated by a Piezo Crystal Audio Transistor (Projects Unlimited, Dayton, OH) situated in the centre of the styrofoam roof of each chamber. The intensity of the tone in the chamber was 97 dB and background noise in the chambers was 44 dB. Sound intensity measures were made with a Bruel Kjaer sound level meter (model 2203; A-scale). Procedure Intracranial Self-Stimulation Mice were anesthetized with sodium pentobarbital (65.0 mg/kg) and were implanted in the lateral hypothalamus with insulated bipolar electrodes (Plastic Products), which had 0.5 mm of the tips separated and scraped. Coordinates for electrode placements determined by preliminary histological analysis were: AP-O.5 mm from Bregma, L + 1.2 mm from midline suture and V 5.7 mm from a flat skull surface. Following a 7 day post-operative period mice were tested for ICSS for a 20 min session on 6 consecutive days. Only mice demonstrating reliable and stable rates of responding were used in the experiment. Mice showing low and irregular rates of responding were replaced. Once baseline rates had stabilized, mice were subdivided on the basis of average equalized baseline rates such that one-half of the animals (N = I0/ group) received 2 daily intraperitoneal (i.p.) injections (lO:O0 a.m. and 4:00 p.m.) of 5.0 mg/kg of d-amphetamine sulfate for lO consecutive days, while the remaining half were treated with an equivalent volume of saline. Twenty-four hours following the last injection of amphetamine mice were placed in the self-stimulation chamber and the number of head-poke responses during a lO min test session were recorded. Forced-swim Test Twenty naive mice were placed into 2 groups (N = lO/group) and injected with either 2 daily i.p. injections of d-amphetamine (5.0 mg/kg) or saline for IO consecutive days. The day after the last injection mice in each group were placed in a glass cylinder (18.5 cm in diameter and 25.0 cm high) filled to within 7.0 cm from the top with warm tap water (30 o C) for 15 min. Following the forced-swim session mice were towel dried and placed in a polypropylene mouse cage lined with paper towels for 30 min. Mice were then re-exposed to the water in the glass cylinder for 5 min. The duration of immobility (i.e., floating) during the 5 min test was recorded.
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Shuttle Escape Task Two groups of mice (N = lO/group) were chronically treated with either amphetamine or saline as previously described. Twenty-four hours after the last injection mice in both groups were exposed to 60 escapable shocks at 60 sec intervals. Shock was presented for 6 sec, after which the gate separating the two compartments opened permitting entry into th~ shock free compartment. The latency to escape into the safe compartment was recorded. If successful escape had not occurred within 24 sec of shock presentation then the trial was terminated and 60 sec later a new trial was initiated. Locomotor A c t i v i t y
and Acoustic Startle
Naive mice were randomly assigned to either the chronic amphetamine or saline conditions. Twenty-four hours after the lO days of 2 daily injections of either 5.0 mg/kg of d-amphetamine or saline mice in each group (N = lO/grou~ were placed into the activity chambers and locomotor activity was monitored for 20 min~. Mice tested for acoustic startle were treated in the same manner with the exception that they were exposed to 160 tone presentations with a I0 sec interval. This procedure has been shown to produce an accurate measure of acoustic startle arousal and changes in activity levels during the tone burst are representative of the startle reflex and not random activity levels (16). Results Following testing mice with electrode placements in the lateral hypothalamus received an overdose of sodium pentobarbital and were perfused with 0.9% saline followed by I0% formalin. Frozen coronal sections were cut at 40 uM and stained with thionin. The histological examination of the sections revealed that in all cases electrode placements were in the vicinity of the lateral hypothalamus. Withdrawal from long-term amphetamine treatment had pronounced effects on ICSS responding. As can be seen in Table l rates of responding 24 hours after withdrawal from amphetamine treatment were significantly depressed relative to ICSS rates of control animals. At the same time interval, withdrawal from amphetamine modified the duration of immobility during the forced-swim test. That is, mice treated with amphetamine spent significantly more time in a floating posture than did saline treated mice. In marked contrast to the effects of amphetamine withdrawal on ICSS and swim-induced immobility, withdrawal from amphetamine did not modify performame in a shuttle escape task. If anything, mice withdrawn from the drug were somewhat faster with respect to escape latencies relative to control animals. Similarly there were no effects of amphetamine withdrawal on the acoustic startle reflex or locomotor activity although consistent with the shuttle escape data mice withdrawn from the drug showed slightly higher rates of locomotor activity than did saline treated mice (see Table l). Discussion Mice exposed to a chronic schedule of amphetamine and then tested of ICSS during withdrawal showed depressed rates of responding for electrical brain stimulation. This observation was consistent with several reports demonstrating the development of post-amphetamine depression of ICSS after amphetamine withdrawal (5, 6, 7, 8, 17). Although this study did not assess the temporal parameters associated with the development of the ICSS depression, several reports have addressed this issue and have shown that the postamphetamine depression persists for several weeks after drug withdrawal (~I0~7~
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Table 1 Effects of Amphetamine Withdrawal on Intracranial Self-Stimulation, Responding, Forced-Swim Immobility, Shuttle Escape Performance, Locomotor Activity and Acoustic Startle Reflex
Saline
Amphetamine
t~Test
ICSS Mean (±S.E.M.) responding in I0 min
468.3 ± 72.1
ll7 ± 89.7
t = 2.98, p < . 0 5
88.7 ± 24.0
165.7 ± 16.9
t = 2.55, p < . 0 5
7.7 ± 1.7
t = 1.33, p >.05
Forced-Swim Mean (±S.E.M.) duration of immobility in 5 min swim test Shuttle Escape Mean (±S.E.M.) escape latency (sec)
10.93 ± 2.1
Locomotor Activity Mean (±S.E.M.) locomotor activity (digitized voltage output) in 20 min 1,372 ± 237.8
1,718 ± 511.8
t =
.54, p >.05
185 ± 70
t =
.32, p >.05
Acoustic Startle Reflex Mean (±S.E.M.) movement ~igitized voltage output) during tone presentation (700 msec)
150 ± 45
A novel finding in this study deals with the effects of amphetamine withdrawal on swim-induced immobility. Specifically, mice exposed to long-term amphetamine treatment spent more time in a floating position during the forced swim test. It has been argued that the forced swim procedure is sensitive to manipulation that alter the motoric ability of the animal (12). It is likely, however, that the effects of amphetamine withdrawal on swim-induced immobility do not involve changes in the motoric capacity of the subjects. This is evident from the observation that drug withdrawal did not modify sensor±motor arousal as measured by the acoustic startle reflex and also had no effect on locomotor activity in an open field situation. Amphetamine withdrawal also did not modify escape performance in a shuttle escape task. In fact, although not significant mice had a tendency to be more active and showed faster escape latencies after withdrawal from amphetamine. These findings are in agreement with previous reports showing that after exposure to long-term amphetamine treatment animals were not less reactive to environmental and stressful stimuli (15, 18). To the contrary, exposure to chronic amphetamine treatment enhanced exploration and prolonged the extinction of well learned responses (19).
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Although acoustic startle, locomotor activity, and escape performance were not affected by drug withdrawal, these negative findings should be considered with a degree of caution. It is possible that changes in these behaviors may have become evident if a different schedule of drug administration had been employed, a higher dose of the drug administered or if the behaviors were assessed at varying intervals following withdrawal. Given these limitations it is not possible to suggest with a degree of certainty that these behaviors are not amenable to change during amphetamine withdrawal. However, the findings of this study indicate that both the ICSS test and the swiminduced immobility paradigm are more sensitive to the effects of drug withdrawal than the other behaviors, clearly ruling out a general motoric or arousal deficit as being responsible for the development of the post-amphetamine depression. Evaluation of stressor-induced behavioral changes, which have been used to model depression, revealed that inescapable shock not only retarded shuttle escape performance (1, 2, 20), but also disrupted responding for electrical brain stimulation from the nucleus accumbens and medial forebrain bundle (21) and interfered with swim escape performance (22, 23), as well as active responding in a forced-swim paradigm (24). Clearly cross generalization of the effects of inescapable shock may be evident in several different paradigms, although such effects may vary with the test and stressor parameters employed (24, 25). In view of the fact that long-term amphetamine treatment like inescapable stressors results in a depletion of norepinephrine (26), serotonin (27), and dopamine (28), it might have been expected that performance in the ICSS, swim and shuttle escape paradigms would be affected by amphetamine withdrawal much in the same way as they are by inescapable shock. The finding that amphetamine withdrawal did not have comparable effects on shuttle escape and forced-swim performance may be related to the demands placed on the organism in the two paradigms. For instance, in the shuttle test mice are exposed to discrete shock trials wherein an appropriate directed response results in stressor termination. In effect, the .increased reactivity engendered by drug withdrawal actually favors efficient performance. In the forced swim test animals are exposed to a single protracted swim ;trial, and hence the task may be better suited to assess disturbances of response maintenance that may stem from drug withdrawal. It will be noted as well that escape in the swim task is not possible, and increased reactivity is of no apparent benefit. In fact, increased floating in this paradigm can be interpreted as an efficient method of dealing with the stressor since it permits energy conservation. Whether these specific factors are responsible for the diverse effects seen after drug withdrawal in the two paradigms cannot be determined on the basis of the present results. However, these considerations reinforce the contention that these paradigms should not be used indiscriminately as animal models of affective disorders. Acknowledqements This research was supported by Grant A7042 from the Natural Sciences and Engineering Council of Canada to Larry Kokkinidis. Requests for reprints should be addressed to Larry Kokkinidis, Department of Psychology,~University of Saskatchewan, Saskatoon, Saskatchewan, Canada, S7N OWO. References I.
MAIER, S. F., and SELIGMAN, M. E. P. (1976).
J. Expt. Psychol. Gen. I05
3-46
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2. 3. 4. 5. 6. 7. 8. 9. lO. II. 12. 13. 14. 15. 16.
17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27.
28.
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ANISMAN, H., KOKKINIDIS, L., and SKLAR, L.S. In: Theory in psychopharmacology, Vol. l, S. Cooper (Ed.). Academic Press, London, pp. 65-IO2 (198]). PORSOLT,R.D., DENIEL, M., and JALFRE, M. Europ. J. Pharmacol. 5__7_7 43]-436 (]979). SHERMAN,A.D., and PETTY, F. Behav. Neural. Biol. 30 119-]34 (]980). KOKKINIDIS, L.,and ZACHARKO, R.M. Psychopharmacology 6__88 73-76 (]980). KOKKINIDIS, L.,and ZACHARKO, R.M. Behav. Neural. Biol. 29493-497 (]980). LEITH, N.J., and BARRETT, R.J. Psychopharmacologia 46 ]9----25 (1976). LEITH, N.J., and BARRETT, R.J. Psychopharmacology 7_2--9-]5 (1980). CASSENS,G., ACTON, C., KLING, M., and SCHILDKRAUT, J.J. Psychopharmacology 73 3]8-322 (]981). KOKKINIDIS, L., ZACHARKO, R.M., and PREDY, P.A. Pharmaco]. Biochem. Behav. 13379-383 (1980). WILLNER, P. Brain Res. Rev. 6 225-236 (1983). BROWNE,R.G. Europ. J. PharmaTol. 58331-334 (1979). LIEBMAN, J.M. Neurosci. Biobehav. Rev. ~ 45-47 (1983). ANTELMAN,S.M., and CHIODO, L.A. In: Stimulants: Neurochemical, behavioral and clinical perspectives, I. Creese (Ed.). Raven Press, New York, pp 269-299 (1983). KOKKINIDIS, t., and MacNEILL, E.P. Pharmacol. Biochem. Behav. I__77 413-417 (1982). KOKKINIDIS, L., and MacNEILL, E.P. Psychopharmacology 7__88 331-334 (1982). ROBERTSON,A., and MOGENSON, G.J. Psychopharmacology 65 149-154 (1979). KOKKINIDIS, L. Pharmacol. Biochem. Behav. 20 367-371--C1984). KOKKINIDIS, L. Pharmacol. Biochem. Behav. l-~ 593-598 (1983). WEISS, J.M., GLAZER, H.I., POHORECKY, L.A., BRICK, J., and MILLER, W.E. Psychosom. Med. 3_~_7522-534 (1975). ZACHARKO,R.M., BOWERS, W.J., KOKKINIDIS, L., and ANISMAN, H. Behav. Brain Res. 9 129-141 (1983). IRWIN, J., SUISSA, A., and ANISMAN, H. J. Expt. Psychol. Anim. Behav. Proc. 6 21-40 (1980). ALTENOR~-A., KAY, E., and RICHTER, M. Learn. Motiv. 8 54-6l (1977). PRINCE, C.R., and ANISMAN, H. Behav. Neural. Biol. 4~ 99-I19 (1984). O'NEILL, K.A., and Valentino, D. Europ. J. Pharmacol. 78379-380 (1982). SEGAL, D.S., WEINBERGER, S.B., CAHILL, J., and McCUNNE~, S.J. Science 2 0 7 9 0 4 - 9 0 7 (1980). TRULSON, M.E., and JACOBS, B.L., J. Pharmacol. Exp. Ther. 211 375-384 (1979). WAGNER,G.C., RICAUSTE, G.A., JOHANSON, C.E., SCHUSTER, C.R., and SEIDEN, L.S. Neurology 30547-550 (]980).
Pergamon Press
AMPHETAMINE WITHDRAWAL: A BEHAVIORAL EVALUATION Larry Kokkinidis, Robert M. Zacharko and Hymie Anisman Department of Psychology, University of Saskatchewan Saskatoon, Saskatchewan S7N OWO and Department of Psychology, Carleton University, Ottawa, Ontario KIS 5B6 Canada (Received in final form February 18, 1986) Summary The effects of withdrawal from long-term amphetamine treatment on intracranial self-stimulation, forced swiminduced immobility, shuttle escape performance, acoustic startle and locomotor activity were evaluated. Mice implanted with stimulating electrodes in the lateral hypothalamus demonstrated stable and reliable rates of self-stimulation responding. After exposure to a chronic schedule of amphetamine treatment response rates were severely depressed. In addition to modifying intracranial self-stimulation responding, amphetamine withdrawal increased the duration of immobility in a forced-swim situation. Although chronic amphetamine exposure induced pronounced behavioral changes in the intracranial self-stimulation and forced swim tasks, drug withdrawal had little effect on shuttle escape performance, acoustic startle and locomotor activity. Based on these findings it was suggested that the development of postamphetamine depression in the self-stimulation and forced swim paradigms was not related to variations in motoric or arousal mechanisms resulting from amphetamine withdrawal, but rather involved drug-induced changes in motivational processes. Several behavioral paradigms currently employed as animal models of clinical depression have proven to be valuable tools in evaluating the potenti~ therapeutic efficacy of antidepressant agents. One behavioral model of depression that has received considerable attention is the development of behavioral deficits in animals following exposure to inescapable shock (1). Specifically, animals exposed to inescapable shock with little or no control of their response to the stressor, developed performance deficits during subsequent escape testing (2). In a somewhat related behavioral situation animals forced to swim with no chance of escape often become immobile and assume a floating posture (3). Tricyclic antidepressant treatment has proven effective in mitigating the stress-induced deficits in escape responding (4), as well as the development of immobility in the forced-swim paradigm (3). Recent work involving the effects of long-term amphetamine administration on intracranial self-stimulation (ICSS) has shown that withdrawal from a chronic drug schedule of amphetamine treatment provoked a pronounced and sustained depression of rates of responding for electrical brain stimulation (5, 6, 7, 8). The available data suggests that the post-amphetamine depression 0024-3205/86 $3.00 + .00 Copyright (c) 1986 Pergamon Press Ltd.
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Post-amphetamine Depression
Vol. 38, No. 17, 1986
of ICSS is not related to a general lethargy induced by drug withdrawal that may exert a non-specific influence on ICSS performance, but rather involves an attenuation of the reinforcing effects of electrical brain stimulation. In accordance with this view it was shown that rats exhibited depressed rates of responding and an elevation of current threshold for several weeks after cessation of amphetamine treatment (8). Using a rate-independent measure of ICSS, Cassens and co-workers (9) found increased reinforcement thresholds after animals were exposed to chronic amphetamine treatment, and these authors suggested as have others, that the post-amphetamine depression of ICSS may have clinical implications when it is considered that anhedonia is characteristic of both amphetamine-induced and naturally occurring depression in humans (8, 9). Consistent with this position we found that chronic administration of the tricyclic antidepressants imipramine and amitriptyline, attenuated the postamphetamine depression of ICSS (lO). Although the ICSS paradigm, the shuttle escape task and the forced swim test have all been offered as potential models of depression and useful predictors of antidepressant drugs (for review see ll), these tasks most likely reflect different features of performance. Indeed, shuttle escape performance and the forced swim procedure appear to be sensitive to manipulations which affect the motoric capacity of the animals (2, 12). In contrast, the ICSS paradigm provides direct evaluation of changes in central reward processes associated with pharmacological treatment, although contribution of performance variables cannot be totally discounted (13). Since it is well established that withdrawal from amphetamine results in a pronounced depression of ICSS, it was of interest to determine whether a schedule of amphetamine administration that impaired ICSS responding would also modify shuttle escape performance and immobility in a forced-swim test. There is reason to believe that this might well be the case given the data showing that stressors and amphetamine are largely interchangeable with respect to their effects on behavior (14, 15). In addition to ICSS, shuttle escape, and swim-induced immobility, the effects of amphetamine withdrawal on locomotor activity and acoustic startle were assessed to determine directly whether amphetamine withdrawal modified arousal processes. Material and Methods Subjects Eighty male CD-I mice, 80 days of age obtained from the Canadian Breeding Laboratories (Quebec, Canada) served as subjects. Mice were housed individually in standard polypropylene cages and allowed free access to food and water. Behavioral testing was carried out during the light portion of the light/dark cycle. Apparatus The ICSS apparatus consisted of a black Plexiglas box 0.6 cm thick and 15.0 cm in width, length and height. A hole 1.5 cm in diameter was located in the centre of the Plexiglas floor of the apparatus. A photobeam unit was situated directly beneath the hold (0.5 cm) such that interruption of the photobeam by a head-poke resulted in electrical brain stimulation (0.3 sec in duration). Brain stimulation (300 uA base to peak) was delivered from an Ortec Dual Channel Stimulator and consisted of monophasic square wave pulses (lO0 Hz) with a pulse duration of O.l msec. Current was determined by monitoring on an oscilloscope the drop in voltage output across a lO K resistor placed in series with the stimulating electrode. LeHigh Valley timers and counters controlled the duration of the brain stimulation and recorded response
Vol. 38, No. 17, 1986
Post-amphetamine Depression
1619
rates. The shuttle boxes used for escape testing consisted of black Plexiglas and measured 26.4 x 9.0 x 15.5 cm. The two compartments of each of the six boxes were separated by a horizontally moveable stainless steel gate. The grid floor of each compartment consisted of stainless steel rods (0.32 cm) spaced l.O cm apart. Footshock (150 uA, AC) was delivered through a 3000 V source. Shuttle boxes were housed in sound attenuated chambers and were controlled by a microcomputer system. Acoustic startle and locomotor activity was recorded in two styrofoam (2.0 cm thick) circular chambers 28.0 cm in diameter and 21.0 cm high. The styrofoam floor of each chamber was covered with a plastic sheath and was positioned over an 8~W speaker (28.0 cm in diameter). Voltages produced by movements on the floor were fed to a Commodore PET Series 2001 computer. The analogue signal from the speaker was amplified and digitized by an 8 bit A/D converter. The digitized output was printed out on a Data Terminal Mart printer. In the case of acoustic startle, only activity during the tone presentation was measured. The 2700 Hz tone (700 msec in duration, 5-msec rise-fall time) was generated by a Piezo Crystal Audio Transistor (Projects Unlimited, Dayton, OH) situated in the centre of the styrofoam roof of each chamber. The intensity of the tone in the chamber was 97 dB and background noise in the chambers was 44 dB. Sound intensity measures were made with a Bruel Kjaer sound level meter (model 2203; A-scale). Procedure Intracranial Self-Stimulation Mice were anesthetized with sodium pentobarbital (65.0 mg/kg) and were implanted in the lateral hypothalamus with insulated bipolar electrodes (Plastic Products), which had 0.5 mm of the tips separated and scraped. Coordinates for electrode placements determined by preliminary histological analysis were: AP-O.5 mm from Bregma, L + 1.2 mm from midline suture and V 5.7 mm from a flat skull surface. Following a 7 day post-operative period mice were tested for ICSS for a 20 min session on 6 consecutive days. Only mice demonstrating reliable and stable rates of responding were used in the experiment. Mice showing low and irregular rates of responding were replaced. Once baseline rates had stabilized, mice were subdivided on the basis of average equalized baseline rates such that one-half of the animals (N = I0/ group) received 2 daily intraperitoneal (i.p.) injections (lO:O0 a.m. and 4:00 p.m.) of 5.0 mg/kg of d-amphetamine sulfate for lO consecutive days, while the remaining half were treated with an equivalent volume of saline. Twenty-four hours following the last injection of amphetamine mice were placed in the self-stimulation chamber and the number of head-poke responses during a lO min test session were recorded. Forced-swim Test Twenty naive mice were placed into 2 groups (N = lO/group) and injected with either 2 daily i.p. injections of d-amphetamine (5.0 mg/kg) or saline for IO consecutive days. The day after the last injection mice in each group were placed in a glass cylinder (18.5 cm in diameter and 25.0 cm high) filled to within 7.0 cm from the top with warm tap water (30 o C) for 15 min. Following the forced-swim session mice were towel dried and placed in a polypropylene mouse cage lined with paper towels for 30 min. Mice were then re-exposed to the water in the glass cylinder for 5 min. The duration of immobility (i.e., floating) during the 5 min test was recorded.
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Shuttle Escape Task Two groups of mice (N = lO/group) were chronically treated with either amphetamine or saline as previously described. Twenty-four hours after the last injection mice in both groups were exposed to 60 escapable shocks at 60 sec intervals. Shock was presented for 6 sec, after which the gate separating the two compartments opened permitting entry into th~ shock free compartment. The latency to escape into the safe compartment was recorded. If successful escape had not occurred within 24 sec of shock presentation then the trial was terminated and 60 sec later a new trial was initiated. Locomotor A c t i v i t y
and Acoustic Startle
Naive mice were randomly assigned to either the chronic amphetamine or saline conditions. Twenty-four hours after the lO days of 2 daily injections of either 5.0 mg/kg of d-amphetamine or saline mice in each group (N = lO/grou~ were placed into the activity chambers and locomotor activity was monitored for 20 min~. Mice tested for acoustic startle were treated in the same manner with the exception that they were exposed to 160 tone presentations with a I0 sec interval. This procedure has been shown to produce an accurate measure of acoustic startle arousal and changes in activity levels during the tone burst are representative of the startle reflex and not random activity levels (16). Results Following testing mice with electrode placements in the lateral hypothalamus received an overdose of sodium pentobarbital and were perfused with 0.9% saline followed by I0% formalin. Frozen coronal sections were cut at 40 uM and stained with thionin. The histological examination of the sections revealed that in all cases electrode placements were in the vicinity of the lateral hypothalamus. Withdrawal from long-term amphetamine treatment had pronounced effects on ICSS responding. As can be seen in Table l rates of responding 24 hours after withdrawal from amphetamine treatment were significantly depressed relative to ICSS rates of control animals. At the same time interval, withdrawal from amphetamine modified the duration of immobility during the forced-swim test. That is, mice treated with amphetamine spent significantly more time in a floating posture than did saline treated mice. In marked contrast to the effects of amphetamine withdrawal on ICSS and swim-induced immobility, withdrawal from amphetamine did not modify performame in a shuttle escape task. If anything, mice withdrawn from the drug were somewhat faster with respect to escape latencies relative to control animals. Similarly there were no effects of amphetamine withdrawal on the acoustic startle reflex or locomotor activity although consistent with the shuttle escape data mice withdrawn from the drug showed slightly higher rates of locomotor activity than did saline treated mice (see Table l). Discussion Mice exposed to a chronic schedule of amphetamine and then tested of ICSS during withdrawal showed depressed rates of responding for electrical brain stimulation. This observation was consistent with several reports demonstrating the development of post-amphetamine depression of ICSS after amphetamine withdrawal (5, 6, 7, 8, 17). Although this study did not assess the temporal parameters associated with the development of the ICSS depression, several reports have addressed this issue and have shown that the postamphetamine depression persists for several weeks after drug withdrawal (~I0~7~
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Table 1 Effects of Amphetamine Withdrawal on Intracranial Self-Stimulation, Responding, Forced-Swim Immobility, Shuttle Escape Performance, Locomotor Activity and Acoustic Startle Reflex
Saline
Amphetamine
t~Test
ICSS Mean (±S.E.M.) responding in I0 min
468.3 ± 72.1
ll7 ± 89.7
t = 2.98, p < . 0 5
88.7 ± 24.0
165.7 ± 16.9
t = 2.55, p < . 0 5
7.7 ± 1.7
t = 1.33, p >.05
Forced-Swim Mean (±S.E.M.) duration of immobility in 5 min swim test Shuttle Escape Mean (±S.E.M.) escape latency (sec)
10.93 ± 2.1
Locomotor Activity Mean (±S.E.M.) locomotor activity (digitized voltage output) in 20 min 1,372 ± 237.8
1,718 ± 511.8
t =
.54, p >.05
185 ± 70
t =
.32, p >.05
Acoustic Startle Reflex Mean (±S.E.M.) movement ~igitized voltage output) during tone presentation (700 msec)
150 ± 45
A novel finding in this study deals with the effects of amphetamine withdrawal on swim-induced immobility. Specifically, mice exposed to long-term amphetamine treatment spent more time in a floating position during the forced swim test. It has been argued that the forced swim procedure is sensitive to manipulation that alter the motoric ability of the animal (12). It is likely, however, that the effects of amphetamine withdrawal on swim-induced immobility do not involve changes in the motoric capacity of the subjects. This is evident from the observation that drug withdrawal did not modify sensor±motor arousal as measured by the acoustic startle reflex and also had no effect on locomotor activity in an open field situation. Amphetamine withdrawal also did not modify escape performance in a shuttle escape task. In fact, although not significant mice had a tendency to be more active and showed faster escape latencies after withdrawal from amphetamine. These findings are in agreement with previous reports showing that after exposure to long-term amphetamine treatment animals were not less reactive to environmental and stressful stimuli (15, 18). To the contrary, exposure to chronic amphetamine treatment enhanced exploration and prolonged the extinction of well learned responses (19).
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Vol. 38, No. 17, 1986
Although acoustic startle, locomotor activity, and escape performance were not affected by drug withdrawal, these negative findings should be considered with a degree of caution. It is possible that changes in these behaviors may have become evident if a different schedule of drug administration had been employed, a higher dose of the drug administered or if the behaviors were assessed at varying intervals following withdrawal. Given these limitations it is not possible to suggest with a degree of certainty that these behaviors are not amenable to change during amphetamine withdrawal. However, the findings of this study indicate that both the ICSS test and the swiminduced immobility paradigm are more sensitive to the effects of drug withdrawal than the other behaviors, clearly ruling out a general motoric or arousal deficit as being responsible for the development of the post-amphetamine depression. Evaluation of stressor-induced behavioral changes, which have been used to model depression, revealed that inescapable shock not only retarded shuttle escape performance (1, 2, 20), but also disrupted responding for electrical brain stimulation from the nucleus accumbens and medial forebrain bundle (21) and interfered with swim escape performance (22, 23), as well as active responding in a forced-swim paradigm (24). Clearly cross generalization of the effects of inescapable shock may be evident in several different paradigms, although such effects may vary with the test and stressor parameters employed (24, 25). In view of the fact that long-term amphetamine treatment like inescapable stressors results in a depletion of norepinephrine (26), serotonin (27), and dopamine (28), it might have been expected that performance in the ICSS, swim and shuttle escape paradigms would be affected by amphetamine withdrawal much in the same way as they are by inescapable shock. The finding that amphetamine withdrawal did not have comparable effects on shuttle escape and forced-swim performance may be related to the demands placed on the organism in the two paradigms. For instance, in the shuttle test mice are exposed to discrete shock trials wherein an appropriate directed response results in stressor termination. In effect, the .increased reactivity engendered by drug withdrawal actually favors efficient performance. In the forced swim test animals are exposed to a single protracted swim ;trial, and hence the task may be better suited to assess disturbances of response maintenance that may stem from drug withdrawal. It will be noted as well that escape in the swim task is not possible, and increased reactivity is of no apparent benefit. In fact, increased floating in this paradigm can be interpreted as an efficient method of dealing with the stressor since it permits energy conservation. Whether these specific factors are responsible for the diverse effects seen after drug withdrawal in the two paradigms cannot be determined on the basis of the present results. However, these considerations reinforce the contention that these paradigms should not be used indiscriminately as animal models of affective disorders. Acknowledqements This research was supported by Grant A7042 from the Natural Sciences and Engineering Council of Canada to Larry Kokkinidis. Requests for reprints should be addressed to Larry Kokkinidis, Department of Psychology,~University of Saskatchewan, Saskatoon, Saskatchewan, Canada, S7N OWO. References I.
MAIER, S. F., and SELIGMAN, M. E. P. (1976).
J. Expt. Psychol. Gen. I05
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2. 3. 4. 5. 6. 7. 8. 9. lO. II. 12. 13. 14. 15. 16.
17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27.
28.
Post-amphetamine Depression
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ANISMAN, H., KOKKINIDIS, L., and SKLAR, L.S. In: Theory in psychopharmacology, Vol. l, S. Cooper (Ed.). Academic Press, London, pp. 65-IO2 (198]). PORSOLT,R.D., DENIEL, M., and JALFRE, M. Europ. J. Pharmacol. 5__7_7 43]-436 (]979). SHERMAN,A.D., and PETTY, F. Behav. Neural. Biol. 30 119-]34 (]980). KOKKINIDIS, L.,and ZACHARKO, R.M. Psychopharmacology 6__88 73-76 (]980). KOKKINIDIS, L.,and ZACHARKO, R.M. Behav. Neural. Biol. 29493-497 (]980). LEITH, N.J., and BARRETT, R.J. Psychopharmacologia 46 ]9----25 (1976). LEITH, N.J., and BARRETT, R.J. Psychopharmacology 7_2--9-]5 (1980). CASSENS,G., ACTON, C., KLING, M., and SCHILDKRAUT, J.J. Psychopharmacology 73 3]8-322 (]981). KOKKINIDIS, L., ZACHARKO, R.M., and PREDY, P.A. Pharmaco]. Biochem. Behav. 13379-383 (1980). WILLNER, P. Brain Res. Rev. 6 225-236 (1983). BROWNE,R.G. Europ. J. PharmaTol. 58331-334 (1979). LIEBMAN, J.M. Neurosci. Biobehav. Rev. ~ 45-47 (1983). ANTELMAN,S.M., and CHIODO, L.A. In: Stimulants: Neurochemical, behavioral and clinical perspectives, I. Creese (Ed.). Raven Press, New York, pp 269-299 (1983). KOKKINIDIS, t., and MacNEILL, E.P. Pharmacol. Biochem. Behav. I__77 413-417 (1982). KOKKINIDIS, L., and MacNEILL, E.P. Psychopharmacology 7__88 331-334 (1982). ROBERTSON,A., and MOGENSON, G.J. Psychopharmacology 65 149-154 (1979). KOKKINIDIS, L. Pharmacol. Biochem. Behav. 20 367-371--C1984). KOKKINIDIS, L. Pharmacol. Biochem. Behav. l-~ 593-598 (1983). WEISS, J.M., GLAZER, H.I., POHORECKY, L.A., BRICK, J., and MILLER, W.E. Psychosom. Med. 3_~_7522-534 (1975). ZACHARKO,R.M., BOWERS, W.J., KOKKINIDIS, L., and ANISMAN, H. Behav. Brain Res. 9 129-141 (1983). IRWIN, J., SUISSA, A., and ANISMAN, H. J. Expt. Psychol. Anim. Behav. Proc. 6 21-40 (1980). ALTENOR~-A., KAY, E., and RICHTER, M. Learn. Motiv. 8 54-6l (1977). PRINCE, C.R., and ANISMAN, H. Behav. Neural. Biol. 4~ 99-I19 (1984). O'NEILL, K.A., and Valentino, D. Europ. J. Pharmacol. 78379-380 (1982). SEGAL, D.S., WEINBERGER, S.B., CAHILL, J., and McCUNNE~, S.J. Science 2 0 7 9 0 4 - 9 0 7 (1980). TRULSON, M.E., and JACOBS, B.L., J. Pharmacol. Exp. Ther. 211 375-384 (1979). WAGNER,G.C., RICAUSTE, G.A., JOHANSON, C.E., SCHUSTER, C.R., and SEIDEN, L.S. Neurology 30547-550 (]980).