BEHAVIORAL AND NEURAL BIOLOGY
53, 269-276 (1990)
Chronic Nicotine and Withdrawal Effects on Radial-Arm Maze Performance in Rats EDWARD D . LEVIN,* CHARLES L E E , t JED E . ROSE,* ANTONIO REYES,~ GAYLORD ELLISON,t MURRAY JARVIK,:~ AND ELLEN GRITZ§ '1
*Nicotine Research Laboratory 151, Veterans Administration Medical Cen~er, Durham, North Carblina 27705; tDepartment of Psychology, University of California, Los Angeles, California 90024; ~;Department of Psychiatry and Biobehavioral Science, University of California, Los Angeles, California 90024; and §Division of Cancer Control, Jonsson Comprehensive Cancer Center, Department of Surgery and Department of Psychiatry and Biobehavioral Science, University of California, Los Angeles, California 90024 Rats were tested for choice accuracy in an eight-arm radial maze during and after chronic administration of nicotine via subcutaneously implanted glass and Silastic capsules. Nicotine administration significantly improved choice accuracy relative to controls. The effect gradually became apparent over the first 2 weeks of exposure and persisted through the third week. Surprisingly, the significant facilitation of the nicotine-treated rats relative to controls continued for 2 weeks after the end of nicotine administration. No effects of nicotine were seen on choice latency or the strategy to make adjacent arm entries. © 1990AcademicPress, Inc.
Nicotine intake has been found to improve cognitive performance in humans (Wesnes & Warburton, 1983), while withdrawal from chronic nicotine has been found to impair it (Hughes & Hatsukami, 1986). Nicotine-induced improvements in cognitive function have also been found in experimental animals (Battig, 1970; Castellano, 1976; Clarke, 1987; Garg, 1969; Hunter, Zornetzer, Jarvik, & McGaugh, 1977). However, since most of these studies used acute nicotine administration, their relevance to the situation of chronic nicotine intake in humans is limited. There have been a few studies which examined chronic nicotine effects on cognitive behavior, but the results are mixed. One study found that chronic nicotine administration improved performance accuracy of rats on a go/no-go task after a few days of impaired responding (Nelsen, 1978). However, another study found that chronic nicotine administration J This study was supported by a grant from the MacArthur Foundation and NIDA Grant DA 02665 awarded to J.E.R. Reprint request should be addressed to Edward Levin. 269
0163-1047/90 $3.00 Copyright © 1990 by Academic Press, Inc. All rights of reproduction in any form reserved.
270
LEVIN ET AL.
impaired learning in a radial-arm maze (Mundy & Iwamoto, 1988). Nicotine withdrawal after chronic administration has been found to impair the performance accuracy of rats in the Sidman avoidance test (Hall & Morrison, 1973). However, this effect was not seen with a less stressful test, indicating that the withdrawal-induced impairment may have been due to increased perception of stress seen after nicotine withdrawal (Hughes & Hatsukami, 1986). The present study was conducted to determine whether chronic nicotine administration would improve choice accuracy in a nonstressful cognitive task, the radial-arm maze test. The radial-arm maze has been found to be sensitive to a wide range of cholinergic manipulations (Levin, 1988), including the adverse effects of the nicotinic antagonist mecamylamine (Levin, Castonguay, & Ellison, 1987). Unlike a previous study of chronic nicotine effects on radial-arm maze performance (Mundy & Iwamoto, 1988) which looked at the effects of daily injections, in the present study, nicotine was administered on a continuous basis via a subcutaneously implanted pellet. In addition, after 3 weeks of administration, withdrawal was instituted to determine if, as with humans, nicotine withdrawal would result in cognitive impairment.
METHODS This experiment was run twice to ensure that the effects were reliable. The same effects of nicotine were seen in both parts of the study. The data from them was combined for analysis with replication as a factor. Except where noted the methods for the two parts were the same. Subjects. Female Sprague-Dawley albino rats were used in this study. Nineteen rats (Simonsen, Gilroy, CA) were used in the first part of the experiment and eighteen rats (UCLA Psychology Department breeding colony) were used in the replication. There were 9 controls and l0 nicotine-treated rats in the first part of the experiment and 9 controls and 9 nicotine-treated rats in the replication. Throughout the experiment, the rats were housed singly and were kept on a 12:12 reverse light-dark cycle. All testing occurred during the dark phase. Apparatus and procedure. The rats were tested on eight-arm mazes patterned after the one developed by Olton and Samuelson (1976). In the first part of the experiment, the maze was made of black painted wood. The center platform was 35 cm across and each of the arms was 10 cm wide and 80 cm long with foodwells located 2 cm from the end. The maze was elevated 30 cm from the floor. In the replication, the maze was the same configuration with different dimensions. This maze was made of white laminated wood. The center platform was 87 cm across and each of the arms was 10 cm wide and 69 cm long with foodwells located 2 cm from the end. The maze was elevated 50 cm
NICOTINE AND RADIAL-ARM MAZE
271
from the floor. Throughout the study testing was conducted in a room with abundant extramaze visual cues under dim illumination. Before each session the maze was wiped with water and one-third to one-half of a piece of sugar-coated cereal (Kellogg's Froot Loops) was placed in the foodwell of each arm. At the beginning of the session, the rat was gently placed inside an opaque Plexiglas ring located on the center platform. After l0 s, the ring was lifted and the rat was flee to roam about the maze. The session lasted until the rat had entered (four paws beyond the threshold) all eight arms or 5 rain had elapsed. Prior to the onset of drug administration the rats underwent at least 20 sessions of training so that they were performing at asymptotic levels during the experiment. Before the start of nicotine administration all of the rats had had at least five sessions with seven or eight correct choices in the first eight arm entries. After acquisition, the rats were tested on the maze twice per week (Tuesday and Friday). They were kept on a restricted diet 5 days per week (Monday-Friday) when they were fed daily in the afternoon. On weekends the rats had ad lib access to food. Choice accuracy was measured by the number of different arms entered before an error was made (entries to repeat). Rats that performed perfectly (i.e., entered all eight arms without making an error) were given scores of eight entries to repeat since any further choices would have inevitably been re-entries. Locomotor speed in the maze was measured by dividing the total time of each session by the number of arms entered (seconds per entry). The maximum seconds per session was 300 s. For sessions when the rat did not enter enough arms for the calculation of the choice accuracy measure, the mean scores for the rest of the rats in the same treatment group were used for statistical analysis. Drug administration. Nicotine was chronically administered by a subcutaneously implanted glass and Silastic pellet (22 mm x 7 ram) filled with 98-100% pure nicotine free base (Sigma, St. Louis, MO). This pellet was originally developed by Erickson, Stavchansky, Koch, and McGinty (1982). It has been found by Erickson et al. (1982) and verified in our laboratory (Morgan & Ellison, 1987) to deliver approximately 3.4 mg of nicotine per day through a Silastic plug at one end. This rate of delivery results in steady-state nicotine blood levels of 400-500 ng/ml (Erickson et al., 1982). The nicotine pellets were implanted in a subcutaneous pocket. The control rats were implanted with identical capsules without nicotine. The pellets were removed 3 weeks after implantation. Rats were randomly assigned to treatment groups with the stipulation that the groups would be balanced for pretreatment entries to repeat scores. Statistics. The choice measure, entries to repeat, and the latency measure, seconds per entry, were evaluated by two-way analyses of variance with nicotine treatment and replication as between-subjects factors and
272
L E V I N ET AL.
weeks as a repeated measure. Significant interactions were followed-up by tests of simple main effects.
RESULTS With body weight there was a significant Nicotine × Weeks interaction (p < .005). Follow-up tests of the simple main effects of nicotine at each week showed a gradual decline in weight followed by a recovery after withdrawal. As shown in Table 1, the nicotine-induced weight loss was marginally significant by the first week of administration (p < .07) and was significant during the second and third weeks of administration (19 < .01). The average weight of the nicotine-treated rats reached a minimum of 239 g during the second week of administration. After nicotine withdrawal, the treated rats regained weight but remained significantly lighter than controls during the first week (p < .025). By the second week there was no longer a significant nicotine-related weight deficit. Nicotine administration caused an improvement in choice accuracy. There was a significant overall effect of nicotine improving entries to repeat scores relative to controls (F(1, 33) = 8.37, p < .01). Significant nicotine-related improvements were also seen when the period during (F(1, 33) = 6.28, p < .025) and after (F(1, 33) = 7.24, p < .025) nicotine administration was separately analyzed. As shown in Fig. 1, the controls showed a slight dip in performance the first 2 weeks after surgery and then returned to pretest levels of performance. On the other hand, the nicotine-treated rats did not show any signs of a dip, but rather after the first week postsurgery showed an improvement which lasted for the rest of nicotine administration and for the 2 weeks after withdrawal. On two occasions rats did not enter enough arms to calculate an entries to repeat score. Analysis of the data excluding these rats did not alter the conclusions. The nicotine-treated group still showed significantly TABLE 1 Weekly Body Weights in Grams (Means ± Standard Error of the Means) during and after Chronic Nicotine Administration Weeks Nicotine
Control Nicotine
Postnicotine
Pre
1
2
3
1
2
269 -+ 7 268 ± 6
258 _+ 6 250 + 6*
261 -+ 6 239 ± 6***
262 ± 5 244 _+ 5***
261 -+ 6 250 ± 5**
260 ± 5 257 ± 4
* p < .07. ** p < .025. *** p < .01.
273
NICOTINE AND RADIAL-ARM MAZE 8.0-
7.57.0no
6.56.0.
W
Control
5.5.
Nicotine 5.0
i
I
i
1
2
1
P re
I
I
2 3 Nicotine
I
i
1
2 Post
F]c. l. Weekly averages for entries to repeat (mean _+ standard error of the mean) before, during and after chronic nicotine administration.
better overall choice accuracy (F(1, 31) = 17.45, p < .001), consisting of significantly better performance both during F(I, 31) = 13.88, p < .005) and after withdrawal (F(1, 31) = 11.64, p < .005). Response latency was not significantly changed by nicotine. Despite its anorectic effects, nicotine treatment did not decrease the number of food reinforcements eaten.
DISCUSSION There were three important findings in the present study. First, chronic nicotine administration was found to improve performance on an appetitively motivated spatial memory test. Second, the appearance of this improvement was delayed until the second week of nicotine administration. Finally, and most surprisingly, this nicotine-induced facilitation persisted after the withdrawal of nicotine administration. The cognitive nature of the nicotine-induced facilitation in choice accuracy is supported by its appearance on an appetitively motivated task. If motivational effects were predominant, the anorectic effect of nicotine should have impaired choice accuracy. The nicotine-treated rats in the present study were not noted to eat fewer of the food rewards. This may have been due to the fact that all of the rats were food deprived for 23 h prior to testing. Another possible "noncognitive" explanation of the nicotine-induced facilitation lies with the stress-reducing characteristics of nicotine. Hall and Morrison (1973) found that withdrawal from chronic nicotine administration impaired performance accuracy in the Sidman avoidance test, but not in a less stressful task. Using an appetitive task, as in the present study, rather than a shock avoidance task substantially reduces the stress associated with testing. There still may be some lower level of stress associated with the testing procedure that is alleviated by nicotine. However, the extensive familiarization of the rats with the task
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LEVIN ET AL.
and testing procedure before drug administration should have tended to reduce such stress. We have previously found that these nicotine pellets are effective in producing a moderate degree of anorexia and weight loss (Levin, Ellison, Salem, Jarvik, & Gritz, 1988; Levin, Morgan, Galvez, & Ellison, 1987; Morgan & Ellison, 1987). The weight loss progressed steadily over the first 9 days to a maximum deficit of about 40 g, after which it gradually abated until the nicotine-treated rats were not significantly different from controls 23 days after the implantation of nicotine pellets (Levin et al., 1987b). The nicotine-induce weight loss in the current study was slightly different in time course, probably because of the feeding restrictions imposed on the rats. The nicotine-induced improvement in choice accuracy was delayed in its appearance in that there was no improvement in choice accuracy during the first week of nicotine administration. A nicotine-induced improvement may have been masked during the first week of exposure because of the hypophagia seen during this period (Levin et al., 1987b). This hypophagia is greatly attenuated by the second week of nicotine administration. However, this explanation is not supported by the observation that the nicotine-treated rats ate their reinforcements just as often during the first week as during subsequent weeks. Another possibility for the delayed effect of chronic nicotine is that it secondarily improves performance by some other neural or behavioral mechanism. This delayed improvement is similar to the finding of Nelson (1978) that chronic nicotine administration improved performance on a go/no-go task after a few days of impaired performance. Chronic administration may be necessary for the cognitive enhancing effects of nicotine in humans as well. It is possible that the nicotine-induced facilitation in choice accuracy was due to the increased use of a response strategy. The simplest strategy to solve the radial-arm maze is to continually make responses to the arm adjacent to one side of the one just chosen. In the current study, the percentage of adjacent arm entries was about 8% higher in the nicotinetreated than control rats during and after administration, but this difference was not even close to being significant (p > .25). Thus, it appears that the effects on choice accuracy were not likely to have been solely the result of differential imposition of the strategy of stringing together adjacent arm entries. However, this analysis does not rule out the development of more complex strategies. The present results provide an interesting extension of those reported by Mundy and Iwamoto (1988). They found that repeated acute nicotine administration (0.45 mg/kg) impaired acquisition of performance in the radial-arm maze, but that in rats who were already performing at asymptotic levels this dose did not cause a significant effect. One major
NICOTINE AND RADIAL-ARM MAZE
275
difference between their study and the current one that may have accounted for the difference in results is that they administered repeated acute doses of nicotine as opposed to continuous administration. Another important dosing factor may have been length of administration. Even with continuous administration in the current study a nicotine-induced facilitation did not become pronounced until the second week of administration. Mundy and Iwamoto (1988) only tested rats treated daily with nicotine for 8 days during acquisition. In their second experiment testing for nicotine effects on asympototic performance, rats were administered nicotine only every fourth day for 16 days. After withdrawal of nicotine in the present study there was no appearance of impairment. In fact, the improved performance accuracy persisted for the 2 weeks of testing after withdrawal. If chronic nicotine administration resulted in a down-regulation of nicotinic receptors one would expect a choice impairment similar to that which has been seen after acute administration of the nicotinic receptor blocker, mecamylamine (Levin et al., 1987a). However, chronic nicotine administration has been found to increase nicotinic receptor binding (Ksir, Hakan, Hall, & Kellar, 1985; Marks, Butch, & Collins, 1983; Schwartz & Kellar, 1983, 1985). Although this increase in receptor binding has been associated with tolerance in a number of behavioral functions including exploratory behavior, body temperature, and heart rate (Marks, Stizel, & Collins, 1985), it is possible that this increase in the number of receptors could also be associated with sensitization to nicotine effects. Marks et al. (1985) found that the increased nicotine receptor binding returned to control levels by 8 days after withdrawal. Behavioral tolerance, however, took varying periods of time to return to normal and with the case of heart rate, tolerance persisted for the entire 20-day withdrawal period. In the present study, the improvement in choice accuracy which began during the second week of nicotine administration persisted at least 2 weeks after withdrawal. This extension of behavioral effects after nicotine withdrawal and after receptor induction has probably returned to normal indicates that other processes induced by chronic nicotine administration must be responsible. One likely candidate is learning. It is possible that chronic nicotine administration caused the rats to undergo an additional period of learning. REFERENCES Battig, K. (1970). The effect of pre- and post-trial application of nicotine on the 12 problems of the Hebb-Williams Test in the rat. Psychopharmacologia (Berlin), 18, 68-76. Castellano, C. (1976). Effects of nicotine on discrimination learning, consolidation and learned behavior in two inbred strains of mice. Psychopharmacology (Berlin), 48, 3743. Clarke, P. B. S. (1987). Nicotine and smoking: A perspective from animal studies. Psychopharmacology (Berlin), 92, 135-143.
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Erickson, C. K., Stavchansky, S. A., Koch, K. I., & McGinty, J. W. (1982). A new subcutaneously-implantable reservoir for sustained release of nicotine in the rat. Pharmacology, Biochemistry & Behavior, 17, 183-185. Garg, M. (1969). The effects of nicotine on two different types of learning. Psychopharmacologia (Berlin), 15, 408-414. Hall, G. H., & Morrison, C. F. (1973). New evidence for a relationship between tobacco smoking, nicotine dependence and stress. Nature (London), 243, 199-201. Hughes, J. RI, & Hatsukami, D. (1986). Signs and symptoms of tobacco withdrawal. Archives of General Psychiatry, 43, 289-294. Hunter, B., Zornetzer, S. F., Jarvik, M. E., & McGaugh, J. L. (1977). Modulation of learning and memory: Effects of drugs influencing neurotransmitters. In L. L. Iverson, S. D. Iverson, & S. H. Snyder (Eds.), Handbook ofpsychopharmacology (Vol. 8, pp. 531-577). New York: Plenum. Ksir, C., Hakan, R., Hall, D. P., & Kellar, K. J. (1985). Exposure to nicotine enhances the behavioral stimulant effect of nicotine and increases binding of 3-H acetylcholine to nicotinic receptors. Neuropharmacology, 24, 527-531. Levin, E. D. (1988). Psychopharmacological effects in the radial-arm maze. Neuroscience & Biobehavioral Reviews, 12, 169-175. Levin, E. D., Castonguay, M., & Ellison, G. D. (1987a). Effects of the nicotinic receptor blocker, mecamylamine, on radial-arm maze performance in rats. Behavioral & Neural Biology, 48, 206-212. Levin, E. D., Ellison, G. D., Salem, C., Jarvik, M., & Gritz, E. (1988). Behavioral effects of acute hexamethonium in rats chronically intoxicated with nicotine. Physiology & Behavior, 44, 355-359. Levin, E. D., Morgan, M. M., Galvez, C., & Ellison, G. D. (1987b). Chronic nicotine and withdrawal effects on body weight and food and water consumption in female rats. Physiology & Behavior, 39, 441-444. Marks, M. J., Burch, J. B., & Collins, A. C. (1983). Effects of chronic nicotine infusion on tolerance development and nicotine receptors. Journal of Pharmacology and Experimental Therapeutics, 226, 817-825. Marks, M. J., Stitzel, J. A., & Collins, A. C. (1985). Time course study of the effects of chronic nicotine infusion on drug response and brain receptors. Journal of Pharmacology and Experimental Therapeutics, 235, 619-628. Morgan, M. M., & Ellison, G. D. (1987). Development of tolerance to nicotine effects in rats. Psychopharmacology (Berlin), 91, 236-238. Mundy, W. R., & Iwamoto, E. T. (1988). Nicotine acquisition of radial maze performance in rats. Pharmacology, Biochemistry & Behavior, 30, 119-122. Nelsen, J. M. (1978). Psychobiological consequences of chronic nicotinization: A focus on arousal. In K. Battig (Ed.), Behavioral effects of nicotine (pp. 1-17). Basel: Karger. Olton, D. S., & Samuelson, R. J. (1976). Rememberance of places passed: Spatial memory in rats. Journal of Experimental Psychology: Animal Behavior Processes, 2, 97-115. Schwartz, R. D., & Kellar, K. J. (1983). Nicotinic cholinergic binding sites in brain: In vivo regulation. Science, 220, 214-216. Schwartz, R. D., & Kellar, K. J. (1985). In vivo regulation of [3-H]acetylcholine recognition sites in brain by nicotinic cholinergic drugs. Journal of Neurochemistry, 45, 427-433. Wesnes, K., & Warburton, D. M. (1983). Smoking, nicotine and human performance. Pharmacology and Therapeutics, 21, 189-208.
53, 269-276 (1990)
Chronic Nicotine and Withdrawal Effects on Radial-Arm Maze Performance in Rats EDWARD D . LEVIN,* CHARLES L E E , t JED E . ROSE,* ANTONIO REYES,~ GAYLORD ELLISON,t MURRAY JARVIK,:~ AND ELLEN GRITZ§ '1
*Nicotine Research Laboratory 151, Veterans Administration Medical Cen~er, Durham, North Carblina 27705; tDepartment of Psychology, University of California, Los Angeles, California 90024; ~;Department of Psychiatry and Biobehavioral Science, University of California, Los Angeles, California 90024; and §Division of Cancer Control, Jonsson Comprehensive Cancer Center, Department of Surgery and Department of Psychiatry and Biobehavioral Science, University of California, Los Angeles, California 90024 Rats were tested for choice accuracy in an eight-arm radial maze during and after chronic administration of nicotine via subcutaneously implanted glass and Silastic capsules. Nicotine administration significantly improved choice accuracy relative to controls. The effect gradually became apparent over the first 2 weeks of exposure and persisted through the third week. Surprisingly, the significant facilitation of the nicotine-treated rats relative to controls continued for 2 weeks after the end of nicotine administration. No effects of nicotine were seen on choice latency or the strategy to make adjacent arm entries. © 1990AcademicPress, Inc.
Nicotine intake has been found to improve cognitive performance in humans (Wesnes & Warburton, 1983), while withdrawal from chronic nicotine has been found to impair it (Hughes & Hatsukami, 1986). Nicotine-induced improvements in cognitive function have also been found in experimental animals (Battig, 1970; Castellano, 1976; Clarke, 1987; Garg, 1969; Hunter, Zornetzer, Jarvik, & McGaugh, 1977). However, since most of these studies used acute nicotine administration, their relevance to the situation of chronic nicotine intake in humans is limited. There have been a few studies which examined chronic nicotine effects on cognitive behavior, but the results are mixed. One study found that chronic nicotine administration improved performance accuracy of rats on a go/no-go task after a few days of impaired responding (Nelsen, 1978). However, another study found that chronic nicotine administration J This study was supported by a grant from the MacArthur Foundation and NIDA Grant DA 02665 awarded to J.E.R. Reprint request should be addressed to Edward Levin. 269
0163-1047/90 $3.00 Copyright © 1990 by Academic Press, Inc. All rights of reproduction in any form reserved.
270
LEVIN ET AL.
impaired learning in a radial-arm maze (Mundy & Iwamoto, 1988). Nicotine withdrawal after chronic administration has been found to impair the performance accuracy of rats in the Sidman avoidance test (Hall & Morrison, 1973). However, this effect was not seen with a less stressful test, indicating that the withdrawal-induced impairment may have been due to increased perception of stress seen after nicotine withdrawal (Hughes & Hatsukami, 1986). The present study was conducted to determine whether chronic nicotine administration would improve choice accuracy in a nonstressful cognitive task, the radial-arm maze test. The radial-arm maze has been found to be sensitive to a wide range of cholinergic manipulations (Levin, 1988), including the adverse effects of the nicotinic antagonist mecamylamine (Levin, Castonguay, & Ellison, 1987). Unlike a previous study of chronic nicotine effects on radial-arm maze performance (Mundy & Iwamoto, 1988) which looked at the effects of daily injections, in the present study, nicotine was administered on a continuous basis via a subcutaneously implanted pellet. In addition, after 3 weeks of administration, withdrawal was instituted to determine if, as with humans, nicotine withdrawal would result in cognitive impairment.
METHODS This experiment was run twice to ensure that the effects were reliable. The same effects of nicotine were seen in both parts of the study. The data from them was combined for analysis with replication as a factor. Except where noted the methods for the two parts were the same. Subjects. Female Sprague-Dawley albino rats were used in this study. Nineteen rats (Simonsen, Gilroy, CA) were used in the first part of the experiment and eighteen rats (UCLA Psychology Department breeding colony) were used in the replication. There were 9 controls and l0 nicotine-treated rats in the first part of the experiment and 9 controls and 9 nicotine-treated rats in the replication. Throughout the experiment, the rats were housed singly and were kept on a 12:12 reverse light-dark cycle. All testing occurred during the dark phase. Apparatus and procedure. The rats were tested on eight-arm mazes patterned after the one developed by Olton and Samuelson (1976). In the first part of the experiment, the maze was made of black painted wood. The center platform was 35 cm across and each of the arms was 10 cm wide and 80 cm long with foodwells located 2 cm from the end. The maze was elevated 30 cm from the floor. In the replication, the maze was the same configuration with different dimensions. This maze was made of white laminated wood. The center platform was 87 cm across and each of the arms was 10 cm wide and 69 cm long with foodwells located 2 cm from the end. The maze was elevated 50 cm
NICOTINE AND RADIAL-ARM MAZE
271
from the floor. Throughout the study testing was conducted in a room with abundant extramaze visual cues under dim illumination. Before each session the maze was wiped with water and one-third to one-half of a piece of sugar-coated cereal (Kellogg's Froot Loops) was placed in the foodwell of each arm. At the beginning of the session, the rat was gently placed inside an opaque Plexiglas ring located on the center platform. After l0 s, the ring was lifted and the rat was flee to roam about the maze. The session lasted until the rat had entered (four paws beyond the threshold) all eight arms or 5 rain had elapsed. Prior to the onset of drug administration the rats underwent at least 20 sessions of training so that they were performing at asymptotic levels during the experiment. Before the start of nicotine administration all of the rats had had at least five sessions with seven or eight correct choices in the first eight arm entries. After acquisition, the rats were tested on the maze twice per week (Tuesday and Friday). They were kept on a restricted diet 5 days per week (Monday-Friday) when they were fed daily in the afternoon. On weekends the rats had ad lib access to food. Choice accuracy was measured by the number of different arms entered before an error was made (entries to repeat). Rats that performed perfectly (i.e., entered all eight arms without making an error) were given scores of eight entries to repeat since any further choices would have inevitably been re-entries. Locomotor speed in the maze was measured by dividing the total time of each session by the number of arms entered (seconds per entry). The maximum seconds per session was 300 s. For sessions when the rat did not enter enough arms for the calculation of the choice accuracy measure, the mean scores for the rest of the rats in the same treatment group were used for statistical analysis. Drug administration. Nicotine was chronically administered by a subcutaneously implanted glass and Silastic pellet (22 mm x 7 ram) filled with 98-100% pure nicotine free base (Sigma, St. Louis, MO). This pellet was originally developed by Erickson, Stavchansky, Koch, and McGinty (1982). It has been found by Erickson et al. (1982) and verified in our laboratory (Morgan & Ellison, 1987) to deliver approximately 3.4 mg of nicotine per day through a Silastic plug at one end. This rate of delivery results in steady-state nicotine blood levels of 400-500 ng/ml (Erickson et al., 1982). The nicotine pellets were implanted in a subcutaneous pocket. The control rats were implanted with identical capsules without nicotine. The pellets were removed 3 weeks after implantation. Rats were randomly assigned to treatment groups with the stipulation that the groups would be balanced for pretreatment entries to repeat scores. Statistics. The choice measure, entries to repeat, and the latency measure, seconds per entry, were evaluated by two-way analyses of variance with nicotine treatment and replication as between-subjects factors and
272
L E V I N ET AL.
weeks as a repeated measure. Significant interactions were followed-up by tests of simple main effects.
RESULTS With body weight there was a significant Nicotine × Weeks interaction (p < .005). Follow-up tests of the simple main effects of nicotine at each week showed a gradual decline in weight followed by a recovery after withdrawal. As shown in Table 1, the nicotine-induced weight loss was marginally significant by the first week of administration (p < .07) and was significant during the second and third weeks of administration (19 < .01). The average weight of the nicotine-treated rats reached a minimum of 239 g during the second week of administration. After nicotine withdrawal, the treated rats regained weight but remained significantly lighter than controls during the first week (p < .025). By the second week there was no longer a significant nicotine-related weight deficit. Nicotine administration caused an improvement in choice accuracy. There was a significant overall effect of nicotine improving entries to repeat scores relative to controls (F(1, 33) = 8.37, p < .01). Significant nicotine-related improvements were also seen when the period during (F(1, 33) = 6.28, p < .025) and after (F(1, 33) = 7.24, p < .025) nicotine administration was separately analyzed. As shown in Fig. 1, the controls showed a slight dip in performance the first 2 weeks after surgery and then returned to pretest levels of performance. On the other hand, the nicotine-treated rats did not show any signs of a dip, but rather after the first week postsurgery showed an improvement which lasted for the rest of nicotine administration and for the 2 weeks after withdrawal. On two occasions rats did not enter enough arms to calculate an entries to repeat score. Analysis of the data excluding these rats did not alter the conclusions. The nicotine-treated group still showed significantly TABLE 1 Weekly Body Weights in Grams (Means ± Standard Error of the Means) during and after Chronic Nicotine Administration Weeks Nicotine
Control Nicotine
Postnicotine
Pre
1
2
3
1
2
269 -+ 7 268 ± 6
258 _+ 6 250 + 6*
261 -+ 6 239 ± 6***
262 ± 5 244 _+ 5***
261 -+ 6 250 ± 5**
260 ± 5 257 ± 4
* p < .07. ** p < .025. *** p < .01.
273
NICOTINE AND RADIAL-ARM MAZE 8.0-
7.57.0no
6.56.0.
W
Control
5.5.
Nicotine 5.0
i
I
i
1
2
1
P re
I
I
2 3 Nicotine
I
i
1
2 Post
F]c. l. Weekly averages for entries to repeat (mean _+ standard error of the mean) before, during and after chronic nicotine administration.
better overall choice accuracy (F(1, 31) = 17.45, p < .001), consisting of significantly better performance both during F(I, 31) = 13.88, p < .005) and after withdrawal (F(1, 31) = 11.64, p < .005). Response latency was not significantly changed by nicotine. Despite its anorectic effects, nicotine treatment did not decrease the number of food reinforcements eaten.
DISCUSSION There were three important findings in the present study. First, chronic nicotine administration was found to improve performance on an appetitively motivated spatial memory test. Second, the appearance of this improvement was delayed until the second week of nicotine administration. Finally, and most surprisingly, this nicotine-induced facilitation persisted after the withdrawal of nicotine administration. The cognitive nature of the nicotine-induced facilitation in choice accuracy is supported by its appearance on an appetitively motivated task. If motivational effects were predominant, the anorectic effect of nicotine should have impaired choice accuracy. The nicotine-treated rats in the present study were not noted to eat fewer of the food rewards. This may have been due to the fact that all of the rats were food deprived for 23 h prior to testing. Another possible "noncognitive" explanation of the nicotine-induced facilitation lies with the stress-reducing characteristics of nicotine. Hall and Morrison (1973) found that withdrawal from chronic nicotine administration impaired performance accuracy in the Sidman avoidance test, but not in a less stressful task. Using an appetitive task, as in the present study, rather than a shock avoidance task substantially reduces the stress associated with testing. There still may be some lower level of stress associated with the testing procedure that is alleviated by nicotine. However, the extensive familiarization of the rats with the task
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and testing procedure before drug administration should have tended to reduce such stress. We have previously found that these nicotine pellets are effective in producing a moderate degree of anorexia and weight loss (Levin, Ellison, Salem, Jarvik, & Gritz, 1988; Levin, Morgan, Galvez, & Ellison, 1987; Morgan & Ellison, 1987). The weight loss progressed steadily over the first 9 days to a maximum deficit of about 40 g, after which it gradually abated until the nicotine-treated rats were not significantly different from controls 23 days after the implantation of nicotine pellets (Levin et al., 1987b). The nicotine-induce weight loss in the current study was slightly different in time course, probably because of the feeding restrictions imposed on the rats. The nicotine-induced improvement in choice accuracy was delayed in its appearance in that there was no improvement in choice accuracy during the first week of nicotine administration. A nicotine-induced improvement may have been masked during the first week of exposure because of the hypophagia seen during this period (Levin et al., 1987b). This hypophagia is greatly attenuated by the second week of nicotine administration. However, this explanation is not supported by the observation that the nicotine-treated rats ate their reinforcements just as often during the first week as during subsequent weeks. Another possibility for the delayed effect of chronic nicotine is that it secondarily improves performance by some other neural or behavioral mechanism. This delayed improvement is similar to the finding of Nelson (1978) that chronic nicotine administration improved performance on a go/no-go task after a few days of impaired performance. Chronic administration may be necessary for the cognitive enhancing effects of nicotine in humans as well. It is possible that the nicotine-induced facilitation in choice accuracy was due to the increased use of a response strategy. The simplest strategy to solve the radial-arm maze is to continually make responses to the arm adjacent to one side of the one just chosen. In the current study, the percentage of adjacent arm entries was about 8% higher in the nicotinetreated than control rats during and after administration, but this difference was not even close to being significant (p > .25). Thus, it appears that the effects on choice accuracy were not likely to have been solely the result of differential imposition of the strategy of stringing together adjacent arm entries. However, this analysis does not rule out the development of more complex strategies. The present results provide an interesting extension of those reported by Mundy and Iwamoto (1988). They found that repeated acute nicotine administration (0.45 mg/kg) impaired acquisition of performance in the radial-arm maze, but that in rats who were already performing at asymptotic levels this dose did not cause a significant effect. One major
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difference between their study and the current one that may have accounted for the difference in results is that they administered repeated acute doses of nicotine as opposed to continuous administration. Another important dosing factor may have been length of administration. Even with continuous administration in the current study a nicotine-induced facilitation did not become pronounced until the second week of administration. Mundy and Iwamoto (1988) only tested rats treated daily with nicotine for 8 days during acquisition. In their second experiment testing for nicotine effects on asympototic performance, rats were administered nicotine only every fourth day for 16 days. After withdrawal of nicotine in the present study there was no appearance of impairment. In fact, the improved performance accuracy persisted for the 2 weeks of testing after withdrawal. If chronic nicotine administration resulted in a down-regulation of nicotinic receptors one would expect a choice impairment similar to that which has been seen after acute administration of the nicotinic receptor blocker, mecamylamine (Levin et al., 1987a). However, chronic nicotine administration has been found to increase nicotinic receptor binding (Ksir, Hakan, Hall, & Kellar, 1985; Marks, Butch, & Collins, 1983; Schwartz & Kellar, 1983, 1985). Although this increase in receptor binding has been associated with tolerance in a number of behavioral functions including exploratory behavior, body temperature, and heart rate (Marks, Stizel, & Collins, 1985), it is possible that this increase in the number of receptors could also be associated with sensitization to nicotine effects. Marks et al. (1985) found that the increased nicotine receptor binding returned to control levels by 8 days after withdrawal. Behavioral tolerance, however, took varying periods of time to return to normal and with the case of heart rate, tolerance persisted for the entire 20-day withdrawal period. In the present study, the improvement in choice accuracy which began during the second week of nicotine administration persisted at least 2 weeks after withdrawal. This extension of behavioral effects after nicotine withdrawal and after receptor induction has probably returned to normal indicates that other processes induced by chronic nicotine administration must be responsible. One likely candidate is learning. It is possible that chronic nicotine administration caused the rats to undergo an additional period of learning. REFERENCES Battig, K. (1970). The effect of pre- and post-trial application of nicotine on the 12 problems of the Hebb-Williams Test in the rat. Psychopharmacologia (Berlin), 18, 68-76. Castellano, C. (1976). Effects of nicotine on discrimination learning, consolidation and learned behavior in two inbred strains of mice. Psychopharmacology (Berlin), 48, 3743. Clarke, P. B. S. (1987). Nicotine and smoking: A perspective from animal studies. Psychopharmacology (Berlin), 92, 135-143.
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Erickson, C. K., Stavchansky, S. A., Koch, K. I., & McGinty, J. W. (1982). A new subcutaneously-implantable reservoir for sustained release of nicotine in the rat. Pharmacology, Biochemistry & Behavior, 17, 183-185. Garg, M. (1969). The effects of nicotine on two different types of learning. Psychopharmacologia (Berlin), 15, 408-414. Hall, G. H., & Morrison, C. F. (1973). New evidence for a relationship between tobacco smoking, nicotine dependence and stress. Nature (London), 243, 199-201. Hughes, J. RI, & Hatsukami, D. (1986). Signs and symptoms of tobacco withdrawal. Archives of General Psychiatry, 43, 289-294. Hunter, B., Zornetzer, S. F., Jarvik, M. E., & McGaugh, J. L. (1977). Modulation of learning and memory: Effects of drugs influencing neurotransmitters. In L. L. Iverson, S. D. Iverson, & S. H. Snyder (Eds.), Handbook ofpsychopharmacology (Vol. 8, pp. 531-577). New York: Plenum. Ksir, C., Hakan, R., Hall, D. P., & Kellar, K. J. (1985). Exposure to nicotine enhances the behavioral stimulant effect of nicotine and increases binding of 3-H acetylcholine to nicotinic receptors. Neuropharmacology, 24, 527-531. Levin, E. D. (1988). Psychopharmacological effects in the radial-arm maze. Neuroscience & Biobehavioral Reviews, 12, 169-175. Levin, E. D., Castonguay, M., & Ellison, G. D. (1987a). Effects of the nicotinic receptor blocker, mecamylamine, on radial-arm maze performance in rats. Behavioral & Neural Biology, 48, 206-212. Levin, E. D., Ellison, G. D., Salem, C., Jarvik, M., & Gritz, E. (1988). Behavioral effects of acute hexamethonium in rats chronically intoxicated with nicotine. Physiology & Behavior, 44, 355-359. Levin, E. D., Morgan, M. M., Galvez, C., & Ellison, G. D. (1987b). Chronic nicotine and withdrawal effects on body weight and food and water consumption in female rats. Physiology & Behavior, 39, 441-444. Marks, M. J., Burch, J. B., & Collins, A. C. (1983). Effects of chronic nicotine infusion on tolerance development and nicotine receptors. Journal of Pharmacology and Experimental Therapeutics, 226, 817-825. Marks, M. J., Stitzel, J. A., & Collins, A. C. (1985). Time course study of the effects of chronic nicotine infusion on drug response and brain receptors. Journal of Pharmacology and Experimental Therapeutics, 235, 619-628. Morgan, M. M., & Ellison, G. D. (1987). Development of tolerance to nicotine effects in rats. Psychopharmacology (Berlin), 91, 236-238. Mundy, W. R., & Iwamoto, E. T. (1988). Nicotine acquisition of radial maze performance in rats. Pharmacology, Biochemistry & Behavior, 30, 119-122. Nelsen, J. M. (1978). Psychobiological consequences of chronic nicotinization: A focus on arousal. In K. Battig (Ed.), Behavioral effects of nicotine (pp. 1-17). Basel: Karger. Olton, D. S., & Samuelson, R. J. (1976). Rememberance of places passed: Spatial memory in rats. Journal of Experimental Psychology: Animal Behavior Processes, 2, 97-115. Schwartz, R. D., & Kellar, K. J. (1983). Nicotinic cholinergic binding sites in brain: In vivo regulation. Science, 220, 214-216. Schwartz, R. D., & Kellar, K. J. (1985). In vivo regulation of [3-H]acetylcholine recognition sites in brain by nicotinic cholinergic drugs. Journal of Neurochemistry, 45, 427-433. Wesnes, K., & Warburton, D. M. (1983). Smoking, nicotine and human performance. Pharmacology and Therapeutics, 21, 189-208.
