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Caffeine promotes ethanol drinking in rats
Alcohol 21 (2000) 271 ± 277
Caffeine promotes ethanol drinking in rats Examination using a limited-access free choice paradigm Daniel Kunin*, Stephane Gaskin, Franc Rogan, Brian R. Smith, Zalman Amit Center for Studies in Behavioral Neurobiology, Department of Psychology, Concordia University, 1455 de Maisonneuve Boulevard, West, H-1013, Montreal, Quebec, Canada H3G 1M8 Received 13 March 2000; received in revised form 1 May 2000; accepted 12 May 2000
Abstract There is growing evidence that caffeine may alter the pattern of intake of a variety of drugs. The present study was designed to assess the effect of caffeine pretreatment on voluntary ethanol consumption. The first experiment examined the effect of caffeine on the acquisition of ethanol intake in a limited-access-choice procedure in which water and ethanol were presented concurrently. Male Wistar rats, exposed to food and water ad lib, were presented with a daily 1-h choice session between water and progressively increasing concentrations of ethanol (2 ± 10%). Each ethanol concentration was made available for 4 ± 6 days for a total of 20 days of access to ethanol. Intraperitoneal injections of caffeine (5 or 10 mg/kg) or saline were administered to the rats 30 min prior to each choice session. Caffeine produced a dose-related facilitation in ethanol drinking whereby the lower caffeine dose produced enhancement in ethanol drinking. The second experiment examined the effect of caffeine on the maintenance of established ethanol consumption. Male Wistar rats, initially acclimatized to increasing concentrations of ethanol (2% ± 10), were presented with an additional 18 ethanol (10%) presentations, comprised of a 6-day baseline period followed by 6 days of treatment where animals were given one of three doses of caffeine (2.5, 5 or 10 mg/kg) or saline prior to ethanol presentation. A final 6-day post-treatment period followed treatment. These results revealed an inverted-U effect of caffeine dose on ethanol ingestion where the low and high caffeine doses produced no effect but the moderate dose of 5 mg/kg enhanced ethanol drinking that persisted throughout the post-treatment period. A third experiment revealed that caffeine did not alter levels of blood ethanol within the time period used for the ethanol drinking session. D 2000 Elsevier Science Inc. All rights reserved. Keywords: Caffeine; Ethanol; Limited access; Rat; Voluntary intake
Multi-drug use for a variety of drugs of abuse is something frequently seen. In fact, the use of alcohol, nicotine and caffeine are known to co-vary (Kozlowski et al., 1993). Recently, much attention has been devoted to studying the interactive effects of these drugs. Several studies have shown that caffeine can increase the behavioral effects of a variety of drugs. It has been shown that the acute co-administration of caffeine and nicotine produced additive effects on locomotion activity (Lee et al., 1987) and on operant responding on a fixed interval 2-in schedule of food reinforcement (White, 1988). Caffeine has also been shown to augment the locomotion-activating effects of both cocaine and amphe-
* Corresponding author. Tel.: +1-514-848-2186; fax: +1-514-8482187. E-mail address: [email protected] (D. Kunin).
tamine (White & Keller, 1984; Misra et al., 1986; Schenk et al., 1989) and has been shown to augment scheduleinduced responding for these drugs (Logan et al., 1989; Evans & Wenger, 1990). Finally, caffeine has been shown to increase the ataxic effects produced by ethanol (Dar et al., 1987). There is growing evidence that the use of caffeine may modify the pattern of intake of a variety of drugs including nicotine, amphetamine, cocaine and ethanol. For example, pretreatment with caffeine has been shown to augment the self-administration of cocaine (Horger et al., 1991; Schenk et al., 1994; Comer & Carroll, 1996), nicotine (Shoaib et al., 1999) and amphetamine (Jaszyna et al., 1998). Studies assessing the effect of caffeine on ethanol intake have been conflicting. Caffeine administered in the diet of malnourished rats has been shown to facilitate ethanol drinking (Gilbert, 1976, 1979). However, interpretation of these findings is problematic as
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they are confounded by the fact that food deprivation may itself enhance the self-administration of drugs (Carroll & Meisch, 1984). In addition, since caffeine was delivered to animals throughout the course of the entire day, it is difficult to determine the exact dose of caffeine delivered at any moment in time. In contrast, an intraperitoneal injection of caffeine at a dose of 50 mg/kg has been shown to decrease ethanol intake but at a dose producing toxic effects (Dietze & Kulkosky, 1991). The aim of the present study was to assess the effect of caffeine pretreatment, at low and moderate doses, on the acquisition and maintenance of ethanol intake in nonfood deprived rats. Experiment 1 was designed to assess the effect of caffeine on the acquisition of ethanol intake in free-feeding, ethanol-naõÈve rats. Experiment 2, assessed the effect of caffeine on the maintenance of ethanol intake. Finally, the effect of caffeine on the overall rates of ethanol metabolism was also assessed. 1. Experiment 1 1.1. Method 1.1.1. Subjects Twenty-one male Wistar rats (Charles River Canada) weighing between 275 and 325 g at the start of the experiment were used. Rats were individually housed in hanging stainless steel wire cages with standard lab chow and water available ad libitum at all times throughout the experiment, except during a 1-h limited-access ethanol session. The animals were maintained on a 12-h on/12-h off light/dark cycle in a room regulated for constant temperature and humidity. All animals used in this and the following experiments were treated in accordance with the guidelines of the Canadian Council for Animal Care. In addition, the care and use, as well as all procedures involving animals in the present study were approved by the Concordia University Animal Care Committee. 1.1.2. Drugs Caffeine (Sigma) was dissolved in 0.9% saline and injected in a volume of 1 ml/kg. Injections were administered IP and injection volumes were constant across doses. Ethanol solutions were prepared from a dilution of a 95% stock solution with tap water and was presented to animals in a 12-ml graduated drinking tube fitted with one-holed rubber stopper with a stainless steel ball-bearing spout. 1.1.3. Procedure Following 7 days of acclimatization to the colony room conditions animals were subjected to an ethanol drinking acquisition regimen. The animals remained in their home cages at all times throughout the study but
had their water bottles removed during a 1-h ethanol presentation period. During this period, animals received two drinking tubes with a choice between water and ethanol. Ethanol was presented to the rats in ascending concentrations beginning with 2% v/v presented for 6 consecutive days followed by an additional 6 days of 5% ethanol, 4 days of 8% culminating with 4 days of 10% ethanol for a total of 20 days (Smith et al., 1999). Prior to the first ethanol exposure session, animals were randomly assigned to one of three groups (n = 7). One group received saline solution at a volume of 1 ml/kg, while the other two groups received injections of caffeine, (5 and 10 mg/kg, respectively). Injections were administered daily, 30 min prior to ethanol exposure sessions for the 20-day duration of the experiment. Ethanol intake was measured to the nearest 0.1 ml. 1.1.4. Data analysis Data from Experiment 1 (acquisition data) that included ethanol intake (g/kg), ethanol intake (ml/kg), water intake (ml/kg) and total fluid intake (ml/kg) were analyzed using a two-way ANOVA (group days). Test of simple main effects and simple main comparisons were used to analyze any significant interaction effects (Keppel et al., 1992). For Experiment 2 (maintenance data), ethanol intake (g/kg), ethanol intake (ml/kg), water intake (ml/kg) and total fluid intake (ml/kg) were also analyzed using a two-way ANOVA (group days). Test of simple main effects and simple main comparisons were used to analyze any significant interaction effects. Data for Experiment 3 was analyzed using a two-way ANOVA (group time) conducted on whole blood levels. 1.2. Results Fig. 1a shows ethanol intake (g/kg) across 20 ethanol presentation days for the caffeine and saline treated animals. A two-way ANOVA with a repeated measure across presentation days for absolute ethanol intake (g/kg) revealed significant effects of group F(2,18) = 1.444, p < 0.05, days F(19,342) = 0.315, p < 0.01 and group days interaction F(38,342) = 0.123, p < 0.01. Test of simple effects followed by test of simple comparisons (a = 0.05) revealed that the caffeine- (5 mg/kg) treated animals increased their ethanol intake relative to the saline- and caffeine- (10 mg/kg) treated animals beginning on the third presentation of 8% ethanol and continuing to the final presentation of 10% ethanol. In contrast, animals treated with caffeine (10 mg/kg) did not differ from saline-treated animals. A two-way ANOVA with a repeated measure across presentation days for ethanol intake (ml/kg) revealed significant effects of group F(2,18) = 4.218, p < 0.05, days F(19,342) = 2.106, p < 0.01 and group days interaction F(38,342) = 1.785, p < 0.01. Further analysis revealed that
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Fig. 1. Effect of caffeine (5 and 10 mg/kg) or saline on ethanol consumed (g ethanol/kg body weight, panel a) and water intake (expressed as ml fluid/kg body weight, panel b). Vertical lines represent SEM.
caffeine-(5 mg/kg) treated animals increased their ethanol intake relative to saline- and caffeine- (10 mg/kg) treated animals beginning from the third presentation of 8% ethanol and continuing to the final presentation of 10% ethanol. Consistent with the data for absolute ethanol intake (g/kg), animals treated with caffeine (10 mg/kg) did not differ from saline-treated animals. Fig. 1b shows water intake expressed as ml/kg across the 20 ethanol presentation days for animals pretreated with either caffeine or saline. Analysis of this data revealed no significant differences in water consumption between the groups across the 20 days. A two-way ANOVA with a repeated measure across presentation days for total fluid data revealed significant effects of group F(2,18) = 5.107, p < 0.05, days F(19,342) = 2.521, p < 0.01 and group days interaction F(38,342) = 1.526, p < 0.05. Subsequent analysis revealed that caffeine- (5 mg/ kg) treated animals increased their total fluid intake relative to saline- and caffeine- (10 mg/kg) treated animals on the third presentation of 8% ethanol and the final two presentations of 10% ethanol.
2. Experiment 2 2.1. Method 2.1.1. Subjects Forty male Wistar rats (Charles River Canada) weighing between 275 and 325 g were used in the present experiment. Housing conditions were identical to those outlined in Experiment 1. 2.1.2. Procedure Animals were first exposed to a free choice between increasing concentrations of ethanol (2 ± 10%) and water for a period of 20 days in a manner identical to the previous experiment but with no drug pretreatment. Following this acclimatization period, animals were presented with ethanol (10%) for an additional 6 days, constituting the baseline phase. During this baseline drinking phase, all animals were given IP saline injections once daily, 30 min prior to choice sessions between ethanol and water. These saline injections served to habituate animals to the
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injection regimen. Following the 6 days of baseline, animals were assigned to one of four treatment conditions based on their mean levels of ethanol intake (g/kg) across the baseline phase, which resulted in similar mean ethanol intake between the four groups. Animals were then given one of three doses of caffeine (2.5, 5 or 10 mg/kg) or saline depending on their group assignment (n = 10 per group) for an additional 6 consecutive days. Injections were administered 30 min prior to choice sessions between ethanol and water. The final 6 days comprised the post-treatment phase, where all animals were administered saline injections, 30 min prior to choice sessions. 2.2. Results Fig. 2a shows the effect of caffeine or saline treatment on the maintenance of ethanol intake (g/kg) across baseline, treatment and post-treatment phases. A two-way ANOVA with a repeated measure across presentation days for absolute ethanol intake (g/kg) revealed significant effects of group F(3,34) = 4.527, p < 0.01, days F(17,578) = 2.114, p < 0.01 and group days interaction F(51,578) = 2.123, p < 0.01. Further analysis revealed that the caffeine- (5
mg/kg) treated animals increased their ethanol intake relative to the saline- and caffeine- (2.5 and 10 mg/kg) treated animals beginning on the fourth treatment day and continuing throughout the post-treatment phase with the exception of day 4. Moreover, animals treated with caffeine (2.5 and 10 mg/kg) did not differ from saline-treated animals on any day. A two-way ANOVA with a repeated measure across presentation days for ethanol intake (ml/kg) revealed significant effects of group F(3,34) = 4.538, p < 0.01, days F(17,578) = 2.087, p < 0.01 and group days interaction F(51,578) = 2.116, p < 0.01. Further analysis demonstrated that the caffeine- (5 mg/kg) treated animals increased their ethanol intake relative to the saline- and caffeine- (2.5 and 10 mg/kg) treated animals beginning on the fourth treatment day and continuing throughout most of the post-treatment phase. Consistent with the data for absolute ethanol intake (g/kg), animals treated with caffeine (2.5 and 10 mg/kg) did not differ from saline-treated animals. Fig. 2b shows water intake expressed as ml/kg across baseline, treatment and post-treatment phases for animals treated with caffeine or saline. These results suggested that there were no systematic differences in water consumption
Fig. 2. Effect of caffeine (2.5, 5, 10 mg/kg) or saline on the maintenance of ethanol intake (g ethanol/kg body weight, panel a) and water intake (expressed as ml fluid/kg body weight, panel b). Vertical lines represent SEM.
D. Kunin et al. / Alcohol 21 (2000) 271±277
between the groups across baseline, treatment and posttreatment phases of the experiment. A two-way ANOVA with a repeated measure across presentation days for total fluid data revealed significant effects of group F(3,34) = 5.248, p < 0.01, days F(17,578) = 2.252, p < 0.01 and group days interaction F(51,578) = 1.526, p < 0.01. Subsequent analysis revealed that caffeine- (5 mg/kg) treated animals increased their total fluid intake relative to saline- and caffeine- (2.5 and 10 mg/ kg) treated animals beginning on the fourth treatment day and continuing throughout most of the post-treatment phase. 3. Experiment 3 3.1. Method 3.1.1. Subjects Twenty-four male Wistar rats (Charles River Canada) weighing between 275 and 325 g were used in the present experiment. Housing conditions were identical to those outlined in Experiment 1. 3.1.2. Procedure After 7 days of adaptation to the laboratory housing conditions, animals were randomly assigned to one of four groups (n = 6). Animals assigned to group caffeine± ethanol and saline± ethanol were injected with caffeine (5 mg/kg) or saline (1 ml/kg) 30 min prior to injections with ethanol (0.8 g/kg; 20% v/v) and sacrificed by decapitation at 15 or 30 min post ethanol injection. The dose of 5 mg/kg of caffeine was chosen since it produced the behavioral effect seen in Experiments 1 and 2. The dose of ethanol selected has been shown to yield blood ethanol levels that are similar to those achieved by animals that voluntarily consume ethanol (Gill et al., 1986). Immediately after its collection, trunk blood was kept on ice in heparinized (100 U) tubes. A volume of 1 ml of heparinized blood was mixed with 0.5 ml of a PCA (perchloric acid) reagent (1 N PCA, 20 mM thiourea). The samples were centrifuged and the supernatant (0.5 ml) was sealed in glass vials for headspace analysis using Perkin-Elmer HS-6B Headspace sampler. Control samples were prepared by replacing blood in an equal volume of saline and alcohol. The vial samples were stored at ÿ 20°C overnight and then subject to analysis the following day. The procedure was based on that of Whitmire and Whitmire (1995). 3.2. Results Table 1 shows the effect of caffeine on blood ethanol levels at different time intervals after ethanol injections. A two-way ANOVA (group time) conducted on whole blood levels revealed no significant effects of group F(1,20) = 0.001, p > 0.05, time F(1,20) = 3.746, p = 0.06 and group time interaction F(1,20) = 0.183, p>0.05. Although, the results
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Table 1 Effect of caffeine (5 mg/kg) on blood ethanol levels (mg/100 ml whole blood) following 1P administration of ethanol (0.8 g/kg) Time after ethanol administration Treatment condition
15 min
30 min
Caffeine ± ethanol Saline ± ethanol
59.098 5.498 61.076 6.748
51.720 3.760 49.515 2.411
revealed no significant effect of time, the effect approached significance. These results suggest there was no significant alteration in blood ethanol levels resulting from caffeine pretreatment within the time period used for the ethanol drinking session. 4. Discussion The present investigation was designed to assess the effect of caffeine pretreatment on the acquisition and maintenance of ethanol consumption in free-feeding laboratory rats. The results of the first experiment revealed that pretreatment with caffeine at a dose of 5 mg/kg but not 10 mg/kg facilitated the ingestion of ethanol solutions at 8% and 10% ethanol concentrations. Moreover, the effects of caffeine appeared to be specific to ethanol and not due to generalized increases in fluid intake as no accompanying increase in water intake was observed during the choice presentation of ethanol and water. The results of Experiment 2 demonstrated a caffeineinduced facilitation in ethanol drinking in animals previously exposed to an ethanol acquisition regimen. The effects of caffeine on ethanol intake followed an invertedU curve progression. Specifically, it was shown that while 5 mg/kg of caffeine augmented ethanol drinking by as much as 400%, caffeine at doses of 2.5 and 10 mg/kg failed to alter ethanol drinking. It is also noteworthy that caffeine- (5 mg/kg) injected animals increased their ethanol drinking after merely four injections, suggesting that acute caffeine (5 mg/kg) treatment is sufficient to augment ethanol consumption. Furthermore, these animals maintained an elevated drinking pattern throughout the post-treatment phase, despite termination of caffeine injections. The present caffeine-induced elevation in ethanol intake merits attention. Specifically, the present findings suggest a very narrow dose range of caffeine that results in elevation in ethanol drinking behavior. One explanation for this phenomenon may relate to the notion that at a dose of 5 mg/kg, caffeine may have enhanced the ability of the animals to associate ethanol-drinking behavior with the perception of ethanol's post-ingestive rewarding effects. Such a view would be consistent with the observation that in Experiment 1 animals treated with caffeine (5 mg/kg) began to diverge from the other treatment conditions at the higher ethanol concentrations where ethanol is believed to be consumed for its' post-ingestive pharmacological effects
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(Gill et al., 1996; Boyle et al., 1997). In fact, the amount of ethanol consumed by these caffeine-injected rats was within a range sufficient to produce psychopharmacological effects (Gill et al., 1986). In contrast, caffeine at a dose of 2.5 may have been below threshold such that it failed to alter the perception of ethanol's post-ingestive rewarding effects. Similarly, caffeine at a dose of 10 mg/kg may have been outside the optimal range of producing an enhancement in ethanol drinking. It is noteworthy that other manipulations have been shown to effect behavior in a fashion resembling an inverted-U shaped curve (e.g., Galina et al., 1983). It is possible that the magnitude of arousal produced by the high dose of caffeine may have interfered with the enhancement of the association between the post-ingestional effects of ethanol and ethanol drinking behavior that appeared to result from exposure to the moderate dose of caffeine. The present findings are in disagreement with previous observations that caffeine administration failed to alter ethanol intake (Potthoff et al., 1983). One explanation for these discrepant results could be related to differences in caffeine delivery (i.e., slow-release pellet vs. an acute injection of caffeine). The present findings are also at odds with results of previous studies showing that caffeine reduced ethanol consumption in laboratory rats (Hederra et al., 1975; Dietze & Kulkosky, 1991). However, these latter studies used high doses of caffeine that were within a range producing toxic effects. The present findings suggest a very narrow dose range of caffeine that results in an elevation in ethanol drinking. It is possible that the dose exposure of caffeine in previous studies assessing the effect of caffeine on ethanol intake may have been outside this narrow effective dose. At this point, the mechanism by which caffeine produces enhancement in ethanol drinking remains unclear. It is unlikely that the alteration in ethanol drinking behavior was the result of a caffeine-induced change in ethanol metabolism. The results of Experiment 3 showed that caffeine, at the dose tested failed to alter blood levels of ethanol following ethanol treatment. This observation suggests that the nature of the interaction between caffeine and ethanol was not the result of a simple change in blood ethanol levels. One mechanism by which caffeine may produce elevations in ethanol drinking may be related to self-medication (Potthoff et al., 1983). Others have demonstrated that increases in ethanol intake may result from treatment with nicotine (Potthoff et al., 1983; Blomqvist et al., 1996; Smith et al., 1999; Leà et al., 2000), amphetamine (Potthoff et al., 1983) and low doses of morphine (Linseman, 1989; Stromberg et al., 1997), drugs which can be classified as having stimulant effects. In the present study, it is shown that caffeine, a drug with stimulant properties, will also augment ethanol intake. It is conceivable that continuous stimulant administration may have induced hyperactivity such that animals may consume more ethanol in order to antagonize this effect (Potthoff et al., 1983). The increase in ethanol drinking seen with 5 mg/kg caffeine may be consistent with
this notion. However, the latter view would predict that animals treated with 10 mg/kg would also increase their ethanol drinking, which was not the case. In addition, a selfmedication explanation cannot account for the observation that in Experiment 2, caffeine- (5 mg/kg) treated animals maintained high levels of ethanol drinking after caffeine injections had terminated. As was previously mentioned, at 5 mg/kg, caffeine may have enhanced the ability of the animals to associate ethanol-drinking behavior with the perception of ethanol's post-ingestive rewarding effects, thus producing elevations in ethanol drinking even upon cessation of caffeine treatment. An alternate hypothesis is that caffeine may have sensitized the animals to ethanol's reinforcing effects and thus produced increases in ethanol drinking. Indeed there is a growing body of data demonstrating that caffeine may potentiate the reinforcing properties of a variety of drugs including cocaine (Horger et al., 1991; Schenk et al., 1994; Comer & Carroll, 1996), nicotine (Shoaib et al., 1999) and amphetamine (Jaszyna et al., 1998). However, if caffeine sensitization were the mechanism by which rats elevated their ethanol drinking, it would be predicted that for both Experiments 1 and 2 a similar number of caffeine injections would enhance ethanol drinking. In fact, examination of the data would reveal that in Experiment 1 rats elevated their ethanol consumption after 14 caffeine injections. In contrast, rats in Experiment 2 elevated their ethanol intake after merely four caffeine injections. Furthermore, if caffeine sensitization were in fact the mechanism by which rats elevated their ethanol consumption, then it would be predicted that at a dose of 10 mg/kg caffeine would also produce elevations in ethanol drinking. These observations may rule out the likelihood that caffeine sensitization produced elevations in ethanol drinking. Despite the fact that a definitive explanation of the mechanisms still remains to be elucidated, the present findings merit attention. The observation that exposure to caffeine may increase the propensity to consume alcohol may be of some relevance for those individuals who concurrently use both substances. Acknowledgments This research was funded by a grant to Z. Amit and B.R. Smith from the Medical Research Council of Canada. References Blomqvist, O., Ericson, M., Johnson, D. H., Engel, J. A., & Sonderpalm, B. (1996). Voluntary ethanol intake in the rat: effects of nicotinic acetylcholine receptor blockade or subchronic nicotine treatment. Eur J Pharmacol 314, 257 ± 267. Boyle, A. E. L., Smith, B. R., & Amit, Z. (1997). A descriptive analysis of the structure and temporal pattern of voluntary ethanol intake within an acquisition paradigm. J Stud Alcohol 58, 382 ± 391.
D. Kunin et al. / Alcohol 21 (2000) 271±277 Carroll, M. E., & Meisch, R. A. (1984). Increased drug self-administration due to food deprivation. In T. Thompson, P. B. Dews, & J. E. Barrett (Eds.), Advances in Behavioral Pharmacology ( pp. 47 ± 88). New York: Academic Press. Comer, S. D., & Carroll, M. E. (1996). Oral caffeine pretreatment produced modest increases in smoked cocaine self-administration in rhesus monkeys. Psychopharmacology 126, 281 ± 285. Dar, M. S., Jones, M., Close, G., Mustafa Jamal, S., & Wooles, W. R. (1987). Behavioral interactions of ethanol and methylxanthines. Psychopharmacology 91, 1 ± 4. Dietze, M. A., & Kulkosky, P. J. (1991). Effects of caffeine and bombesin on ethanol and food intake. Life Sci 48, 1837 ± 1844. Evans, E. B., & Wenger, G. R. (1990). The effects of cocaine in combination with other drugs of abuse on schedule-controlled behavior in the pigeon. Pharmacol Biochem Behav 37, 349 ± 357. Galina, Z. H., Sutherland, C. J., & Amit, Z. (1983). Effects of heat-stress on behavior and the pituitary adrenal axis in rats. Pharmacol Biochem Behav 19, 251 ± 256. Gilbert, R. M. (1976). Dietary caffeine and EtOH consumption by rats. J Stud Alcohol 37, 11 ± 18. Gilbert, R. M. (1979). Augmentation of EtOH-consumption by caffeine in malnourished rats. J Stud Alcohol 40, 19 ± 27. Gill, K., France, C., & Amit, Z. (1986). Voluntary ethanol consumption in rats: an examination of blood/brain ethanol levels and behavior. Alcohol Clin Exp Res 10, 457 ± 462. Gill, K., Smith, B. R., & Amit, Z. (1996). Alcohol as a food: a commentary on Richter. Physiol Behav 60, 1485 ± 1490. Hederra, A., Aldunate, J., Segovia-Riquelme, N., & Mardones, J. (1975). Effects of caffeine on the voluntary alcohol intake of rats. In J. D. Sinclair, & K. Kiianmaa (Eds.), The Effects of Centrally Active Drugs on Voluntary Alcohol Consumption ( pp. 9 ± 13). Helsinki: Finnish Foundation for Alcohol Studies. Horger, B. A., Wellman, P. J., Morien, A., Davies, B. T., & Schenk, S. (1991). Caffeine exposure sensitizes rats to the reinforcing effects of cocaine. NeuroReport 2, 53 ± 56. Jaszyna, M., Gasior, M., Shoaib, M., Yasar, S., & Goldberg, S. (1998). Behavioral effects of nicotine, amphetamine and cocaine under a fixed-interval schedule of food reinforcement in rats chronically exposed to caffeine. Psychopharmacology 140, 257 ± 271. Keppel, G., Sauffley, W. H., & Tokunaga, H. (1992). Introduction to Design and Analysis (2nd ed.). New York: Freeman. Kozlowski, L. T., Henningfield, J. E., Keenan, R. M., Lei, H., Leigh, G., Jelinek, L. C., Pope, M. A., & Haertzen, C. A. (1993). Patterns of
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alcohol, cigarette, and caffeine and other drug use in two drug abusing populations. J Subst Abuse Treat 10, 171 ± 179. LeÃ, A. D., Corrigall, W. A., Harding, J. W. S., Juzytsch, W., & Li, T. K. (2000). Involvement of nicotinic receptors in alcohol self-administration. Alcohol Clin Exp Res 24, 155 ± 163. Lee, E. Y., Tsai, M. J., Tang, Y. P., & Chai, C. Y. (1987). Differential biochemical mechanisms mediate locomotor stimulation effects by caffeine and nicotine in rats. Pharmacol Biochem Behav 26, 427 ± 430. Linseman, M. A. (1989). Central vs. peripheral mediation of opioid effects on alcohol consumption in free-feeding rats. Pharmacol Biochem Behav 33, 407 ± 413. Logan, L., Carney, J. M., Holloway, F. A., & Seale, T. W. (1989). Effects of caffeine, cocaine and their combination on fixed-interval behavior in rats. Pharmacol Biochem Behav 33, 99 ± 104. Misra, A. L., Vadlamani, N. L., & Pontani, R. B. (1986). Effect of caffeine on cocaine locomotor stimulant activity in rats. Pharmacol Biochem Behav 24, 761 ± 764. Potthoff, A. D., Ellison, G., & Nelson, L. (1983). Ethanol intake increases during continuous administration of amphetamine and nicotine, but not several other drugs. Pharmacol Biochem Behav 18, 489 ± 493. Schenk, S., Horger, B. A., & Snow, S. (1989). Caffeine preexposure sensitizes rats to the motor activating effects of cocaine. Behav Pharmacol 1, 447 ± 451. Schenk, S., Valades, A., Horger, B. A., Snow, S., & Wellman, P. J. (1994). Interactions between caffeine and cocaine in tests of self-administration. Behav Pharmacol 5, 153 ± 158. Shoaib, M., Swanner, L. S., Yasar, S., & Goldberg, S. (1999). Chronic caffeine exposure potentiates nicotine self-administration in rats. Psychopharmacology 142, 327 ± 333. Smith, B. R., Horan, J. T., Gaskin, S., & Amit, Z. (1999). Exposure to nicotine enhances acquisition of ethanol drinking by laboratory rats in a limited access paradigm. Psychopharmacology 142, 408 ± 412. Stromberg, M. F., Meister, S., Volpicelli, J. R., & Ulm, R. (1997). Morphine enhances selection of both sucrose and ethanol in a two-bottle test. Alcohol 14, 55 ± 62. White, B. C., & Keller, G. E. (1984). Caffeine pretreatment: enhancement and attenuation of D-amphetamine-induced activity. Pharmacol Biochem Behav 20, 383 ± 386. White, J. M. (1988). Behavioral interactions between nicotine and caffeine. Pharmacol Biochem Behav 29, 63 ± 66. Whitmire, D., & Whitmire, P. S. (1995). Analysis of ethanol and acetaldehyde recovery from perchloric acid-treated blood. Alcohol Alcohol 5, 623 ± 628.
Caffeine promotes ethanol drinking in rats Examination using a limited-access free choice paradigm Daniel Kunin*, Stephane Gaskin, Franc Rogan, Brian R. Smith, Zalman Amit Center for Studies in Behavioral Neurobiology, Department of Psychology, Concordia University, 1455 de Maisonneuve Boulevard, West, H-1013, Montreal, Quebec, Canada H3G 1M8 Received 13 March 2000; received in revised form 1 May 2000; accepted 12 May 2000
Abstract There is growing evidence that caffeine may alter the pattern of intake of a variety of drugs. The present study was designed to assess the effect of caffeine pretreatment on voluntary ethanol consumption. The first experiment examined the effect of caffeine on the acquisition of ethanol intake in a limited-access-choice procedure in which water and ethanol were presented concurrently. Male Wistar rats, exposed to food and water ad lib, were presented with a daily 1-h choice session between water and progressively increasing concentrations of ethanol (2 ± 10%). Each ethanol concentration was made available for 4 ± 6 days for a total of 20 days of access to ethanol. Intraperitoneal injections of caffeine (5 or 10 mg/kg) or saline were administered to the rats 30 min prior to each choice session. Caffeine produced a dose-related facilitation in ethanol drinking whereby the lower caffeine dose produced enhancement in ethanol drinking. The second experiment examined the effect of caffeine on the maintenance of established ethanol consumption. Male Wistar rats, initially acclimatized to increasing concentrations of ethanol (2% ± 10), were presented with an additional 18 ethanol (10%) presentations, comprised of a 6-day baseline period followed by 6 days of treatment where animals were given one of three doses of caffeine (2.5, 5 or 10 mg/kg) or saline prior to ethanol presentation. A final 6-day post-treatment period followed treatment. These results revealed an inverted-U effect of caffeine dose on ethanol ingestion where the low and high caffeine doses produced no effect but the moderate dose of 5 mg/kg enhanced ethanol drinking that persisted throughout the post-treatment period. A third experiment revealed that caffeine did not alter levels of blood ethanol within the time period used for the ethanol drinking session. D 2000 Elsevier Science Inc. All rights reserved. Keywords: Caffeine; Ethanol; Limited access; Rat; Voluntary intake
Multi-drug use for a variety of drugs of abuse is something frequently seen. In fact, the use of alcohol, nicotine and caffeine are known to co-vary (Kozlowski et al., 1993). Recently, much attention has been devoted to studying the interactive effects of these drugs. Several studies have shown that caffeine can increase the behavioral effects of a variety of drugs. It has been shown that the acute co-administration of caffeine and nicotine produced additive effects on locomotion activity (Lee et al., 1987) and on operant responding on a fixed interval 2-in schedule of food reinforcement (White, 1988). Caffeine has also been shown to augment the locomotion-activating effects of both cocaine and amphe-
* Corresponding author. Tel.: +1-514-848-2186; fax: +1-514-8482187. E-mail address: [email protected] (D. Kunin).
tamine (White & Keller, 1984; Misra et al., 1986; Schenk et al., 1989) and has been shown to augment scheduleinduced responding for these drugs (Logan et al., 1989; Evans & Wenger, 1990). Finally, caffeine has been shown to increase the ataxic effects produced by ethanol (Dar et al., 1987). There is growing evidence that the use of caffeine may modify the pattern of intake of a variety of drugs including nicotine, amphetamine, cocaine and ethanol. For example, pretreatment with caffeine has been shown to augment the self-administration of cocaine (Horger et al., 1991; Schenk et al., 1994; Comer & Carroll, 1996), nicotine (Shoaib et al., 1999) and amphetamine (Jaszyna et al., 1998). Studies assessing the effect of caffeine on ethanol intake have been conflicting. Caffeine administered in the diet of malnourished rats has been shown to facilitate ethanol drinking (Gilbert, 1976, 1979). However, interpretation of these findings is problematic as
0741-8329/00/$ ± see front matter D 2000 Elsevier Science Inc. All rights reserved. PII: S 0 7 4 1 - 8 3 2 9 ( 0 0 ) 0 0 1 0 1 - 4
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they are confounded by the fact that food deprivation may itself enhance the self-administration of drugs (Carroll & Meisch, 1984). In addition, since caffeine was delivered to animals throughout the course of the entire day, it is difficult to determine the exact dose of caffeine delivered at any moment in time. In contrast, an intraperitoneal injection of caffeine at a dose of 50 mg/kg has been shown to decrease ethanol intake but at a dose producing toxic effects (Dietze & Kulkosky, 1991). The aim of the present study was to assess the effect of caffeine pretreatment, at low and moderate doses, on the acquisition and maintenance of ethanol intake in nonfood deprived rats. Experiment 1 was designed to assess the effect of caffeine on the acquisition of ethanol intake in free-feeding, ethanol-naõÈve rats. Experiment 2, assessed the effect of caffeine on the maintenance of ethanol intake. Finally, the effect of caffeine on the overall rates of ethanol metabolism was also assessed. 1. Experiment 1 1.1. Method 1.1.1. Subjects Twenty-one male Wistar rats (Charles River Canada) weighing between 275 and 325 g at the start of the experiment were used. Rats were individually housed in hanging stainless steel wire cages with standard lab chow and water available ad libitum at all times throughout the experiment, except during a 1-h limited-access ethanol session. The animals were maintained on a 12-h on/12-h off light/dark cycle in a room regulated for constant temperature and humidity. All animals used in this and the following experiments were treated in accordance with the guidelines of the Canadian Council for Animal Care. In addition, the care and use, as well as all procedures involving animals in the present study were approved by the Concordia University Animal Care Committee. 1.1.2. Drugs Caffeine (Sigma) was dissolved in 0.9% saline and injected in a volume of 1 ml/kg. Injections were administered IP and injection volumes were constant across doses. Ethanol solutions were prepared from a dilution of a 95% stock solution with tap water and was presented to animals in a 12-ml graduated drinking tube fitted with one-holed rubber stopper with a stainless steel ball-bearing spout. 1.1.3. Procedure Following 7 days of acclimatization to the colony room conditions animals were subjected to an ethanol drinking acquisition regimen. The animals remained in their home cages at all times throughout the study but
had their water bottles removed during a 1-h ethanol presentation period. During this period, animals received two drinking tubes with a choice between water and ethanol. Ethanol was presented to the rats in ascending concentrations beginning with 2% v/v presented for 6 consecutive days followed by an additional 6 days of 5% ethanol, 4 days of 8% culminating with 4 days of 10% ethanol for a total of 20 days (Smith et al., 1999). Prior to the first ethanol exposure session, animals were randomly assigned to one of three groups (n = 7). One group received saline solution at a volume of 1 ml/kg, while the other two groups received injections of caffeine, (5 and 10 mg/kg, respectively). Injections were administered daily, 30 min prior to ethanol exposure sessions for the 20-day duration of the experiment. Ethanol intake was measured to the nearest 0.1 ml. 1.1.4. Data analysis Data from Experiment 1 (acquisition data) that included ethanol intake (g/kg), ethanol intake (ml/kg), water intake (ml/kg) and total fluid intake (ml/kg) were analyzed using a two-way ANOVA (group days). Test of simple main effects and simple main comparisons were used to analyze any significant interaction effects (Keppel et al., 1992). For Experiment 2 (maintenance data), ethanol intake (g/kg), ethanol intake (ml/kg), water intake (ml/kg) and total fluid intake (ml/kg) were also analyzed using a two-way ANOVA (group days). Test of simple main effects and simple main comparisons were used to analyze any significant interaction effects. Data for Experiment 3 was analyzed using a two-way ANOVA (group time) conducted on whole blood levels. 1.2. Results Fig. 1a shows ethanol intake (g/kg) across 20 ethanol presentation days for the caffeine and saline treated animals. A two-way ANOVA with a repeated measure across presentation days for absolute ethanol intake (g/kg) revealed significant effects of group F(2,18) = 1.444, p < 0.05, days F(19,342) = 0.315, p < 0.01 and group days interaction F(38,342) = 0.123, p < 0.01. Test of simple effects followed by test of simple comparisons (a = 0.05) revealed that the caffeine- (5 mg/kg) treated animals increased their ethanol intake relative to the saline- and caffeine- (10 mg/kg) treated animals beginning on the third presentation of 8% ethanol and continuing to the final presentation of 10% ethanol. In contrast, animals treated with caffeine (10 mg/kg) did not differ from saline-treated animals. A two-way ANOVA with a repeated measure across presentation days for ethanol intake (ml/kg) revealed significant effects of group F(2,18) = 4.218, p < 0.05, days F(19,342) = 2.106, p < 0.01 and group days interaction F(38,342) = 1.785, p < 0.01. Further analysis revealed that
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Fig. 1. Effect of caffeine (5 and 10 mg/kg) or saline on ethanol consumed (g ethanol/kg body weight, panel a) and water intake (expressed as ml fluid/kg body weight, panel b). Vertical lines represent SEM.
caffeine-(5 mg/kg) treated animals increased their ethanol intake relative to saline- and caffeine- (10 mg/kg) treated animals beginning from the third presentation of 8% ethanol and continuing to the final presentation of 10% ethanol. Consistent with the data for absolute ethanol intake (g/kg), animals treated with caffeine (10 mg/kg) did not differ from saline-treated animals. Fig. 1b shows water intake expressed as ml/kg across the 20 ethanol presentation days for animals pretreated with either caffeine or saline. Analysis of this data revealed no significant differences in water consumption between the groups across the 20 days. A two-way ANOVA with a repeated measure across presentation days for total fluid data revealed significant effects of group F(2,18) = 5.107, p < 0.05, days F(19,342) = 2.521, p < 0.01 and group days interaction F(38,342) = 1.526, p < 0.05. Subsequent analysis revealed that caffeine- (5 mg/ kg) treated animals increased their total fluid intake relative to saline- and caffeine- (10 mg/kg) treated animals on the third presentation of 8% ethanol and the final two presentations of 10% ethanol.
2. Experiment 2 2.1. Method 2.1.1. Subjects Forty male Wistar rats (Charles River Canada) weighing between 275 and 325 g were used in the present experiment. Housing conditions were identical to those outlined in Experiment 1. 2.1.2. Procedure Animals were first exposed to a free choice between increasing concentrations of ethanol (2 ± 10%) and water for a period of 20 days in a manner identical to the previous experiment but with no drug pretreatment. Following this acclimatization period, animals were presented with ethanol (10%) for an additional 6 days, constituting the baseline phase. During this baseline drinking phase, all animals were given IP saline injections once daily, 30 min prior to choice sessions between ethanol and water. These saline injections served to habituate animals to the
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injection regimen. Following the 6 days of baseline, animals were assigned to one of four treatment conditions based on their mean levels of ethanol intake (g/kg) across the baseline phase, which resulted in similar mean ethanol intake between the four groups. Animals were then given one of three doses of caffeine (2.5, 5 or 10 mg/kg) or saline depending on their group assignment (n = 10 per group) for an additional 6 consecutive days. Injections were administered 30 min prior to choice sessions between ethanol and water. The final 6 days comprised the post-treatment phase, where all animals were administered saline injections, 30 min prior to choice sessions. 2.2. Results Fig. 2a shows the effect of caffeine or saline treatment on the maintenance of ethanol intake (g/kg) across baseline, treatment and post-treatment phases. A two-way ANOVA with a repeated measure across presentation days for absolute ethanol intake (g/kg) revealed significant effects of group F(3,34) = 4.527, p < 0.01, days F(17,578) = 2.114, p < 0.01 and group days interaction F(51,578) = 2.123, p < 0.01. Further analysis revealed that the caffeine- (5
mg/kg) treated animals increased their ethanol intake relative to the saline- and caffeine- (2.5 and 10 mg/kg) treated animals beginning on the fourth treatment day and continuing throughout the post-treatment phase with the exception of day 4. Moreover, animals treated with caffeine (2.5 and 10 mg/kg) did not differ from saline-treated animals on any day. A two-way ANOVA with a repeated measure across presentation days for ethanol intake (ml/kg) revealed significant effects of group F(3,34) = 4.538, p < 0.01, days F(17,578) = 2.087, p < 0.01 and group days interaction F(51,578) = 2.116, p < 0.01. Further analysis demonstrated that the caffeine- (5 mg/kg) treated animals increased their ethanol intake relative to the saline- and caffeine- (2.5 and 10 mg/kg) treated animals beginning on the fourth treatment day and continuing throughout most of the post-treatment phase. Consistent with the data for absolute ethanol intake (g/kg), animals treated with caffeine (2.5 and 10 mg/kg) did not differ from saline-treated animals. Fig. 2b shows water intake expressed as ml/kg across baseline, treatment and post-treatment phases for animals treated with caffeine or saline. These results suggested that there were no systematic differences in water consumption
Fig. 2. Effect of caffeine (2.5, 5, 10 mg/kg) or saline on the maintenance of ethanol intake (g ethanol/kg body weight, panel a) and water intake (expressed as ml fluid/kg body weight, panel b). Vertical lines represent SEM.
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between the groups across baseline, treatment and posttreatment phases of the experiment. A two-way ANOVA with a repeated measure across presentation days for total fluid data revealed significant effects of group F(3,34) = 5.248, p < 0.01, days F(17,578) = 2.252, p < 0.01 and group days interaction F(51,578) = 1.526, p < 0.01. Subsequent analysis revealed that caffeine- (5 mg/kg) treated animals increased their total fluid intake relative to saline- and caffeine- (2.5 and 10 mg/ kg) treated animals beginning on the fourth treatment day and continuing throughout most of the post-treatment phase. 3. Experiment 3 3.1. Method 3.1.1. Subjects Twenty-four male Wistar rats (Charles River Canada) weighing between 275 and 325 g were used in the present experiment. Housing conditions were identical to those outlined in Experiment 1. 3.1.2. Procedure After 7 days of adaptation to the laboratory housing conditions, animals were randomly assigned to one of four groups (n = 6). Animals assigned to group caffeine± ethanol and saline± ethanol were injected with caffeine (5 mg/kg) or saline (1 ml/kg) 30 min prior to injections with ethanol (0.8 g/kg; 20% v/v) and sacrificed by decapitation at 15 or 30 min post ethanol injection. The dose of 5 mg/kg of caffeine was chosen since it produced the behavioral effect seen in Experiments 1 and 2. The dose of ethanol selected has been shown to yield blood ethanol levels that are similar to those achieved by animals that voluntarily consume ethanol (Gill et al., 1986). Immediately after its collection, trunk blood was kept on ice in heparinized (100 U) tubes. A volume of 1 ml of heparinized blood was mixed with 0.5 ml of a PCA (perchloric acid) reagent (1 N PCA, 20 mM thiourea). The samples were centrifuged and the supernatant (0.5 ml) was sealed in glass vials for headspace analysis using Perkin-Elmer HS-6B Headspace sampler. Control samples were prepared by replacing blood in an equal volume of saline and alcohol. The vial samples were stored at ÿ 20°C overnight and then subject to analysis the following day. The procedure was based on that of Whitmire and Whitmire (1995). 3.2. Results Table 1 shows the effect of caffeine on blood ethanol levels at different time intervals after ethanol injections. A two-way ANOVA (group time) conducted on whole blood levels revealed no significant effects of group F(1,20) = 0.001, p > 0.05, time F(1,20) = 3.746, p = 0.06 and group time interaction F(1,20) = 0.183, p>0.05. Although, the results
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Table 1 Effect of caffeine (5 mg/kg) on blood ethanol levels (mg/100 ml whole blood) following 1P administration of ethanol (0.8 g/kg) Time after ethanol administration Treatment condition
15 min
30 min
Caffeine ± ethanol Saline ± ethanol
59.098 5.498 61.076 6.748
51.720 3.760 49.515 2.411
revealed no significant effect of time, the effect approached significance. These results suggest there was no significant alteration in blood ethanol levels resulting from caffeine pretreatment within the time period used for the ethanol drinking session. 4. Discussion The present investigation was designed to assess the effect of caffeine pretreatment on the acquisition and maintenance of ethanol consumption in free-feeding laboratory rats. The results of the first experiment revealed that pretreatment with caffeine at a dose of 5 mg/kg but not 10 mg/kg facilitated the ingestion of ethanol solutions at 8% and 10% ethanol concentrations. Moreover, the effects of caffeine appeared to be specific to ethanol and not due to generalized increases in fluid intake as no accompanying increase in water intake was observed during the choice presentation of ethanol and water. The results of Experiment 2 demonstrated a caffeineinduced facilitation in ethanol drinking in animals previously exposed to an ethanol acquisition regimen. The effects of caffeine on ethanol intake followed an invertedU curve progression. Specifically, it was shown that while 5 mg/kg of caffeine augmented ethanol drinking by as much as 400%, caffeine at doses of 2.5 and 10 mg/kg failed to alter ethanol drinking. It is also noteworthy that caffeine- (5 mg/kg) injected animals increased their ethanol drinking after merely four injections, suggesting that acute caffeine (5 mg/kg) treatment is sufficient to augment ethanol consumption. Furthermore, these animals maintained an elevated drinking pattern throughout the post-treatment phase, despite termination of caffeine injections. The present caffeine-induced elevation in ethanol intake merits attention. Specifically, the present findings suggest a very narrow dose range of caffeine that results in elevation in ethanol drinking behavior. One explanation for this phenomenon may relate to the notion that at a dose of 5 mg/kg, caffeine may have enhanced the ability of the animals to associate ethanol-drinking behavior with the perception of ethanol's post-ingestive rewarding effects. Such a view would be consistent with the observation that in Experiment 1 animals treated with caffeine (5 mg/kg) began to diverge from the other treatment conditions at the higher ethanol concentrations where ethanol is believed to be consumed for its' post-ingestive pharmacological effects
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(Gill et al., 1996; Boyle et al., 1997). In fact, the amount of ethanol consumed by these caffeine-injected rats was within a range sufficient to produce psychopharmacological effects (Gill et al., 1986). In contrast, caffeine at a dose of 2.5 may have been below threshold such that it failed to alter the perception of ethanol's post-ingestive rewarding effects. Similarly, caffeine at a dose of 10 mg/kg may have been outside the optimal range of producing an enhancement in ethanol drinking. It is noteworthy that other manipulations have been shown to effect behavior in a fashion resembling an inverted-U shaped curve (e.g., Galina et al., 1983). It is possible that the magnitude of arousal produced by the high dose of caffeine may have interfered with the enhancement of the association between the post-ingestional effects of ethanol and ethanol drinking behavior that appeared to result from exposure to the moderate dose of caffeine. The present findings are in disagreement with previous observations that caffeine administration failed to alter ethanol intake (Potthoff et al., 1983). One explanation for these discrepant results could be related to differences in caffeine delivery (i.e., slow-release pellet vs. an acute injection of caffeine). The present findings are also at odds with results of previous studies showing that caffeine reduced ethanol consumption in laboratory rats (Hederra et al., 1975; Dietze & Kulkosky, 1991). However, these latter studies used high doses of caffeine that were within a range producing toxic effects. The present findings suggest a very narrow dose range of caffeine that results in an elevation in ethanol drinking. It is possible that the dose exposure of caffeine in previous studies assessing the effect of caffeine on ethanol intake may have been outside this narrow effective dose. At this point, the mechanism by which caffeine produces enhancement in ethanol drinking remains unclear. It is unlikely that the alteration in ethanol drinking behavior was the result of a caffeine-induced change in ethanol metabolism. The results of Experiment 3 showed that caffeine, at the dose tested failed to alter blood levels of ethanol following ethanol treatment. This observation suggests that the nature of the interaction between caffeine and ethanol was not the result of a simple change in blood ethanol levels. One mechanism by which caffeine may produce elevations in ethanol drinking may be related to self-medication (Potthoff et al., 1983). Others have demonstrated that increases in ethanol intake may result from treatment with nicotine (Potthoff et al., 1983; Blomqvist et al., 1996; Smith et al., 1999; Leà et al., 2000), amphetamine (Potthoff et al., 1983) and low doses of morphine (Linseman, 1989; Stromberg et al., 1997), drugs which can be classified as having stimulant effects. In the present study, it is shown that caffeine, a drug with stimulant properties, will also augment ethanol intake. It is conceivable that continuous stimulant administration may have induced hyperactivity such that animals may consume more ethanol in order to antagonize this effect (Potthoff et al., 1983). The increase in ethanol drinking seen with 5 mg/kg caffeine may be consistent with
this notion. However, the latter view would predict that animals treated with 10 mg/kg would also increase their ethanol drinking, which was not the case. In addition, a selfmedication explanation cannot account for the observation that in Experiment 2, caffeine- (5 mg/kg) treated animals maintained high levels of ethanol drinking after caffeine injections had terminated. As was previously mentioned, at 5 mg/kg, caffeine may have enhanced the ability of the animals to associate ethanol-drinking behavior with the perception of ethanol's post-ingestive rewarding effects, thus producing elevations in ethanol drinking even upon cessation of caffeine treatment. An alternate hypothesis is that caffeine may have sensitized the animals to ethanol's reinforcing effects and thus produced increases in ethanol drinking. Indeed there is a growing body of data demonstrating that caffeine may potentiate the reinforcing properties of a variety of drugs including cocaine (Horger et al., 1991; Schenk et al., 1994; Comer & Carroll, 1996), nicotine (Shoaib et al., 1999) and amphetamine (Jaszyna et al., 1998). However, if caffeine sensitization were the mechanism by which rats elevated their ethanol drinking, it would be predicted that for both Experiments 1 and 2 a similar number of caffeine injections would enhance ethanol drinking. In fact, examination of the data would reveal that in Experiment 1 rats elevated their ethanol consumption after 14 caffeine injections. In contrast, rats in Experiment 2 elevated their ethanol intake after merely four caffeine injections. Furthermore, if caffeine sensitization were in fact the mechanism by which rats elevated their ethanol consumption, then it would be predicted that at a dose of 10 mg/kg caffeine would also produce elevations in ethanol drinking. These observations may rule out the likelihood that caffeine sensitization produced elevations in ethanol drinking. Despite the fact that a definitive explanation of the mechanisms still remains to be elucidated, the present findings merit attention. The observation that exposure to caffeine may increase the propensity to consume alcohol may be of some relevance for those individuals who concurrently use both substances. Acknowledgments This research was funded by a grant to Z. Amit and B.R. Smith from the Medical Research Council of Canada. References Blomqvist, O., Ericson, M., Johnson, D. H., Engel, J. A., & Sonderpalm, B. (1996). Voluntary ethanol intake in the rat: effects of nicotinic acetylcholine receptor blockade or subchronic nicotine treatment. Eur J Pharmacol 314, 257 ± 267. Boyle, A. E. L., Smith, B. R., & Amit, Z. (1997). A descriptive analysis of the structure and temporal pattern of voluntary ethanol intake within an acquisition paradigm. J Stud Alcohol 58, 382 ± 391.
D. Kunin et al. / Alcohol 21 (2000) 271±277 Carroll, M. E., & Meisch, R. A. (1984). Increased drug self-administration due to food deprivation. In T. Thompson, P. B. Dews, & J. E. Barrett (Eds.), Advances in Behavioral Pharmacology ( pp. 47 ± 88). New York: Academic Press. Comer, S. D., & Carroll, M. E. (1996). Oral caffeine pretreatment produced modest increases in smoked cocaine self-administration in rhesus monkeys. Psychopharmacology 126, 281 ± 285. Dar, M. S., Jones, M., Close, G., Mustafa Jamal, S., & Wooles, W. R. (1987). Behavioral interactions of ethanol and methylxanthines. Psychopharmacology 91, 1 ± 4. Dietze, M. A., & Kulkosky, P. J. (1991). Effects of caffeine and bombesin on ethanol and food intake. Life Sci 48, 1837 ± 1844. Evans, E. B., & Wenger, G. R. (1990). The effects of cocaine in combination with other drugs of abuse on schedule-controlled behavior in the pigeon. Pharmacol Biochem Behav 37, 349 ± 357. Galina, Z. H., Sutherland, C. J., & Amit, Z. (1983). Effects of heat-stress on behavior and the pituitary adrenal axis in rats. Pharmacol Biochem Behav 19, 251 ± 256. Gilbert, R. M. (1976). Dietary caffeine and EtOH consumption by rats. J Stud Alcohol 37, 11 ± 18. Gilbert, R. M. (1979). Augmentation of EtOH-consumption by caffeine in malnourished rats. J Stud Alcohol 40, 19 ± 27. Gill, K., France, C., & Amit, Z. (1986). Voluntary ethanol consumption in rats: an examination of blood/brain ethanol levels and behavior. Alcohol Clin Exp Res 10, 457 ± 462. Gill, K., Smith, B. R., & Amit, Z. (1996). Alcohol as a food: a commentary on Richter. Physiol Behav 60, 1485 ± 1490. Hederra, A., Aldunate, J., Segovia-Riquelme, N., & Mardones, J. (1975). Effects of caffeine on the voluntary alcohol intake of rats. In J. D. Sinclair, & K. Kiianmaa (Eds.), The Effects of Centrally Active Drugs on Voluntary Alcohol Consumption ( pp. 9 ± 13). Helsinki: Finnish Foundation for Alcohol Studies. Horger, B. A., Wellman, P. J., Morien, A., Davies, B. T., & Schenk, S. (1991). Caffeine exposure sensitizes rats to the reinforcing effects of cocaine. NeuroReport 2, 53 ± 56. Jaszyna, M., Gasior, M., Shoaib, M., Yasar, S., & Goldberg, S. (1998). Behavioral effects of nicotine, amphetamine and cocaine under a fixed-interval schedule of food reinforcement in rats chronically exposed to caffeine. Psychopharmacology 140, 257 ± 271. Keppel, G., Sauffley, W. H., & Tokunaga, H. (1992). Introduction to Design and Analysis (2nd ed.). New York: Freeman. Kozlowski, L. T., Henningfield, J. E., Keenan, R. M., Lei, H., Leigh, G., Jelinek, L. C., Pope, M. A., & Haertzen, C. A. (1993). Patterns of
277
alcohol, cigarette, and caffeine and other drug use in two drug abusing populations. J Subst Abuse Treat 10, 171 ± 179. LeÃ, A. D., Corrigall, W. A., Harding, J. W. S., Juzytsch, W., & Li, T. K. (2000). Involvement of nicotinic receptors in alcohol self-administration. Alcohol Clin Exp Res 24, 155 ± 163. Lee, E. Y., Tsai, M. J., Tang, Y. P., & Chai, C. Y. (1987). Differential biochemical mechanisms mediate locomotor stimulation effects by caffeine and nicotine in rats. Pharmacol Biochem Behav 26, 427 ± 430. Linseman, M. A. (1989). Central vs. peripheral mediation of opioid effects on alcohol consumption in free-feeding rats. Pharmacol Biochem Behav 33, 407 ± 413. Logan, L., Carney, J. M., Holloway, F. A., & Seale, T. W. (1989). Effects of caffeine, cocaine and their combination on fixed-interval behavior in rats. Pharmacol Biochem Behav 33, 99 ± 104. Misra, A. L., Vadlamani, N. L., & Pontani, R. B. (1986). Effect of caffeine on cocaine locomotor stimulant activity in rats. Pharmacol Biochem Behav 24, 761 ± 764. Potthoff, A. D., Ellison, G., & Nelson, L. (1983). Ethanol intake increases during continuous administration of amphetamine and nicotine, but not several other drugs. Pharmacol Biochem Behav 18, 489 ± 493. Schenk, S., Horger, B. A., & Snow, S. (1989). Caffeine preexposure sensitizes rats to the motor activating effects of cocaine. Behav Pharmacol 1, 447 ± 451. Schenk, S., Valades, A., Horger, B. A., Snow, S., & Wellman, P. J. (1994). Interactions between caffeine and cocaine in tests of self-administration. Behav Pharmacol 5, 153 ± 158. Shoaib, M., Swanner, L. S., Yasar, S., & Goldberg, S. (1999). Chronic caffeine exposure potentiates nicotine self-administration in rats. Psychopharmacology 142, 327 ± 333. Smith, B. R., Horan, J. T., Gaskin, S., & Amit, Z. (1999). Exposure to nicotine enhances acquisition of ethanol drinking by laboratory rats in a limited access paradigm. Psychopharmacology 142, 408 ± 412. Stromberg, M. F., Meister, S., Volpicelli, J. R., & Ulm, R. (1997). Morphine enhances selection of both sucrose and ethanol in a two-bottle test. Alcohol 14, 55 ± 62. White, B. C., & Keller, G. E. (1984). Caffeine pretreatment: enhancement and attenuation of D-amphetamine-induced activity. Pharmacol Biochem Behav 20, 383 ± 386. White, J. M. (1988). Behavioral interactions between nicotine and caffeine. Pharmacol Biochem Behav 29, 63 ± 66. Whitmire, D., & Whitmire, P. S. (1995). Analysis of ethanol and acetaldehyde recovery from perchloric acid-treated blood. Alcohol Alcohol 5, 623 ± 628.