Center for Alcohol and Drug Abuse Studies & Department of Pharmacology, Brody School of Medicine, East Carolina University, Greenville, NC 27858, USA
Received 12 July 2001; in revised form 6 February 2002; accepted 15 February 2002
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ABSTRACT |
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INTRODUCTION |
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A relationship between taste aversion conditionability and ethanol acceptance was demonstrated within rats selectively bred for high and low acceptance of ethanol. Alcohol-non-preferring (NP) rats were more susceptible to developing a taste aversion to saccharin than alcohol-preferring (P) rats when ethanol was used as the aversive-conditioning agent (Froehlich et al., 1988). On the other hand, selective breeding for taste aversion conditionability produces rat lines that differ in ethanol preference, as seen when comparing TAP (taste aversion prone) and TAR (taste aversion resistant) line preferences with respect to ethanol free-choice consumption. Thus, rats genetically selected for high and low CTA acquisition, based on an emetic type of unconditioned stimulus and a saccharin solution conditioned stimulus, consume relatively low and high levels of ethanol, respectively, in a free-choice paradigm (Orr et al., 1997
). The amount of ethanol consumed by the TAR rats was also comparable with that of the P rats, which were reported to drink regularly >5 g/kg body wt/day in a free-choice paradigm (Gatto et al., 1987
; Orr et al., 1997
). TAP rats had higher serotonin levels than TAR rats in whole brain tissue, which was in harmony with the relationship between the NP and P rats (Murphy et al., 1982
, 1987
; Orr et al., 1993
). The behaviour of the TAP and TAR lines also resembles that of the NP and P rat lines in that both models display line differences in response to CTA paradigms in which ethanol is used as the aversive conditioning stimulus (Froehlich et al., 1988
; Elkins et al., 1991
, 1992
). This behaviour has also been observed more recently in other high- and low-ethanol drinking inbred lines, like the HAD/LAD and the UChA/UChB rats (Badia-Elder et al., 2000
; Quintanilla et al., 2001
). Therefore, it is possible that genetically mediated substrates of individual differences in taste aversion conditionability contribute to individual differences in propensities for ethanol consumption.
The mHEP (Myers high-ethanol-preferring) rat was recently developed at East Carolina University by crossing female SpragueDawley rats selected for high consumption of ethanol after a 10-day step-up procedure with male P rats. mHEP rats consume larger than normal quantities of ethanol and prefer high concentrations of ethanol. Similar to the P rat, they maintain their preference for ethanol in the presence of other palatable drinks. They have also demonstrated increased levels of anxiety in behaviour tests, such as the elevated plus-maze (Myers et al., 1998).
The selectively bred P rat does not exhibit a taste aversion to ethanol at low to moderate concentrations, and TAR rats selectively bred for their inability to learn the taste aversion paradigm consume high quantities of ethanol, but where does the mHEP rat fall? Does the mHEP rat respond differently from an outbred rat in the conditioned taste aversion paradigm? Also, with the determination of the ability or inability of the mHEP rat to learn the taste aversion paradigm, the role of taste in high ethanol consumption may be further elucidated. In the CTA paradigm, solutions of ethanol and saccharin were used as the conditioned stimulus (CS) since they represented a novel taste to the animals. An aversive dose of LiCl for the animal was used as the unconditioned stimulus (US).
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MATERIALS AND METHODS |
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Taste aversion
Two days prior to the beginning of the experiment, animals were placed in suspended wire cages with unlimited access to water and food. Approximately 24 h before beginning training for limited access to fluids, all fluids were removed (McMillen and Williams, 1998). Animals were randomly assigned to one of three treatment groups. On days 1 and 2, animals were adapted to a 2 h presentation of water beginning in the early afternoon between 13:00 and 14:00. On day 3, the animals received either 0.05% (w/v) saccharin or 7% (v/v) ethanol for 1 h and the amount consumed was noted. At the end of 1 h, either 0.5 M NaCl or 0.5 M LiCl was injected intraperitoneally at a volume of 0.2 ml/100 g body weight. This dose is equivalent to 42 mg LiCl/kg. Novel solutions were removed immediately following each injection, so that each rat had no further access to solutions that day. The presentation of the novel solutions was limited to 1 h, as our experience with limited access has shown that most of the consumption occurs during the first hour and the injection of a noxious stimulus should be paired with consumption. On day 4, water was presented for 2 h. On the final day, the novel solutions were presented again for 1 h and consumption noted. The decrease in consumption was taken as a measurement of the strength of the aversion learned. Data were analysed using a three-way, repeated measures analysis of variance (ANOVA) and Newman Keuls post hoc analysis, using GB-STAT (Dynamic Microsystems, Inc., Silver Spring, MD, USA).
Screening for ethanol preference and consumption
Those male and female mHEP rats which received ethanol or saccharin as the novel solutions followed by injection of LiCl for CTA were later screened, using a 10-day step-up procedure (Myers et al., 1998). Animals were housed individually in suspended metal cages. Three calibrated drinking tubes were attached to each cage: one for ethanol, one for water, and a blank. Drinking tubes were rotated daily in a semi-random manner. Animals received 3% (v/v) ethanol, which was increased daily to 30% over the 10-day period. Body weight, fluid intake, and food intake were measured daily. The data at each concentration were analysed by a two-way repeated measures ANOVA and NewmanKeuls post hoc analysis.
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RESULTS |
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Analysis of data for ethanol consumption with a repeated measures three-way ANOVA showed a significant interaction between strain, sex, and treatment received [F(2,65) = 26.12, P < 0.001]. The interaction of strain with sex was not significant [F(2,32) = 1.16, P > 0.05], but there was a significant interaction of sex with treatment [F(1,33) = 5.70, P < 0.05]. Table 2 shows that when animals were exposed to ethanol as the novel solution followed by injection of 0.5 M NaCl, consumption of ethanol tended to increase with the second ethanol exposure. The increase was significant for the male Wistar rat (41%) and the male mHEP rat (69%), but not for female SpragueDawley or female mHEP rats. This result demonstrated that the handling and injection of solution is not aversive to any of these strains and sexes of rat. In contrast, injection of 0.5 M LiCl produced an aversive response with the female SpragueDawley rat exhibiting the most dramatic decrease (-88.5%). Male Wistar, male mHEP and female mHEP rats also significantly decreased their consumption, by 45, 52 and 30%, respectively.
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DISCUSSION |
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One explanation for the difference in the ability of the male mHEP rat to learn the taste aversion paradigm and the female mHEP rat not to learn the paradigm is that taste aversion is not related to alcohol drinking, but rather has been passed along during the selective breeding of various lines. That is, as one behavioural phenotype is selected for during the breeding process, unselected behavioural differences between the selected lines may develop, and the possibility that the unselected line differences are due to the inadvertent fixation of trait-irrelevant genes cannot therefore be ruled out (Orr et al., 1997). Another more likely line of reasoning is that the male and female mHEP rats consume high quantities of ethanol for different reasons. Earlier generations of mHEP rats were already characterized as having biochemical and behavioural differences between the male and female mHEP rats (Myers et al., 1998
). The female mHEP rats not only consume more ethanol when considering g/kg intake and proportion to total fluid intake, compared with their male counterparts, but they also prefer a higher concentration of ethanol. Perhaps most interesting are the differences in brain chemistry between mHEP male and female rats (Lucas and McMillen, 2002
). Male mHEP rats have lower than normal brain concentrations of serotonin, similar to the P rat. Low serotonin concentrations have long been implicated as causes of high alcohol consumption in humans (Olson et al., 1960
; Thomson and McMillen, 1987
), but males represent the majority of subjects in these studies. The female mHEP rat with the exception of the nucleus accumbens, however, has normal concentrations of serotonin (Lucas and McMillen, 2002
). It is possible that the male mHEP rat is disposed to consuming high amounts of ethanol partly because of a deficiency in brain serotonin, whereas something else related to taste reactivity disposes the female mHEP rat to consume high quantities of ethanol.
The present data also demonstrate that both male and female mHEP rats continue to consume high amounts of ethanol when screened with a 10-day step-up procedure, regardless of whether they received the taste aversion paradigm with ethanol or saccharin as the novel solution. With the female mHEP rat, there appears to be no significant difference in g/kg intake or preference between either treatment group throughout the step-up procedure. On the other hand, after learning a taste aversion to 7% (v/v) ethanol, the male mHEP rat will still consume high amounts of ethanol. At lower concentrations (3, 4, 5 and 7%), however, the amount consumed is significantly less in the group that received the ethanol/LiCl treatment in the taste aversion paradigm than the group that received the saccharin/LiCl treatment. This difference disappeared at ethanol concentrations >9%, when consumption by the two groups became approximately the same. This would suggest that, after several days of ethanol presentation in the step-up procedure, the male mHEP rats that had learned a taste aversion to ethanol are able to extinguish the aversion and begin to consume quantities of ethanol similar to male mHEP rats which did not receive a taste aversion to ethanol. Another possibility is that there is something different in how the ethanol taste is perceived, so that the taste of higher concentrations of ethanol is perceived differently from lower concentrations. Therefore, animals that received a taste aversion to 7% ethanol would consume less ethanol at concentrations of 7%, but at higher concentrations would consume more ethanol because it presents a different taste than the one an aversion was learned to. This is in agreement with human studies, which show that different concentrations of ethanol exert different orosensory responses (Scinska et al., 2000
; Mattes and DiMeglio, 2001).
In summary, the male mHEP rat will learn and exhibit a CTA with either saccharin or ethanol as the novel solution. The learning from CTA with ethanol carries over into the ethanol preference step-up procedure, but only at the lower concentrations of ethanol. The female mHEP rat exhibits neither response. These data indicate that the basis for high ethanol consumption by the male mHEP rat is different from the female mHEP rat.
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ACKNOWLEDGEMENTS |
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FOOTNOTES |
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REFERENCES |
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Elkins, R. L. (1986) Separation of taste-aversion-prone and taste-aversion-resistant rats through selective breeding: implications for individual differences in conditionability and aversion-therapy alcoholism treatment.Behavioral Neuroscience 100, 121124.[ISI][Medline]
Elkins, R. L. (1991) An appraisal of chemical aversion (emetic therapy) approaches to alcoholism treatment.Behaviour Research and Therapy 29, 387413.[ISI][Medline]
Elkins, R. L., Walters, P. A., Orr, T. E., Kolbe, E. F., Ritch, J. E., Hess, D. L. and Hobbs, S. H. (1991) Taste aversion, hypnotic and hypothermic effects of alcohol in rats genetically predisposed (selectively bred) to differ in taste aversion conditionability.Alcoholism: Clinical and Experimental Research 15, 321.
Elkins, R. L., Walters, P. A. and Orr, T. E. (1992) Continued development and unconditioned stimulus characterization of selectively bred lines of taste aversion prone and resistant rats.Alcoholism: Clinical and Experimental Research 16, 928934.[ISI][Medline]
Froehlich, J. C., Harts, J., Lumeng, L. and Li, T.-K. (1988) Differences in response to the aversive properties of ethanol in rats selectively bred for oral ethanol preference.Pharmacology, Biochemistry and Behavior 31, 215222.[ISI][Medline]
Gatto, G. J., Murphy, J. M., Waller, M. B., McBride, W. J., Lumeng, L. and Li, T.-K. (1987) Chronic ethanol tolerance through free-choice drinking in the P line of alcohol-preferring rats.Pharmacology, Biochemistry and Behavior 28, 111115.[ISI][Medline]
Lucas, L. A. C. and McMillen, B. A. (2002) Differences in brain area concentrations of dopamine and serotonin in Myers high ethanol preferring (mHEP) and outbred rats.Journal of Neural Transmission 109, 279292.[ISI]
Lush, I. E. (1991) The genetics of bitterness, sweetness, and saltiness in strains of mice. In Chemical Senses, Wysocki, C. J. and Kare, M. R. eds, Vol. 3, pp. 227241. Marcel Dekker, New York.
Mattes, R. D. and DiMeglio, D. (2000) Ethanol perception and ingestion.Physiology and Behavior 72, 217229.[ISI]
McMillen, B. A. and Williams, H. L. (1998) Role of taste and calories in the selection of ethanol by C57bl/6NHsd and Hsd:ICR mice.Alcohol 15, 193198.[ISI][Medline]
Murphy, J. M., McBride, W. J., Lumeng, L. and Li, T.-K. (1982) Regional brain levels of monoamines in alcohol-preferring and -nonpreferring lines of rats.Pharmacology, Biochemistry and Behavior 16, 145149.[ISI][Medline]
Murphy, J. M., McBride, W. J., Lumeng, L. and Li, T.-K. (1987) Contents of monoamines in forebrain regions of alcohol-preferring (P) and -nonpreferring (NP) lines of rats.Pharmacology, Biochemistry and Behavior 26, 389392.[ISI][Medline]
Myers, R. D., Robinson, D. E., West, M. W., Biggs, T. A. and McMillen, B. A. (1998) Genetics of alcoholism: rapid development of a new high-ethanol-preferring (HEP) strain of female and male rats.Alcohol 16, 343357.[ISI][Medline]
Olson, R. E., Gursey, D. and Vester, J. (1960) Evidence for a defect in tryptophan metabolism in chronic alcoholism.New England Journal of Medicine 263, 11691174.[ISI]
Orr, T. E., Walters, P. A., Carl, G. F. and Elkins, R. L. (1993) Brain levels of amines and amino acids in taste aversion-prone and -resistant rats.Physiology and Behavior 53, 495500.[ISI][Medline]
Orr, T. E., Walters, P. A. and Elkins, R. L. (1997) Differences in free-choice ethanol acceptance between taste aversion-prone and taste aversion-resistant rats.Alcoholism: Clinical and Experimental Research 21, 14911496.[ISI][Medline]
Quintanilla, M. E., Callejas, O. and Tampier, L. (2001) Differences in sensitivity to the aversive effects of ethanol in low-alcohol drinking (UChA) and high-alcohol drinking (UChB) rats.Alcohol 23, 177182.[ISI][Medline]
Scinska, A., Koros, E., Habrat, B., Kukwas, A., Kostowski, W. and Bienkowski, P. (2000) Bitter and sweet components of ethanol taste in human.Drug and Alcohol Dependence 60, 199206.[ISI][Medline]
Thomson, S. M. Jr and McMillen, B. A. (1987) Test for decreased serotonin/tryptophan metabolite ratios in abstinent alcoholics.Alcohol 4, 15.[ISI][Medline]