Evidence that oral and nutrient reinforcers differentially condition appetitive and consummatory responses to flavors

MYERS, K. P. AND W. G. HALL. Evidence that oral and nutrient reinforcers differentially condition appetitive and consummatory responses to flavors. PHYSIOL BEHAV 64(4) 493–500, 1998.—Rats tend to increase their intake of a flavor that has previously been paired with either sweet taste or with caloric repletion. However, it is unclear whether such a change in intake is caused by changes in appetitive behaviors such as orienting and approach, or changes in consummatory behaviors and oral responsiveness. Also, it is unclear whether oral reinforcers (sweetness) and postingestive reinforcers (nutrients) lead to the same kinds of behavioral change. In the current experiments, weanling rats with oral and gastric cannulas repeatedly experienced a flavor paired with either sweetness, high caloric density, or neither. Rats were then tested for differences in appetitive olfactory orienting and consummatory oral responsiveness elicited by the flavor. Results suggest that oral reinforcement (sweetness) produces conditioning of appetitive responding to the flavor, while postingestive reinforcement produces conditioning of consummatory responding. A second experiment indicates that these behavioral changes are specific increases in responsiveness conditioned by flavor + unconditioned stimulus (US) pairing, and are unlikely to be nonspecific effects of daily unconditioned stimulus exposure.     

Flavor learning in weanling rats and its retention

The present study examined whether weanling animals can acquire associative memory for reward and retain it several weeks later. Three-week-old Wistar male rats were trained in a flavor learning task. Half of the rats received unsweetened grape-flavored water on odd-numbered days and sweetened (sucrose) cherry-flavored solution on even-numbered days. The remaining rats received sweetened grape-flavored solution on odd-numbered days and unsweetened cherry-flavored water on even-numbered days. During the acquisition session, the liquid was presented to each rat for 15 min daily for 6 consecutive days. In the following test session, each rat was presented with unsweetened cherry-flavored water and grape-flavored water simultaneously for 15 min daily for 4 consecutive days. The rats showed significant preferences for the flavor previously associated with 2% and 10% sucrose, significant aversion to the flavor associated with 30% sucrose, and no particular preference or aversion to the flavor associated with 20% sucrose, indicating a hedonic shift from positive to negative with an increasing concentration of sucrose. The association learning acquired at the age of 3 weeks was retained when re-tested in adulthood at the age of 20 weeks. In contrast to the conditioned flavor aversion associated with 30% sucrose, 20-week-old rats showed a preference for this flavor. In accordance with these learning effects, 3-week-old rats preferred 2% sucrose to 30% sucrose, and the reverse was true in 20-week-old rats. The reasons for rejection of high-concentration sucrose by weanling rats are also discussed. The present study showed that weanling rats established a conditioned flavor preference or aversion depending on the concentration of associated sucrose and retained it in adulthood, indicating that feeding experience in the weanling period is important in influencing later dietary preferences.     

Flavor preferences conditioned by sucrose depend upon training and testing methods: two-bottle tests revisited

In confirmation of prior work, rats given one-bottle training with flavored 5% and 30% sucrose solutions (CS5 and CS30) strongly preferred the CS5 when both flavors were presented in intermediate 17.5% sucrose solutions. The CS5 preference has been attributed to a conditioned satiety response to the CS30 flavor, but equal intakes of CS5 and CS30 in one-bottle tests did not support this view. To determine if sweetness differences between training and test solutions contributed to the CS5 preference, new rats were trained and tested with flavored 10% sucrose solutions. One flavor (CS5) was paired with matched intragastric (ig) water infusions (=net 5% solution) and another flavor (CS30) was paired with matched infusions of 50% sucrose (=net 30% solution) during one-bottle training. In two-bottle tests with both flavors paired with an intermediate infusion (25%=net 17.5%), the rats initially showed no overall preference for the CS5 or CS30. Following additional training, the rats significantly preferred the CS30 to the CS5. The intragastric data suggested that a change in sweet taste context between training and testing might have accounted for the strong CS5 preference obtained in the first experiment. This was confirmed in a third experiment in which rats were trained with flavored 5% and 30% sucrose solutions and then given two-bottle tests with both flavors presented either in 5% sucrose or 30% sucrose. Rats tested with 30% sucrose strongly preferred the CS5 flavor, whereas rats tested with 5% sucrose significantly preferred the CS30 flavor. Thus, the outcome of two-bottle flavor preference tests and presumably other tests of conditioned flavor reward may be greatly influenced by the solutions used in the tests. The impact of this variable may be greatest when the training solutions do not substantially differ in their net postingestive reinforcing actions. This appears to be the case with 5% and 30% sucrose solutions because the satiating effect of the concentrated solution tends to counteract its nutrient reinforcing action.     

Muscarinic receptor antagonism of the nucleus accumbens core causes avoidance to flavor and spatial cues

Pharmacological blockade of muscarinic receptors in the nucleus accumbens reduces food intake and instrumental behaviors that are reinforced by food delivery. Nucleus accumbens muscarinic antagonism may specifically suppress the hedonic or reinforcing effects of food, thus blocking its capacity to direct behavior. Alternatively, muscarinic receptor blockade may cause a negative hedonic state that interferes with appetitive learning and food intake. In these experiments, rats received infusions of scopolamine methyl bromide (10 microg/0.5 microl) into the nucleus accumbens core, following exposure to a novel flavor of liquid diet (Experiment 1) or prior to being placed into a place preference apparatus (Experiment 2). In both experiments, nucleus accumbens muscarinic receptor antagonism caused subsequent avoidance of the paired cue (flavor or spatial location). This effect was specific to cholinergic manipulation; no conditioned taste avoidance was observed after pairing the novel flavor with nucleus accumbens core antagonism of N-methyl-D-aspartate, dopamine D-sub-1, or opioid receptors (Experiment 3). These experiments confirm previous reports of a critical role for striatal acetylcholine in modulating goal-directed behaviors, but suggest caution when interpreting behavioral effects of pharmacological manipulation of striatal acetylcholine.     

Effect of lithium chloride-induced aversion on appetitive and consummatory behavior

The effect of lithium chloride-induced conditioned taste aversions on appetitive and consummatory behavior was determined. Rats were given access to a 0.1% saccharin solution for 15 min either in bottles or by infusion through an intraoral cannula. Bottle-fed rats given postprandial injections of lithium chloride showed greater aversion to saccharin than cannula-fed rats. During extinction, cannula-fed rats gradually recovered to control levels of intake, whereas bottle-fed rats continued to avoid the saccharin. These results suggest that lithium chloride affects appetitive behavior to a greater extent than it affects consummatory behavior.     

Conditioned acceptance and preference but not altered taste reactivity responses to bitter and sour flavors paired with intragastric glucose infusion

Nutrient-conditioned flavors preferences are thought to involve an increase in flavor palatability (hedonic evaluation). Consistent with this view is the recent finding that a sweet flavor paired with intragastric glucose infusions elicited more hedonic taste reactivity (TR) responses than did an alternative sweet flavor paired with intragastric water. The generality of this finding was examined by conditioning preferences for inherently avoided nonsweet flavors. Rats were trained in 20 h/day and then 30 min/day sessions with a CS+ flavor (sour citric acid or bitter sucrose octaacetate) paired with intragastric 16% glucose infusion, and the opposite flavor (CS-) paired with intragastric water. Glucose conditioning increased the CS+ acceptance in one-bottle tests and produced a 95% CS+ preference in two-bottle sessions. Yet, TR responses to brief intraoral infusions of the two CS flavors did not differ, even after extensive testing. Subsequent choice tests revealed that a 1% fructose solution was preferred to the CS-, whereas the CS+ was preferred to 1% and 2% fructose and equally preferred to 4%, 8%, and 16% fructose. These results indicate that strong nutrient-conditioned flavor preferences are not always associated with increased flavor palatability as measured by TR tests. Therefore, nonhedonic processes, perhaps increased incentive salience, appear to mediate the enhanced preference and acceptance conditioned by postingestive nutrient actions.     

Ethanol-conditioned flavor preferences compared with sugar- and fat-conditioned preferences in rats

Rats can learn to prefer flavors paired with ethanol and various nutrients. The present study examined the relative strengths of flavor preferences conditioned by 5% ethanol and isocaloric solutions of 7.18% sucrose, 7.18% fructose, or 3.26% corn oil. In three experiments, nondeprived rats were trained with different flavored solutions (conditioned stimuli, CS) paired with intragastric (IG) infusions: a CS+E flavor paired with ethanol infusion, a second CS+ paired with a nutrient infusion, and a CS- paired with water infusion. In two-bottle tests, rats strongly preferred a sucrose-paired CS+S over the CS- and over the CS+E. The preference for the CS+E over CS- was weaker. These effects occurred when the rats drank substantially more CS+S than CS+E in training and when training intakes were matched. Similar results were obtained when the nutrient infusion was fructose or corn oil, except that preferences for the CS+F or CS+O over the CS+E were less pronounced than with CS+S. Consistent with the IG results, rats trained to drink flavored sucrose and ethanol solutions preferred the CS+S to CS+E in a flavored water test. These results confirm prior reports of ethanol-conditioned preferences but show that ethanol is less effective than other nutrients at isocaloric concentrations. The marked individual differences in ethanol-conditioned preferences may be related to the impact of the sugar or fat infusions on the reward evaluation of the ethanol-paired flavor.     

Conditioned flavor preference learning by intragastric administration of l-glutamate in rats

The preference for foods or fluids in rats is partly dependent on its postingestive consequences. Many studies have investigated postingestive effect of high caloric substances, such as carbohydrate or fat. In this study, we examined postingestive effect of l-glutamate at the preferable concentration using conditioned flavor preference paradigm. Adult male rats with chronic intragastric (IG) cannula were trained to drink a flavored solution (conditioned stimulus; CS+) paired with IG infusion of nutrient solution and another flavored solution (CS−) with IG distilled water infusion on alternate days. The nutrient solution was 60 mM monosodium l-glutamate, sodium chloride or glucose. Before and after conditioning, rats received 30 min two-bottle choice tests for CS+ and CS− solution. All groups exhibited no significant preference for CS+ in pre-test period. By the last half of conditioning period, intake of CS+ solution was significantly higher than that of CS− in MSG group, but not in NaCl and glucose groups. After conditioned, the MSG group showed significantly higher intake and preference for CS+ solution (69.9%), while the NaCl and glucose group did not show any significant intake and preference for CS+ solution (50.9%, 43.5%, respectively). These results indicate that the amino acid l-glutamate at a preferable concentration has a positive postingestive effect as demonstrated by its ability to condition a flavor preference. The mechanism(s) for this positive effect could be through a direct effect on gut Glu receptors rather than the provision of calories or glucose from metabolized Glu; Further studies are needed to test these hypotheses.     

The amygdala and flavour preference conditioning: Crossed lesions and inactivation

Current studies examined whether temporary inactivation of the amygdala influenced the learning and/or expression of conditioned flavour preferences and whether interactions between the amygdala and the nucleus accumbens contribute to this learning. Experiments 1A and 1B examined temporary inactivation of the amygdala in rats, by the administration of muscimol through chronically implanted cannulae, given during acquisition and/or expression of flavour preferences based on a sucrose reinforcer. Despite differences in the number of training trials and control procedures, in both of Experiments 1A and 1B inactivation during training attenuated, but did not totally prevent, the acquisition of a preference for the CS+ (conditioned stimulus) flavour over the CS−. Inactivation during testing had no effect on the preference for the CS+. In Experiment 2A rats were given access to a CS+ flavour paired with fructose and a CS− flavour without fructose prior to testing the preference for the CS+ over the CS− in the absence of the reinforcer. In Experiment 2B the same rats were tested for their preference with another set of CS+ and CS− flavours and maltodextrin as the reinforcing solution. Contralateral unilateral lesions of the amygdala and nucleus accumbens attenuated, but did not totally prevent, flavour preference learning based on either fructose or maltodextrin compared to either ipsilateral or sham lesioned animals. These results suggest that the amygdala plays a role in the learning, but not expression, of flavour preferences and that this role is partially dependent on interactions with the nucleus accumbens.     

Oral and postoral determinants of food reward

Tasty high-fat and high-sugar foods induce overeating and obesity in animals. To separate the roles of oral and postoral factors in food preference and consumption, recent studies have used an "electronic esophagus" preparation. With this system, when an animal drinks a flavored solution, it controls the infusion of nutrients directly into its stomach through an implanted catheter. Improving the palatability of the flavored solution increases solution intake, infused nutrient intake, and weight gain. Changing the composition of the infused nutrient (e.g., fat-carbohydrate ratio) can also alter energy intake when flavor cues are held constant. In addition, animals learn to associate flavor cues with the postoral effects of nutrients, which can significantly increase flavor preference and acceptance. In some but not all cases, conditioned preferences are associated with increased hedonic evaluation of the nutrient-paired flavor. Flavor preference conditioning by nutrients can occur in chow-sated animals and is not dependent upon an energy-depleted state. Thus, food reward is determined by oral and postoral stimuli and may override the homeostatic regulation of energy balance.     

Age-related differences in avoidance behavior in rats following CS preexposure

Weanling and adult rats were exposed to either 0 or 10 conditioned stimulus (CS) presentations prior to 1-way active-avoidance (AA) training. Although CS exposure retarded avoidance acquisition in the adults, it produced no effect in the pups during avoidance learning. Neither adult nor young rats demonstrated preexposure effects in extinction. A general extinction analysis showed that pups had less resistance to extinction than the adults, despite a comparable avoidance learning criterion between age groups. The lack of preexposure influences on performance by the pups was compared to previous findings of response inhibitory deficits in immature rats. The results were considered in light of selective attention interpretations of latent inhibition.     

Nutrient-conditioned flavor preference and incentive value measured by progressive ratio licking in rats

Rats develop strong preferences for flavors associated with the postingestive actions of glucose and other nutrients. This preference may involve changes in both the hedonic appeal and incentive salience of the nutrient-paired flavor. The present study used a progressive ratio (PR) operant licking task to evaluate the degree to which nutrient conditioning changes the incentive value of flavors. Food restricted rats were trained to associate one flavored saccharin solution (CS+) with intragastric glucose infusions and another flavored solution (CS-) with water infusions. The rats subsequently showed a strong preference for the CS+ in two-bottle tests and also licked more for the CS+ than CS- in PR tests. PR licking for the CS+ was similar to that for an 8% fructose solution. Together with earlier data indicating that the CS- is isopreferred to a 3% fructose solution, these findings indicate that IG glucose conditioning enhances the hedonic and incentive value of the CS+ solution so that the animal responds as if the solution had a sweeter taste.     

Generalization of appetitive conditioned responses

A stimulus (conditioned stimulus, CS) associated with an appetitive unconditioned stimulus (US) acquires positive properties and elicits appetitive conditioned responses (CR). Such associative learning has been examined extensively in animals with food as the US, and results are used to explain psychopathologies (e.g., substance‐related disorders or obesity). Human studies on appetitive conditioning exist, too, but we still know little about generalization processes. Understanding these processes may explain why stimuli not associated with a drug, for instance, can elicit craving. Forty‐seven hungry participants underwent an appetitive conditioning protocol during which one of two circles with different diameters (CS+) became associated with an appetitive US (chocolate or salty pretzel, according to participants’ preference) but never the other circle (CS?). During generalization, US were delivered twice and the two CS were presented again plus four circles (generalization stimuli, GS) with gradually increasing diameters from CS? to CS+. We found successful appetitive conditioning as reflected in appetitive subjective ratings (positive valence, higher contingency) and physiological responses (startle attenuation and larger skin conductance responses) to CS+ versus CS?, and, importantly, both measures confirmed generalization as indicated by generalization gradients. Small changes in CS‐US contingency during generalization may have weakened generalization processes on the physiological level. Considering that appetitive conditioned responses can be generalized to non‐US‐associated stimuli, a next important step would be to investigate risk factors that mediate overgeneralization.     

Reinforcer-specificity of appetitive and consummatory behavior of rats after Pavlovian conditioning with food reinforcers

We examined the reinforcer-specificity of Pavlovian conditioning in the control of appetitive and consummatory behaviors in Pavlovian-to-instrumental transfer, cue-potentiated eating, and devaluation procedures. Rats received pairings of one conditioned stimulus with sucrose and another conditioned stimulus with maltodextrin. In Experiment 1, rats were also trained to earn sucrose for one instrumental response and maltodextrin for another. In a transfer test, the Pavlovian cues enhanced the rate of instrumental responding more when the food reinforcer predicted by the instrumental response and the Pavlovian cue were consistent than when they were inconsistent, but both cues enhanced both responses. In Experiment 2, sated rats' consumption of each food was potentiated in the presence of a cue for that food, but not in the presence of a cue for the other food. In Experiment 3, one food was devalued by pairing it with lithium chloride, prior to testing food consumption and food-cup directed behaviors. The food cues selectively controlled food-cup related behaviors, regardless of the presence of the devalued or nondevalued foods in the food cup. Together, these results are consistent with the view that conditioned cues modulate appetitive and consummatory behaviors with increasing levels of specificity. The closer an action comes to ingestion, the more it is controlled by sensory properties conveyed by learned cues. These data are discussed in the context of allostatic regulation of food foraging and intake.     

Flavor preferences conditioned by intragastric polycose infusions: a detailed analysis using an electronic esophagus preparation

The present series of experiments examined the rat's conditioned preference for flavors associated with intragastric (IG) Polycose infusions. Adult female rats were fitted with two chronic intragastric catheters and were trained to drink flavored water (CS+; e.g., cherry-water) paired with IG infusions of 32% Polycose. On alternate days a different flavor (CS-; grape-water) was paired with IG water infusions. The flavored water and chow were available 23 hr/day. In subsequent two-choice tests the rats displayed strong preferences for the CS+ flavor (up to 98%). The CS+ preference persisted for several weeks during extinction tests when both the CS+ and CS- were paired with IG water or with no infusions. The rats also preferred the CS+ to plain water which contrasts with the mild aversion naive rats display to the flavored water. The acquired preference for the CS+ flavor was not as strong, however, as the rats' innate preferences for the taste of saccharin or Polycose. Also, unlike their response to saccharin and Polycose, the rats' acceptance (absolute intake) of the CS+ flavor was not elevated. CS+ intake was increased, though, when saccharin was added to the flavored water. The rats reversed their flavor preference when the reinforcement contingencies were reversed. Also, in the absence of unique flavor cues, the rats learned to prefer, apparently based on somatosensory cues, the sipper tube that was paired with IG Polycose infusions. The effects of flavor and IG Polycose infusions on drinking patterns and caloric intake are also described.     

Cerebral glucose utilization during conditioned sexual arousal.

Local cerebral glucose utilization was investigated in male rats during conditioned sexual arousal. Increased glucose utilization was found in three amygdaloid nuclei after exposure to a stimulus associated with exposure to a sexually active female. No changes were observed in areas known to be of crucial importance for the expression of consummatory aspects of sexual behavior. These results corroborate and extend previous results showing a dissociation between the expression of appetitive and consummatory aspects of sexual behavior at a neural level.     

Flavor preferences conditioned by high-fat versus high-carbohydrate diets vary as a function of session length.

Intragastric (i.g.) infusions of fat and carbohydrate condition flavor preferences in rats, but different results have been obtained in studies using pure and mixed nutrient infusions. This experiment compared the preference conditioning effects of mixed high-carbohydrate (HC) and high-fat (HF) diet infusions during short-term and long-term sessions. In Experiment 1 food-deprived rats were given one flavored saccharin solution (CS+HC) paired with i.g. infusions of an HC liquid diet, a second flavor (CS+HF) paired with HF diet infusions, and a third flavor (CS-) paired with i.g. water infusions during 30-min one-bottle training sessions. In subsequent two-bottle tests (30 min/day), the rats preferred both CS+s to the CS- and preferred the CS+HC to the CS+HF. In Experiment 2, the same rats were trained and tested with the CSs and paired infusions during 22 h/day sessions with chow available ad lib. Both CS+s were again preferred to the CS-, but now the CS+HF was preferred to the CS+HC. When given additional 30-min choice sessions in Experiment 3 the rats showed no reliable preference for the CS+HC versus CS+HF under food-deprived or ad lib conditions. In Experiment 4, the rats were given 22-h CS+HC versus CS+HF choice sessions every other day. They showed no reliable CS preference during the first 30 min of each session, but reliably preferred the CS+HF during the remaining 21.5 h. These findings indicate that previously reported differences in preferences conditioned by pure versus mixed nutrient infusions are due to training procedures (session length, deprivation state) rather than to the type of nutrient infusions per se. The rats displayed different CS+HF versus CS+HC preferences as a function of test duration even after being given both short- and long-term training. Thus, short-term choice tests do not always predict the long-term intakes and preferences for high-fat and high-carbohydrate foods.     

The influence of natural preference for tactile stimuli on appetitive learning in rat pups

Stimuli of the same modality tend to be organized along a “natural preference scale.” This study examined the ability of six- and nine-day-old rat pups to acquire appetitive learning, when the CS was one of two differently “naturally preferred” tactile stimuli (floor textures: rug and plywood). In Experiment 1, all pups showed a relative natural preference for the rug texture over the plywood texture. Pups conditioned on the plywood texture (exposed to a sibling pup as the US) showed a robust increase in preference for the conditioned texture whereas pups conditioned on the rug texture showed a different and more moderate pattern of acquisition. Developmental differences were found only in extinction of the conditioned response: six-day-old (but not nine-day-old) pups displayed extinction of the response over four trials. Experiment 2 indicated that preexposure to the rug CS prior to conditioning (latent inhibition) did not interfere with the learning process on the rug texture. Two major alternative explanations for the differential learning patterns are discussed: motivational and catecholaminergic influences on learning. The results suggest that natural preferences may modulate early learning and memory processes. © 1997 John Wiley & Sons, Inc. Dev Psychobiol 30: 29–30, 1997     

Flavor preferences conditioned by postingestive effects of nutrients in preweanling rats

The purpose of these experiments was to determine if preweanling rats, like adults, are capable of learning to associate an arbitrary flavor with the postingestive effects of nutrients, and then demonstrate a preference for that flavor after weaning. In Experiment 1, preweanlings were trained daily from postnatal day (P) 16 through P19 with intraoral (IO) infusions of a grape or cherry flavor (CS+) mixed with 20% glucose (US), and the opposite flavor (CS-) mixed with 0.05% saccharin. After weaning, rats were given a 4-h two-bottle choice between the CS+ and CS- flavors both presented in 0.05% saccharin. Rats preferred the flavor previously paired with glucose. In Experiment 2 using similar methods, rats learned to prefer a flavor (CS+G) paired with a glucose US over a flavor (CS+S) paired with a sweeter but less nutritive sucrose US, indicating involvement of postingestive reinforcement. In Experiment 3 preweanling rats with IO and intragastric (IG) catheters were trained with a CS+ flavor paired with IG nutrient infusion, and a CS- flavor paired with no IG infusion. These rats showed no flavor preference 3 days after weaning, but did significantly prefer the CS+ flavor over the CS- flavor 10 days after weaning. Together these experiments demonstrate that neural mechanisms for flavor-nutrient associations are developed before weaning, allowing young rats to learn associations between arbitrary flavors and nutritive consequences. Thus nutrient conditioning may be one way that early experience (such as flavors from the maternal diet transmitted in milk) influences later dietary preferences.     

Conditioned enhancement of flavor evaluation reinforced by intragastric glucose: I. Intake acceptance and preference analysis.

Rats acquire strong preferences for flavors that are paired with intragastric (IG) glucose infusions. The present study assessed the magnitude of this nutrient-conditioned change in flavor evaluation by comparing the strength of the rats' conditioned preference to their unconditioned preference for the sweet taste of sugar. Food-restricted rats were first trained (20 h/day) to associate one flavor (CS+) with IG 16% glucose infusions and another flavor (CS-) with IG water infusions. Then, in a series of 30 min/day two-bottle tests, the CS+ and CS- were separately offered in a choice vs. fructose of varying concentrations. The CS- was preferred to 2% fructose, but not 4% fructose. The CS+ was preferred to 2%, 4%, and 8% fructose and was isopreferred to 16% fructose. This CS+ vs. fructose preference profile was replicated and found to be independent of deprivation state. The intake-stimulating efficacy of the CS+ flavor and of fructose taste was also compared in one-bottle sham-feeding tests. Intakes of CS+ and of 16% fructose increased by similar amounts in the first sham-feeding test. However, with repeated testing, fructose intake continued to increase, whereas CS+ intake declined. This profile suggests extinction of the conditioned satiation response to fructose and extinction of the conditioned acceptance response to the CS+, respectively. These findings provide novel evidence for a robust learned shift in flavor evaluation reinforced by the postingestive actions of glucose.