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Neuroscience Program and Department of Psychology, University of Western Ontario, London, Ontario N6A 5C2, Canada
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ABSTRACT |
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 TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
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The differential effects of CCK and lipopolysaccharide (LPS) on sucrose intake and palatability were examined. Rats were injected with LPS (200 µg/kg ip) or NaCl (0.9%, vehicle) and 2 h later received a second injection of either CCK (8 µg/kg ip) or NaCl. In experiment 1, sucrose (0.3 M) intake was monitored for 1 h on three different test days 72 h apart, while in experiment 2, palatability was assessed by means of the taste reactivity test (TRT) on two separate days (72 h apart). In the TRT, orofacial and somatic responses to brief (30 s) intraoral infusions of sucrose were recorded and analyzed for response frequency. Singly, LPS and CCK reduced sucrose intake, with a more pronounced effect from combined LPS and CCK. LPS by itself did not alter sucrose palatability, as evidenced by continuous high levels of ingestive responding. In contrast, CCK-treated rats displayed a pattern of responding indicative of satiety, as did the combined LPS-CCK-treated rats. These results suggest that LPS does not induce hypophagia by altering palatability.
taste reactivity test; endotoxin; acute phase response; anorexia; food intake
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INTRODUCTION |
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TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
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BACTERIAL LIPOPOLYSACCHARIDES (LPSs) are the active components of the cell walls of gram-negative bacteria and are known to induce many of the symptoms associated with the acute phase response (25). These symptoms include fever, lethargy, and hyperalgesia, as well as reductions in activity, grooming, and feeding (15, 16). Initiation of the host's immune response by LPS occurs with the production of proinflammatory cytokines, including interleukin (IL)-1, IL-6, and tumor necrosis factor (TNF) (8). Results of several studies have demonstrated a pronounced reduction in food intake after peripheral administration of LPS (e.g., Refs. 18, 19, 21-23). However, the mechanism(s) by which LPS induces hypophagia are still poorly understood.
It has been suggested that the discomfort and illness resulting from activation of the immune system could induce the formation of a conditioned taste aversion (CTA) (e.g., Refs. 19, 37). Langhans et al. (19) have shown that association of the effects of LPS with a novel saccharin taste results in a pronounced reduction in the subsequent preference for the saccharin. More recently Goehler et al. (13) showed that pairing an IL-1 injection with exposure to a novel saccharin taste produces a CTA to the saccharin taste. As well, peripheral administration of TNF has been shown to produce a strong conditioned aversion to a novel diet, an effect that was significantly reduced in rats with area postrema lesions (2). Area postrema lesions have been shown to abolish or attenuate CTAs induced with a variety of toxic agents (30). However, in some CTA paradigms, such lesions have been shown to enhance the subsequent aversions (e.g., Ref. 31). In this context it is interesting to note that lesioning of the area postrema and the adjacent nucleus of the solitary tract was found to augment LPS-induced hypophagia (39). Another study found that LPS could only induce a CTA under certain conditions, which included a novel tasting diet and no food deprivation before testing (39). Thus issues of whether LPS is able to induce a CTA and whether the formation of a CTA is responsible for the observed hypophagia remain to be clarified.
CCK is an endogenous gut-brain hormone that is involved in the physiological control of feeding in a variety of species, including rats and humans (24). Systemic administration of CCK has been shown to produce a dose-dependent reduction in food intake within minutes of administration (e.g., Ref. 11). CCK is known to reduce food intake via satiety-related mechanisms rather than the production of aversive internal cues (9, 34). For instance, it has been shown that rats with open gastric fistulas reduce sham feeding after administration of CCK (12), as well as exhibit stereotypical behaviors associated with satiety (1).
There is increasing evidence suggesting that CCK may play a role in immune system-to-brain communication and that CCK is involved in mediating the neural effects of the peripherally activated immune system. Bucinskaite and colleagues (5) have found an increased sensitivity of gastric vagal afferent activity to CCK after administration of IL-1 in the rat. In addition, the blockade of CCK type A receptors resulted in an attenuation of this IL-1-induced increase in gastric vagal afferent activity (17). An in vitro study found an increased release of CCK from isolated hypothalamic cells after exogenous administration of IL-1 (29). It was also shown that CCK antagonists significantly attenuated an LPS-induced fever in rats (36). However, more importantly, Daun and McCarthy (7) found that activation of the immune system via IL-1 resulted in significantly elevated levels of plasma CCK within 1 h, reduced food intake, and a decrease in gastric emptying. Furthermore, they found that pretreatment with a CCK receptor antagonist significantly attenuated the reduction in food intake and gastric stasis, clearly suggesting that CCK may mediate the hypophagia observed during the acute phase response. The exact mechanisms and interactions between immune activation and CCK still need to be elucidated.
The taste reactivity test (TRT), developed by Grill and Norgren (14), differs from traditional intake measures in that it provides a behavioral measure of the palatability of an infused tastant. The procedure involves quantifying the orofacial and somatic responses of a rat to an intraorally infused tastant. The species-specific behaviors produced by the animal are videotaped and later analyzed for frequency of occurrence. The stereotypical behaviors are clustered into three distinct categories (14, 30). First, ingestive responses typically occur during infusions of palatable substances such as sucrose and facilitate the ingestion of a tastant. Second, active aversive responses, which are normally produced during infusions of an unpalatable taste, such as quinine, aid in the removal of the tastant from the oral cavity. Third, the passive aversive response (passive drip), is normally indicative of satiety and occurs when the rat simply lets the infused tastant passively fall from the oral cavity (14, 30).
The TRT is useful in that it allows for a detailed examination of shifts in palatability and the real-time rapid formation of CTAs. This procedure has been used successfully to track within-session changes in palatability of a sucrose solution after exposure to LiCl and the development of a CTA to the sucrose (e.g., Refs. 4, 10, 30, 35). The TRT also has been used as a tool for deciphering subtle differences between satiety and aversion, differences that cannot be distinguished using the traditional intake measures (9). By means of the TRT, it has been shown that exogenous administration of CCK results in satiety as opposed to aversion (9).
The present study systematically examined the effects of LPS and CCK, alone and in combination, on sucrose intake. Previous research, based on traditional intake measures, has shown that LPS and CCK reduce sucrose intake on their own. However, because CCK has been suggested to be a mediator of the hypophagia experienced during illness, the effects of both agents in combination was also examined. To gain additional information about the mechanisms involved in LPS-induced hypophagia, this study also examined the effects of LPS and CCK on sucrose palatability by means of the taste reactivity paradigm. If LPS administration induces aversive internal cues, we would predict the development of a CTA to the sucrose taste as evidenced by a decrease in ingestive responses and an increase in active aversive responses, reflecting a shift in sucrose palatability. In contrast, if LPS reduces food intake by signaling satiety, we would expect to see reductions in ingestive responses and increases in passive drips but no increases in active aversive responses.
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MATERIALS AND METHODS |
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TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
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Animals
Sixty-four naive, male adult Long Evans hooded rats (Charles River, Quebec), weighing between 325 and 375 g at the start of each experiment, were used as subjects. The rats were housed individually in either polypropylene cages (experiment 1) or stainless steel wire mesh cages (experiment 2) and were kept in a temperature-controlled room (21 ± 1°C) under a 12:12-h light-dark cycle (lights on between 0700 and 1900). Rat chow (Agway) and tap water were available ad libitum unless otherwise specified. Different animals were used for each experiment (n = 32 for each experiment).Drug Administration Schedule
On each of the test days (both experiments 1 and 2), each rat received two separate intraperitoneal injections. The first injection was either LPS (200 µg/kg, from Escherichia coli 0111:B4, L-2630; Sigma, St. Louis, MO) that was dissolved in 0.9% saline or a control injection of the 0.9% saline vehicle (NaCl, 1 ml/kg). Studies that have examined the effects of LPS on food intake have used doses ranging from 50 to 1,000 µg/kg (20, 41). The particular dose of LPS used here (200 µg/kg) was chosen to ensure that the previously reported aversive effects induced by LPS on behavior would be observed. After this first injection, animals were returned to their home cages for a period of 2 h, during which there was no access to food or water. This interval was chosen on the basis of prior results showing that treatment with LPS produced several behavioral effects, including hypophagia, within 2 h of peripheral administration (19). The second injection consisted of either the synthetic sulfated octapeptide form of CCK (8 µg/kg, Sincalide; Squibb, Princeton, NJ) dissolved in 0.9% saline, or the 0.9% saline vehicle (1 ml/kg). This dose of CCK has been shown previously to induce a reasonable suppression in ingestive responding (9). Immediately after the second injection (CCK or saline) rats were tested for sucrose intake (experiment 1) or sucrose palatability (experiment 2). A 0.3 M sucrose solution was chosen as it has been shown to elicit high levels of ingestive responding (30). Thus four experimental groups were examined (n = 8/group) in each of the two experiments: NaCl-NaCl, NaCl-CCK, LPS-NaCl, and LPS-CCK, with group designation denoted by the first and second injections, respectively, that an animal received before behavioral testing.Procedures
Experiment 1. Rats were food and water deprived for 24 h before each of three test days. On the first test day, rats were randomly assigned to one of four experimental groups. Immediately after the second injection (either CCK or NaCl), the animals were returned to their home cage, in which a graduated drinking tube containing 0.3 M sucrose solution was available, and cumulative fluid intake was measured (±0.5 ml) after 5, 10, 20, 30, 40, 50, and 60 min. Immediately after the intake determination, food and water were again made available. Test days 2 and 3 involved exactly the same injection and handling procedures, and all test days were separated by a 72-h interval. Body weights were recorded immediately before each testing period and then 24 h later to determine any change in body weight. All testing was carried out between 0900 and 1200.Experiment 2. INTRAORAL CANNULATION. One week after arriving in the laboratory all of the rats received surgical implantation of intraoral cannulas according to the procedure of Parker (32). After 24 h of food deprivation animals were anesthetized by an intraperitoneal injection of pentobarbital sodium (Somnotol, 50 mg/kg ip). A 15-gauge stainless steel needle was inserted through the rat's skin in the dorsal midneck region and was guided subcutaneously below the ear and along the cheek, where it exited the mouth just rostral to the first maxillary molar. Once the needle was in place, a 10-cm piece of polyethylene tubing (PE-90) was threaded through the barrel of the needle, and the needle was then removed. The tubing was secured at the neck with a 20-gauge intramedic adapter cap, and in the mouth by a smooth plastic washer (5 mm in diameter), which was held in place by heat flaring the end of the tubing with a soldering iron. The skin around the puncture sites was swabbed with alcohol, and the rats were given 3 days to recover from surgery before testing began. Throughout the entire experiment, the cannulas were flushed daily with distilled water to prevent blockage.
TASTE REACTIVITY TEST CHAMBERS. TRTs were conducted in transparent Plexiglas chambers (29 × 25 × 29 cm). At the start of each testing period, rats were placed individually in a testing chamber, and intraoral infusions were delivered via an infusion hose (PE-90 tubing, 1 m in length) attached to the rat's cannula and to an infusion pump (model 341-A; Sage Instruments, Cambridge, MA). Infusions were delivered at a constant rate of 0.78 ml/min. A mirror was mounted beneath the transparent floor of the chamber at a 45° angle to facilitate videotaping the facial region (ventral view) of the rat. The orofacial and somatic responses of the rat, elicited during the intraoral infusions, were videotaped with a video camera (Panasonic AG-195; London, ON, Canada), located ~1 m from the mirror, and a videocassette recorder (Panasonic AG-1970).
TASTE REACTIVITY TEST PROCEDURE. All rats were habituated to the general testing procedure on three consecutive days before the first test day. During this habituation phase animals were placed in the testing chamber for 15 min followed by a 1-min infusion (0.78 ml/min) of distilled water.
On each of the two test days, rats received two separate intraperitoneal injections as described previously (NaCl-NaCl, NaCl-CCK, LPS-NaCl, and LPS-CCK). Immediately after the second injection (CCK or NaCl), the rats were placed individually in the TRT chamber and received a series of 30-s intraoral sucrose infusions at 0, 3, 6, 9, and 12 min post-second injection. Behaviors displayed during each intraoral infusion were video recorded. Each rat received two identical test days separated by 72 h. Body weight was recorded immediately before each test session and then again 24 h later to note changes. All testing occurred between 0900 and 1200.
BEHAVIORAL SCORING. Once the experiment was completed, videotapes of the behaviors occurring during the TRTs were examined in slow motion (one-fifth the regular speed) to facilitate the scoring of the discrete taste reactivity responses. The specific behaviors, grouped into three main categories, ingestive, aversive, and passive aversive, are described by Grill and Norgren (14) and elaborated on by Ossenkopp and Eckel (30). The ingestive composite score consists of the combined frequency of tongue protrusions (which include both midline and lateral extensions of the tongue) and rhythmical mouth movements (small-amplitude movements of the mouth without the extension of the tongue). The aversive composite score consisted of the combined frequency of four behaviors: head shakes with fluid expulsion, forelimb flails, chin rubs, and gapes. The passive-aversive response consisted of the frequency of passive drips (number of fluid drops).
Data Analysis
The data from both sets of experiments were analyzed with a mixed-design ANOVA procedure. Post hoc examinations of significant main effects and interactions were done with Tukey's honestly significant difference test. All hypothesis tests used
= 0.05 as the criterion
for significance.
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RESULTS |
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TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
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Experiment 1
Body weight. Body weight analysis was carried out with a two-factor mixed-design ANOVA with treatment as the between-subjects factor (at four levels: NaCl-NaCl, NaCl-CCK, LPS-NaCl, and LPS-CCK) and test day as the within-subjects factor (at three levels: test days 1, 2, and 3). The changes in body weight across a 24-h period (after 24 h food and water deprivation) for all four experimental groups are depicted in Fig. 1. Significant main effects of treatment [F(3,20) = 21.98, P < 0.001] and day [F(2,56) = 152.53, P < 0.001] were observed. There was also a significant treatment × day interaction [F(6,56) = 13.36, P < 0.001]. Post hoc analysis revealed that both of the groups of rats that were injected with LPS (LPS-NaCl and LPS-CCK) showed a significant decrease in body weight (P < 0.05) relative to the other two groups (NaCl-NaCl and NaCl-CCK) after the first test day. After the second test day all groups exhibited an increase in body weight, with the LPS-NaCl group displaying a significantly smaller increase (P < 0.05) than all other groups. By the third test day, all animals showed similar increases in body weight over the 24-h period.
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Sucrose intake. Statistical analysis
of cumulative sucrose intake was conducted with a three-factor
mixed-design ANOVA with treatment as the between-subjects factor (at
four levels: NaCl-NaCl, NaCl-CCK, LPS-NaCl, and LPS-CCK) and both time
(at 7 levels: t = 5, 10, 20, 30, 40, 50, and 60 min) and test day (at 3 levels: test
days
1, 2,
and 3) as within-subjects factors.
Analysis of cumulative sucrose intake across the test hour and across
the days of testing (see Fig. 2,
A, B,
and C) yielded significant main
effects of treatment [F(3,28) = 20.06, P < 0.001], day
[F(2,56) = 32.21, P < 0.001], and time
[F(6,168) = 57.19, P < 0.001]. There was both a significant treatment × day
interaction [F(6,56) = 4.72, P < 0.001] and treatment × day × time interaction
[F(36,336) = 2.37, P < 0.001]. Post hoc
comparisons of group mean total sucrose intake over the test days (Fig.
3) revealed that the control rats (NaCl-NaCl) consumed significantly more sucrose
(P < 0.05) than all other groups on
the first test day, with LPS-CCK-treated rats consuming significantly
less sucrose (P < 0.05) than both
LPS-NaCl and NaCl-CCK groups. However, on the second test day, the
NaCl-NaCl rats did not differ significantly from the LPS-NaCl rats in
the total amount of sucrose consumed, but the NaCl-NaCl group did drink
significantly more sucrose (P < 0.05) than both the CCK-treated groups (NaCl-CCK and LPS-CCK). On the
final test day, both NaCl-NaCl and LPS-NaCl groups consumed
significantly more sucrose (P < 0.05) than did the two groups of rats that received CCK.
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Experiment 2
Body weight. The change in body weight was analyzed by means of a two-factor mixed-design ANOVA with treatment as the between-subjects factor (at 4 levels) and test day (at 2 levels: test days 1 and 2) as the within-subjects factor. Analysis of body weight changes after administration of LPS and/or CCK across the two test days (see Fig. 4) revealed significant main effects of treatment [F(3,30) = 28.07, P < 0.01] and day [F(1,30) = 37.25, P < 0.01] and a significant treatment × day interaction [F(3,30) = 14.15, P < 0.01]. Both experimental groups that were treated with LPS displayed significantly greater (P < 0.05) decreases in body weight after the first test day relative to the other two groups (NaCl-NaCl and NaCl-CCK). However, these differences were not obtained after the second test day.
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Taste reactivity responses. Only the NaCl-CCK-treated group displayed any aversive responses (head shakes, forelimb flails, chin rubs, or gapes), and these responses were too infrequent to warrant any statistical analyses. The taste reactivity responses (ingestive composite score as well as each individual behavior alone) were analyzed by means of a three-factor mixed-design ANOVA with treatment as the between-subjects factor (at 4 levels) and both infusion time (at 5 levels: 0, 3, 6, 9, and 12 min) and day (at 2 levels) as within-subjects factors.
Ingestive composite score. Group mean
frequencies of total ingestive responses (mouth movements and tongue
protrusions) elicited by the intraoral sucrose infusions on both test
days are depicted in Fig. 5. Statistical
analysis revealed significant main effects of treatment
[F(3,28) = 17.52, P < 0.001] and
time [F(4,112) = 4.74, P = 0.01], but not day. A
significant treatment × time interaction [F(12,112) = 2.73, P < 0.005] was obtained. Post
hoc analysis showed that on the first test day, LPS-NaCl and NaCl-NaCl
rats did not differ significantly in the total number of ingestive responses at any point during testing. However, the LPS-NaCl and NaCl-NaCl groups did exhibit significantly
(P < 0.05) higher levels of
ingestive responding than did both groups of rats that received CCK
injections. On the second test day, rats treated with NaCl-CCK displayed significantly (P < 0.05)
fewer ingestive responses relative to all other groups at several time
points (t = 6, 9, and 12 min).
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Tongue protrusions. The frequency of
tongue protrusions across the two test days for each of the four
experimental groups is shown in Fig. 6.
Analysis of tongue protrusion data yielded a significant main effect of
treatment [F(3,28) = 19.42, P < 0.001] and day
[F(1,28) = 5.89, P < 0.025]. A significant
treatment × day interaction
[F(3,28) = 3.34, P < 0.05] as well as a
treatment × time interaction
[F(12,112) = 4.13, P < 0.001] were also obtained. Post hoc analysis revealed that on the first test day the two groups
that had received CCK (NaCl-CCK and LPS-CCK) displayed significantly
(P < 0.05) fewer tongue protrusions
relative to the other two groups (NaCl-NaCl and LPS-NaCl) at several
time points (t = 3, 6, and 9 min).
Again, on the second test day the NaCl-NaCl and LPS-NaCl groups
exhibited significantly (P < 0.05) higher levels of tongue protrusions than the two CCK-treated groups (P < 0.05). Inspection of Fig. 6
suggests a trend for the LPS-NaCl rats to display increased number of
tongue protrusions relative to the NaCl-NaCl group. This was supported
by post hoc comparisons that revealed significant differences
(P < 0.05) at both
time 0 and
9 min.
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Mouth movements. Analysis of the
frequency of mouth movements (see Fig. 7)
revealed significant main effects of treatment [F(3,28) = 6.34, P < 0.01], day
[F(1,28) = 4.45, P < 0.05], and time
[F(4,112) = 6.99, P < 0.001]. The only
significant interaction was time × day
[F(4,112) = 4.53, P < 0.002]. Post hoc
comparisons revealed that LPS-CCK rats displayed significantly
(P < 0.05) more mouth
movements relative to NaCl-NaCl rats at the start of the first test day
(t = 0 and 3 min). On the second test
day LPS-CCK-treated rats showed significantly
(P < 0.05) higher levels of mouth
movements than all other groups (LPS-NaCl, NaCl-CCK, and NaCl-NaCl) at
several time points (t = 0, 3, and 9 min).
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Passive drip. The frequency of the
passive aversive response, passive drip, is shown in Fig.
8. Statistical analysis revealed significant main effects of treatment
[F(3,28) = 5.96, P < 0.005] and time
[F(4,112) = 5.64, P < 0.001] as well
as significant treatment × time
[F(12,112) = 3.03, P < 0.001] and treatment × time × day [F(12,112) = 2.06, P < 0.025] interactions. Post
hoc analysis revealed that on the first test day LPS-CCK-treated
animals produced significantly (P < 0.05) more passive drips than both LPS-NaCl and NaCl-NaCl groups at
several times (t = 3 and 6 min).
However, on the second test day the NaCl-CCK-treated rats exhibited
significantly higher number of passive drips relative to LPS-NaCl and
NaCl-NaCl rats (at t = 6, 9, and 12 min).
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DISCUSSION |
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TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
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Experiment 1
The present findings show that both LPS and CCK, when administered alone, significantly reduce sucrose intake. The results also revealed that tolerance develops to the effects of LPS, but not to those of CCK, and that there is an interactive effect of LPS and CCK. Rats treated with both LPS and CCK showed a more pronounced reduction in sucrose intake than all other groups after the first test day.Results of previous studies have reported reductions in food intake and food-motivated behavior after LPS administration at doses and time courses that are comparable to those used in the present experiment (e.g., Ref. 18). The present observation, that LPS-treated rats showed decreased levels of sucrose intake relative to saline-treated animals, is also consistent with the work of Yirmiya (41), demonstrating similar reductions in nonnutritive saccharin consumption, a result hypothesized to reflect anhedonia or a disinterest in pleasure. The present finding that sucrose intake was reduced after CCK treatment is also consistent with previous reports of reduced food intake in animals treated with CCK (11). It has been suggested that these pronounced effects of CCK result from satiety rather than reflecting an aversive effect of this hormone (11).
The present study is, to date, the first examination of the interactive effects of LPS and CCK on sucrose intake. The results show that on the first test day, rats treated with both LPS and CCK exhibited a greater reduction in sucrose intake than after either agent alone. It is difficult to distinguish whether this reduction is a result of an additive effect of LPS and CCK or a potentiation, because the LPS-CCK rats consumed almost no sucrose on the first test day. Further studies using subthreshold doses of LPS and CCK are necessary to characterize the nature of the interaction that occurs after systemic administration of LPS and CCK.
The rats treated with CCK alone showed reduced levels of sucrose intake across all days of testing. This is consistent with previous findings, which have demonstrated that CCK-treated rats do not exhibit any apparent tolerance to the anorexic effects of CCK with repeated exposures (40). In contrast, LPS-treated rats increased their sucrose intake after subsequent exposures to LPS, suggestive of the development of tolerance. By the final test day, sucrose consumption of the LPS-NaCl rats was similar to that of saline control animals. This is consistent with results of previous studies that have found significant tolerance to the effects of LPS on food intake (18, 21, 22). The levels of sucrose intake seen in rats treated with both LPS and CCK were also subject to the development of tolerance, as these animals showed an increase in intake on subsequent exposures to LPS. Further studies, using an increased number of test days, would be able to determine whether LPS can completely block the sustained anorectic effects seen in animals treated with only CCK. In the present study, both groups of rats treated with LPS exhibited significant body weight reductions relative to the other two groups (NaCl-NaCl and NaCl-CCK) after the first test day, an effect that was no longer significant on subsequent exposures to LPS. This finding is also consistent with the development of tolerance, and the results of prior investigations that demonstrated tolerance effects to LPS to various physiological measures, including fever (28).
Experiment 2
The results from the TRT demonstrated that systemic administration of LPS on its own does not alter the palatability of sucrose. LPS-treated animals did not differ from the saline controls in their pattern of taste reactivity responding to the sucrose. In addition, the present study demonstrated a pronounced reduction in ingestive responding in rats treated with CCK, evidence of a change in palatability and reflective of a satiety effect. Furthermore, rats treated with both LPS and CCK produced a pattern of responding that was similar to that shown by rats treated with only CCK, suggesting no interaction between LPS and CCK on sucrose palatability.The present study is, to the best of our knowledge, the first examination of the effects of LPS on palatability changes. The findings demonstrate that systemic administration of LPS does not alter the palatability of sucrose. LPS-NaCl rats displayed a high level of ingestive responding without showing any active aversive responses or passive drips throughout both testing sessions. The pattern of taste reactivity responses exhibited by the LPS-treated rats did not differ from that of the saline-treated controls. This result provides evidence against the hypothesis that LPS administration induces a state of anhedonia (41). Moreover, animals treated with LPS alone actually showed a nonsignificant trend for increased preference for sucrose, as evidenced by higher levels of ingestive responses relative to the saline-treated animals. It is likely that the increased ingestive response frequencies were limited by a ceiling effect. Maximal levels of ingestive responses that can be produced by a rat during a 30-s infusion, given a typical lick rate of 6-7 licks/s (14), would be ~210 licks. Thus further studies are necessary to determine whether LPS-treated animals would show an enhanced preference for sucrose compared with saline-treated rats, at concentrations of sucrose that do not elicit maximal ingestive responding.
The data from the present experiment are consistent with previous demonstrations of reduced levels of ingestive responding to sucrose after systemic administration of CCK (6, 9). Animals treated with CCK alone (NaCl-CCK) exhibited an immediate reduced level of ingestive responding that was maintained across the testing periods on both test days. Increased levels of passive drip and almost no active aversive responses accompanied this decreased ingestive responding, a pattern of taste reactivity responding that is suggestive of satiety as opposed to an aversive effect (9). Indication of aversion is evidenced by a decrease in ingestive responses and an increase in active aversive responses, a pattern of responding that occurs during infusions of quinine (14). Thus our results are consistent with the hypothesis that CCK produces its feeding suppression via satiety-related mechanisms.
A conditioned aversion in the TRT is evidenced by a gradual decrease in ingestive responses and concomitant increase in active aversive responses. This pattern of responding has been shown repeatedly after systemic administration of LiCl (4, 14, 30, 35). The high levels of ingestive responses and absence of active aversive response seen in the LPS-treated rats are not indicative of a conditioned aversion. This suggests that the acute phase response induced by administration of LPS does not induce internal aversive cues similar to those of LiCl, cues that are necessary to induce a CTA. The present data are also consistent with a previous study in which an antiemetic, trimethobenzamide, failed to attenuate the feeding suppressive effects of LPS (39). Taken together, these data suggest that a learned taste aversion may not contribute to the normal anorectic effect of LPS.
The finding that LPS did not immediately produce any active or passive rejection responses, nor immediately suppress ingestive responses, suggests that LPS does not result in an unconditioned reduction of sucrose palatability. Rejection responses are normally elicited in animals infused with an unpalatable tastant, such as a quinine solution. Unconditioned palatability shifts do not rely on associative mechanisms, and result in immediate decreases in ingestive responding and increases in active and/or passive aversive responses.
When LPS and CCK were administered in combination, a significant reduction in ingestive responding and an increase in passive drips similar to that observed in the NaCl-CCK rats was evident. These reductions in ingestive responses were significantly lower than those seen in either the saline-treated controls or rats treated with LPS alone. These data suggest that CCK is responsible for the reduced level of ingestive responding that is apparent in the LPS-CCK-treated rats. However, on the second test day the LPS-CCK group displayed significantly more ingestive responding than the NaCl-CCK group, suggesting that LPS tolerance may have attenuated the normal anorectic effects of CCK with repeated injections of LPS-CCK. This result is similar to the findings reported in experiment 1, in which the LPS-CCK group exhibited increased intake of sucrose solution on repeated exposures to LPS relative to the CCK-only group.
Contrary to our original hypothesis, we failed to find evidence of augmentation by CCK of the normal hypophagic effects of LPS using the TRT. Past research has demonstrated that activation of the immune system results in both the activation of gastric vagal afferents via CCK receptors (17) as well as an increased sensitivity to CCK (5). These previous studies also found that blocking the actions of CCK with a CCK receptor antagonist resulted in an attenuation of the hypophagia induced by systemic administration of IL-1 (7). These findings were the basis for our original hypothesis that CCK and LPS together would result in an augmentation of either satiety or aversion, both of which could account for the hypophagia experienced during the acute phase response. Clearly this hypothesis was not supported, as LPS-treated rats showed an enhanced preference for sucrose, whereas CCK-treated rats displayed responses indicative of satiety. LPS and CCK together did not produce patterns or levels of taste reactivity responses that differed from those produced by rats treated with only CCK. This result provides further support against the hypothesis that LPS and CCK induce similar internal states, states that would account for the observed reductions in food intake during the acute phase response. In this context, the present data are consistent with a previous report in which CCK receptor antagonists did not alter LPS-induced reductions in social exploration in mice, suggesting that CCK may not be a mediator of the behavioral effects of LPS (3). Although further research is necessary to determine whether CCK plays a role in other behavioral responses, the present results suggest that CCK does not play a role in LPS-induced hypophagia.
Perspectives
The results from the present study have shown a clear dissociation between the effects of LPS on intake measures and taste reactivity behaviors, measures that, in untreated rats, show a strong positive correlation with one another (33). LPS treatment clearly reduced sucrose intake, yet animals displayed no evidence of a change in palatability to sucrose after treatment with LPS. This finding necessitates a consideration of the differences between intake and taste reactivity measures. The TRT is a forced-choice procedure and involves direct measurement of consummatory behaviors. These are behaviors that are involved in the actual consumption of food, such as licking, chewing, and swallowing. In contrast, traditional intake measures require the animal to recognize that food is available and make subsequent approach movements followed by consumption of the food. These behaviors that occur before food consumption are categorized as appetitive behaviors. As such, the present study suggests that LPS may only affect the appetitive component but not the consummatory behaviors.The deprivation state of the animals is another important procedural difference between intake measures and the TRT. Because the TRT is a forced-choice procedure, animals need not be food nor water deprived before testing. Conversely, to motivate an animal to ingest food during an intake test, animals are usually food and/or water deprived to ensure adequate consumption. Several studies have also suggested that the hypophagic effects induced by LPS may be dependent on the state of hydration of the animal. Langhans et al. (19) found that peripherally administered LPS reduced food and water intake when both food and water were available and inhibited food intake during water deprivation but failed to reduce drinking during food deprivation. Although it is unlikely that food and water deprivation before behavioral testing in the TRT would cause LPS animals to decrease their ingestive responding, this possibility cannot be completely discounted. Further studies are necessary to determine whether deprivation state would affect the results obtained from the TRT.
The majority of previous studies have found a decrease in water intake during the acute phase response, induced either by LPS or cytokine administration (e.g., Refs. 26, 27). Wang et al. (38), however, reported an increase in water intake in water-replete animals after LPS administration. This increase in drinking was suggested to represent an adaptive mechanism whereby an animal could increase water intake at a time when metabolic changes, such as fever, sweating, and diarrhea, could cause dehydration. Although our data from experiment 1 do not support this hypothesis, the results from experiment 2, showing an increase in ingestive responding by LPS-treated animals, could reflect such an adaptive mechanism to compensate for dehydration.
In summary, this study showed that treatment with LPS produces a reduction in sucrose intake without changing its palatability. Systemic administration of LPS resulted in elevated levels of ingestive responding despite a reduction in sucrose intake. The present study also showed that LPS-induced reductions in sucrose intake are not the result either of an unconditioned or conditioned aversive effect or the result of enhanced satiety. Indeed, LPS may enhance sucrose palatability. The results of the TRT also revealed that systemic administration of LPS or CCK did not induce similar internal states, even though both resulted in reductions in sucrose intake. Finally, the present results demonstrated that CCK does not augment the hypophagic effects produced by administration of LPS, as has been previously hypothesized. The present study underscores the importance of and necessity for examination of palatability changes, reflected by taste reactivity responses, in addition to traditional intake measures.
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ACKNOWLEDGEMENTS |
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This research was supported by Natural Sciences and Engineering Research Council of Canada operating grants to K.-P. Ossenkopp and M. Kavaliers.
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FOOTNOTES |
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Portions of these data were presented at the 28th annual meeting of the Society for Neuroscience, Los Angeles, CA, November, 1998.
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.
Address for reprint requests and other correspondence: S. K. Cross-Mellor, Neuroscience Program, c/o Dept. of Psychology, Univ. of
Western Ontario, London, Ontario N6A 5C2, Canada
(E-mail:
skcross{at}julian.uwo.ca).
Received 18 February 1999; accepted in final form 18 May 1999.
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REFERENCES |
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TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
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