INTRODUCTION

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NUMEROUS PHYSIOLOGICAL SIGNALS have been implicated in the control of ingestive behavior in rats. Among the first signals proposed, and later demonstrated, to affect ingestion are changes related to the utilization of energy (17). For example, in adult rats, decreasing the utilization of glucose with agents such as 2-deoxyglucose or 5-thioglucose (5-TG) can produce a significant increase in food intake (18, 22, 23, 27). Similarly, decreasing glucose availability by administration of insulin can also enhance ingestion (21, 27). In addition, blocking the metabolism of fats by inhibiting the oxidation of fatty acids with 2-mercaptoacetate (MA) or methyl palmorixate (MP) can produce increases in food intake (5, 6, 12, 26). These increases after alteration of fatty acid metabolism are particularly effective in adult rats that have been maintained on high-fat diets and are presumably more reliant on fatty acid oxidation for energy than animals maintained on standard, low-fat chow diets (6, 26). In addition, simultaneous blockade of both fat and glucose metabolism produces synergistic effects on food intake (5, 6, 29), suggesting that these signals are integrated to control intake. Studies of the mechanisms by which changes in energy utilization affect food intake suggest that signals related to glucose regulation may have direct central effects, whereas signals related to fat utilization are conveyed centrally by vagal fibers (12, 24, 25).

These data from studies of energy metabolism in adult rats suggest that the control of food intake is sensitive to current utilization of energy. However, the development of ingestive responsiveness to these energy-related signals is currently unclear. Early studies examining the control of intake in rat pups focused on suckling behavior and demonstrated that changes in glucose utilization fail to affect suckling intake in rat pups until at least the third week of life (10, 16, 32). However, it appears that suckling behavior may not represent the developmental precursor to adult ingestion (9). Instead, adult ingestive behavior is more closely related to the ingestive behavior that rat pups demonstrate independent of the dam and the suckling situation. Studies examining adultlike ingestion in rat pups measure intake either of a diet spread on the floor of a test container or of a diet infused through an oral cannula placed at the front of the pup's mouth. More recent studies using these independent ingestive tests have also demonstrated that pups do not respond to glucoprivation produced by intracerebroventricular administration of 5-TG until 30 days of age, even though changes in circulating glucose levels are noted in pups as young as 9 days of age (13).

Fewer studies have examined the development of ingestive responding to signals related to fat metabolism. In one series of studies, Leshem et al. (13) examined the effects of blocking fatty acid oxidation on intake from the dam in a suckling situation. Their data suggest that blocking fatty acid oxidation using MA or MP fails to affect suckling intake in rat pups, but the ontogeny of independent ingestive responding to changes in fatty acid oxidation in rat pups remains unclear. It is logical for pups to be particularly sensitive to fat metabolism since oxidation of fatty acids is an important source of energy for rat pups before weaning. Their primary food source, mother's milk, provides ~60-70% of its calories from fat (3, 31). In addition, compared with adult rats, pups have higher capacities for the oxidation of fatty acids and appear to preferentially use products of fatty acid oxidation in the brain (14, 15, 31, 33). The present series of experiments was designed to examine the development of independent ingestive responding to blockade of fatty acid oxidation in rat pups.

	GENERAL METHODS

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Subjects. Subjects were progeny of primiparous and multiparous female Sprague-Dawley rats (Harlan, Indianapolis, IN) bred in the laboratory. Animals were housed in plastic maternity cages lined with aspen shavings and allowed ad libitum access to food (Lab Diets chow no. 5012) and water. Temperature was maintained at 25°C, and the animals were on a 14:10 light-dark cycle. Female rats were placed in pairs with male rats for 1 wk for mating. Females were then separated from the males and housed in pairs until the week of parturition, when they were separated into individual plastic maternity cages. Cages were checked daily for births; pups present at 1700 were considered to be born that day, designated as day 0. Litters were culled to 10 pups (5 male, 5 female where possible) on the day after birth. Except for routine maintenance, litters remained undisturbed with the dam until the day of testing.

Testing. On the day of testing, pups were removed from the dam, stimulated to urinate and defecate by stroking with an artist's brush wetted with warm water, and weighed. The dose of drug to be delivered was calculated, and the drug was delivered by intraperitoneal injection (MA) or gavage (MP). Pups were then placed into a warm moist glass aquarium incubator (7) maintained at 31-34°C. Pups receiving MA were tested 1 h after administration, whereas pups receiving MP were tested 6.5 h after administration. These delays, doses, and routes of administration were chosen based on those previously demonstrated to affect intake in adult rats (4, 26). Immediately before testing, pups were removed from the incubator, stimulated to urinate and defecate, and reweighed. They were then placed inside plastic test containers inside the incubator. The test containers were lined with paper towels wetted with a commercial half-and-half diet or a 15% glucose solution warmed to 31-33°C. Pups were allowed to consume the diet for 30 min; the paper towels were rewetted with warm diet every 10 min during testing as necessary. Pups were then removed from the test containers, dried carefully, and reweighed. Because pups at these ages do not readily urinate and defecate spontaneously, particularly after being stimulated to urinate and defecate, the amount of weight gained during the intake was used as a reliable measure of intake (8). Pups were tested once at 6, 9, 12, or 15 days of age as described in individual experiments. All animal care and experiments conform to the guidelines of and were approved by the Purdue University Animal Care and Use Committee.

Statistical analysis. Intake was expressed as a percentage of the pretest body weight. Results were first analyzed with a two-way (age × dose) analysis of variance (ANOVA) (GLM, SAS Institute, Cary, NC). Individual ANOVAs were then performed as necessary at each age. Where appropriate, post hoc tests using Tukey's highly significant difference test (HSD) were performed. A level of P < 0.05 was used to determine significance.

	EXPERIMENT 1A

When administered to adult rats, MA produces a reliable increase in ingestive responding commencing after 1 h and continuing for at least 6 h (26). In this experiment, the development of ingestive responding following administration of MA was examined in 6-, 9-, 12-, and 15-day-old pups.

Methods. Pups were removed from the dam and received an intraperitoneal injection of a dose of 0, 100, 200, 400, or 800 µmol/kg MA dissolved in a 0.135 M NaCl vehicle at a concentration of 100 µM. This dose range was chosen to span the typical doses known to increase intake in adult rats (24, 26). Because the volumes to be delivered were so small, the concentration delivered was one-half of that typically used in adult rats. After injection, pups were placed in an incubator for 1 h to allow the MA to produce its effects. They were then given a 30-min intake test with a diet of commercial half-and-half. No more than two pups from each of 6 or 7 litters were tested at each dose at each age (n = 12-14/group).

Results and discussion. When rats were tested with a diet of half-and-half, MA affected intake at some ages and doses [main effect of age: F(3,220) = 13.05, P < 0.0001; main effect of dose: F(4,220) = 15.25, P < 0.0001; age × dose interaction: F(12,220) = 5.48, P < 0.0001]. In 6-day-old pups, no dose of MA affected intake [Fig. 1A; F(4,55) = 0.52, P < 0.72]. Similarly, in 9-day-old pups, MA was ineffective at altering intake at any dose compared with saline [Fig. 1B; F(4,55) = 0.34, P < 0.85]. However, in 12-day-old pups, MA significantly affected intake [Fig. 1C; F(4,55) = 17.64, P < 0.0001]. Low doses of MA increased intake of a half-and-half diet compared with saline, whereas the highest dose tested significantly suppressed intake compared both with saline and with the other doses of MA. In addition, MA significantly affected intake in 15-day-old pups [F(4,55) = 10.17, P < 0.0001]. Low doses of 200 or 400 µmol/kg increased intake compared with saline, whereas the highest dose (800 µmol/kg) significantly suppressed intake compared with the other doses of MA, but not compared with the saline control. Thus pups aged 12 or 15 days are capable of demonstrating changes in ingestive responding to blockade of fatty acid oxidation after administration of MA, whereas younger animals fail to show changes in ingestive responding to any of the doses tested.

View larger version (34K): [in this window] [in a new window]   Fig. 1.   Intake (expressed as %pup's body wt) of a commercial half-and-half diet during a 30-min intake test from the floor of test containers 1 h after administration of 2-mercaptoacetate (MA). A: 6-day-old pups; B: 9-day-old pups; C: 12-day-old pups; and D: 15-day-old pups. Note that scales are different for pups at different ages. * P < 0.05 compared with saline control; # P < 0.05 compared with 100, 200, and 400 µmol/kg MA.

These data suggest that, when pups are given a high-fat diet to ingest during testing, ingestive responding to MA emerges between 9 and 12 days of age in rat pups. This time frame is significantly earlier than that of ingestive responding to changes in glucose metabolism (13, 16). These results therefore suggest that changes in fatty acid oxidation may be the first metabolic signal that influences ingestion in rat pups.

	EXPERIMENT 1B

The diet used in experiment 1a, half-and-half, provides ~75% of its calories from fat, comparable to the fat percentage available in the dam's milk. MA may produce its effects by specifically enhancing intake of diets high in fat; the pup senses a decrease in utilization of fats and therefore responds by consuming more fat. Alternatively, MA may increase intake more generally. This experiment was designed to examine whether MA produces a general increase in ingestive responding or a specific enhancement of fat intake by testing ingestion of a glucose diet in 9- and 12-day-old pups.

Methods. The results of experiment 1a demonstrate an emergence of ingestive responding to MA between 9 and 12 days of age. Therefore, in this experiment, pups were tested once at 9 or 12 days of age. As in experiment 1a, pups received an intraperitoneal injection of MA 1 h before testing at a dose of 0, 100, 200, 400, or 800 µmol/kg. However, in this experiment, pups received a 30-min intake test with a 15% solution (wt/vol) of glucose instead of half-and-half. No more than two pups from each of five litters were tested at each dose at each age (n = 9 or 10/group).

Results and discussion. The results again demonstrate that MA affects intake differentially at different ages and doses [main effect of age: F(9,89) = 10.46, P < 0.0001]. In 9-day-old pups, there was no effect of any dose of MA compared with saline on intake of a 15% glucose solution [Fig. 2A; F(4,44) = 1.10, P < 0.37]. However, in 12-day-old pups, MA significantly affected intake [Fig. 2B; F(4,45) = 13.58, P < 0.0001]. The low doses of MA significantly increased intake compared with saline, whereas the highest dose of MA significantly suppressed intake compared with the other doses of MA but not compared with saline.

View larger version (20K): [in this window] [in a new window]   Fig. 2.   Intake of a 15% (wt/vol) glucose solution from the floor of test containers 1 h after administration of MA. A: 9-day-old pups. B: 12-day-old pups. * P < 0.05 compared with saline control; # P < 0.05 compared with 100, 200, and 400 µmol/kg MA.

These data demonstrate that the effects of MA on intake are not specific to high-fat diets. Pups 9 days of age or younger failed to respond to any dose of MA when tested with either a high-fat or no-fat diet. On the other hand, low doses of MA enhance intake of both diets in older pups, whereas the highest test dose suppresses intake. The general intake-enhancing effects of MA in pups are consistent with the effects seen in adults (2). Taken together, the results of these experiments support a role for changes in fatty acid oxidation in a generalized control of intake that emerges between 9 and 12 days of age in rat pups.

	EXPERIMENT 2

The results of experiments 1a and 1b suggest that administration of MA to pups produces alterations in intake by 12 days of age. These behavioral effects presumably result from the blockade of fatty acid oxidation produced by MA. Although it is currently unclear why pups younger than 12 days of age fail to respond to MA, one possibility is that the doses of MA used in these experiments fail to effectively impair fatty acid utilization in pups younger than 12 days of age. Thus this experiment was designed to assess the effects of MA on physiological measurements related to oxidation of fatty acids, including free fatty acids (FFA), ketone bodies [specifically -hydroxybutyrate (-HBA)], and blood glucose.

Methods. Pups were tested once at 9, 12, or 15 days of age; no more than two pups from each of six litters were tested at each dose at each age (n = 11 or 12/group). Pups received an intraperitoneal injection of 0, 400, or 800 µmol/kg MA and were placed into an incubator for 1 h. After the 1-h delay, pups were killed with an overdose of Brevital sodium, and blood was collected from the inferior vena cava into nonheparinized microhematocrit tubes and centrifuged immediately. The plasma was separated, and plasma samples from the same animal were combined. Plasma was frozen at 80°C until assay. At the same time that blood was collected, stomachs were also removed for measurement of gastric emptying. After removal, stomachs were weighed, gastric contents were removed, and stomachs were reweighed. The weight difference was used as an estimate of gastric volume. Plasma FFA and -HBA levels were measured enzymatically (NEFA-C kit, Wako Chemicals for FFA and -HBA kit, Sigma Chemicals), and plasma glucose levels were measured using a Beckman glucometer.

Statistical analysis. Separate two-way (age × dose) ANOVAs were run on stomach contents, FFA, -HBA, and blood glucose measurements. Then, separate one-way repeated-measure ANOVAs were run at each age where appropriate. Post hoc tests using Tukey's HSD were performed, with P < 0.05 taken as significant.

Results and discussion. Administration of MA resulted in changes in -HBA levels within 1 h (Fig. 3A). Levels of -HBA were higher in 12-day-old animals than in 9-day-old pups [main effect of age: F(2,98) = 3.71, P < 0.05]. In addition, across ages, levels of -HBA were significantly reduced in pups that received 400 or 800 µmol/kg MA compared with controls [main effect of dose: F(2,98) = 55.22, P < 0.0001]. In addition, animals receiving 800 µmol/kg MA had significantly lower -HBA levels than pups receiving 400 µmol/kg. These differences in -HBA levels were also observed within ages [9 day olds: F(2,33) = 16.76, P < 0.0001; 12 day olds: F(2,32) = 15.86, P < 0.0001; 15 day olds: F(2,33) = 26.72, P < 0.0001]. At all ages, -HBA levels were significantly lower after a dose of 800 µmol/kg MA than after a dose of 400 or 0 µmol/kg MA, and a dose of 400 µmol/kg MA resulted in significantly lower -HBA levels than a dose of 0 µmol/kg. Although -HBA levels were significantly affected by administration of MA, FFA levels were unaffected by age or dose of MA (Fig. 3B). Blood glucose levels were also unaffected by administration of MA; however, the age of the pup did affect blood glucose [main effect of age: F(2,98) = 20.72, P < 0.0001]. Blood glucose levels were significantly higher in 15-day-old pups than in 9 and 12 day olds (Fig. 3C). Finally, stomach contents were significantly affected by administration of MA and by the age of the pup [main effect of age: F(2,98) = 9.02, P < 0.001; main effect of dose: F(2,98) = 10.68, P < 0.0001; age × dose interaction, F(4,98) = 2.78, P < 0.05]. In 9-day-old pups, there was no effect of administration of MA (Fig. 3D). However, in 12-day-old pups, administration of 800 µmol/kg MA resulted in significantly greater gastric volume after 1 h [F(2,32) = 5.32, P < 0.01]. In addition, gastric volume in 15-day-old pups was significantly greater after administration of 400 or 800 µmol/kg MA [F (2,33) = 15.78, P < 0.0001].

View larger version (43K): [in this window] [in a new window]   Fig. 3.   Physiological measurements determined 1 h after administration of MA. A: plasma -hydroxybutyrate (-HBA) levels. B: plasma free fatty acid (FFA) levels. C: plasma glucose levels. D: stomach contents. BW, body weight. * P < 0.05 compared with saline control; # P < 0.05 compared with 800 µmol/kg MA.

These results indicate that MA produces physiological changes in rat pups within 1 h of administration. Decreases in -HBA levels were observed at all ages tested; these decreases in a ketone body are consistent with the proposed inhibitory action of MA on oxidation of fatty acids to ketone bodies. Thus such results suggest that the effects of MA on intake may be mediated through changes in the utilization of lipids. In addition, these results suggest that the failure of 9-day-old pups to alter intake after administration of MA does not result from a failure of MA to significantly alter fatty acid oxidation in young pups. Significant decreases in -HBA were observed in 9-day-old pups, and the magnitude of change after each dose was similar in pups at the three ages tested. Thus it appears that 9-day-old pups may fail to alter intake after administration of MA because they fail to detect a signal or signals related to the alteration of fat utilization. Although -HBA levels were significantly altered by administration of MA, neither FFA levels nor glucose levels were affected. FFA levels are typically (but not always, e.g., Ref. 26) increased in adult animals 1 h after administration of MA. However, FFA levels in control adult animals are also typically lower than those observed in control rat pups (26). Thus it is possible that no increases in FFA levels were observed in pups because of the high values demonstrated in controls. Finally, administration of 800 µmol/kg MA appears to significantly suppress gastric emptying in 12- and 15-day-old pups. This suppression of gastric emptying may contribute to the decreased intake produced by 800 µmol/kg MA in 12- and 15-day-old pups. Because these pups begin the ingestive test with larger stomach volumes, they may stop eating sooner than pups receiving lower doses. It is unclear how MA produces decreased gastric emptying, but it does not appear to be a direct result of a decrease in lipid utilization since 9-day-old pups show changes in -HBA levels but not changes in gastric emptying. Although decreased gastric emptying may contribute to decreased intake after administration of 800 µmol/kg MA, additional mechanisms may also be operating since administration of 400 µmol/kg MA suppresses gastric emptying in 15-day-old pups, but increases intake.

	EXPERIMENT 3

Blockade of fatty acid oxidation using MA appears to occur through a decrease in activity of acyl-coenzyme A dehydrogenases (1). However, MA is not the only agent used to block fatty acid oxidation that affects intake in adult rats. MP operates by blocking the enzyme carnitine palmitoyltransferase I (CPT-I), thus inhibiting the transport of long-chain fatty acids into mitochondria (30). Although rat pups appear to develop responsiveness to MA between 9 and 12 days of age, it is possible that responsiveness to blockade of fatty acid oxidation at a different step in the biochemical cascade using MP has a different developmental time course. This experiment was designed to assess the age at which responding to administration of MP emerges.

Methods. Because experiments 1a and 1b had demonstrated that pups first respond to MA at 12 days of age, pups were tested at 12 days of age or 15-18 days of age. Each pup received a gavage load of 1.25, 2.5, 5, or 10 mg/kg MP or its vehicle (0.5% methylcellulose). The volume of the gavage load was held constant, and the concentration was varied between 0.375 and 3 mg/ml. Pups were allowed 6.5 h for the MP to take effect (n = 6 pups from 3 litters/group) and were then given a 30-min intake test with a half-and-half diet.

Results and discussion. In 12-day-old pups, no dose of MP tested affected intake [Fig. 4A; F(4,24) = 1.45, P < 0.25]; however, administration of MP significantly suppressed intake in pups aged 15-18 days [Fig. 4B; F(4,34) = 2.99, P < 0.04].

View larger version (22K): [in this window] [in a new window]   Fig. 4.   Intake of a commercial half-and-half diet from the floor of test containers 6 h after administration of methyl palmoxirate. A: 12-day-old pups. B: 15- to 18-day-old pups. * P < 0.05 compared with saline control.

The results of this experiment suggest that MP fails to increase intake in pups aged 12-18 days of age. Thus the developmental emergence of ingestive responding to MP does not appear to coincide with that of MA. One explanation of the failure of MP to influence intake in the present study derives from the source of the fat that the pups are metabolizing. MP operates by blocking the transport of long-chain fatty acids into mitochondria, and the effects of MP are diminished in adult rats that are maintained on diets high in medium-chain triglycerides (4). Although pups have high levels of fatty acid oxidation and receive high levels of fat from maternal milk, rat milk is unusually high in medium-chain triglycerides (50-70% of triglycerides present; see Refs. 3, 12). Thus it is likely that MP fails to produce changes in intake in pups at the ages tested because it is not effective at blocking oxidation of the highly prevalent medium-chain fatty acids available to pups. Indeed, recent biochemical data support the idea that, in rat pups 10-15 days of age, the enzyme CPT-I, which MP blocks, is not the rate-limiting step in oxidation of fatty acids (11). Finally, a dose of 10 mg/kg MP failed to affect ketone body levels in 15-day-old rat pups when measured 1, 2, 4, or 6 h after administration (31). These results suggest that the failure of MP to affect intake in pups at the ages tested may result from a failure of MP to significantly affect fatty acid oxidation and not from the differences in testing procedures between the two groups (e.g., gavage vs. intraperitoneal injection; 1-h delay vs. 6-h delay).

	GENERAL DISCUSSION

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The results from these experiments demonstrate that MA affects intake in rat pups aged 12 or 15 days, but not in younger pups. One interesting aspect of these data is the U-shaped dose-response curve seen in the older animals; low doses of MA enhance intake, whereas the highest dose tested (800 µmol/kg) suppresses intake. Previous studies in adult rats have demonstrated a similar U-shaped dose-response curve under some circumstances. For example, adult rats maintained on a high-fat (23%) diet and tested with a high-fat diet showed suppressed intake 1 h after 800 µmol/kg MA (2); in fact, these authors report significant mortality after this dose of MA. In addition, this dose of MA significantly suppressed 12-h, but not 2-h, intake in both intact and hepatic vagotomized adult rats maintained and tested with an 18% fat diet (12). However, animals maintained on a lower fat diet (e.g., 3.3%), tested with a low-fat diet, or tested at other delays demonstrate increases in intake following the same 800 µmol/kg MA dose (2, 12, 24). In the present study, pups were maintained on a high-fat diet of dam's milk and tested with both high and nonfat diets. A dose of 800 µmol/kg MA suppressed intake of both diets 1 h after administration in older pups. One potential influence is the effect of MA on gastric emptying in 12- and 15-day-old pups. However, it is currently unclear why a dose of 800 µmol/kg MA suppresses intake in some situations and enhances intake in others. There are suggestions that the intake-enhancing and intake-suppressing properties of MA may be differentially mediated. For example, whereas hepatic vagotomy eliminates the increases in intake produced by 800 µmol/kg MA in a 2-h test, it fails to affect the suppression of intake by the same dose in a 12-h test (12). Interestingly, both the intake-enhancing effects and intake-suppressing effects of MA emerge developmentally at the same age. Neither suppression nor enhancement of intake are observed in 6- or 9-day-old pups, but 12- and 15-day-old pups show increased intake after low doses and decreased intake following the 800 µmol/kg dose of MA. Finally, as with any suppression of intake, malaise may be responsible. Further study may reveal the mechanisms mediating the intake-enhancing as well as the intake-suppressing effects of MA in pups.

The data suggest that a metabolic signal related to changes in the oxidation of fatty acids first affects independent intake between 9 and 12 days of age in rat pups. Previous studies have described the time between 9 and 12 days of age as a period during which independent ingestive responding to a number of physiological signals emerges. Pups begin to be responsive to caloric properties of a gastric preload at this time and begin to respond to caloric privation following an overnight fast (19, 20, 28). There is evidence that changes in fatty acid oxidation are produced by deprivation in pups at these ages (13), and the present data support the ability of pups to respond behaviorally to changes in fatty acid oxidation. Thus the signal that informs pups of caloric changes between 9 and 12 days of age may be related to oxidation of fatty acids. Dam's milk contains high levels of fat, and high levels of fatty acid oxidation are observed in rat pups compared with adult rats maintained on chow (14). Pups' preferential use of ketone bodies for energy may make them particularly sensitive to changes in fatty acid oxidation, and thus responsiveness to changes in fatty acid oxidation makes sense as an early metabolic signal to affect intake in rats.

The data from the present experiments support the emergence of ingestive responding to MA by 12 days of age. As mentioned above, previous results have suggested that pups at this age are insensitive to MA (13). This difference in findings between the two reports likely results both from differences in the ingestive system studied and from differences in the doses of MA tested. First, in the present experiment, pups were tested independent of the mother and the suckling situation. The previous report primarily explored development of responsiveness to MA in pups attached to mothers and suckling. Although suckling is the principal mode of ingestion for young rats, evidence supports independent ingestion as the developmental precursor of adult ingestion (9). Thus although independent ingestion is unlikely to be displayed by preweaning pups in a natural environment, studies of pups' ingestive capacities away from the dam can inform us about neural and behavioral maturation that would be masked in studies of suckling behavior. Second, the doses of MA tested in the previous study were 600-1,000 µmol/kg. Given the intake-suppressing effect of a dose of 800 µmol/kg MA shown in the present study, the previous failure to demonstrate increased intake after administration of MA may have also resulted from the doses previously used.

Although MA appears to produce changes in ingestive behavior early in a rat pup's life, development of responding to MA does not parallel the development of responding to administration of MP. In fact, the exact age at which responsiveness to MP emerges is still unclear. The present experiments examined pups up to 18 days of age. Testing the effects of MP on pups older than 18 days of age is complicated by the fact that, by this age, pups have begun to supplement their diet of mother's milk by sampling chow. Consequently, the fat composition of their diet begins to decrease. MP most consistently increases intake in animals that have been maintained on high-fat diets. Thus if ingestive responsiveness to MP actually does not emerge until 18 days or later, determining its development may require manipulation of the diet to which pups have access. An alternative possibility is that the failure of pups to respond to MP results not from decreased levels of fat in the diet late in the preweaning period, but from the type of fat available to them throughout lactation. If pups fail to respond to MP because of high availability of medium-chain fatty acids, altering the fatty acid composition of the dam's milk through dietary changes may alter the developmental profile of ingestive responding to MP. In fact, dams maintained on a high-fat diet produce milk with a significantly lower medium chain triglyceride concentration than dams on standard chow (31). In these pups, a 10 mg/kg dose of MP of the dam does lower plasma ketone levels within 1 h of administration (31) at 15 days of age. Thus administration of MP to pups reared by a dam maintained on a high-fat diet may reveal that the effects of MP emerge at an age similar to that of the emergence of effects of MA.

Perspectives