Hyperphagia and obesity of OLETF rats lacking CCK1 receptors: developmental aspects

Otsuka Long Evans Tokushima Fatty (OLETF) rats have a deletion in the gene encoding the cholecystokinin-1 (CCK1) receptor. This deletion prevents protein expression, making the OLETF rat a CCK1 receptor knockout model. Consistent with the absence of CCK1 receptors, OLETF rats do not reduce their food intake in response to exogenously administered CCK and consume larger than normal meals. This deficit in within-meal feedback signaling is evident in liquid as well as solid meals. Neonatal OLETF rats show similar differences in independent ingestion tests. Intake is higher and is reflected in greater licking behavior. Neonatal OLETF rats also have diminished latencies to consume and higher initial ingestion rats. Adult OLETF rats are hyperphagic and obese. Although arcuate nucleus peptide gene expression is apparently normal in OLETF rats, when obesity is prevented through pair-feeding to amounts consumed by control Long Evans Tokushima Otsuka (LETO) rats, dorsomedial hypothalamic NPY mRNA expression is significantly elevated in OLETF rats. NPY overexpression is also evident in preobese, juvenile OLETF rats suggesting a causal role for this overexpression in the hyperphagia and obesity. Running wheel exercise normalizes food intake and body weight in OLETF rats. When access to exercise is provided at a time when OLETF rats are obese, the effects are limited to the period of exercise. When running wheel access is available to younger, preobese OLETF rats, exercise results in long lasting reductions in food intake and body weight and improved glucose regulation. These lasting metabolic effects of exercise may be secondary to an exercise induced reduction in DMH NPY mRNA expression.     

Roles of dorsomedial hypothalamic cholecystokinin signaling in the controls of meal patterns and glucose homeostasis

A role for dorsomedial hypothalamus (DMH) cholecystokinin (CCK) signaling in feeding control has been proposed. Administration of CCK into the DMH reduces food intake and OLETF rats lacking CCK1 receptors (CCK1R) become hyperphagic and obese. We hypothesized that site specific replenishment of CCK1R in the DMH of OLETF rats would attenuate aspects of their feeding deficits. Recombinant vectors of adeno-associated viral (AAV)-mediated expression of CCK1R (AAVCCK1R) were bilaterally delivered into the DMH of OLETF. OLETF rats with AAVCCK1R injections demonstrated a 65% replenishment of Cck1r mRNA expression in the DMH relative to lean LETO control rats. Although this level of replenishment did not significantly affect overall food intake or body weight through 14 weeks following viral injections, meal patterns were partially normalized in OLETF rats receiving AAVCCK1R with a significant decrease in dark cycle meal size and a small but significant decrease in daily food intake in the meal analysis chambers. Importantly, the elevation in blood glucose level of OLETF rats was attenuated by the AAVCCK1R injections (p=0.03), suggesting a role for DMH CCK signaling in glucose homeostasis. In support of this role, administration of CCK into the DMH of intact rats enhanced glucose tolerance, as this occurred through activation of CCK1R but not CCK2R signaling. In conclusion, partial replenishment of CCK1R in the DMH of OLETF rats, although insufficient for altering overall food intake and body weight, normalizes meal pattern changes and reduces blood glucose levels. Our study also shows a novel role of DMH CCK signaling in glucose homeostasis.     

Perinatal undernutrition-induced obesity is independent of the developmental programming of feeding

Protein or calorie restriction during gestation and/or suckling induces hyperphagia and increases the susceptibility to develop obesity, glucose intolerance and hypertension in adulthood. The mechanisms by which early nutrient restriction affects the normal physiological regulation of feeding as well as to what extent the metabolic programming of hyperphagia contributes to the long-term risk of obesity and insulin resistance remain, however, to be determined. Here the temporal pattern of food intake and the behavioural satiety sequence were investigated in the offspring of Sprague-Dawley rats fed a control (C) or a low-protein (LP) diet throughout pregnancy and lactation. During the first two months of their post-natal life, protein-restricted animals exhibited hyperphagia characterized by a delayed appearance of satiety, an increase in meal size and reduced latency to eat. Protein-restricted pups also exhibited an enhanced expression of the orexigenic peptides Agouti-related protein and neuropeptide Y and decreased hypothalamic levels of the anorexigenic peptide pro-opiomelanocortin. At 8 months, LP rats still consumed larger meals than their control counterparts but they ingested daily the same amount of food as control offspring and exhibited enhanced abdominal fat and increased levels of triglycerides and fatty acids in serum. These observations indicate that the hyperphagia observed in young LP rats results from a decreased action of negative feedback signals critical to meal termination and an enhanced function of the positive signals that initiate and maintain eating. These results also suggest that perinatal malnutrition programmes obesity through a mechanism independent of its effects on feeding behaviour.     

Responsivity to NPY and melanocortins in obese OLETF rats lacking CCK-A receptors

Otsuka Long-Evans Tokushima Fatty (OLETF) rat lacking CCK-A receptors are hyperphagic and obese. Previous work has demonstrated alterations in neuropeptide Y (NPY) and proopiomelanocortin (POMC) mRNA expression in ad libitum fed OLETF rats compared to lean Long-Evans Tokushima Otsuka (LETO) controls. In order to determine whether alterations in sensitivity to central peptides involved in overall feeding control may contribute to the hyperphagia and obesity in OLETF rats, we assessed OLETF and LETO rats feeding responses to lateral ventricular infusions of NPY (1 and 3.2 nmol), the melanocortin 3/4 agonist MTII (0.1 and 0.32 nmol) and the melanocortin receptor antagonist SHU-9119 (0.25 and 0.5 nmol). At a 3-h time point, NPY increased food intake in both OLETF and LETO rats. OLETF rats were more sensitive, having significant increases at both NPY doses and a greater increase at the higher dose. The melanocortin agonist MTII decreased intake in both LETO and OLETF rats. At the 20-h time point, the magnitude of suppression was greater in OLETF rats. SHU-9119 increased food intake in both groups. OLETF rats were more sensitive with larger relative increase and longer-lasting effects at the lower dose. These results are consistent with demonstrated alterations in neuropeptide gene expression in OLETF rats and indicate that alterations in responsivity to NPY and melanocortin signaling are unlikely to contribute to their hyperphagia and obesity.     

Response to acute food deprivation in OLETF rats lacking CCK-A receptors

Otsuka Long-Evans Tokushima Fatty (OLETF) rats lacking cholecystokinin (CCK)-A receptors are hyperphagic and obese, and exhibit deficits in meal size control and in neuropeptide Y (NPY) gene expression in the dorsomedial hypothalamus (DMH). The present study was intended to determine whether these deficits would affect OLETF rat's response to an acute 24-h period of food deprivation. OLETF rats lost more body weight in response to deprivation but recovered their weight more quickly during refeeding than did lean Long-Evans Tokushima Otsuka (LETO) rats. Food deprivation decreased plasma glucose and leptin levels to a similar degree in both strains. Both groups increased intake during refeeding but the magnitude of increase was significantly greater in OLETF rats. Deprivation resulted in a significant elevation in arcuate NPY gene expression (approximately 47%) in LETO rats but only produced a small nonsignificant increase in the already decreased level of expression in OLETF rats (approximately 24%, P>.05). DMH NPY gene expression was not changed by deprivation in either OLETF or LETO rats. Although paraventricular corticotropin-releasing factor (CRF) expression was decreased by deprivation in LETO rats, CRF expression was not affected in OLETF rats. Together, these data suggested that OLETF rats lacking CCK-A receptors are not only capable of increasing their food intake in response to food deprivation, but also exhibit differential sensitivity to the effects of deprivation during both the food deprivation and refeeding periods.     

Trypsin inhibitor effects on food intake and weight gain in Zucker rats

Decreased body weight and increased pancreas weight which occur in rats fed raw soybeans are thought to be due to the presence of trypsin inhibitors in the soybeans (SBTI). Since trypsin is postulated to be a negative feedback signal for cholecystokinin (CCK) secretion, SBTI may have these effects by increasing secretion of CCK. CCK is a putative satiety signal; thus, increased secretion of CCK could decrease food intake, and, if maintained over a period of time, body weight. In these experiments the effects of a trypsin inhibitor [N,N-dimethyl-carbamoyl 4-(4-guanidino-benzylyloxy)-phenyl acetate methane-sulfate (DGPM)]on feeding pattern were investigated in Zucker obese and lean rats. Administration of 25-200 mg/kg DGPM to 6-hr fasted rats decreased daily food intake by dose-dependently decreasing average meal size in both obese and lean rats, but the response was greater in obese rats. Administration of 100 mg/kg DGPM twice daily for 7 days decreased food intake and body weight in obese but not lean rats. Thus, these results suggest that decreased body weight associated with SBTI is due to decreased food intake partly as a result of increased secretion of the putative satiety peptide CCK.     

Feeding and drinking behavior responses of adult Zucker obese rats to cholecystokinin

Increased meal size in obese animals may occur because of decreased sensitivity to satiety factors. Feeding and drinking behavior responses of Zucker obese and lean rats to two forms of cholecystokinin (CCK), a putative satiety factor, were compared in these experiments. Octapeptide of CCK (CCK-8) in both obese and lean rats decreased meal size (47 and 65%) and rate of eating but not meal duration, and increased satiety ratio but not postmeal interval. Impure CCK decreased first meal size after a 6-hr fast similarly in obese and lean rats (33 and 40%, respectively); however, meal duration was decreased only in lean rats, and rate of eating was decreased only in obese rats. Postmeal interval was decreased in lean rats, while satiety ratio was increased only in obese rats. In spite of decreased first meal size daily food intakes were increased by both impure CCK and CCK-8 in lean rats. In 6-hr water-deprived rats injected with CCK-8, decreased water intake was associated with decreased food intake. However, while impure CCK in lean rats elicited similar responses as CCK-8, impure CCK in obese rats decreased food but not water intake. Feeding behavior response to the putative satiety agent, cholecystokinin, depended on form of the peptide administered and phenotype.     

Sleep deprivation of rats: the hyperphagic response is real

STUDY OBJECTIVES: Chronic sleep deprivation of rats causes hyperphagia without body weight gain. Sleep deprivation hyperphagia is prompted by changes in pathways governing food intake; hyperphagia may be adaptive to sleep deprivation hypermetabolism. A recent paper suggested that sleep deprivation might inhibit ability of rats to increase food intake and that hyperphagia may be an artifact of uncorrected chow spillage. To resolve this, a palatable liquid diet (Ensure) was used where spillage is insignificant. DESIGN: Sleep deprivation of male Sprague Dawley rats was enforced for 10 days by the flowerpot/platform paradigm. Daily food intake and body weight were measured. On day 10, rats were transcardially perfused for analysis of hypothalamic mRNA expression of the orexigen, neuropeptide Y (NPY). SETTING: Morgan State University, sleep deprivation and transcardial perfusion; University of Maryland, NPY in situ hybridization and analysis. MEASUREMENTS AND RESULTS: Using a liquid diet for accurate daily measurements, there was no change in food intake in the first 5 days of sleep deprivation. Importantly, from days 6-10 it increased significantly, peaking at 29% above baseline. Control rats steadily gained weight but sleep-deprived rats did not. Hypothalamic NPY mRNA levels were positively correlated to stimulation of food intake and negatively correlated with changes in body weight. CONCLUSION: Sleep deprivation hyperphagia may not be apparent over the short term (i.e., < or = 5 days), but when extended beyond 6 days, it is readily observed. The timing of changes in body weight and food intake suggests that the negative energy balance induced by sleep deprivation prompts the neural changes that evoke hyperphagia.     

Spontaneous meal patterns in female rats with and without access to running wheels

Rats display strong behavioral rhythms during the ovarian cycle. During estrus, food intake is minimal due to a decrease in meal size, and locomotor activity is maximal. To investigate how activity influences feeding patterns across the ovarian cycle, we used a computerized system to monitor spontaneous meal patterns in intact, cycling female rats with and without access to running wheels. We found that running wheel access decreased dark meal frequency, increased dark meal size, and increased 24-h water intake during each phase of the ovarian cycle. In contrast, body weight, 24-h food intake, and the ovarian rhythms of reduced food intake, meal size, and body weight during estrus were not affected by running wheel access. In particular, the reduction in food intake during estrus was due to a selective reduction in dark meal size, not dark meal frequency, and this occurred independent of wheel access. These data indicate that estrus-related changes in spontaneous meal patterns and locomotor activity are independently controlled and that the reduction in food intake during estrus involves a selective change in the neurobiological controls of meal size.     

Lack of satiety effect of cholecystokinin (CCK) in a new rat model not expressing the CCK-A receptor gene

This work expands recent observations that Otsuka Long-Evans Tokushima Fatty (OLETF) rats show little or no pancreatic expression of the cholecystokinin (CCK)-A receptor gene. We examined whether the CCK-A and -B receptor genes were expressed in the brain (hypothalamus) of OLETF rats in comparison with control (Long-Evans Tokushima Otsuka = LET) rats. CCK-A receptor mRNA was detected in the hypothalamus of LETO rats but not OLETF rats. The CCK-B receptor gene was expressed in the hypothalamus in both strains. Cerebroventricular administration of CCK-8 sulfate inhibited daily food intake in LETO rats, but not in OLETF rats. These results show that in OLETF rats the absence of CCK-A receptor gene expression in the hypothalamus results in hyperphagia because of lack of satiety.     

Patterns of eating as a function of the cost of the meal

Rats were required to complete fixed ratio schedules (FR 20-FR 2560) of wheel turns to obtain access to food. By decreasing meal frequency and increasing meal size directly as functions of the fixed ratio requirement, animals controlled total daily food intake and body weight relatively constant until the highest ratio requirement was introduced. These functional changes in feeding patterns provide experimental support for theoretical models of optimal feeding strategies. At the highest ratio requirement, as animals lost weight, they increased running and therefore opportunities to feed, however, food intake continued to decrease with increasing exposure to this schedule. As rats on this schedule initiated feeding each time food became available, but did not eat large enough meals to maintain body weight, it is suggested that activity may interact with satiety mechanisms to produce termination of meals.     

Decreased hypothalamic concentration of neuropeptide Y correlates with onset of hyperphagia in fa/fa rats on postnatal day 12

An increased action of hypothalamic neuropeptide Y (NPY) has been proposed as a major factor in the pathophysiology of the obesity syndrome in Zucker (fa/fa) rats. Using a developmental strategy to test this hypothesis, we showed previously that significantly more arcuate NPY was expressed in fa/fa pups than in lean littermates on postnatal day (P) 2 and throughout the preweaning period [Physiol. Behav. 67 (1999) 521], and that hyperphagia first appeared on P12 [Am. J. Physiol. 275 (1998) R1106]. To test the hypothesis further, we used a specific radioimmunoassay to measure the concentration of hypothalamic NPY peptide in lean (+/+ and +/fa) and obese fa/fa Zucker rat pups on P9, P10, and P12. The concentration of NPY in fa/fa pups was not significantly different from that of the other genotypes. There was, however, a significant decrease in NPY concentration from P9 to P12 in fa/fa pups, but not in lean pups. The combination of increased NPY message and decreasing concentration of NPY peptide in fa/fa pups with age is consistent with, but does not prove, increased release of hypothalamic NPY in fa/fa pups just before and on P12 when hyperphagia emerges. These results provide further support for the importance of hypothalamic NPY in the phenotypic expression of hyperphagia in the fa/fa pups during the second postnatal week.     

Development of, and recovery from, activity-based anorexia in female rats

Activity-based anorexia occurs in rats maintained on a restricted-feeding schedule while given free access to running wheels. These conditions induce high levels of wheel running and rapid weight loss. Although this procedure was developed as an animal model of anorexia nervosa, it has been studied primarily in male rats. Our goal was to examine the development of, and recovery from, activity-based anorexia in female rats. Food intake, wheel running, body weight, and phase of the estrous cycle were monitored daily prior to, during, and after a period of restricted feeding in which access to food was limited to 2 h/day. Food intake, body weight, and estrous cyclicity were also monitored in a control group housed without access to running wheels. Prior to food restriction, rats with wheels displayed high levels of wheel running and consumed more food than rats without wheels. Despite that both groups consumed similar amounts of food during the restricted-feeding phase, only rats with wheels developed symptoms of activity-based anorexia, including increased wheel running, rapid weight loss, and disruptions in estrous cyclicity. Recovery from activity-based anorexia was associated with hypoactivity and hyperphagia. Resumption of estrous cycles occurred when the weight lost during food restriction was regained. Hyperphagia, but not hypoactivity, was maintained following resumption of estrous cycles; however, this hyperphagia was limited to nonestrous phases. Our findings suggest that recovery from activity-based anorexia is mediated primarily by an increase in orexigenic signaling that promotes pronounced hyperphagia, and that the increase in satiogenic signaling during estrus abolishes this compensatory hyperphagia.     

Estrogens inhibit food intake in CCK-1 receptor-deficient rats

In human and many other animals, estrogens inhibit food intake and increases spontaneous activity. Previous studies hypothesized that the anorexigenic effect of estrogens is mediated by the cholecystokinin (CCK)-induced satiety effect. In the present study, we investigated whether estrogens-induced anorexigenic and hyper-active effects are present in Otsuka-Long-Evans-Tokushima-Fatty (OLETF) rat, which is deficient in the CCK1 receptor. In OLETF rats with a regular 4-day estrous cycle, food intake decreased and spontaneous activity increased significantly more during estrus than diestrus as compared to control Long-Evans-Tokushima-Otsuka (LETO) rats. Subcutaneous injection of estradiol benzoate into ovariectomized OLETF rats significantly decreased feeding and increased spontaneous activity to the same extent as in LETO rats. These results suggest that the anorexigenic and hyper-active effects of estrogen can be mediated via pathways other than CCK-CCK1 receptor signaling pathway in CCK1 receptor-deficient rats.     

Paraventricular hypothalamic clonidine increases rather than decreases susceptibility to activity-based anorexia in the rat.

Anorexia has been related to reduced activity of the paraventricular hypothalamic (PVN) noradrenergic-feeding system. In this study we determined whether clonidine (an alpha 2-adrenergic agonist) infused into the PVN reduced susceptibility to activity-based anorexia (ABA) in the rat. In Experiment 1, clonidine (6 doses) was chronically infused into the PVN of male Sprague-Dawley rats. All animals were exposed to ABA (1.5 hr/day food access; 22.5 hr/day running wheel access) until a 25% body weight loss was reached. Dose-related increases in susceptibility to ABA and decreases in food intake were observed. In Experiment 2, for which heavier animals and 3 doses of clonidine were used, we found no difference in food intake and wheel activity but increased susceptibility to ABA. Chronic clonidine infused into the PVN does not produce hyperphagia and exacerbates rather than attenuates susceptibility to ABA.     

Wheel running, food intake, and body weight in male rats

The acquisition of wheel running, its effects on food intake and body weight, and the effects of wheel deprivation, were examined in male rats. Running increased during the first 15 days of access, then plateaued. When wheels were unlocked after 10 days of deprivation, running was reduced, but quickly recovered to original levels. Animals first given wheel access 49 days into the study ran little, with no increase over days. Food intake dropped each time with wheel access, but recovered to control levels over 10-14 days. Wheel deprivation resulted in a temporary hyperphagia. With wheel access, weight initially dropped and was then maintained at a reduced percentage of homecage-housed animals. In male rats wheel access appears to have temporary effects on food intake, and long term effects on weight. Marked differences in the activity of same-age rats suggest that wheel running is in part a function of housing history.     

Oral preloads of liquid diet and hormonal satiety mechanisms

A hormonal short-term satiety mechanism involving cholecystokinin (CCK) has recently been proposed. We attempted to manipulate endogenous CCK levels in rats by administering oral preloads at varying time intervals before a test meal. Support for the CCK hypothesis was equivocal. In addition, there was little support for other hypothesized satiety mechanisms, such as regulation of volume or calories, or glucose-related mechanisms. The results are discussed in terms of the necessity of considering the total “ecological” situation (diet composition, diet palatability, diet availability, and training) when studying food intake and body weight.     

Appetite suppression through delayed fat digestion

High-fat diets are often associated with greater caloric intake and weight gain. Since satiety during fat intake is induced by fat in the intestine we investigated the efficiency of a lipid compound that retards fat digestion to regulate fat intake. We found this compound to reduce high-fat food intake, body weight and blood lipids in Sprague-Dawley rats, without causing steatorrhea. The absence of steatorrhea is explained by an increased pancreatic lipase/colipase secretion, compensating the impaired lipolysis by the added compound. The animals also had an elevated CCK secretion. The satiety for fat may be the consequence of elevated CCK and procolipase/enterostatin levels. We conclude that compounds can be found that delay intestinal fat digestion and control high-fat food intake through the release of satiety signals, without causing steatorrhea. The absence of steatorrhea makes such compounds advantageous over lipase inhibitors in the treatment of obesity.     

Repeated hypothalamic stimulation with neuropeptide Y increases daily carbohydrate and fat intake and body weight gain in female rats

Neuropeptide Y (NPY), repeatedly injected in the hypothalamic paraventricular nucleus (PVN), produces dramatic obesity and overeating in female rats maintained on a single nutritionally complete diet. In the present study, we investigated whether these effects could also be obtained in animals with a choice of three pure macronutrients: protein, carbohydrate, and fat. Female rats with indwelling PVN cannulas were injected with NPY (235 pmol) or its saline vehicle every 8 hr for 6 days. A third group was left undisturbed. Consumption of each macronutrient and body weight were measured every 24 hr for 6 days preinjection, 6 days during injections, and 21 days after the injections were terminated. Relative to vehicle or preinjection rates of body weight gain (approximately 1.5 g/day), NPY dramatically enhanced weight gain to a rate of 9.3 g/day and more than doubled total daily food intake. This augmentation was accounted for by increases in carbohydrate intake (+26.4 kcal/day) and fat intake (+48.5 kcal/day), with no significant potentiation of protein consumption. When the NPY injections were terminated, body weight and macronutrient intake returned to control levels within 1 or 2 weeks. These findings are consistent with a role for NPY in hypothalamic mechanisms of macronutrient intake and body weight regulation and suggest that disturbances in brain NPY may contribute to the development of eating and weight disorders.     

Interrelationships among activity,food intake and weight gain in genetically obese and lean Zucker rats

In Experiment 1, food deprivation resulting in a 30% reduction in body weight produced significant increases in wheel running in both obese and lean female Zucker rats. In Experiment 2, a new technique, food contingent activity (FR, VI), dramatically increased wheel running in both obese and lean female Zucker rats. This increase in activity was achieved primarily during the dark period. Regardless of changes in activity levels, food intake and body weight gain remained similar to controls. When food was again available ad lib, activity levels rapidly decreased for obese but not lean rats. These results indicate that behavioral interventions alone are not sufficient to correct the obesity of the genetically obese rat.