Additive feeding inhibitory and aversive effects of naltrexone and exendin-4 combinations.

OBJECTIVE: One developing strategy for obesity treatment has been to use combinations of differently acting pharmacotherapies to improve weight loss with fewer adverse effects. The purpose of this study was to determine whether the combination of naltrexone (Nal), an opioid antagonist acting on the reward system, and exendin-4 (Ex-4), a glucagon-like peptide 1 agonist acting on satiety signaling, would produce larger reductions in food intake than either alone in rats. Because the anorectic potencies of both compounds have been associated with nausea and malaise, the influence of these drug combinations on the acquisition of a conditioned taste aversion (CTA) was also determined. METHODS: In Experiment 1, the acute anorectic effects of Nal (0.32-3.2 mg kg(-1); intraperitoneally (i.p.)) and Ex-4 (1-10 µg kg(-1); i.p.) were assessed alone or in combination. Combinational doses were further investigated by the repeated daily administration of 1 mg kg(-1) Nal+3.2 µg kg(-1) Ex-4 for 4 days. In Experiment 2, both compounds alone or in combination were used as unconditioned stimuli in a series of CTA tests. RESULTS: Nal and Ex-4, alone or in combination, suppressed food intake in a dose-dependent manner, and the interaction on food intake between Nal and Ex-4 was additive. In the CTA paradigm, Nal (1 mg kg(-1)) alone did not support acquisition, whereas a CTA was evident with doses of Ex-4 (1 or 3.2 µg kg(-1)). Combinations of Nal and Ex-4 also resulted in a more rapid and robust acquisition of a CTA. CONCLUSION: Given that the Nal and Ex-4 combination produces additive effects on not only food intake reduction but also food aversion learning, this specific drug combination does not have the benefit of minimizing the adverse effects associated with each individual drug. These data suggest that it is necessary to evaluate both the positive and adverse effects at early stages of combinational drug development.

A mouse model for Down syndrome exhibits learning and behaviour deficits.

Trisomy 21 or Down syndrome (DS) is the most frequent genetic cause of mental retardation, affecting one in 800 live born human beings. Mice with segmental trisomy 16 (Ts65Dn mice) are at dosage imbalance for genes corresponding to those on human chromosome 21q21-22.3--which includes the so-called DS 'critical region'. They do not show early-onset of Alzheimer disease pathology; however, Ts65Dn mice do demonstrate impaired performance in a complex learning task requiring the integration of visual and spatial information. The reproducibility of this phenotype among Ts65Dn mice indicates that dosage imbalance for a gene or genes in this region contributes to this impairment. The corresponding dosage imbalance for the human homologues of these genes may contribute to cognitive deficits in DS.

Synphilin-1 alters metabolic homeostasis in a novel Drosophila obesity model.

AIMS: The pathogenesis of obesity remains incompletely understood. Drosophila have conserved neuroendocrine and digestion systems with human and become an excellent system for studying energy homeostasis. Here, we reported a novel obesity Drosophila model, in which expression of human protein, synphilin-1 (SP1), in neurons fosters positive energy balance. SUBJECTS AND METHODS: To further understand the actions of SP1 in energy balance control, the upstream activation sequence UAS/GAL4 system was used to generate human SP1 transgenic Drosophila. We characterized a human SP1 transgenic Drosophila by assessing SP1 expression, fat lipid deposition, food intake and fly locomotor activity to determine the major behavioral changes and their consequences in the development of the obesity-like phenotype. RESULTS: Overexpression of SP1 in neurons, but not peripheral cells, increased the body weight of flies compared with that of non-transgenic controls. SP1 increased food intake but did not affect locomotor activity. SP1 increased the levels of triacylglycerol, and the size of fat body cells and lipid droplets, indicating that SP1 increased lipid-fat disposition. Survival studies showed that SP1 transgenic flies were more resistant to food deprivation. SP1 regulated lipin gene expression that may participate in SP1-induced fat deposition and starvation resistance. CONCLUSION: These studies demonstrate that SP1 expression affects energy homeostasis in ways that enhance positive energy balance and provide a useful obesity model for future pathogenesis and therapeutic studies.

A novel obesity model: synphilin-1-induced hyperphagia and obesity in mice.

AIMS: The pathogenesis of obesity remains incompletely understood and the exploration of the role of novel proteins in obesity may provide important insights into its causes and treatments. Here, we report a previously unidentified role for synphilin-1 in the control of food intake and body weight. Synphilin-1, a cytoplasmic protein, was initially identified as an interaction partner of alpha-synuclein, and has implications in Parkinson's disease pathogenesis related to protein aggregation. SUBJECTS AND METHODS: To study the in vivo role of synphilin-1, we characterized a human synphilin-1 transgenic mouse (SP1) by assessing synphilin-1 expression, plasma parameters, food intake and spontaneous activity to determine the major behavioral changes and their consequences in the development of the obesity phenotype. RESULTS: Expression of human synphilin-1 in brain neurons in SP1 mice resulted in increased food intake, body weight and body fat. SP1 mice also displayed hyperinsulinemia, hyperleptinemia and impaired glucose tolerance. Pair-feeding SP1 mice to amounts consumed by non-transgenic mice prevented the increased body weight, adiposity, hyperinsulinemia and hyperleptinemia demonstrating that these were all the consequences of increased food intake. Transgenic expression of synphilin-1 was enriched in hypothalamic nuclei involved in feeding control, and fasting-induced elevated endogenous synphilin-1 levels at these sites, suggesting that synphilin-1 is an important player in the hypothalamic energy balance regulatory system. CONCLUSION: These studies identify a novel function of synphilin-1 in controlling food intake and body weight, and may provide a unique obesity model for future studies of obesity pathogenesis and therapeutics.

Physiology: does gut hormone PYY3-36 decrease food intake in rodents?

Batterham et al. report that the gut peptide hormone PYY3-36 decreases food intake and body-weight gain in rodents, a discovery that has been heralded as potentially offering a new therapy for obesity. However, we have been unable to replicate their results. Although the reasons for this discrepancy remain undetermined, an effective anti-obesity drug ultimately must produce its effects across a range of situations. The fact that the findings of Batterham et al. cannot easily be replicated calls into question the potential value of an anti-obesity approach that is based on administration of PYY3-36.

Gut peptides in the control of food intake.

Multiple gut peptides are involved in the overall control of food intake. Plasma levels of gut peptides are differentially affected by food intake, and the different patterns of release around meals provides an indication of a peptide's specific role in feeding control. Ghrelin is a gastric peptide whose plasma levels are high before meals and are suppressed in response to food intake. Consistent with this pattern, ghrelin has been shown to stimulate food intake by hastening meal initiations. Cholecystokinin (CCK) is released from upper intestinal sites in response to food intake. CCK inhibits eating in a manner consistent with a role in satiety. Pancreatic glucagon and amylin play similar roles in meal termination. In contrast, the lower gut peptides, peptide YY (3-36) and glucagon-like peptide 1, are released more slowly in response to food intake and levels remain elevated for hours after a meal. This pattern of release suggests effects across multiple meals, and these peptides have been shown to inhibit food intake by both decreasing meal size and increasing the satiating potency of consumed nutrients. Together, these actions indicate multiple roles for gut peptides in feeding control.

Satiety: a graded behavioural phenomenon regulating caloric intake.

Rhesus monkeys inhibited their feeding in response to intragastric nutrient preloads, so as to maintain a constant caloric intake. There is a definite dose- response relationship to calories which does not depend on the volume, caloric concentration, or nature of the nutrient. This accurate control is accomplished even though some of the preload remains in the stomach throughout the feeding period.

Intracranial self-stimulation in 3-day-old rat pups.

Three-day-old rat pups with electrodes directed at the medial forebrain bundle at the level of the lateral hypothalamic area were trained to push a paddle to receive electrical brain stimulation. Pups receiving stimulation that was contingent on lifting the paddle responded more frequently than did control pups and also learned a two-choice spatial discrimination task that was rewarded with brain stimulation. The experiments indicate that a neural substrate in the area of the medial forebrain bundle is involved in the central mediation of reinforcement in the rat pup.

Adaptation to lactation in OLETF rats lacking CCK-1 receptors: body weight, fat tissues, leptin and oxytocin.

OBJECTIVE: To understand the adaptation to lactation of obese rats, by studying the interplay among the gut hormone cholecystokinin (CCK), the adiposity hormone leptin and the affiliation hormone oxytocin in modulating body mass and fat storage. DESIGN: Strain differences were examined between Otsuka Long Evans Tokushima Fatty (OLETF) rats lacking expression of functional CCK-1 receptors and Long Evans Tokushima Otsuka (LETO) controls, tested as nulliparous dams, at the 7 and 15th lactation day, at weaning (lactation day 22) or 8 weeks postweaning. MEASUREMENTS: We measured body mass, fat pads (brown, retroperitoneal and inguinal) and inguinal adipocytes. Plasma levels of leptin and oxytocin were determined. RESULTS: Fat depots of LETO female rats were larger during lactation compared to the levels found in postweaning and nulliparous female rats. LETO female rats gained weight and accumulated fat during pregnancy and lactation, returning to their normal fat levels postweaning. In contrast, OLETF female rats presented lower body weight and fat depots during the lactation period than nulliparous dams, and regained the weight and fat postweaning. Plasma leptin and oxytocin were highly correlated and followed the same pattern. OLETF leptin levels were highly correlated with fat depot and inguinal cell surface. No significant correlation was found for LETO parameters. CONCLUSIONS: Pregnancy and lactation are energy-consuming events, which naturally induce female rats to increase food intake and accumulate fat. When challenged by the demands of rapidly growing preobese OLETF pups, OLETF dams' fat stores are reduced to lean, LETO levels. During lactation, sensitivity of the oxytocinergic neurons descending from the paraventricular nuclei to the nucleus of the solitary tract to CCK is reduced. We theorized that this pathway is not available to OLETF female rats that lack functional CCK-1 receptors to mediate the signal. The current study contributes to the understanding of the female body's adaptation to lactation.

Characterization of the feeding inhibition and neural activation produced by dorsomedial hypothalamic cholecystokinin administration.

Within the dorsomedial hypothalamus (DMH), cholecystokinin (CCK) has been proposed to modulate neuropeptide Y (NPY) signaling to affect food intake. However, the neural circuitry underlying the actions of this CCK-NPY signaling system in the controls of food intake has yet to be determined. We sought to characterize the feeding inhibition and brain neural activation produced by CCK administration into the DMH of rats. We determined the time course of feeding inhibitory effects of exogenous DMH CCK, assessed NPY gene expression in the DMH in response to DMH CCK administration, and characterized c-Fos activation in the entire brain induced by CCK injection into the DMH using c-Fos like immunohistochemistry. We found that parenchymal injection of CCK into the DMH decreased food intake during the entire 22 h observation period, with a primary effect in the first 4 h, and down-regulated NPY gene expression in the DMH. c-Fos immunohistochemistry revealed that DMH CCK increased the number of c-Fos positive cells in the paraventricular nucleus (PVN), arcuate nucleus, suprachiasmatic nucleus and retrochiasmatic area as well as in the contralateral DMH. This pattern of activity is different from that produced by peripherally administered CCK which is short acting and primarily activates neurons in the nucleus of the solitary tract and area postrema, as well as the PVN and DMH. Together, these data suggest that DMH CCK plays an important role in the control of food intake, and does so by activating different pathways from those activated by peripheral CCK.

Effects of meal composition on postprandial incretin, glucose and insulin responses after surgical and medical weight loss.

BACKGROUND: Meal tolerance tests are frequently used to study dynamic incretin and insulin responses in the postprandial state; however, the optimal meal that is best tolerated and suited for hormonal response following surgical and medical weight loss has yet to be determined. OBJECTIVE: To evaluate the tolerability and effectiveness of different test meals in inducing detectable changes in markers of glucose metabolism in individuals who have undergone a weight loss intervention. METHODS: Six individuals who underwent surgical or medical weight loss (two Roux-en-Y gastric bypass, two sleeve gastrectomy and two medical weight loss) each completed three meal tolerance tests using liquid-mixed, solid-mixed and high-fat test meals. The tolerability of each test meal, as determined by the total amount consumed and palatability, as well as fasting and meal-stimulated glucagon-like peptide, glucose-dependent insulinotropic polypeptide, insulin and glucose were measured. RESULTS: Among the six individuals, the liquid-mixed meal was better and more uniformly tolerated with a median meal completion rate of 99%. Among the four bariatric surgical patients, liquid-mixed meal stimulated on average a higher glucagon-like peptide (percent difference: 83.7, 89), insulin secretion (percent difference: 155.1, 158.7) and glucose-dependent insulinotropic polypeptide (percent difference: 113.5, 34.3) compared with solid-mixed and high-fat meals. CONCLUSIONS: The liquid-mixed meal was better tolerated with higher incretin and insulin response compared with the high-fat and solid-mixed meals and is best suited for the evaluation of stimulated glucose homeostasis.

Search for common targets of lithium and valproic acid identifies novel epigenetic effects of lithium on the rat leptin receptor gene.

Epigenetics may have an important role in mood stabilizer action. Valproic acid (VPA) is a histone deacetylase inhibitor, and lithium (Li) may have downstream epigenetic actions. To identify genes commonly affected by both mood stabilizers and to assess potential epigenetic mechanisms that may be involved in their mechanism of action, we administered Li (N = 12), VPA (N = 12), and normal chow (N = 12) to Brown Norway rats for 30 days. Genomic DNA and mRNA were extracted from the hippocampus. We used the mRNA to perform gene expression analysis on Affymetrix microarray chips, and for genes commonly regulated by both Li and VPA, we validated expression levels using quantitative real-time PCR. To identify potential mechanisms underlying expression changes, genomic DNA was bisulfite treated for pyrosequencing of key CpG island 'shores' and promoter regions, and chromatin was prepared from both hippocampal tissue and a hippocampal-derived cell line to assess modifications of histones. For most genes, we found little evidence of DNA methylation changes in response to the medications. However, we detected histone H3 methylation and acetylation in the leptin receptor gene, Lepr, following treatment with both drugs. VPA-mediated effects on histones are well established, whereas the Li effects constitute a novel mechanism of transcriptional derepression for this drug. These data support several shared transcriptional targets of Li and VPA, and provide evidence suggesting leptin signaling as an epigenetic target of two mood stabilizers. Additional work could help clarify whether leptin signaling in the brain has a role in the therapeutic action of Li and VPA in bipolar disorder.

Sub-diaphragmatic vagal afferent integration of meal-related gastrointestinal signals.

We have established a method to investigate the range of mechanical, nutrient chemical and peptidergic meal-related stimuli t hat may generate vagal afferent neurophysiological signals critical to the negative feedback control of food intake in the rat. We have identified populations of fibers that respond with increased neurophysiological discharge rates to gastric loads, duodenal loads, and close celiac arterial administration of a brain-gut peptide, cholecystokinin. Load-sensitive fibers with gastric and duodenal mechanoreceptive fields are able to integrate information arising from mechanical and peptidergic stimulation, where cholecystokinin octapeptide (CCK) administration potentiates subsequent responses to distending loads, and synergizes with distending loads to produce greater excitation than either load stimulus alone or peptide stimulation alone. In addition, we have identified situations where the duodenal presence of nutrients modifies the vagal afferent activity of gastric load-sensitive fibers. Thus, our approach can mimic the temporal and spatial distribution of meal-related stimuli in the gut, and reveals the potential for nutrients in one gastrointestinal compartment to affect neutral signals arising from another gut compartment.

Caloric compensation for lunches varying in fat and carbohydrate content by humans in a residential laboratory.

Two groups of three subjects participated in a residential study that assessed the effects of varying the macronutrient and caloric content of a required lunch meal on subsequent food choice and intake. Lunches contained 431 or 844 kcal, with the caloric differential created by manipulating the calories derived from either fat or carbohydrate (CHO). Each lunch condition (high-fat, high-CHO, low-fat, and low-CHO) was examined for 3 consecutive days. Subjects controlled their own patterns of food intake and could consume any item or number of items at any time during the day or night. There were no significant differences in total daily caloric intake across conditions, indicating that subjects compensated for the caloric content of the lunch regardless of the macronutrient content. Total daily caloric intake under the high-fat and high-CHO conditions was 2824 +/- 151 (mean +/- SEM) and 2988 +/- 187 kcal, respectively, whereas intake under the low-fat and low-CHO conditions was 2700 +/- 131 and 2890 +/- 247 kcal, respectively.

Satiety after preloads with different amounts of fat and carbohydrate: implications for obesity.

High intake of dietary fat may be key in both the etiology and maintenance of obesity. Because a reduction in the proportion of energy derived from fat will be accompanied by an increase in the proportion of energy derived from carbohydrate, this study compared the effects of these macronutrients on eating behavior in obese and lean individuals. The effects of different amounts of fat and carbohydrate, covertly incorporated into yogurt preloads, on subsequent food intake, hunger, and satiety were assessed. A group of 12 normal-weight men, unconcerned about eating and body weight (unrestrained), accurately compensated for the energy in the preloads regardless of the nutrient composition. Other groups (n = 12 per group), including normal-weight restrained men and normal-weight and obese restrained and unrestrained females, did not show such orderly energy compensation; joule-for-joule, the high-fat preloads suppressed intake at lunch less than did high-carbohydrate preloads. These results suggest that a relative insensitivity to the satiating effect of fat could be involved in the development and maintenance of obesity.

Caloric, but not macronutrient, compensation by humans for required-eating occasions with meals and snack varying in fat and carbohydrate.

Six subjects participated in a residential study assessing the effects of covert macronutrient and energy manipulations during three required-eating occasions (breakfast, lunch, and afternoon snack) on total macronutrient and energy intakes. Overall, energy content of the occasions varied between approximately 3000 and approximately 7000 kJ (approximately 700 and approximately 1700 kcal) with the majority of the differential derived from either fat or carbohydrate (CHO). Each condition (high, medium, and low fat; high, medium, and low CHO; and no required eating) was examined for 2 d. Subjects compensated for the energy content of the required occasions such that only under the low-CHO condition (11,297 +/- 3314 kJ) was total daily energy intake lower than that observed in the absence of required occasions (13,297 +/- 1356 kJ). Only total energy intake under the high-fat condition (12,326 +/- 2548 kJ) was significantly different from its matched CHO condition (high-CHO condition: 14,665 +/- 2686 kJ). In contrast to the clear evidence for caloric compensation, there were no differential effects of condition on macronutrient intake, ie, there was no macronutrient compensation.

Potent and sustained satiety actions of a cholecystokinin octapeptide analogue.

The relative ability of a norleucine substituted cholecystokinin (CCK) analogue, U-67827E, to interact with CCK receptors and to inhibit food intake was examined across a variety of paradigms. U-67827E and CCK had identical in vitro potencies as demonstrated by their ability to induce pyloric contractions or competitively inhibit [125I]CCK-8 binding to type A and B CCK receptors. However, the in vivo potency of U-67827E was significantly greater than that of CCK-8. In rats, U-67827E inhibited food intake with 10-100 times the potency of CCK. In rhesus monkeys, U-67827E produced significantly greater inhibitions of daily food intake and did so in a dose-dependent manner with no evidence of compensation or tolerance. U-67827E also inhibited gastric emptying for significantly longer durations than CCK. Together these results demonstrate that CCK analogues with increased in vivo bioavailability can affect food intake beyond a single meal.

Food intake, hunger, and satiety after preloads in women with eating disorders.

Food intake, food selection, macronutrient intake, sensory-specific satiety, and ratings of hunger and satiety were measured after high- and low-energy salad preloads (2414 kJ, or 172 kJ) or no preload to determine whether patients with eating disorders compensate appropriately for different energy intakes. Subjects were female patients with a DSM-III-R diagnosis of anorexia nervosa with bulimic features or bulimia nervosa, or non patient, normal-weight, nondieters (n = 9/group). At a self-selected lunch 30 min after the preloads, all of the groups reduced intake after the high-energy preload, with the bulimics showing the best compensation. The anorexics chose low-energy foods and in some conditions ate a smaller proportion of fat than did the other groups. The bulimics ate more high-energy foods than did the anorexics. The anorexics demonstrated sensory-specific satiety only after the high-energy salad and the bulimics only after the low-energy salad. Overall, these data suggest that while many of their responses to food are abnormal, patients with eating disorders have some capacity to respond to physiological hunger and satiety cues.