Time-dependent changes in feeding behavior and energy balance associated with weight gain in mice fed obesogenic diets.

OBJECTIVE: Obesity is characterized by dysregulated homeostatic mechanisms resulting in positive energy balance; however, when this dysregulation occurs is unknown. We assessed the time course of alterations to behaviors promoting weight gain in male and female mice switched to an obesogenic high-fat diet (HFD). METHODS: Male and female C57BL/6J mice were housed in metabolic chambers and were switched from chow to a 60% or 45% HFD for 4 and 3 weeks, respectively. Food intake, meal patterns, energy expenditure (EE), and body weight were continuously measured. A separate cohort of male mice was switched from chow to a 60% HFD and was given access to locked or unlocked running wheels. RESULTS: Switching mice to obesogenic diets promotes transient bouts of hyperphagia during the first 2 weeks followed by persistent caloric hyperphagia. EE increases but not sufficiently enough to offset increased caloric intake, resulting in a sustained net positive energy balance. Hyperphagia is associated with consumption of calorically larger meals (impaired satiation) more frequently (impaired satiety), particularly during the light cycle. Running wheel exercise delays weight gain in male mice fed a 60% HFD by enhancing satiation and increasing EE. However, exercise effects on satiation are no longer apparent after 2 weeks, coinciding with weight gain. CONCLUSIONS: Exposure to obesogenic diets engages homeostatic regulatory mechanisms for ~2 weeks that ultimately fail, and consequent weight gain is characterized by impaired satiation and satiety. Insights into the etiology of obesity can be obtained by investigating changes to satiation and satiety mechanisms during the initial ~2 weeks of HFD exposure.

Time dependent changes in feeding behavior and energy balance associated with weight gain in mice fed obesogenic diets.

Obesity is characterized by dysregulated homeostatic mechanisms resulting in positive energy balance, yet when this dysregulation occurs is unknown. We assessed the time course of alterations to behaviors promoting weight gain in male and female mice switched to obesogenic 60% or 45% high fat diet (HFD). Switching mice to obesogenic diets promotes transient bouts of hyperphagia during the first 2 weeks followed by persistent caloric hyperphagia. Energy expenditure increases but not sufficiently to offset increased caloric intake, resulting in a sustained net positive energy balance. Hyperphagia is associated with consumption of calorically larger meals (impaired satiation) more frequently (impaired satiety) particularly during the light-cycle. Running wheel exercise delays weight gain in 60% HFD-fed male mice by enhancing satiation and increasing energy expenditure. However, exercise effects on satiation are no longer apparent after 2 weeks, coinciding with weight gain. Thus, exposure to obesogenic diets engages homeostatic regulatory mechanisms for ∼2 weeks that ultimately fail, and consequent weight gain is characterized by impaired satiation and satiety. Insights into the etiology of obesity can be obtained by investigating changes to satiation and satiety mechanisms during the initial ∼2 weeks of HFD exposure. What is already known about this subject?: Obesity is associated with dysregulated homeostatic mechanisms.Increased caloric consumption contributes to obesity.Obese rodents tend to eat larger, more frequent meals. What are the new findings in your manuscript?: Exposure to obesogenic diets promotes transient attempts to maintain weight homeostasis.After ∼2 weeks, caloric hyperphagia exceeds increased energy expenditure, promoting weight gain.This is associated with consumption of larger, more frequent meals. How might your results change the direction of research or the focus of clinical practice?: Our findings suggest that molecular studies focusing on mechanisms that regulate meal size and frequency, particularly those engaged during the first ∼2 weeks of obesogenic diet feeding that eventually fail, can provide unique insight into the etiology of obesity.

Fibroblast growth factor-21 is required for weight loss induced by the glucagon-like peptide-1 receptor agonist liraglutide in male mice fed high carbohydrate diets.

OBJECTIVE: Glucagon-like peptide-1 receptor (GLP-1R) agonists (GLP-1RA) and fibroblast growth factor-21 (FGF21) confer similar metabolic benefits. GLP-1RA induce FGF21, leading us to investigate mechanisms engaged by the GLP-1RA liraglutide to increase FGF21 levels and the metabolic relevance of liraglutide-induced FGF21. METHODS: Circulating FGF21 levels were measured in fasted male C57BL/6J, neuronal GLP-1R knockout, ß-cell GLP-1R knockout, and liver peroxisome proliferator-activated receptor alpha knockout mice treated acutely with liraglutide. To test the metabolic relevance of liver FGF21 in response to liraglutide, chow-fed control and liver Fgf21 knockout (LivFgf21-/-) mice were treated with vehicle or liraglutide in metabolic chambers. Body weight and composition, food intake, and energy expenditure were measured. Since FGF21 reduces carbohydrate intake, we measured body weight in mice fed matched diets with low- (LC) or high-carbohydrate (HC) content and in mice fed a high-fat, high-sugar (HFHS) diet. This was done in control and LivFgf21-/- mice and in mice lacking neuronal ß-klotho (Klb) expression to disrupt brain FGF21 signaling. RESULTS: Liraglutide increases FGF21 levels independently of decreased food intake via neuronal GLP-1R activation. Lack of liver Fgf21 expression confers resistance to liraglutide-induced weight loss due to attenuated reduction of food intake in chow-fed mice. Liraglutide-induced weight loss was impaired in LivFgf21-/- mice when fed HC and HFHS diets but not when fed a LC diet. Loss of neuronal Klb also attenuated liraglutide-induced weight loss in mice fed HC or HFHS diets. CONCLUSIONS: Our findings support a novel role for a GLP-1R-FGF21 axis in regulating body weight in a dietary carbohydrate-dependent manner.

Liver Fibroblast Growth Factor 21 (FGF21) is Required for the Full Anorectic Effect of the Glucagon-Like Peptide-1 Receptor Agonist Liraglutide in Male Mice fed High Carbohydrate Diets.

Glucagon-like peptide-1 receptor (GLP-1R) agonists and fibroblast growth factor 21 (FGF21) confer similar metabolic benefits. Studies report that GLP-1RA induce FGF21. Here, we investigated the mechanisms engaged by the GLP-1R agonist liraglutide to increase FGF21 levels and the metabolic relevance of liraglutide-induced FGF21. We show that liraglutide increases FGF21 levels via neuronal GLP-1R activation. We also demonstrate that lack of liver Fgf21 expression confers partial resistance to liraglutide-induced weight loss. Since FGF21 reduces carbohydrate intake, we tested whether the contribution of FGF21 to liraglutide-induced weight loss is dependent on dietary carbohydrate content. In control and liver Fgf21 knockout (Liv Fgf21 -/- ) mice fed calorically matched diets with low- (LC) or high-carbohydrate (HC) content, we found that only HC-fed Liv Fgf21 -/- mice were resistant to liraglutide-induced weight loss. Similarly, liraglutide-induced weight loss was partially impaired in Liv Fgf21 -/- mice fed a high-fat, high-sugar (HFHS) diet. Lastly, we show that loss of neuronal ß-klotho expression also diminishes liraglutide-induced weight loss in mice fed a HC or HFHS diet, indicating that FGF21 mediates liraglutide-induced weight loss via neuronal FGF21 action. Our findings support a novel role for a GLP-1R-FGF21 axis in regulating body weight in the presence of high dietary carbohydrate content.

High fat diet blunts stress-induced hypophagia and activation of Glp1r dorsal lateral septum neurons in male but not in female mice.

OBJECTIVE: While stress typically reduces caloric intake (hypophagia) in chow-fed rodents, presentation of palatable, high calorie substances during stress can increase caloric consumption (i.e. "comfort feeding") and promote obesity. However, little is known about how obesity itself affects feeding behavior in response to stress and the mechanisms that can influence stress-associated feeding in the context of obesity. METHODS: We assessed food intake and other metabolic parameters in lean and obese male and female mice following acute restraint stress. We also measured real-time activity of glucagon-like peptide-1 (Glp1) receptor (Glp1r)-expressing neurons in the dorsal lateral septum (dLS) during stress in lean and obese mice using fiber photometry. Glp1r activation in various brain regions, including the dLS, promotes hypophagia in response to stress. Finally, we used inhibitory Designer Receptors Activated Exclusively by Designer Drugs (DREADDs) to test whether activation of Glp1r-expressing neurons in the LS is required for stress-induced hypophagia. RESULTS: Lean male mice display the expected hypophagic response following acute restraint stress, but obese male mice are resistant to this acute stress-induced hypophagia. Glp1r-positive neurons in the dLS are robustly activated during acute restraint stress in lean but not in obese male mice. This raises the possibility that activation of dLS Glp1r neurons during restraint stress contributes to subsequent hypophagia. Supporting this, we show that chemogenetic inhibition of LS Glp1r neurons attenuates acute restraint stress hypophagia in male mice. Surprisingly, we show that both lean and obese female mice are resistant to acute restraint stress-induced hypophagia and activation of dLS Glp1r neurons. CONCLUSIONS: These results suggest that dLS Glp1r neurons contribute to the hypophagic response to acute restraint stress in male mice, but not in female mice, and that obesity disrupts this response in male mice. Broadly, these findings show sexually dimorphic mechanisms and feeding behaviors in lean vs. obese mice in response to acute stress.

Chemospecific deficits in taste sensitivity following bilateral or right hemispheric gustatory cortex lesions in rats.

Our prior studies showed bilateral gustatory cortex (GC) lesions significantly impair taste sensitivity to salts (NaCl and KCl) and quinine ("bitter") but not to sucrose ("sweet"). The range of qualitative tastants tested here has been extended in a theoretically relevant way to include the maltodextrin, Maltrin, a preferred stimulus by rats thought to represent a unique taste quality, and the "sour" stimulus citric acid; NaCl was also included as a positive control. Male rats (Sprague-Dawley) with histologically confirmed neurotoxin-induced bilateral (BGCX, n = 13), or right (RGCX, n = 13) or left (LGCX, n = 9) unilateral GC lesions and sham-operated controls (SHAM, n = 16) were trained to discriminate a tastant from water in an operant two-response detection task. A mapping system was used to determine placement, size, and symmetry (when bilateral) of the lesion. BGCX significantly impaired taste sensitivity to NaCl, as expected, but not to Maltrin or citric acid, emulating our prior results with sucrose. However, in the case of citric acid, there was some disruption in performance at higher concentrations. Interestingly, RGCX, but not LGCX, also significantly impaired taste sensitivity, but only to NaCl, suggesting some degree of lateralized function. Taken together with our prior findings, extensive bilateral lesions in GC do not disrupt basic taste signal detection to all taste stimuli uniformly. Moreover, GC lesions do not preclude the ability of rats to learn and perform the task, clearly demonstrating that, in its absence, other brain regions are able to maintain sensory-discriminative taste processing, albeit with attenuated sensitivity for select stimuli.

Extensive Gustatory Cortex Lesions Significantly Impair Taste Sensitivity to KCl and Quinine but Not to Sucrose in Rats.

Recently, we reported that large bilateral gustatory cortex (GC) lesions significantly impair taste sensitivity to salts in rats. Here we extended the tastants examined to include sucrose and quinine in rats with ibotenic acid-induced lesions in GC (GCX) and in sham-operated controls (SHAM). Presurgically, immediately after drinking NaCl, rats received a LiCl or saline injection (i.p.), but postsurgical tests indicated a weak conditioned taste aversion (CTA) even in controls. The rats were then trained and tested in gustometers to discriminate a tastant from water in a two-response operant taste detection task. Psychometric functions were derived for sucrose, KCl, and quinine. Our mapping system was used to determine placement, size, and symmetry of the lesions (~91% GC damage on average). For KCl, there was a significant rightward shift (ΔEC50 = 0.57 log10 units; p<0.001) in the GCX psychometric function relative to SHAM, replicating our prior work. There was also a significant lesion-induced impairment (ΔEC50 = 0.41 log10 units; p = 0.006) in quinine sensitivity. Surprisingly, taste sensitivity to sucrose was unaffected by the extensive lesions and was comparable between GCX and SHAM rats. The fact that such large bilateral GC lesions did not shift sucrose psychometric functions relative to SHAM, but did significantly compromise quinine and KCl sensitivity suggests that the neural circuits responsible for the detection of specific taste stimuli are partially dissociable. Lesion-induced impairments were observed in expression of a postsurgical CTA to a maltodextrin solution as assessed in a taste-oriented brief-access test, but were not reflected in a longer term 46-h two-bottle test. Thus, deficits observed in rats after extensive damage to the GC are also dependent on the test used to assess taste function. In conclusion, the degree to which the GC is necessary for the maintenance of normal taste detectability apparently depends on the chemical and/or perceptual features of the stimulus.

Extensive lesions in rat insular cortex significantly disrupt taste sensitivity to NaCl and KCl and slow salt discrimination learning.

While studies of the gustatory cortex (GC) mostly focus on its role in taste aversion learning and memory, the necessity of GC for other fundamental taste-guided behaviors remains largely untested. Here, rats with either excitotoxic lesions targeting GC (n = 26) or sham lesions (n = 14) were assessed for postsurgical retention of a presurgically LiCl-induced conditioned taste aversion (CTA) to 0.1M sucrose using a brief-access taste generalization test in a gustometer. The same animals were then trained in a two-response operant taste detection task and psychophysically tested for their salt (NaCl or KCl) sensitivity. Next, the rats were trained and tested in a NaCl vs. KCl taste discrimination task with concentrations varied. Rats meeting our histological inclusion criterion had large lesions (resulting in a group averaging 80% damage to GC and involving surrounding regions) and showed impaired postsurgical expression of the presurgical CTA (LiCl-injected, n = 9), demonstrated rightward shifts in the NaCl (0.54 log10 shift) and KCl (0.35 log10 shift) psychometric functions, and displayed retarded salt discrimination acquisition (n = 18), but eventually learned and performed the discrimination comparable to sham-operated animals. Interestingly, the degree of deficit between tasks correlated only modestly, if at all, suggesting that idiosyncratic differences in insular cortex lesion topography were the root of the individual differences in the behavioral effects demonstrated here. This latter finding hints at some degree of interanimal variation in the functional topography of insular cortex. Overall, GC appears to be necessary to maintain normal taste sensitivity to NaCl and KCl and for salt discrimination learning. However, higher salt concentrations can be detected and discriminated by rats with extensive damage to GC suggesting that the other resources of the gustatory system are sufficient to maintain partial competence in these tasks, supporting the view that such basic sensory-discriminative taste functions involve distributed processes among central gustatory structures.

Induction of salivary proteins modifies measures of both orosensory and postingestive feedback during exposure to a tannic acid diet.

There are hundreds of proteins in saliva. Although it has long been hypothesized that these proteins modulate taste by interacting with taste receptors or taste stimuli, the functional impact of these proteins on feeding remains relatively unexplored. We have developed a new technique for saliva collection that does not interfere with daily behavioral testing and allows us to explore the relationship between feeding behavior and salivary protein expression. First, we monitored the alterations in salivary protein expression while simultaneously monitoring the animals' feeding behavior and meal patterns on a custom control diet or on the same diet mixed with 3% tannic acid. We demonstrated that six protein bands increased in density with dietary tannic acid exposure. Several of these bands were significantly correlated with behaviors thought to represent both orosensory and postingestive signaling. In a follow-up experiment, unconditioned licking to 0.01-3% tannic acid solutions was measured during a brief-access taste test before and after exposure to the tannic acid diet. In this experiment, rats with salivary proteins upregulated found the tannin solution less aversive (i.e., licked more) than those in the control condition. These data suggest a role for salivary proteins in mediating changes in both orosensory and postingestive feedback.

High-resolution lesion-mapping strategy links a hot spot in rat insular cortex with impaired expression of taste aversion learning.

Gustatory cortex (GC), an assemblage of taste-responsive neurons in insular cortex, is widely regarded as integral to conditioned taste aversion (CTA) retention, a link that has been primarily established using lesion approaches in rats. In contrast to this prevailing view, we found that even the most complete bilateral damage to GC produced by ibotenic acid was insufficient to disrupt postsurgical expression of a presurgical CTA; nor were such lesions sufficient to disrupt postsurgical acquisition and initial expression of a second CTA. However, some rats with lesions were significantly impaired on these tests. Further examination of all conditioned rats with lesions, regardless of the lesion topography, revealed a significant positive association between damage in the posterior portion of GC and especially within adjacent posterior regions of insular cortex. Accordingly, we developed a high-resolution lesion-mapping program that permitted the overlay of the individual lesion maps from rats with CTA impairments to produce a groupwise aggregate lesion map. Comparison of this map with one derived from the unimpaired counterparts indicated a specific lesion "hot spot" associated with CTA deficits that included the most posterior end of GC and overlying granular layer and encompassed an area provisionally referred to in the literature as visceral cortex. Thus, the detailed mapping of the lesion in behaviorally defined subgroups of rats allowed us to exploit the variability in performance to uncloak an important potential component of the functional topography of insular cortex; such an approach could have general applicability to other brain structure-function endeavors as well.

Water restriction and fluid temperature alter preference for water and sucrose solutions.

The role of diet temperature in ingestive behavior is poorly understood. We examined the importance of stimulus temperature and water-restriction state on the preference for and intake of water and sucrose. Using custom-designed equipment that allows us to monitor and maintain solution temperatures during testing (±0.1 °C), we conducted a series of 2-bottle preference tests (10 °C water vs. sucrose 10-40 °C) and brief access tests (10-40 °C water and sucrose). Water-restricted rats preferred cold water over any sucrose concentration (0.0-1.0 M) if the sucrose was 30 or 40 °C, whereas the same rats preferred sucrose at all concentrations and temperatures when unrestricted suggesting that the water-restriction state interacts with temperature preference. In a series of brief-access tests using a Davis Rig (MS-180), rats reduced licking to cold sucrose compared with 20 °C sucrose, suggesting that unlike water, cold temperature reduced the palatability of sucrose.