Paul R. Burghardt

Spontaneous activity, economy of activity, and resistance to diet-induced obesity in rats bred for high intrinsic aerobic capacity.

Though obesity is common, some people remain resistant to weight gain even in an obesogenic environment. The propensity to remain lean may be partly associated with high endurance capacity along with high spontaneous physical activity and the energy expenditure of activity, called non-exercise activity thermogenesis (NEAT). Previous studies have shown that high-capacity running rats (HCR) are lean compared to low-capacity runners (LCR), which are susceptible to cardiovascular disease and metabolic syndrome. Here, we examine the effect of diet on spontaneous activity and NEAT, as well as potential mechanisms underlying these traits, in rats selectively bred for high or low intrinsic aerobic endurance capacity. Compared to LCR, HCR were resistant to the sizeable increases in body mass and fat mass induced by a high-fat diet; HCR also had lower levels of circulating leptin. HCR were consistently more active than LCR, and had lower fuel economy of activity, regardless of diet. Nonetheless, both HCR and LCR showed a similar decrease in daily activity levels after high-fat feeding, as well as decreases in hypothalamic orexin-A content. The HCR were more sensitive to the NEAT-activating effects of intra-paraventricular orexin-A compared to LCR, especially after high-fat feeding. Lastly, levels of cytosolic phosphoenolpyruvate carboxykinase (PEPCK-C) in the skeletal muscle of HCR were consistently higher than LCR, and the high-fat diet decreased skeletal muscle PEPCK-C in both groups of rats. Differences in muscle PEPCK were not secondary to the differing amount of activity. This suggests the possibility that intrinsic differences in physical activity levels may originate at the level of the skeletal muscle, which could alter brain responsiveness to neuropeptides and other factors that regulate spontaneous daily activity and NEAT.

Individual Differences in Cue-Induced Motivation and Striatal Systems in Rats Susceptible to Diet-Induced Obesity

Pavlovian cues associated with junk-foods (caloric, highly sweet, and/or fatty foods), like the smell of brownies, can elicit craving to eat and increase the amount of food consumed. People who are more susceptible to these motivational effects of food cues may have a higher risk for becoming obese. Further, overconsumption of junk-foods leading to the development of obesity may itself heighten attraction to food cues. Here, we used a model of individual susceptibility to junk-foods diet-induced obesity to determine whether there are pre-existing and/or diet-induced increases in attraction to and motivation for sucrose-paired cues (ie, incentive salience or ‘wanting’). We also assessed diet- vs obesity-associated alterations in mesolimbic function and receptor expression. We found that rats susceptible to diet-induced obesity displayed heightened conditioned approach prior to the development of obesity. In addition, after junk-food diet exposure, those rats that developed obesity also showed increased willingness to gain access to a sucrose cue. Heightened ‘wanting’ was not due to individual differences in the hedonic impact (‘liking’) of sucrose. Neurobiologically, Mu opioid receptor mRNA expression was lower in striatal ‘hot-spots’ that generate eating or hedonic impact only in those rats that became obese. In contrast, prolonged exposure to junk-food resulted in cross-sensitization to amphetamine-induced locomotion and downregulation of striatal D2R mRNA regardless of the development of obesity. Together these data shed light on individual differences in behavioral and neurobiological consequences of exposure to junk-food diets and the potential contribution of incentive sensitization in susceptible individuals to greater food cue-triggered motivation.

Dietary n-3:n-6 fatty acid ratios differentially influence hormonal signature in a rodent model of metabolic syndrome relative to healthy controls

Dietary ratios of omega-3 (n-3) to omega-6 (n-6) polyunsaturated fatty acids (PUFAs) have been implicated in controlling markers of the metabolic syndrome, including insulin sensitivity, inflammation, lipid profiles and adiposity. However, the role of dietary PUFAs in regulating energy systems in healthy relative to metabolic diseased backgrounds has not been systematically addressed. We used dietary manipulation of n-3 to n-6 PUFA ratios in an animal model of metabolic syndrome and a related healthy line to assay feeding behavior and endocrine markers of feeding drive and energy regulation. Two related lines of rodents with a healthy and a metabolic syndrome phenotype were fed one of two isocaloric diets, comprised of either a 1:1 or a 1:30 n-3 to n-6 ratio, for 30 days. Food intake and weight gain were monitored; and leptin, ghrelin, adiponectin and a suite of hypothalamic neuropeptides involved in energy regulation were assayed following the dietary manipulation period. There was no difference in caloric intake or weight gain between diet groups, however there was a significant interaction between diet and phenotypic line on central and peripheral markers of energy homeostasis. Thus serum levels of leptin, acylated-ghrelin and adiponectin, and mRNA levels of the anorexigenic hypothalamic neuropeptide, cocaine-amphetamine related transcript (CART), showed differential, dietary responses with HCR rats showing an increase in anorexigenic signals in response to unbalanced n-3:6 ratios, while LCR did not. These data are the first to demonstrate that a rodent line with a metabolic syndrome-like phenotype responds differentially to dietary manipulation of n-3 and n-6 fatty acids relative to a related healthy line with regard to endocrine markers of energy homeostasis. The dietary n-3:n-6 ratios used in this experiment represent extreme points of natural human diets, however the data suggest that optimal recommendations regarding omega-3 and omega-6 intake may have differing effects in healthy subjects relative to metabolic syndrome patients. Further research is necessary to establish these responses in human populations.

The influence of metabolic syndrome, physical activity and genotype on catechol-O-methyl transferase promoter-region methylation in schizophrenia

The catechol-O-methyl transferase (COMT) 158Val/Met variant has been suggested to play a role in COMT function. Epigenetic regulation of COMT may further influence the prevalence of metabolic syndrome in these patient populations. This study examined the correlation between COMT promoter methylation and metabolic syndrome in schizophrenia patients receiving atypical antipsychotic (AAP) therapy. DNA was extracted from peripheral blood samples of schizophrenia subjects screened for metabolic syndrome. Pyrosequencing was used to analyze two methylation sites of the soluble COMT (COMT-s) promoter region. Associations between AAP use, lifestyle variables, metabolic syndrome and COMT genotype with peak methylation values were analyzed. Data are reported in 85 subjects. Methylation on CpG site 1 had a mean of 79.08% (±4.71) and it was 12.43% (±1.19) on site 2. COMT genotype proved to be an indicator of COMT methylation status on site 1 (F(2, 84)=5.78, P=0.0044) and site 2 (F(2, 84),=3.79, P=0.027). A significant negative correlation between physical activity and COMT promoter region methylation was found in Val/Val homozygous patients (site 1: P=0.013 and site 2: P=0.019). Those homozygous for Met/Met showed a positive correlation between promoter site methylation and physical activity (site 1: P=0.027, site 2: P=0.005), and between CpG site methylation and metabolic syndrome (site 1: P=0.002; site 2: P=0.001). The results of this study suggest that COMT promoter region methylation is largely influenced by COMT genotype and that physical activity plays a significant role in epigenetic modulation of COMT.

Microinjection of naltrexone into the central, but not the basolateral, amygdala blocks the anxiolytic effects of diazepam in the plus maze

The amygdala is involved in behavioral and physiological responses to fear, and the anxiolytic properties of several drugs are localized to this region. Activation of endogenous opioid systems is known to occur in response to stress and a growing body of literature suggests that opioid systems regulate the properties of anxiolytic drugs. These experiments sought to elucidate the role of opioid receptors in the central (CeA) and basolateral (BLA) nuclei of the amygdala in regulating the anxiolytic properties of ethanol and diazepam. Male rats fitted with cannula received bilateral microinjections of the nonselective opioid receptor antagonist naltrexone (NAL) immediately followed by systemic delivery of either ethanol (1 g/kg) or diazepam (2 mg/kg) in the elevated plus maze. Both diazepam and ethanol decreased anxiety-like behavior. Delivery of NAL into the CeA blocked the anxiolytic properties of diazepam. Delivery of NAL into the BLA slightly increased open arm avoidance, but had no effect on the anxiolytic properties of diazepam. Microinjection of NAL into either nucleus failed to block the effects of ethanol. These results were specific to the anxiolytic properties of diazepam, since baseline behaviors were unaffected by microinjection of NAL. Microinjection of lidocaine produced results distinct from NAL and failed to block the anxiolytic actions of diazepam. These studies indicate distinct roles for opioid receptor systems in the CeA and BLA in regulating the anxiolytic properties of diazepam in the elevated plus maze. Further, opioid receptor systems in the CeA and BLA do not regulate the anxiolytic properties of ethanol in this test.

Endogenous Opioid Mechanisms Are Implicated in Obesity and Weight Loss in Humans

CONTEXT Successful long-term weight loss is challenging. Brain endogenous opioid systems regulate associated processes; however, their role in the maintenance of weight loss has not been adequately explored in humans. OBJECTIVE In a preliminary study, the objective was to assess central μ-opioid receptor (MOR) system involvement in eating behaviors and their relationship to long-term maintenance of weight loss. DESIGN This was a case-control study with follow-up of the treatment group at 1 year after intervention. SETTING The study was conducted at a tertiary care university medical center. PARTICIPANTS Lean healthy (n = 7) and chronically obese (n = 7) men matched for age and ethnicity participated in the study. INTERVENTIONS MOR availability measures were acquired with positron emission tomography and [(11)C]carfentanil. Lean healthy men were scanned twice under both fasted and fed conditions. Obese men were placed on a very low-calorie diet to achieve 15% weight loss from baseline weight and underwent two positron emission tomography scans before and two after weight loss, incorporating both fasted and fed states. MAIN OUTCOME MEASURES Brain MOR availability and activation were measured by reductions in MOR availability (nondisplaceable binding potential) from the fed compared with the fasted-state scans. RESULTS Baseline MOR nondisplaceable binding potential was reduced in obese compared with the lean and partially recovered obese after weight loss in regions that regulate homeostatic, hedonic, and emotional responses to feeding. Reductions in negative affect and feeding-induced MOR system activation in the right temporal pole were highly correlated in leans but not in obese men. A trend for an association between MOR activation in the right temporal pole before weight loss and weight regain 1 year was found. CONCLUSIONS Although these preliminary studies have a small sample size, these results suggest that obesity and diet-induced weight loss impact central MOR binding and endogenous opioid system function. MOR system activation in response to an acute meal may be related to the risk of weight regain.

Risk-assessment and Coping Strategies Segregate with Divergent Intrinsic Aerobic Capacity in Rats

Metabolic function is integrally related to an individuals susceptibility to, and progression of, disease. Selective breeding for intrinsic treadmill running in rats has produced distinct lines of high- or low-capacity runners (HCR and LCR, respectively) that exhibit numerous physiological differences. To date, the role of intrinsic aerobic capacity on behavior and stress response in these rats has not been addressed and was the focus of these studies. HCR and LCR rats did not differ in their locomotor response to novelty or behavior in the light/dark box. In contrast, immobility in the forced swim test was higher in LCR rats compared with HCR rats, regardless of desipramine treatment. Although both HCR and LCR rats responded to cat odor with decreased exploration and increased risk assessment, HCR rats showed greater contextual conditioning to cat odor. HCR rats exhibited higher expression of corticotropin-releasing hormone in the central nucleus of the amygdala, as well as heavier adrenal and thymus weight. Corticosterone was comparable among HCR and LCR rats at light/dark transitions, and in response to unavoidable cat odor. HCR rats, however, exhibited a greater corticosterone response following the light/dark box. These experiments show that the LCR phenotype associates with decreased risk assessment in response to salient danger signals and passive coping. In contrast, HCR rats show a more naturalistic strategy in that they employ active coping and a more vigilant and cautious response to environmental novelty and salient danger signals. Within this context, we propose that intrinsic aerobic capacity is a central feature mechanistically linking complex metabolic disease and behavior.

Upregulation of GAD65 mRNA in the medulla of the rat model of metabolic syndrome.

Metabolic syndrome is characterized by obesity, elevated blood pressure (BP), insulin resistance, and hypercholesterolemia. Recently an animal model of this disorder has been proposed in rats selectively bred based on their performance on a treadmill-running task. Accordingly, low capacity runner (LCR) rats exhibited all of the diagnostic criteria for metabolic syndrome, including elevated BP, as compared to their high capacity runner (HCR) counterparts [U. Wisløff, S.M. Najjar, O. Ellingsen, P.M. Haram, S. Swoap, Q. Al-Share, M. Fernstrom, K. Rezaei, S.J. Lee, L.G. Koch, S.L. Britton, Cardiovascular risk factors emerge after artificial selection for low aerobic capacity, Science 307 (2005) 418-420]. Previous studies have highlighted the importance of GABAergic neurotransmission in the medullary cardiovascular-regulatory areas in the central control of BP. Thus, we hypothesized a dysregulation in GABAergic transmission in the medullary cardiovascular-regulatory nuclei of LCR rats. To begin testing this hypothesis we carried out experiments examining expression of the GABA synthetic enzymes, GAD65 and GAD67, mRNAs in the two rat strains via radioactive in situ hybridization. Our results showed GAD65 and GAD67 mRNAs were widely expressed throughout the brainstem; quantification revealed increased GAD65 mRNA expression in LCR animals in the caudal nucleus tractus solitarius (NTS) and rostral ventrolateral medulla (VLM) as compared to HCR rats. Conversely, no differences in the expression of GAD67 were detected in these regions. These data are consistent with the notion of altered GABAergic neurotransmission in the NTS and VLM in metabolic syndrome, and point to the importance of these regions in cardiovascular regulation.

Atypical antipsychotics, insulin resistance and weight; a meta-analysis of healthy volunteer studies

&NA; Atypical antipsychotics increase the risk of diabetes and cardiovascular disease through their side effects of insulin resistance and weight gain. The populations for which atypical antipsychotics are used carry a baseline risk of metabolic dysregulation prior to medication which has made it difficult to fully understand whether atypical antipsychotics cause insulin resistance and weight gain directly. The purpose of this work was to conduct a systematic review and meta‐analysis of atypical antipsychotic trials in healthy volunteers to better understand their effects on insulin sensitivity and weight gain. Furthermore, we aimed to evaluate the occurrence of insulin resistance with or without weight gain and with treatment length by using subgroup and meta‐regression techniques. Overall, the meta‐analysis provides evidence that atypical antipsychotics decrease insulin sensitivity (standardized mean difference = −0.437, p < 0.001) and increase weight (standardized mean difference = 0.591, p < 0.001) in healthy volunteers. It was found that decreases in insulin sensitivity were potentially dependent on treatment length but not weight gain. Decreases in insulin sensitivity occurred in multi‐dose studies <13 days while weight gain occurred in studies 14 days and longer (max 28 days). These findings provide preliminary evidence that atypical antipsychotics cause insulin resistance and weight gain directly, independent of psychiatric disease and may be associated with length of treatment. Further, well‐designed studies to assess the co‐occurrence of insulin resistance and weight gain and to understand the mechanisms and sequence by which they occur are required. HighlightsAtypical antipsychotics may cause insulin resistance and weight gain independent of psychiatric disease.Decreases in insulin sensitivity occurred both with and without significant weight gain when analyzed by subgrouping.Insulin resistance and weight gain may be associated with treatment length.The mechanisms and interplay of insulin resistance and weight gain with atypical antipsychotics need future investigation.

Brain Activation Patterns at Exhaustion in Rats That Differ in Inherent Exercise Capacity

In order to further understand the genetic basis for variation in inherent (untrained) exercise capacity, we examined the brains of 32 male rats selectively bred for high or low running capacity (HCR and LCR, respectively). The aim was to characterize the activation patterns of brain regions potentially involved in differences in inherent running capacity between HCR and LCR. Using quantitative in situ hybridization techniques, we measured messenger ribonuclease (mRNA) levels of c-Fos, a marker of neuronal activation, in the brains of HCR and LCR rats after a single bout of acute treadmill running (7.5–15 minutes, 15° slope, 10 m/min) or after treadmill running to exhaustion (15–51 minutes, 15° slope, initial velocity 10 m/min). During verification of trait differences, HCR rats ran six times farther and three times longer prior to exhaustion than LCR rats. Running to exhaustion significantly increased c-Fos mRNA activation of several brain areas in HCR, but LCR failed to show significant elevations of c-Fos mRNA at exhaustion in the majority of areas examined compared to acutely run controls. Results from these studies suggest that there are differences in central c-Fos mRNA expression, and potential brain activation patterns, between HCR and LCR rats during treadmill running to exhaustion and these differences could be involved in the variation in inherent running capacity between lines.