Links Between Glucocorticoid Responsiveness and Obesity

C H A P T E R

23 Links Between Glucocorticoid Responsiveness and Obesity: Involvement of Food Intake and Energy Expenditure Belinda A. Henry1, Iain J. Clarke2 1

Metabolism, Diabetes and Obesity Program, Monash Biomedical Discovery Institute, Department of Physiology, Monash University, Clayton, VIC, Australia; 2Neuroscience Program, Monash Biomedical Discovery Institute, Department of Physiology, Monash University, Clayton, VIC, Australia

O U T L I N E Introduction Nexus Between Body Weight, Obesity and Activation of the HPA Axis

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Physiological Determinants of Glucocorticoid Responsiveness: Selection of LR and HR Individuals 311 Cortisol Responsiveness and Innate Predisposition to Weight Gain

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Activation of the hypothalamoepituitarye adrenal (HPA) axis and the secretion of glucocorticoids influence energy homeostasis via effects on food intake and energy expenditure. Furthermore, abdominal obesity is associated with hyperactivity of the HPA axis, which is driven by impaired glucocorticoid negative feedback. Recent data, however, suggest that altered glucocorticoid secretion may precede the onset of obesity. In any given population, there is great

Stress: Physiology, Biochemistry, and Pathology https://doi.org/10.1016/B978-0-12-813146-6.00023-0

Cortisol Responsiveness and the Neural Control of Food Intake

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Cortisol Responsiveness and Thermogenesis 315 Neuroendocrine Determinants of Altered Thermogenesis in LR and HR

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Cortisol Responsiveness, Coping Strategies, and Physical Activity 317 Future Perspective 319 References

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variation in the glucocorticoid secretory response to stress, with individuals identified as either high responders (HR) or low responders (LR). Sheep characterized as HR have relatively increased food intake in response to stress, impaired satiety in response to melanocortin treatment, and reduced energy expenditure. The reduction in energy expenditure in HR is due to diminished thermogenesis in skeletal muscle and decreased physical activity.

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Copyright © 2019 Elsevier Inc. All rights reserved.

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This complex physiological phenotype is associated with increased propensity to become obese in HR animals. This chapter describes the metabolic, neuroendocrine, and behavioral phenotype in ewes characterized as LR and HR. We propose that altered expression of key hypothalamic genes underpin the metabolic, thermogenic, and behavioral sequelae associated with increased weight gain and increased susceptibility to become obese in HR compared with LR animals. The work described herein suggests that innate variation in cortisol responsiveness precedes weight gain and may be a marker to identification individuals at increased risk of becoming obese.

KEY POINTS • Glucocorticoid secretory responses to stress vary greatly. • Animals can be selected for high and low cortisol responsiveness using a simple adrenocorticotropic hormone challenge. • Animals characterized as high cortisol responders have increased propensity to become obese when fed a high-energy diet compared with low cortisol responders. • Increased propensity to develop obesity in high responders is associated with a distinct metabolic, neuroendocrine, and behavioral phenotype. Including: • reduced postprandial thermogenesis in skeletal muscle. • relatively increased food intake in response to stress. • impaired melanocortin signaling and resistance to the satiety effect of a-melanocyteestimulating hormone. • reduced physical activity. • Characterization of cortisol responses in humans may be a marker for identifying individuals with increased susceptibility to gain weight and to strategize weight loss therapies.

INTRODUCTION It is well recognized that, once obese, it is extremely difficult to lose weight and maintain weight loss, due to homeostatic defense mechanisms that reset hunger and energy expenditure. Body weight is determined by the balance between energy intake and the rate of energy expenditure. The latter is comprised of three major facets, including basal metabolic rate, physical activity, and adaptive thermogenesis. Reduced energy expenditure and in particular reduced thermogenesis is a key homeostatic component that confounds weight loss and long-term weight maintenance. The recent identification of functional brown adipose tissue (BAT) in adults has created a surge of interest in harnessing thermogenesis to develop novel weight loss strategies. A common practice is to use a combination of calorie restriction, exercise, and pharmacotherapies to achieve weight loss, but large cohort studies show that only 2% of subjects maintain weight loss at 2 years postintervention.1,2 A “one-size-fits-all” approach is ineffective because people vary greatly in their weight loss response to lifestyle and/or pharmacotherapy interventions. It is hypothesized that formulation of personalized weight loss strategies is the key to long-term weight loss in obese individuals. This chapter addresses the use of cortisol responsiveness to personalize weight loss. In response to any stressor, the amount of cortisol secreted is highly variable, and a proportion of individuals can be identified as either high (HR) or low (LR) cortisol responders. Animal studies have demonstrated that in response to a single injection of adrenocorticotropic hormone (ACTH), which can be performed with ease in large animals, where repeated blood sampling is simple and relatively non-invasive, approximately 10% of individuals can be characterised as LR or HR. Sheep selected as HR have greater propensity to become obese than those characterized as LR, which is associated with innate differences in the “set-point” of genes in the hypothalamus. Studies have demonstrated that high cortisol responsiveness is associated with

PHYSIOLOGICAL DETERMINANTS OF GLUCOCORTICOID RESPONSIVENESS: SELECTION OF LR AND HR INDIVIDUALS

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a distinct neuroendocrine, metabolic and behavioral phenotype that ultimately leads to increased food intake (in response to stress) and reduced energy expenditure. This review will detail the relationship between cortisol responses and weight control, highlighting the possible use of cortisol responsiveness as a marker to identify individuals that are highly susceptible to become obese and personalized weight loss strategies.

mammals, including sheep and humans, it is unclear as to whether obesity precedes the dysregulation of cortisol secretion and thus the causative relationship between HPA axis function and obesity remains to be elucidated. Recent work suggests that altered glucocorticoid responsiveness actually precedes the development of obesity and thus may be a marker to identify individuals with increased susceptibility to weight gain and greater propensity to become obese.

NEXUS BETWEEN BODY WEIGHT, OBESITY AND ACTIVATION OF THE HPA AXIS

PHYSIOLOGICAL DETERMINANTS OF GLUCOCORTICOID RESPONSIVENESS: SELECTION OF LR AND HR INDIVIDUALS

Various animal models show increased glucocorticoid levels in obesity. For example, in mice naturally occurring mutations in the gene encoding leptin (ob/ob) results in morbid obesity and elevated corticosterone levels.3e5 Furthermore, adrenalectomy reverses the obese phenotype of ob/ob mice, and glucocorticoid replacement therapy reinstates this.5 In sheep, hypothalamopituitary disconnection (HPD) increases basal secretion of cortisol,6 and this is associated with increased body weight and adiposity.7 It is important to note, that HPD disrupts the entire hypothalamo-pituitary axis and thus dysregulation of cortisol secretion is not the sole endocrine feature of this model. Nonetheless, diet-induced obesity in female sheep leads to hyperactivation of the HPA axis and greater cortisol secretion in response to isolation restraint stress compared to lean animals.8 Thus suggesting, an interconnection between enhanced glucocorticoid secretion and obesity. Similarly, in humans, it is generally thought that obesity leads to hyperactivity of the HPA axis,9,10 but this is somewhat controversial since various studies have produced contradictory data.11 Cortisol secretion follows a diurnal pattern, and obesity may cause a subtle shift to increase overnight secretion.12,13 Abdominal or visceral obesity, however, is associated with impaired glucocorticoid negative feedback as demonstrated by the dexamethasone suppression test14,15; this leads to enhanced cortisol secretion in response to stress.16 In large

Increased glucocorticoid secretion is fundamental to the physiological and behavioral responses to stress. In any given population, however, the glucocorticoid secretory response to stress varies between individuals. Rats exhibit strain differences in the corticosterone response to stress with the Fisher 344 strain displaying relatively higher corticosterone levels across the circadian period and in response to various stressors compared to the LOU/C strain.17 Furthermore, in outbred animals such as sheep, individuals can be identified as low (LR) or high (HR) glucocorticoid responders18 in response to ACTH treatment. Indeed, selection for LR and HR in response to stress or ACTH challenge has been demonstrated in humans,19 sheep,20,21 fish,22 mice,23 and pigs.24 A key feature of LR and HR individuals, irrespective of species, is that basal glucocorticoid levels are similar, and divergence in cortisol/corticosterone concentration is only evident in response to stress-related stimuli. Various physiological factors impact on glucocorticoid responses to stress, including sex,25 seasonality,26 lactation,27,28 and pregnancy.29 Innate variation in glucocorticoid responses, however, can manifest independent of the aforementioned factors and are determined by both genetic and environmental factors.30,31 Inherent differences in HPA axis responsiveness can manifest at various levels including impaired negative feedback at the level of the

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brain and anterior pituitary gland or altered steroidogenesis at the level of the adrenal gland. Characterization of the HPA axis in LR and HR ewes demonstrate innate differences in the “setpoint” of the HPA axis (Fig. 23.1).32 HR ewes have elevated expression of CRF and AVP, as

well as reduced expression of oxytocin in the paraventricular nucleus of the hypothalamus.32 Furthermore, the expression of proopiomelanocortin (POMC), the precursor for ACTH, is also greater in the anterior pituitary gland of HR than LR ewes.32 Thus HR have increased

FIGURE 23.1 Cortisol responsiveness is measured by an adrenocorticotropin (ACTH) challenge. Serial blood samples

are collected before and after the administration of a standardized dose of ACTH (0.2 mg/kg body weight). This allows for identification and selection of high and low cortisol responding animals. Representative profiles of high responders (HR) are shown in green and low responders (LR) in purple. HR and LR animals exhibit differences in the setpoint of the hypothalamoepituitaryeadrenal axis. Expression of corticotropin-releasing factor (CRF) and arginine vasopressin (AVP) is increased in the paraventricular nucleus (PVN) of HR compared with LR. While the expression of oxytocin in the PVN is reduced in HR compared with LR. Furthermore, expression of proopiomelanocortin (POMC) is increased in the anterior pituitary (AP) of HR. This shows that selection for high cortisol responses identifies animals with an innate upregulation of key genes within the hypothalamoepituitaryeadrenal axis.

CORTISOL RESPONSIVENESS AND INNATE PREDISPOSITION TO WEIGHT GAIN

expression of key neuroendocrine factors that act to stimulate the HPA axis, including CRF, AVP, and ACTH. Reduced expression of oxytocin in HR would promote increased stress responsiveness since oxytocin is known to dampen the HPA axis and exert beneficial effects on anxiety and social bonding behaviors.33e36 It is of importance to highlight that these differences in gene expression are observed under basal, nonstressed conditions, emphasizing the fact that HR ewes have an innate upregulation in the “setpoint” of the HPA axis. It has been postulated that individual differences in glucocorticoid responses may be driven by impaired negative feedback through altered expression and/or function of the glucocorticoid receptor (GR) and mineralocorticoid receptor (MR). Glucocorticoids bind to the MR with high affinity, and thus this receptor subtype is important in regulating basal glucocorticoid secretion.37 In contrast, glucocorticoids bind GR with low affinity, but the receptor is engaged when glucocorticoid levels are high, during stress. As such, GR is ubiquitous and is considered to be the primary receptor mediating glucocorticoid negative feedback in the brain and anterior pituitary gland. Indeed, GR in the paraventricular nucleus mediates fast, nongenomic negative feedback on CRF neurons and thus controls stress-induced activation of the HPA axis.38 Despite this, expression of GR and MR in the paraventricular nucleus of the hypothalamus was similar in LR and HR animals.32 The neural pathways that relay glucocorticoid negative feedback have been extensively characterized in rodents,37 showing that the parvocellular neurons of the PVN receive input from a number of brain structures including the nucleus of the solitary tract, raphe nucleus, bed nucleus of the stria terminalis, prefrontal cortex, amygdala, and the hippocampus.39 Indeed, long-term negative feedback effects of corticosterone are thought to be relayed to the CRF neurons in the paraventricular nucleus of the hypothalamus via the medial amygdala and hippocampus.40,41 It remains possible that impaired negative glucocorticoid negative feedback in HR manifests upstream to the paraventricular nucleus, resulting in an upregulation in the steady-state expression of key genes involved in activation of the HPA axis.

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Irrespective of altered gene expression within the hypothalamus and the anterior pituitary gland, HR animals also display greater adrenal responsiveness to ACTH than LR ewes. LR and HR ewes can be identified or selected using a low-dose ACTH challenge (Fig. 23.1). Prior to screening, estrous cycles are synchronized to control for fluctuating levels of ovarian steroids. Studies show that 10% of animals are consistently identified as either low or high cortisol responders (Fig. 23.1). In this model, animal selection is based on innate differences in adrenal responsiveness to ACTH. Despite this, the molecular or cellular mechanisms underpinning altered adrenal responses are unknown. Gene analyses have shown that expression of key steroidogenic enzymes (StAR, 11bHSD1, CYP11A, and CYP17) as well as the melanocortin 2 receptor (MC2R) is similar in the adrenal cortex of LR and HR animals.32 Furthermore, the basal concentration of cortisol in the adrenal gland is similar in LR and HR as are adrenal gland weights and the cortex:medulla ratio.32 It is possible that differences in adrenal gland function will only be evident in response to stress or ACTH challenge, as exemplified by similar basal secretion of cortisol in LR and HR ewes.42 Further work is required to characterize the role of the adrenal cortex in determining innate differences in cortisol responsiveness in LR and HR. Irrespective of the mechanisms that underpin differences in cortisol responsiveness, strong evidence suggests that the glucocorticoid responses are important in determining subsequent metabolic sequelae and may be useful to predict propensity to become obese.

CORTISOL RESPONSIVENESS AND INNATE PREDISPOSITION TO WEIGHT GAIN As outlined previously, in humans, abdominal obesity is associated with increased cortisol secretion in response to stress or stress-related stimuli (CRF/ACTH).10,14,16 Furthermore, high cortisol responsiveness to ACTH in rams is correlated to reduced feed efficiency and increased levels of adiposity.20 This demonstrates an association between obesity and

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dysregulation of the HPA axis. The aforementioned studies, however, do not establish causal links between weight gain and cortisol responsiveness and, importantly, these studies do not address whether differences in cortisol responses precede the onset of obesity. In sheep of normal body weight, chronic elevation in cortisol via daily injection of Synacthen Depot (long-acting synthetic ACTH) causes weight gain.43 Furthermore, the degree of weight gain is correlated to the cortisol response as determined by measuring the area under the curve,43 suggesting that differences in adrenal gland response precede obesity. Indeed, HR and LR ewes have similar body weights at baseline, but HR gain greater adiposity in response to feeding a high energy diet than LR.21 This clearly supports the notion that innate differences in HPA axis function precede the onset of obesity and in fact can be a physiological marker to identify individuals that have greater propensity to gain weight. The inherent differences in predisposition to become obese is associated with a suite of neuroendocrine, metabolic, and behavioral differences that ultimately lead to increased food intake and reduced energy expenditure in HR animals. These mechanisms will be the focus of the second half of this chapter.

that HR individuals eat relatively more in response to stressful stimuli. Indeed, this is further supported by altered expression of key appetite-regulating genes in the hypothalamus of LR and HR ewes46 (Fig. 23.2). Food intake is tightly controlled by the hypothalamus. Endocrine factors including leptin, insulin, and ghrelin modulate food intake via hypothalamic appetite-regulating peptides. These blood-borne factors act primarily at the arcuate nucleus of the hypothalamus to regulate orexigenic and satiety neurons. POMC neurons are activated by leptin47,48 and insulin,49 causing release of a-melanocyteestimulating hormone (aMSH), which elicits satiety via the MC4R in the paraventricular nucleus.50 A second population of neurons contain neuropeptide Y (NPY) and agouti-related protein (AgRP), which are activated by ghrelin but inhibited by leptin and insulin; NPY/AgRP neurons are orexigenic and

CORTISOL RESPONSIVENESS AND THE NEURAL CONTROL OF FOOD INTAKE At baseline, LR and HR individuals eat similar amounts,21 but differences in food intake are unmasked in response to stressful stimuli. In women, stress typically increases food intake, and only around 10% of individuals show a reduced appetite.44,45 Interestingly, in response to psychosocial stress, women characterized as HR show greater preference for “comfort” foods high in fat and sugar.19 Likewise, LR ewes reduce food intake in response to a barking dog (psychosocial/predator stress) with no effect in HR animals.42 Immune challenge caused by lipopolysaccharide administration decreases food intake in both LR and HR ewes, but this effect is amplified in the former.42 This demonstrates

FIGURE 23.2 High (HR) and low (LR) cortisol responders exhibit differential neuroendocrine control of food intake. At baseline, food intake is similar between the two groups; however, HR eat relatively more than LR in response to stressful stimuli. In addition, HR animals are resistant to the satiety effect of a-melanocyte stimulating hormone (aMSH). Despite differences in feeding behavior, expression of genes encoding key appetite-regulating peptides including neuropeptide Y (NPY), agouti-related protein (AgRP), and proopiomelanocortin (POMC) are similar in LR and HR. Thus differences in the neuroendocrine control of food intake manifest downstream to the arcuate nucleus (ARC) at the level of the receptors; expression of the melanocortin 4 receptor (MC4R) in the paraventricular nucleus (PVN) of the hypothalamus is lower in HR than LR.

CORTISOL RESPONSIVENESS AND THERMOGENESIS

increase food intake.51 The primary role of NPY/ AgRP neurons is to protect against starvation since targeted ablation of these neurons reduces food intake and causes wasting, eventually leading to starvation and death.52 Innate susceptibility to become obese in HR animals is not associated with early onset leptin resistance,21 and the expression of NPY, AgRP, and POMC is similar in LR and HR animals46 (Fig. 23.2). Despite this, HR animals exhibit a marked reduction in the expression of the MC4R in the paraventricular nucleus, and this coincides with impaired satiety in response to intracerebroventricular infusion of aMSH.46 Previous studies in sheep demonstrate that decreased food intake in response to immune stress is relayed via the central melanocortin pathway.53 It is therefore hypothesized that reduced expression of MC4R in HR animals is a fundamental neuroendocrine determinant of altered food intake in response to stressful stimuli, and this directly relates to increased susceptibility to become obese in HR individuals (Fig. 23.2).

CORTISOL RESPONSIVENESS AND THERMOGENESIS Body weight is not only determined by food intake but also the rate at which energy is expended. Energy expenditure is comprised of three major facets including basal metabolic rate, physical activity, and adaptive thermogenesis. The latter is defined as the dissipation of energy through specialized production of heat and is well characterized in BAT of rodents. Brown adipocytes are rich in mitochondria and express uncoupling protein 1 (UCP1),54,55 which when activated creates a proton leak across the inner mitochondrial membrane, directing protons away from ATP synthesis and resulting in futile heat production. Furthermore, BAT is highly vascularized and receives profuse innervation from the sympathetic nervous system (SNS),54,55 which is essential to the activation of thermogenesis. Earlier dogma stipulated that BAT was exclusively found in neonates, where it was essential for the maintenance of core body temperature, but recent advances demonstrate that

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BAT is retained in adults. Landmark imaging studies in humans revealed the presence of BAT in the clavicular, neck, and sternal regions of adults,56e58 and as such this has become a significant therapeutic target for the development of weight loss pharmacotherapies. In large mammals, including sheep and humans, BAT does not account for total thermogenic capacity, and additional tissues contribute to thermogenesis. In this regard, skeletal muscle may play an important role. Prior to the identification of functional BAT by positron emission tomography-computed tomography (PETeCT) imaging in adult humans, skeletal muscle was thought to be the primary thermogenic tissue. Initial work demonstrated that muscle accounts for up to 50% of ephedrine-induced thermogenesis, whereas adipose tissue accounts for approximately 5%.59 It is important to emphasize that this earlier work did not study adipose tissue in the neck and supra-clavicular regions, sites where most brown/beige adipocytes are found. More recent work using PETeCT scanning has shown that acute low doses of ephedrine have no effect on BAT activity, but high doses increase BAT activity in lean humans. Furthermore, chronic low-dose ephedrine treatment actually reduces BAT activity,60 and isoprenaline (a nonspecific bAR) treatment increases energy expenditure without an associated activation of BAT.61 Similarly, blockade of the bAR with propranolol had no effect on cold-induced BAT thermogenesis in humans.62 This lack of effect, however, is likely due to receptor specificity, as both isoprenaline and propranolol show preferential agonistic and antagonistic affinity to the b1AR and b2AR, respectively. A64 Trp/Arg genetic polymorphism in the b3AR is linked to the decline in BAT function with ageing in men,63 suggesting a predominant role for this receptor subtype in controlling BAT function in humans. Indeed, in healthy lean men, administration of the b3AR-specific agonist, mirabegron, activates BAT and causes a concurrent increase in resting metabolic rate.64 These studies highlight that, in humans, the b3AR is essential to the SNS-driven activation of BAT thermogenesis. It remains possible, however, that the effects of isoprenaline to increase energy expenditure via

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the b1/b2 AR61 are mediated via skeletal muscle thermogenesis.65 Skeletal muscle thermogenesis occurs via two distinct cellular pathways, including UCP3mediated mitochondrial uncoupling and futile calcium cycling. Myocytes express UCP3, which is capable of uncoupling oxidative phosphorylation in isolated yeast mitochondria.66 Individual variation in UCP3 expression and mitochondrial uncoupling in skeletal muscle is linked to the ability to successfully lose weight and maintain weight loss. In obese women, reduced expression of UCP3 in skeletal muscle causes a weakened proton leak in mitochondria and impaired ability to lose weight.67 Furthermore, innate variation in basal mitochondrial uncoupling in skeletal muscle accounts for 20%e50% of the variation in basal metabolic rate. Thus skeletal muscle appears to be an important site of thermogenesis in humans, contributing to total energy expenditure and long-term regulation of body weight. In addition to mitochondrial uncoupling, futile calcium cycling can drive adaptive thermogenesis in skeletal muscle. In this regard, calcium exits the sarcoendoplasmic reticulum (SR) via the ryanodine receptor (RyR). To maintain cytosolic calcium levels, activation of the sarcoendoplasmic reticulum ATPases (SERCA) propel calcium back into the SR; this effect is driven by the hydrolysis of ATP and results in heat production.68,69 In rodents, sarcolipin is an endogenous activator of SERCA, which uncouples calcium transport from the hydrolysis of ATP, leading to an increase in the futile cycling of calcium and heat production. In the absence of BAT (surgical removal) or UCP1 gene knockout animals, sarcolipin increases muscle thermogenesis and is essential for cold adaptation.68,70 Furthermore, overexpression of sarcolipin in skeletal muscle increases oxygen consumption and fatty acid oxidation, which is associated with resistance to weight gain in mice fed a high-fat diet.71 The role of sarcolipin in thermogenesis in larger mammals, however, is relatively unexplored and requires closer investigation. Nonetheless, in sheep, postprandial thermogenesis is associated with increased expression of RyR1 and SERCA,72 indicative of a role of futile calcium cycling in adaptive thermogenesis in skeletal muscle.

Skeletal muscle accounts for approximately 40% of total body mass, so in large mammals at least, even small differences in muscle thermogenesis could contribute substantially to heat production and total energy expenditure. Consistent with this notion, at normal body weight, HR ewes have reduced postprandial thermogenesis in skeletal muscle, which is likely to significantly impact on energy expenditure. Sheep are a grazing species, so they do not typically display any meal-associated changes in various endocrine factors (e.g., ghrelin) or peripheral heat production. Despite this, if animals are entrained to a fixed feeding regime, whereby food is provided at set “meal times” across a number of days, excursions in both ghrelin73 and thermogenesis can be engendered (Fig. 23.3). In ewes, postprandial heat production in skeletal muscle is not related to any change in femoral artery blood flow but is associated with increased uncoupled or state 4 respiration in mitochondria isolated from skeletal muscle as well as increased expression of UCP3 and key markers of the futile calcium cycling pathway (Fig. 23.3). Postprandial skeletal muscle heat production is greater in HR than LR, and this effect is enhanced with central administration of leptin21 (Fig. 23.3). Thus not only do LR and HR animals display innate differences in the neuroendocrine control of food intake there are clear differences in skeletal muscle thermogenesis. It is proposed that reduced postprandial thermogenesis in skeletal muscle leads to a reduction in energy expenditure and thus contributes to the increased susceptibility of HR to become obese. It is important to note that inherent variation in adaptive thermogenesis manifests specifically in skeletal muscle of HR and LR animals. Sheep are unlike rodents in that they do not have a defined or demarcated brown fat depot, but brown adipocytes are interspersed throughout typically white fat depots.74e76 Consistent with this, discrete adipose tissues display marked variation in the expression of UCP177 and thermogenic potential.74,75 The primary sites of adipose thermogenesis in sheep are sternal and retroperitoneal tissues, both of which show abundant expression of UCP1 and pronounced

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muscle accounts for approximately 40% of total body mass, differences in thermogenesis in this tissue are likely to impact on total energy expenditure and long-term regulation of body weight.

NEUROENDOCRINE DETERMINANTS OF ALTERED THERMOGENESIS IN LR AND HR

FIGURE 23.3 Skeletal muscle thermogenesis is lower in high cortisol responders (HR) than low cortisol responders. Skeletal muscle thermogenesis occurs via two distinct cellular mechanisms, including uncoupling protein 3 (UCP3)-driven mitochondrial uncoupled respiration and futile calcium cycling. Futile calcium cycling occurs across the sarcoendoplasmic reticulum (SR), where calcium (Ca2þ) is extricated from the SR via the ryanodine 1 receptor (RyR1). To maintain cytosolic Ca2þ levels, the sarcoendoplasmic reticulum ATP-dependent ATPases (SERCA) are activated, pumping Ca2þ back into the SR. The activation of SERCA is dependent on the hydrolysis of the ATP, which is a thermogenic process. Reduced skeletal muscle thermogenesis in HR is associated with altered expression of key genes in the hypothalamus including decreased melanocortin 4 receptor (MC4R) and prepro-orexin expression. Both the melanocortin pathway and orexin act within the brain to increase thermogenesis, and thus lowered expression is associated with attenuated postprandial thermogenesis in skeletal muscle of HR compared with LR.

thermogenic responses.74e76 Temperature recordings in retroperitoneal adipose tissue show that postprandial thermogenesis is similar in BAT of LR and HR sheep.21 Thus, in conclusion, reduced thermogenesis is a key component of altered propensity to gain weight in LR and HR sheep; however, this innate divergence manifests primarily in skeletal muscle. Given that skeletal

In addition to controlling food intake, the hypothalamus exerts reciprocal control on energy expenditure and in particular acts to control adaptive thermogenesis. Neuropeptides that regulate food intake have dual effect to regulate thermogenesis, whereby factors that increase food intake typically reduce thermogenesis and vice versa77a. For example, aMSH acts to reduce food intake and increases thermogenesis in both BAT77b and skeletal muscle77c. In contrast, orexin is produced in the lateral hypothalamus (LH) and increases both food intake and energyexpenditure via physical activity and adaptive thermogenesis.78,79 A subpopulation of orexin neurons in the LH project to the raphe pallidus and microinjection of orexin into the latter increased BAT thermogenesis.80 Furthermore, ablation of the orexin neurons in the LHA eliminates cold-, stress- and immune-induced BAT thermogenesis.81,82 A recent study demonstrated that obesity in orexin-knockout mice is associated with reduced adaptive thermogenesis in BAT due to an inability of brown preadipocytes to differentiate into mature brown adipocytes.83 In addition to reduced MC4R expression in the PVN, expression of prepro-orexin is lower in the LH of HR than LR46. Thus increased expression of orexin in the LH of LR animals may be important in mediating the enhanced skeletal muscle thermogenesis.

CORTISOL RESPONSIVENESS, COPING STRATEGIES, AND PHYSICAL ACTIVITY A number of animal models have shown that low cortisol responsiveness is associated with enhanced aggressive behaviors.23,24,84 Similarly,

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in humans, cortisol responses and obesity are associated with distinct behavioral traits. In obese subjects, emotional eating is correlated to impulsiveness and depression, whereas restrained eating is correlated with openness, conscientiousness, and extraverted personality traits.85 Furthermore, low cortisol responses are associated with increased incidence of

neuroticism in women, low extraversion in males, and reduced openness in both sexes.86 Cortisol responsiveness is also associated with differences in coping strategies. In LR and HR ewes, behavioral responses and coping strategies have been assessed using the open field test, arena test, and food competition test (Fig. 23.4). Low cortisol responding ewes have enhanced

FIGURE 23.4 Behavioral differences in high and low cortisol responders (HR and LR) have been assessed by the open field, arena, and food competition tests. The open field test measures the degree of locomotion an animal exhibits in response to isolation in an enclosed field. The arena test measures fear and avoidance behaviors where an animals desire to be with their flockmates is assessed in relation to their fear of approaching a human. Finally, the food competition test measures initiative behaviors by assessing the latency for the test animal (LR or HR) to reach a food reward when in competition with a control flockmate. HR animals show reduced physical activity and initiative, but increased fear that is indicative of a reactive behavioral coping style. In contrast, LR animals exhibit increased physical activity and initiative, but reduced fear, and this is indicative of a proactive coping style. Increased physical activity in the LR animals coincides with greater expression of prepro-orexin in LR than HR. Thus innate differences in the orexin system may also underpin the behavioral phenotypes of LR and HR animals.

REFERENCES

sympathoadrenal activation in response to stress and exhibit proactive coping strategies.87 Ewes selected for low cortisol responsiveness show greater physical activity in response to isolation stress (open field), reduced fear toward humans (arena test), reduced freezing, and increased initiative to reach a food reward (food competition test)42 (Fig. 23.4). It is hypothesized that a proactive coping style rather than a reactive style may be more likely to expend energy. A proactive coping strategy is typically associated with increased aggression and physical activity, which are behaviors that are consistent with increased energy expenditure. Importantly, mutations in the orexin system result in narcolepsy, and orexin is known to increase physical activity.78,88e90 Thus increased expression of prepro-orexin in LR does not only align with increased thermogenesis but also the relative increase in physical activity in LR compared with HR (Fig. 23.4). It is therefore proposed that altered expression of both MC4R and preproorexin are key neuroendocrine determinants of the increased propensity to become obese in HR animals.

Future Perspective There is variation in the susceptibility to gain weight and become obese. In addition, weight loss is also vastly variable across populations. Despite our understanding of the physiological control of body weight, weight loss remains elusive to the vast majority. Long-term studies have shown that approximately 2% of subjects maintain weight loss 5 years after weight loss intervention.1,2 A key component that determines individual differences in either weight gain or the ability to successfully lose weight is influenced by inherent variation in energy expenditure. Collectively, a net increase in food intake and reduced energy expenditure culminate in an obesity prone phenotype. One means in which we may improve weight loss is to strategize treatment through a personalized medicine approach. It may be proposed that characterization of cortisol responsiveness may be a means to identify individuals with increased susceptibility to become obese. Gene

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expression analyses within the hypothalamus indicate that reduced expression of MC4R in the PVN and/or reduced expression of orexin in the LH underpin the physiological and metabolic phenotype in obesity-prone HR animals. A number of the new generation of antiobesity drugs are known to target the melanocortin pathway, including Contrave, Lorcaserin, and Liraglutide.91e93 Our experimental data suggest that HR individuals may be less responsive to these pharmacotherapies and thus would greatly benefit from alternative weight loss strategies and therapies. Future studies need to address whether cortisol responsiveness may be a beneficial clinical marker for improving and maintaining long-term weight loss.

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