Zachary R. Patterson

Central Ghrelin Signaling Mediates the Metabolic Response of C57BL/6 Male Mice to Chronic Social Defeat Stress

Chronic stressors promote metabolic disturbances, including obesity and metabolic syndrome. Ghrelin, a peptide that promotes appetite and the accumulation of adipose tissue, is also secreted in response to stressors to protect the brain and peripheral tissues from the effects of these stressors. Here we demonstrate that elevated ghrelin levels produced by chronic exposure to social stress are associated with increased caloric intake and body weight gain in male C57BL mice. In contrast, stressed mice lacking ghrelin receptors (GHSR KO mice) or C57BL mice receiving chronic intracerebroventricular delivery of the ghrelin receptor antagonist [d-Lys(3)]-GHRP-6 show attenuated weight gain and feeding responses under the same social stress paradigm. Interestingly, stressed GHSR KO mice showed depleted sc and intrascapular brown fat depots, whereas stressed young wild-type mice did not. In old wild-type mice, chronic social defeat increased visceral and intrascapular brown fat depots in association with increases in obesity markers like hyperleptinemia and hyperinsulinemia along with increased hypothalamic expression of neuropeptide Y and Agouti related peptide. Importantly, the elevated expression of these peptides persisted least for 2 weeks after cessation of the stressor regimen. In contrast, old GHSR KO mice did not show these alterations after chronic social defeat. These results suggest that ghrelin plays an important role in the metabolic adaptations necessary to meet the energetic demands posed by stressors, but chronic exposure to stress-induced ghrelin elevations ultimately could lead to long lasting metabolic dysfunctions.

Organizational Effects of Perinatal Exposure to Bisphenol-A and Diethylstilbestrol on Arcuate Nucleus Circuitry Controlling Food Intake and Energy Expenditure in Male and Female CD-1 Mice

The endocrine disrupting compound bisphenol-A (BPA) has been reported to act as an obesogen in rodents exposed perinatally. In this study, we investigated the effects of early-life BPA exposure on adult metabolic phenotype and hypothalamic energy balance circuitry. Pregnant and lactating CD-1 dams were exposed, via specially prepared diets, to 2 environmentally relevant doses of BPA. Dams consumed an average of 0.19 and 3.49 μg/kg per day of BPA in the low and high BPA treatments prenatally and an average of 0.36 and 7.2 μg/kg per day of BPA postnatally. Offspring were weaned initially onto a normal (AIN93G) diet, then as adults exposed to either a normal or high-fat diet (HFD). Males exposed to the high dose of BPA showed impaired glucose tolerance on both diets. They also showed reduced proopiomelanocortin fiber innervation into the paraventricular nucleus of the hypothalamus, and when exposed to HFD, they demonstrated increased neuropeptide Y and Agouti-related peptide expression in the arcuate nucleus (ARC). Females exposed to the high BPA dose were heavier, ate more, and had increased adiposity and leptin concentrations with reduced proopiomelanocortin mRNA expression in the ARC when consuming a HFD. BPA-exposed females showed ARC estrogen receptor α expression patterns similar to those seen in males, suggesting a masculinizing effect of BPA. These results demonstrate that early-life exposure to the obesogen BPA leads to sexually dimorphic alterations in the structure of hypothalamic energy balance circuitry, leading to increased vulnerability for developing diet-induced obesity and metabolic impairments, such as glucose intolerance.

Altered metabolic and neurochemical responses to chronic unpredictable stressors in ghrelin receptor-deficient mice

Ghrelin, a hormone produced by the stomach, is generally associated with feeding responses and the regulation of food intake. Recent evidence, however, suggests that ghrelin is also a stress hormone, given that it is released following acute and chronic stressors. The present study examined the role of ghrelin in producing normal metabolic and neurochemical responses to chronic stress. This was achieved by examining these responses in mice with targeted deletions of the ghrelin receptor gene (GHSR KO mice), and comparing them with the same responses in their wild‐type (WT) littermates. As expected, WT stressed mice decreased their caloric intake, body weight gain and caloric efficiency while maintaining adiposity. GHSR KO mice, however, did not show these alterations despite having normal glucocorticoid responses to stress. In parallel with these changes, chronic unpredictable stress caused changes in norepinephrine, dopamine and serotonin in a number of brain regions. Of these, norepinephrine neurotransmission in the arcuate nucleus and prefrontal cortex was differentially altered in GHSR KO mice. Within the nucleus acumbens, dopamine utilization was increased in WT mice but not in GHSR KO mice. Finally, there were strain differences in serotonin neurotransmission that may explain interstrain body weight and adiposity differences. These results suggest that the metabolic changes necessary to deal with the energetic challenge presented by repeated exposure to stressors do not occur in GHSR KO mice, and they are discussed within the context of the potential vulnerability to stress‐induced pathology.

Stress induced obesity: lessons from rodent models of stress

Stress was once defined as the non-specific result of the body to any demand or challenge to homeostasis. A more current view of stress is the behavioral and physiological responses generated in the face of, or in anticipation of, a perceived threat. The stress response involves activation of the sympathetic nervous system and recruitment of the hypothalamic-pituitary-adrenal (HPA) axis. When an organism encounters a stressor (social, physical, etc.), these endogenous stress systems are stimulated in order to generate a fight-or-flight response, and manage the stressful situation. As such, an organism is forced to liberate energy resources in attempt to meet the energetic demands posed by the stressor. A change in the energy homeostatic balance is thus required to exploit an appropriate resource and deliver useable energy to the target muscles and tissues involved in the stress response. Acutely, this change in energy homeostasis and the liberation of energy is considered advantageous, as it is required for the survival of the organism. However, when an organism is subjected to a prolonged stressor, as is the case during chronic stress, a continuous irregularity in energy homeostasis is considered detrimental and may lead to the development of metabolic disturbances such as cardiovascular disease, type II diabetes mellitus and obesity. This concept has been studied extensively using animal models, and the neurobiological underpinnings of stress induced metabolic disorders are beginning to surface. However, different animal models of stress continue to produce divergent metabolic phenotypes wherein some animals become anorexic and lose body mass while others increase food intake and body mass and become vulnerable to the development of metabolic disturbances. It remains unclear exactly what factors associated with stress models can be used to predict the metabolic outcome of the organism. This review will explore a variety of rodent stress models and discuss the elements that influence the metabolic outcome in order to further extend our understanding of stress-induced obesity.

Understanding the neuroinflammatory response following concussion to develop treatment strategies

Mild traumatic brain injuries (mTBI) have been associated with long-term cognitive deficits relating to trauma-induced neurodegeneration. These long-term deficits include impaired memory and attention, changes in executive function, emotional instability, and sensorimotor deficits. Furthermore, individuals with concussions show a high co-morbidity with a host of psychiatric illnesses (e.g., depression, anxiety, addiction) and dementia. The neurological damage seen in mTBI patients is the result of the impact forces and mechanical injury, followed by a delayed neuroimmune response that can last hours, days, and even months after the injury. As part of the neuroimmune response, a cascade of pro- and anti-inflammatory cytokines are released and can be detected at the site of injury as well as subcortical, and often contralateral, regions. It has been suggested that the delayed neuroinflammatory response to concussions is more damaging then the initial impact itself. However, evidence exists for favorable consequences of cytokine production following traumatic brain injuries as well. In some cases, treatments that reduce the inflammatory response will also hinder the brains intrinsic repair mechanisms. At present, there is no evidence-based pharmacological treatment for concussions in humans. The ability to treat concussions with drug therapy requires an in-depth understanding of the pathophysiological and neuroinflammatory changes that accompany concussive injuries. The use of neurotrophic factors [e.g., nerve growth factor (NGF)] and anti-inflammatory agents as an adjunct for the management of post-concussion symptomology will be explored in this review.

Ghrelin-deficient mice have fewer orexin cells and reduced cFOS expression in the mesolimbic dopamine pathway under a restricted feeding paradigm.

Ghrelin is an orexigenic stomach peptide previously found to be important for the full display of anticipatory locomotor activity and hypothalamic neuronal activation that precedes a daily scheduled meal in mice. Ghrelin is also important for food-related motivation and seems to have direct effects in the mesocorticolimbic dopamine reward system. Here we hypothesized that neuronal activation in reward-related areas in anticipation of a scheduled meal could be mediated by elevated ghrelin induced by scheduled feeding, and therefore this would be attenuated in ghrelin receptor knock-out (GHSR KO) animals. We found that this was indeed the case for the ventral tegmental area and the shell, but not the core, of the nucleus accumbens. In addition, our results show a reduction in the proportion of activated orexin-immunoreactive (IR) neurons in GHSR KO animals in anticipation of the scheduled meal in comparison to the proportion of activated orexin neurons in wild type (WT) mice. Interestingly we observed that both GHSR and ghrelin KO mice had fewer orexin-IR cells than their WT littermates suggesting that lack of ghrelin or sensitivity to ghrelin may play a role in the development of the orexin system. Our data also suggest that ghrelin may mediate food anticipation, in part, by stimulating both the orexin system and the mesolimbic reward system.

Circadian Mechanisms of Food Anticipatory Rhythms in Rats Fed Once or Twice Daily: Clock Gene and Endocrine Correlates

Circadian clocks in many brain regions and peripheral tissues are entrained by the daily rhythm of food intake. Clocks in one or more of these locations generate a daily rhythm of locomotor activity that anticipates a regular mealtime. Rats and mice can also anticipate two daily meals. Whether this involves 1 or 2 circadian clocks is unknown. To gain insight into how the circadian system adjusts to 2 daily mealtimes, male rats in a 12∶12 light-dark cycle were fed a 2 h meal either 4 h after lights-on or 4 h after lights-off, or a 1 h meal at both times. After 30 days, brain, blood, adrenal and stomach tissue were collected at 6 time points. Multiple clock genes from adrenals and stomachs were assayed by RT-PCR. Blood was assayed for corticosterone and ghrelin. Bmal1 expression was quantified in 14 brain regions by in situ hybridization. Clock gene rhythms in adrenal and stomach from day-fed rats oscillated in antiphase with the rhythms in night-fed rats, and at an intermediate phase in rats fed twice daily. Corticosterone and ghrelin in 1-meal rats peaked at or prior to the expected mealtime. In 2-meal rats, corticosterone peaked only prior the nighttime meal, while ghrelin peaked prior to the daytime meal and then remained elevated. The olfactory bulb, nucleus accumbens, dorsal striatum, cerebellum and arcuate nucleus exhibited significant daily rhythms of Bmal1 in the night-fed groups that were approximately in antiphase in the day-fed groups, and at intermediate levels (arrhythmic) in rats anticipating 2 daily meals. The dissociations between anticipatory activity and the peripheral clocks and hormones in rats anticipating 2 daily meals argue against a role for these signals in the timing of behavioral rhythms. The absence of rhythmicity at the tissue level in brain regions from rats anticipating 2 daily meals support behavioral evidence that circadian clock cells in these tissues may reorganize into two populations coupled to different meals.

Perinatal Exposure to Low-Dose Bisphenol-A Disrupts the Structural and Functional Development of the Hypothalamic Feeding Circuitry

Bisphenol-A (BPA) is a component of polycarbonate and other plastics to which humans are regularly exposed at low levels. BPA is characterized as an endocrine disruptor because of observations of its estrogenic activity in various experimental models. We have previously shown evidence of disrupted hypothalamic feeding circuitry and leptin sensitivity in adult BPA-exposed animals subjected to a high-fat diet, but because these animals were already exhibiting a diet-induced obese phenotype, we could not rule out the possibility that these observations were simply consequences of the obesity, not a preexisting phenotype produced by BPA exposure. Here, we studied leptin sensitivity and hypothalamic structure in young BPA-exposed animals before the onset of a body weight or metabolic phenotype. Pregnant and lactating CD-1 mice were exposed to either BPA or diethylstilbestrol (DES) at low, environmentally relevant doses via their diet. Studies of leptin function and neurobiology were conducted on offspring at several time points. Young adult offspring from this experiment were resistant to leptin-induced suppression of food intake, body weight loss, and hypothalamic pro-opiomelanocortin (POMC) upregulation. Both male and female BPA-exposed mice showed a reduced density of POMC projections into the paraventricular nucleus of the hypothalamus (PVN). BPA- and DES-exposed pups had respectively delayed and blunted postnatal leptin surges, and POMC projections into the PVN were rescued in female BPA-exposed animals given daily injections of supplemental leptin. Our findings suggest that BPA, a putative obesogen, may exert its effects through developmental programming of the hypothalamic melanocortin circuitry, permanently altering the neurobiology of metabolic homeostasis.

Interruption of ghrelin signaling in the PVN increases high-fat diet intake and body weight in stressed and non-stressed C57BL6J male mice.

Chronic social stress has been associated with increased caloric intake and adiposity. These effects have been linked to stress induced changes in the secretion of ghrelin, a hormone that targets a number of brain regions to increase food intake and energy expenditure and promote increased body fat content. One of the brain sites targeted by ghrelin is the hypothalamic paraventricular nucleus (PVN), a region critical for both the regulation of the stress response and the regulation of energy balance. Given these data, we examined the contribution of ghrelin receptors in the PVN to the metabolic and behavioral changes that are seen during chronic social stress in mice. To do this, mice were implanted with cannulae attached to osmotic minipumps and delivering either vehicle or the ghrelin receptor (growth hormone secretagogue receptor) antagonist [D-Lys-3]-GHRP-6 (20 nmol/day/mouse). Following a week of recovery, half of the animals in each group were exposed to chronic social defeat stress for a period of 3 weeks whereas the other half were left undisturbed. During this time, all animals were given ad libitum access to standard laboratory chow and presented a high-fat diet for 4 h during the day. Results showed that the ghrelin receptor antagonism did not decrease stressed induced caloric intake, but paradoxically increased the intake of the high fat diet. This would suggest that ghrelin acts on the PVN to promote the intake of carbohydrate rich diets while decreasing fat intake and blockade of ghrelin receptors in the PVN leads to more consumption of foods that are high in fat.

Rats perinatally exposed to food restriction and high-fat diet show differences in adipose tissue gene expression under chronic caloric restriction

The aim of this study is to analyze how maternal diet during the lactational period influences the adipose tissue response to chronic caloric restriction in offspring. Lactating dams were subjected to one of three treatments: 50% food restriction (FR), ad lib standard chow (AL), or ad lib high-fat diet (HF). Juveniles were first weaned onto standard chow, then in adulthood 50% calorically restricted and maintained at 90% of normal body weight for 60 d. HF animals showed increased percent body fat compared with AL and FR animals despite equivalent body weights. HF animals showed alterations in the balance of adipose tissue lipogenic (FAS, LPL) and lipolytic (HSL) gene expression that may underlie their propensity to maintain fat stores under caloric restriction.