Yvonne M. Ulrich-Lai - US grants

DESCRIPTION (provided by applicant): Obesity is a major public health problem affecting 22% of adults in the United States. Despite public health efforts to combat obesity, it continues to increase in incidence along with obesity-related health costs. Obesity increases susceptibility for cardiovascular disease and non-insulin dependent diabetes, and has been associated with depression and anxiety. Chronic stress plays a role in the development of obesity, and has also been linked with depression and anxiety disorders. Moreover, obese individuals often have disruptions in the activity of the stress-responsive hypothalamic-pituitary-adrenal (HPA) axis. Thus, the literature supports strong links between stress, stress-related illnesses and the dysregulation of body weight. To determine the nature of the interactions between chronic stress and obesity, this proposal will use a rat model of high fat diet-induced obesity (DIO) combined with chronic variable stress (CVS) exposure to address three principal hypotheses: (1) chronic stress potentiates obesity in rats consuming high fat diet; (2) chronic stress responses are potentiated by short term consumption of high fat diet; and (3) chronic stress responses are attenuated by obesity induced by long term consumption of high fat diet.

DESCRIPTION (provided by applicant): The literature clearly demonstrates that food intake and energy balance are profoundly influenced by stress, and vice versa. However, the neural mechanisms underlying these effects are largely unknown. My career goal is to establish an independent academic research program focused on pursuing the mechanisms by which stress and food/energy balance interact. To begin addressing these interactions, I have developed a model of limited palatable food intake in which rats with free access to food and water are given additional twice daily access to a small amount of sucrose versus water. In this model, sucrose rats show attenuated hypothalamic-pituitary-adrenal (HPA) axis responses to stress and diminished stress-induced neuronal activation in brain reward regions. Moreover, the calories and other post-ingestive consequences of sucrose are neither sufficient nor necessary for the HPA dampening. I hypothesize that activation of brain reward pathways by the highly-palatable sucrose mediates the reduced stress responsiveness. The present proposal tests this hypothesis by using (1) molecular approaches to determine whether a history of limited sucrose intake alters brain reward pathways similar to other types of reward, (2) pharmacological activation of brain reward pathways to determine whether their activation is sufficient for HPA dampening, and (3) excitotoxic lesions of principal brain reward regions to determine whether activity in these regions is necessary for sucrose-mediated HPA dampening. Success in this area of inquiry necessitates a solid background in stress, energy balance, and reward research. My background in stress regulation is strong, and additional expertise in the areas of energy balance and reward, as provided by the proposed research project, coursework, training environment, and career development activities, would greatly enhance my ability to obtain independent funding and pursue an independent research career at the juncture of these highly-interconnected fields. Lastly, the proposed work has important implications for public health. When under stress, many people increase their intake of highly palatable (e.g. tasty high-sugar or high-fat) foods, presumably to help calm or comfort themselves. However, this behavior can increase body weight and contribute to the development of obesity. This project seeks to understand how palatable foods act to decrease the response to stress, thereby providing insight into the motivation to consume these foods and potentially offering new strategies for the prevention and treatment of obesity.

DESCRIPTION (provided by applicant): Food intake and energy balance are profoundly influenced by stress, and vice versa. However, the neural mechanisms underlying these effects are largely unknown. The long-term objective of this project is to pursue the mechanisms by which stress and food/energy balance interact. More specifically, this proposal seeks to understand how highly palatable foods provide stress relief. These mechanisms are studied using a model of limited palatable food intake in which rats with free access to food and water are given additional twice-daily access to a small amount of sucrose solution or water as a control. In this model, sucrose rats have attenuated hypothalamic-pituitary-adrenal (HPA) axis responses to stress and diminished stress-induced neuronal activation in brain reward regions. Moreover, the calories and other post-ingestive consequences of sucrose are neither sufficient nor necessary for the HPA dampening, suggesting that brain reward regions may mediate the response. The basolateral amygdala (BLA) is a key brain reward region that is also implicated in driving stress responses. Moreover, neural activity in the BLA is necessary for stress-dampening by sucrose, and genes related to synaptic plasticity are up-regulated in the BLA following a history of sucrose intake. Collectively, this suggests that palatable foods may dampen stress responses via synaptic remodeling in the BLA. Thus, the current proposal addresses the hypothesis palatable food causes synaptic reorganization in the BLA. We predict that sucrose intake increases BLA inhibitory tone, leading to less stress-excitatory output. Furthermore, we hypothesize that enhanced inhibitory tone in the BLA is due to increased drive to the intra-BLA inhibitory interneurons. The proposal tests this hypothesis in two specific aims. The first specific aim will test the hypothesis that sucrose intake increases synaptic appositions in a manner consistent with enhanced inhibition of BLA output. This aim will use anatomical approaches to identify and quantify synaptic appositions onto BLA neurons following sucrose intake. The second specific aim will test that hypothesis that sucrose-induced BLA synaptic reorganization is mediated by AMPA receptor activation of pCREB. This aim will use intra-BLA blockade of AMPA receptor activity and CREB/pCREB expression to determine whether this signaling pathway mediates sucrose-induced synaptic reorganization. PUBLIC HEALTH RELEVANCE: When under stress, many people increase their intake of highly palatable (e.g., tasty, high-sugar or high-fat) foods, presumably to help calm or comfort themselves. However, this behavior can increase body weight and contribute to the development of obesity. This project seeks to understand how palatable foods act to decrease the response to stress, thereby providing insight into the motivation to consume these foods and potentially offering new strategies for the prevention and treatment of obesity.

DESCRIPTION (provided by applicant): Individuals often engage in rewarding behaviors, including consuming highly-palatable, calorically- dense comfort foods or taking drugs of abuse, as a means of self-medication for stress relief, but the neural mechanisms underlying stress relief by palatable foods are largely unknown. We propose to study these mechanisms using a model in which rats with free access to food and water are given twice-daily access to a small amount of palatable sucrose solution or water as a control. Using this model, we have found that sucrose rats have attenuated hypothalamic-pituitary-adrenal (HPA) axis and behavioral-anxiety responses to stress and diminished stress-induced neuronal activation in brain reward regions. Moreover, the calories and other post- ingestive consequences of sucrose are neither sufficient nor necessary for the HPA dampening, suggesting that brain reward per se may mediate the response. The basolateral amygdala (BLA) is a key brain reward region that is also implicated in driving stress responses. Moreover, neural activity in the BLA is necessary for stress-dampening by sucrose, and genes related to structural and functional plasticity are up-regulated in the BLA following a history of sucrose intake. In support of this idea, immunolabeling for synaptophysin (a marker of presynaptic terminals), phosphorylated cAMP response element-binding protein (pCREB; a postsynaptic marker associated with synaptic plasticity), and gephyrin (a postsynaptic marker of inhibitory postsynaptic densities) are all increased in the BLA following sucrose. The current proposal addresses the hypothesis that palatable food dampens stress responses via pCREB-dependent synaptic remodeling in the BLA. We predict that sucrose intake increases BLA inhibitory tone, leading to attenuated stress-excitatory output. We will test this hypothesis in three specific aims. The first aim will use intra-BLA blockade of CREB/pCREB expression to determine whether this signaling pathway mediates sucrose-induced synaptic reorganization and stress- dampening. The second aim will assess structural and functional plasticity in the BLA after sucrose (using dual immunolabeling with confocal microscopy to quantify synaptic appositions onto BLA neurons, as well as whole- cell electrophysiological recordings from BLA slice preparations) to test the hypothesis that sucrose-induced synaptic remodeling results in increased inhibitory tone in BLA. The third aim will combine tract-tracing and lesion approaches to test the hypothesis that medial prefrontal cortex (mPFC) projections to BLA are necessary for sucrose-mediated synaptic remodeling and stress dampening.

Project Summary The majority of people report eating highly palatable, calorically-dense ?comfort? foods as a means of stress relief. Indeed, individuals with a history of eating palatable foods have improved mood and reduced physiological and emotional responses to stress. However, comfort feeding comes at a significant cost to metabolic health, as stress-related eaters have higher BMIs and have more difficulty losing weight. Despite clear evidence that comfort feeding is a primary cause of obesity for many people, we know very little about why this is the case. This proposal addresses two critical gaps in our knowledge. First, it determines the mechanisms by which comfort feeding gives stress relief in normal weight individuals. Second, it identifies the extent that this stress relief is impaired during diet-induced obesity (DIO). This has important health implications, as it suggests a vicious cycle whereby obese individuals continually increase their consumption of palatable foods to maintain effective stress relief at the cost of worsening metabolic health. We propose to study these relationships using a palatable ?snacking? paradigm in which rats are given twice-daily access to a small amount of palatable sucrose solution, or water as a control. Rats given this limited sucrose intake (LSI) paradigm reduce their chow intake to compensate for the calories in the sucrose and maintain normal body weight, allowing us to isolate the mechanisms by which LSI reduces stress responses in normal weight individuals. Indeed, LSI rats have attenuated neuroendocrine (HPA axis), behavioral, and metabolic (energy mobilization) responses to a stress challenge. Moreover, LSI is unable to provide stress-blunting in DIO rats, suggesting that obesity may increase the amount of palatable food needed to obtain stress relief. The LSI paradigm therefore provides the unique opportunity to determine the mechanism underlying effective stress relief in normal weight rodents, how these mechanisms are disrupted by DIO, and whether DIO escalates sucrose intake thereby restoring effective stress relief at the cost of worsening metabolic health. Furthermore, our prior work implicates the basolateral amygdala (BLA) as a key site for LSI stress relief, though the intra- BLA mechanism that mediates this stress-blunting is not known. Our new preliminary data point to endocannabinoids (eCB; endogenous marijuana-like molecules) acting within the BLA as this mechanism. For instance, restraint stress rapidly increases BLA eCB (anandamide) content in LSI rats, but not water controls, and eCB signaling in the BLA is known to blunt stress responses. This proposal therefore addresses the hypothesis that palatable foods curb stress responses by increasing eCB signaling in the BLA during stress. The proposed experiments determine the contribution of BLA eCB-signaling to LSI stress relief in vivo, and to post-stress neuronal function and plasticity in BLA slices in vitro. Experiments also determine the extent to which DIO interferes with BLA eCB signaling after LSI, as well as the ability of escalated sucrose consumption to recover BLA eCB signaling and stress-blunting during DIO.

Project Summary While the physiological importance of lymph chylomicron transport is well-documented, the physiological roles of other lymph factors remain poorly understood. This is a critical area for additional new research, as indicated in the RFA, which requests that applications determine ?how the signals lymphatics are receiving may affect their function and, thus overall intestinal function?. In order to study these relationships, we will use the rat lymph-fistula model, a novel and powerful paradigm that is routine in our lab, to study the secretion of hormones and other GI factors in vivo. Using this paradigm, we initially demonstrated that lymphatic concentrations of incretin hormones (GLP-1, GIP) are markedly higher than those in the portal or systemic circulation. Our new preliminary data indicate the intriguing possibility that the lymph also transports locally-produced glucocorticoid hormones in response to dietary lipids, and that the lymphatic transport of dietary lipids and hormones varies in a sex- and/or estrous cycle-dependent manner. The present proposal addresses the overall hypothesis that the transport and signaling of hormones and related factors within the GI lymphatic system is necessary for normal functioning of the GI tract and for overall metabolic health. Aim 1 determines the physiological relevance of high incretin concentrations in intestinal lymph. We assess the impact of lymph diversion on glucose absorption and distribution, and determine the mechanisms by which the stomach regulates incretin secretion during nutrient ingestion. Aim 2 determines the physiological relevance of nutrient-induced glucocorticoid hormones in intestinal lymph. We identify the source of the lymphatic glucocorticoid response to lipids, and determine the extent that lymphatic glucocorticoids regulate lipid absorption and its associated proinflammatory response. Aim 3 determines the role of sex and the estrous cycle in intestinal lymph physiology. We evaluate the extent that lymphatic transport functions (for chylomicrons, incretins, inflammatory mediators, and glucocorticoids) vary with sex and/or the estrous cycle, and test the mechanistic role of ovarian hormones in these processes. This multi-PI proposal leverages the complementary expertise of a highly collaborative team that includes Dr. Patrick Tso (expert in GI lymphatics and metabolic hormones), Dr. Yvonne Ulrich-Lai (expert in the HPA axis and corticosterone signaling), and Dr. Min Liu (expert in ovarian hormone signaling and energy balance).

Project Summary Engaging in pleasurable pastimes (e.g., hobbies, sports, and other leisure activities) can improve mood and reduce perceived stress, suggesting that these activities are an effective means to confer stress resilience. Chronic stress is often unavoidable, making the development of strategies to enhance stress resilience a clear priority for the prevention or amelioration of stress-related diseases. Since beneficial behaviors likely promote stress buffering via activation of brain pleasure and reward circuitry, we have developed and characterized a rat model of stress buffering using intermittent access to a natural reward, limited sucrose intake (LSI). LSI reduces the adverse behavioral effects of chronic stress (e.g., diminished sociability and threat appraisal) and decreases hypothalamic-pituitary-adrenocortical axis reactivity. The stress-buffering provided by LSI is reproduced by a noncaloric sweetener and other naturally rewarding behaviors (sexual activity), but not by intragastric gavage of sucrose, supporting that the stress-protective effects of LSI are primarily due to its rewarding properties. Our preliminary data suggest that LSI acts by altering top-down regulation of the basolateral amygdala (BLA) by the prelimbic medial prefrontal cortex (PL mPFC). In addition, BLA projection neurons can be divided into multiple subsets based on their distinct efferent projection sites, and can play distinct roles in BLA-related behaviors. Thus, while LSI reduces total stress-induced neuronal activation (cFos) in the BLA, the impact on distinct BLA PN populations will likely underlie its role in stress resilience. In support of this idea, LSI reduces post-stress cFos expression in the ventral hippocampus (vHPC) and increases it in the anterodorsal bed nucleus of the stria terminalis (adBST) ? two regions that have receive direct BLA input and exert opposing effects on stress-related behaviors. This suggests that LSI may provide stress resilience by reducing the activity of direct BLA-vHPC projections, and increasing the activity of direct BLA-adBST projections. This proposal therefore uses the LSI model to test the hypothesis that chronic engagement in naturally rewarding experiences promotes behavioral resilience to chronic stress by altering a stress-reward neurocircuitry linking the mPFC, BLA, vHPC and adBST. The first aim tests the contribution of PL top-down regulation of the BLA, while the second aim tests the contribution of specific BLA projections to the vHPC and adBST. Chemogenetic (DREADD) technology is combined with a retrograde viral approach to obtain circuit- specific modulation of neural activity. The effects of circuit manipulation (activation and inhibition) on sociability and threat appraisal behaviors is assessed in the context of chronic stress and/or reward (LSI). This work has important implications, suggesting the presence of endogenous neurocircuits for stress buffering that can be recruited by engaging in naturally-rewarding behaviors. An improved understanding of these neurocircuit mechanisms may be leveraged to develop therapeutic strategies that minimize the adverse effects of chronic stress on mental health, and may guide the optimization of alternative interventions for stress relief.