Gary J. Schwartz - US grants

DESCRIPTION (provided by applicant): Understanding normal and dysfunctional energy balance and body weight regulation requires neural evaluation of the signals involved in the control of food intake within a meal, as well as signals related to the availability of stored fuels. Because ingested nutrients stimulate multiple gastrointestinal sites simultaneously, evaluating the neural representation of combinations of meal-related stimuli is critical to the characterization of putative negative feedback signals that mediate the control of food intake. In the proposed studies, we outline neurophysiological experiments designed to elucidate the neuro-humoral basis of energy balance. These experiments will: 1) identify and characterize short term meal-related gut neurophysiological signals, 2) determine their representation and integration at peripheral vagal, non-vagal and central brainstem nervous system sites, and 3) evaluate how they are interpreted in the context of the neuro-humoral signals related to the long-term control and mobilization of stored fuels. We will focus on the sensory vagus and splanchnic nerves and their central nervous system projections as the main neuroanatomical pathways linking the upper gastrointestinal sites exposed to nutrients during a meal and the central nervous system sites mediating the control of food intake. We will characterize vagal and splanchnic afferent responses to meal-related stimulation of the stomach and duodenum, and determine the extent to which these responses are modulated by administration of gut-brain peptides and neurotransmitters normally released by the duodenal presence of nutrients. Using single unit extracellular neurophysiological recordings, we will assess the neural brainstem representation of signals arising from meal-related stimulation of multiple alimentary tract compartments, including the stomach and duodenum. To characterize the ability of the brainstem to integrate long- and short-term energy balance signals, we will also determine how central administration of putative long-term energy balance peptides modifies meal-related brainstem neurophysiological signals from the gut. This systematic assessment of meal-related peripheral neural activity, its brainstem neural representation, and its integration with putative energy balance peptide signals represent a synthetic evaluation that will significantly advance our understanding of neuro-humoral interactions in the metabolic control of food intake.

DESCRIPTION (provided by applicant): Understanding energy balance and body weight regulation requires an understanding of the behavioral and neural evaluation of the oral and post-oral sensory signals involved in the control of food intake within a meal, as well as signals related to the availability of stored fuels. Obesity represents an important dysfunctional state of energy balance, and is frequently accompanied by hyperphagia that is manifested by increased meal size. Two mouse models of obesity, the ob/ob mouse lacking leptin, and the db/db mouse, lacking functional leptin receptors (LEPR-B), are also hyperphagic, and exhibit increased meal size without altered meal frequency relative to wild type lean controls. In the proposed studies, we outline behavioral and immunocytochemical studies designed to elucidate the role of leptin signaling in determining the oral and post-oral sensory influences on ingestion in obesity. These experiments will: 1) characterize the ability of oral and upper gastrointestinal (GI) food stimuli to affect food intake within a meal, 2) assess the degree to which genetic leptin signaling deficiency interferes with the feedback potency of oral and upper GI signals in the control of meal size, and 3) characterize the patterns of central nervous system activation excited by oral and GI stimuli that affect meal size in mice with alterations in leptin signaling. We will focus on the C57B6J mouse as the background strain for these studies because: 1) it is has a well-described tendency toward dietary obesity and, 2) it is the background strain for ob/ob and db/db mice. We will evaluate: 1) the ability of leptin to modify the feedback potency of oral and GI food stimuli in ob/ob mice, 2) the ability of transgenic neuron-specific replacement of LEPR-B to restore normal processing of oral and GI food stimuli in db/db mice, and 3) the effect of central vs. peripheral inducible LEPR-B deficiency on eating in mice. To identify central neuronal regions important in leptin's ability to modulate the processing of food stimuli, we will also evaluate the central nervous system patterns of c-Fos expression in these strains in response to selective oral and/or GI food stimuli. This systematic assessment of meal-related stimuli, their central neural representation, and their integration with energy balance peptide signals will significantly advance our understanding of neuro-humoral interactions in the metabolic control of food intake.

? DESCRIPTION (provided by applicant): Central nutrient sensing of the essential amino acid l-leucine is a critical determinant of food intake and meal size. We have shown that: 1) endogenous central levels of leucine are rapidly elevated after a meal, 2) blocking endogenous leucine catabolism within the mediobasal hypothalamus (MBH), thereby promoting local leucine availability, reduces food intake, 3) MBH leucine administration reduces food intake by reducing meal size, 4) blocking downstream intracellular cascades of leucine signaling in the MBH promote feeding, while 5) chronic activation of these downstream pathways in the MBH limit high fat diet hyperphagia and associated weight gain. These actions appear to be mediated by two intracellular signaling pathways: the mammalian target of rapamycin (mTOR) - serine/threonine kinase p70S6K (S6K) pathway, and the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway. MBH leucine at feeding inhibitory doses also activates the brainstem dorsal vagal complex of the caudal brainstem, particularly the caudomedial region of the nucleus of the solitary tract (cmNTS), where meal-related gut negative feedback signals converge and are integrated to mediate the neural control of meal size. Our recent published and preliminary results support the identification of the cmNTS as a site where local leucine acts to reduce food intake by limiting meal size and by increasing the feeding inhibitory potency of CCK. These actions appear to be mediated by both mTOR-S6K and ERK pathways as well. Furthermore, diet induced obesity (DIO) attenuates cmNTS leucine's feeding inhibitory actions. Taken together, these data suggest a new brainstem nutrient sensing capability, and its novel integration with direct controls of meal size. Studies in this proposal will apply a coordinated combination of behavioral, neurophysiological, pharmacological, immunohistochemical and molecular genetic approaches to identify and characterize the neural and molecular mechanisms underlying brainstem nutrient sensing in the control of feeding, how it is disrupted in DIO, and how it can be targeted to control food intake and body weight in obesity.

The Animal Physiology Core (APC) employs sophisticated research methodologies to assist Einstein-Mount Sinai Diabetes Research Center (ES-DRC) investigators in the in vivo assessment of glucose and fatty acid metabolism, insulin sensitivity and energy homeostasis in mice and rats. Through collaborative efforts with the other Cores of the ES-DRC, the APC enables investigators to thoroughly characterize the effects of defined pharmacological, dietary, environmental and genetic alterations on glucose and lipid homeostasis, insulin action, and metabolism. The Core performs studies of adiposity distribution and facilitates NMR spectroscopy, fMRI and microPET analysis of experimental animal models of diabetes undergoing metabolic studies. The Core also provides specialized rodent surgeries for investigator laboratories and several cardiac functional assessments related to diabetic complications. To accomplish these goals, the Animal Physiology Core will: 1) advise investigators in the design of metabolic studies relevant to the control of glucose homeostasis and insulin action in rodents; 2) make available to investigators specialized measurements of whole body and tissue-specific glucose sensitivity and insulin action including, but not limited to insulin, pancreatic and hyperglycemic clamp studies; 3) provide specialized surgical models for the study of insulin sensitivity, energy balance, and glucose and fatty acid metabolism; 4) offer instruction to students, postdoctoral fellows, investigators and technical staff in performing surgical and physiological techniques necessary to evaluate the controls of glucose homeostasis and insulin action; 5) provide analysis of whole body carbohydrate/fatty acid oxidation, energy expenditure, feeding behavior, and locomotor activity using specialized metabolic (indirect calorimetry) and behavioral rodent cages; 6) provide assessment of the effects of spontaneous or scheduled exercise on glucose homeostasis and metabolism; 7) make available to investigators specialized measurements of rodent adipose tissue distribution using microCT and measurements of glycogen in liver and muscle, intrahepatic lipids and intramyocellular lipids using NMR; 8) make available to investigators specialized measurements of brain energy and glucose utilization by functional MRI (fMRI) and microPET scanning; and 9) coordinate these efforts with other ES-DRC and Institutional Core facilities at Einstein and Mount Sinai. All these services are available to investigators new to diabetes research, as well as to investigators working on diabetes-related projects that can be enriched and extended by the use of the expertise and facilities of this Core.

ABSTRACT Phenotyping animal models of disease is a critical element in our Center's goal to understand, effectively treat, and when possible, prevent intellectual and developmental disabilities in children. Accordingly, the mission of the Animal Phenotyping Core (Core E, AP) is to assist investigators seeking to discover behavioral, physiological, structural and metabolic phenotypes in diverse rodent models of intellectual and developmental disabilities. The AP Core performs studies primarily in mice and rats to identify the functional alterations resulting from genetic, developmental or environmental manipulations that may impair neural and behavioral development. These include changes in developmental milestones, sensorimotor function, cognitive function, affective and social behaviors, feeding and activity patterns, body composition and/or energy expenditure, patterns of brain activity as assessed by EEG and brain imaging in MRI/DTI (diffusion tensor imaging) and PET scans. The AP Core accomplishes its goals through SubCores focused on (1) metabolism, (2) behavior, (3) brain imaging and (4) electroencephalography. By combining existing capabilities and highly experienced faculty we have established an animal phenotyping facility uniquely suited to plan, perform and evaluate coordinated behavioral, metabolic, and functional neuroimaging and electrophysiological assessments in developing and adult rodents. Through close collaborative efforts with the Neural Cell Engineering and Imaging (NCEI) Core and the Human Clinical Phenotyping (HCP) Core, as well as the Neurogenomics (NGEN) Core, the consequences of defined genetic, environmental and/or physiological alterations are thoroughly characterized to determine their impact in the context of measures most relevant and translatable to the human disease phenotype. The AP Core also makes critical contributions to Aim 2 of the IDDRC Research Project focused on the origins of ID in 22q11.2 deletion syndrome. In addition to the wide range of expertise of its leadership and the resources they bring to this effort, the AP Core also emphasizes the importance of integration across measurement and analytical capabilities, i.e., it facilitates a combination of methodological approaches such as pursuit of brain imaging simultaneously with behavioral studies. We also emphasize phenotyping techniques that mesh well with the types of studies conducted in children with IDD. In pursuit of these scientific objectives, the AP Core leadership, in concert with ADM Core oversight, also carefully monitors IDDRC investigator Core access and user satisfaction, cost effectiveness and quality control.

Animal Phenotyping Core PROJECT SUMMARY/ABSTRACT Rodent models of obesity have provide invaluable to the study of obesity and to the work carried out by NYONRC investigators. To meet the growing needs of Center researchers, the proposed NYONRC Animal Phenotyping Core (APC) reflects a significant expansion of the Animal Energy Balance Core (Einstein) and integration with the animal component of the Adipose Tissue Core (Columbia). During the past 4 years of the current cycle, these components of the NYONRC have processed more than 14,000 service requests for 38 NYONRC members supported by 45 grants, and provided additional services to 9 NIH-funded non-NYONRC members. The services offered by the APC have been instrumental in obtaining 19 new or renewed grants, and in the generation of 213 publications. The proposed APC concentrates and coordinates significantly expanded technical and research capabilities of this amalgamated Core. The incorporation of additional animal resources at both the Einstein and Columbia campuses, will significantly increase the user and research base of the NYONRC overall, and of the APC Core in particular. By coordinated application of NYONRC dedicated facilities, and resources and expertise at both Columbia and Einstein, the proposed APC Core will combine sophisticated qualitative and quantitative measurements of ingestive behavior with concurrent assessments of energy expenditure, physical activity, body temperature, and repeatable noninvasive body composition analysis by MRI and MicroCT. Also available are a range of metabolic surgeries, including gastrointestinal bypass, autonomic denervation procedures, and adipose tissue services including: 1) quantifying adipocyte subpopulations (brown, white, beige), in terms of size, variation, and cellular ultrastructure; 2) primary cell isolation and analysis; 3) differentiation of adipocytes and adipose tissue macrophages; and 4) adipose tissue transplantation. This expanded array of services stems from the growing realization that our understanding of the biology of obesity requires simultaneous assessment of energy intake and energy expenditure, and of the means to determine their net effects on energy balance repeatedly in individual animals. The Specific Aims of the APC are to provide state-of-the-art assessments of: 1) Adipose Tissue Morphology & Function, in terms of cellular, histologic and functional analyses; 2) Body Composition, in terms of the total relative fat and lean mass, and whole body anatomical distribution of adipose tissue; 3) Energy intake and Expenditure, including oxygen and CO2 consumption, respiratory quotient, and thermogenesis in brown adipose tissue; and 4) consequences of Metabolic Procedures and Surgeries, designed to illuminate the physiology of weight regulation and obesity pathophysiology, including gastrointestinal bypass, autonomic denervations and central nervous system targeting using viral, optogenetic and chemogenetic approaches..

Abstract The induction of obesity results from an increase in energy storage relative to energy expenditure and until a new energy equilibrium is established at higher pathophysiologic weight. Obesity is increasing at an alarming rate in the USA with approximately one-third of adults and one-fifth of children classified as obese. In addition, to the direct health care costs, work loss and morbidly, obesity is the primary factor driving insulin resistance and type 2 diabetes. At the experimental level over the past decade there has been a substantial effort focused on increasing energy expenditure through the development/activation of brown/beige adipose tissue thermogenesis. In contrast to adipose tissue, collectively skeletal muscle accounts for approximately 50% of body mass, is the primary determinant of basal metabolic rate and is the major driver of increased energy expenditure that occurs during physical activity. In addition, involuntary skeletal muscle contractions or voluntary activity based skeletal muscle contractions accounts for the majority of heat production during cold induced thermogenesis that can reach 15-20 times the resting basal metabolic rate. During our phenotypic characterization of the TIGAR knockout mice, to our surprise these mice display a remarkable cold resistant phenotype that is independent of brown and beige adipocyte function and is a result of increased skeletal muscle thermogenesis. We plan to use mouse genetics, metabolic profiling and physiologic assessments to determine the molecular basis for the cold resistance that occurs due to TIGAR deficiency. The specific novel aspects of our current findings are that: i) TIGAR deficiency does not affect energy production at room temperature but markedly enhances cold induced thermogenesis, ii) the increased thermogenic response is due to a direct activation of skeletal muscle ATP turnover through increased contractile activity and iii) the increased skeletal muscle contractile activity results from TIGAR deficiency in cholinergic neurons at the neuromuscular junction. A schematic representation of the mechanism(s) responsible for cold resistance in TIGAR knockout mice is illustrated and described in Figure 1. The specific novel aspects of the proposed research plan are: 1) to genetically determine whether TIGAR deficiency enhances skeletal muscle contraction based thermogenesis through increased cholinergic tone, 2) to determine whether the enhanced activation of skeletal muscle thermogenesis results from a loss of TIGAR protein dependent binding interaction(s) and/or due to a loss of TIGAR phosphatase activity, 3) to map the resultant changes in metabolic flux that occurs between thermoneutrality, room temperature and cold exposure in skeletal muscle, and 4) to identify the novel TIGAR-dependent molecular pathway responsible for the increase in cholinergic neuron signaling.

The Animal Physiology Core (APC) employs sophisticated research methodologies to assist Einstein-Mount Sinai Diabetes Research Center (ES-DRC) investigators in the in vivo assessment of glucose and fatty acid metabolism, insulin sensitivity and energy homeostasis in mice and rats. Through collaborative efforts with the other Cores of the ES-DRC, the APC enables investigators to thoroughly characterize the effects of defined pharmacologic, dietary, environmental and genetic alterations on glucose and lipid homeostasis, insulin action, and metabolism. To accomplish these goals, the Animal Physiology Core will: 1) Offer advice and instruction to students, postdoctoral fellows, investigators and technical staff in the design and performance of physiologic approaches and techniques necessary to evaluate the control of glucose homeostasis and insulin action in rodents, 2) Make available to investigators specialized measurements of whole body and tissue-specific glucose metabolism and insulin action in rodent models including insulin, pancreatic and hyperglycemic clamps and spontaneous glucose monitoring, 3) Provide specialized gastrointestinal, neurosurgical and histological models for the study of insulin sensitivity, energy balance, and glucose and fatty acid metabolism, including gastric bypass and adipose and hepatic tissue denervation, imaging and photo-stimulation, 4) Provide analysis of whole body carbohydrate/fatty acid oxidation, energy expenditure, thermogenesis, food intake, and locomotor activity using specialized metabolic (indirect calorimetry) rodent cages, 5) Provide assessments of the effects of diet, exercise, light/dark cycle and environmental temperature on continuous and acute assessments of glucose homeostasis, metabolism, and shivering via electromyography, 6) Make available to investigators specialized measurements of rodent adipose tissue distribution using magnetic resonance spectroscopy, microCT, as well as measurements of glycogen in liver and muscle, intrahepatic lipids and intramyocellular lipids using nuclear magnetic resonance (NMR), 7) Make available to investigators specialized measurements of brain energy and glucose utilization by functional magnetic resonance imaging (fMRI) and microPET scanning, 8) Assist investigators in the interpretation of data and to design further experimental approaches to reveal the molecular and physiological bases of metabolically relevant rodent phenotypes, and 9) Facilitate and integrate the functional assessments provided by the APC with assays provided by other ES-DRC Biomedical Cores. All these services are available to investigators new to diabetes research, as well as to investigators working on diabetes- related projects that can be enriched and extended by the use of the expertise and facilities of this Core.

PROJECT SUMMARY/ABSTRACT ? ANIMAL PHENOTYPING Phenotyping animal models of disease is a critical element in our Center?s goal to understand, effectively treat, and when possible, prevent intellectual and developmental disabilities in children. Accordingly, the mission of the Animal Phenotyping Core (Core E, AP) is to assist investigators seeking to discover behavioral, physiological, structural and metabolic phenotypes in diverse rodent models of intellectual and developmental disabilities. The AP Core performs studies primarily in mice and rats to identify the functional alterations resulting from genetic, developmental or environmental manipulations that may impair neural and behavioral development. These include changes in developmental milestones, sensorimotor function, cognitive function, affective and social behaviors, feeding and activity patterns, body composition and/or energy expenditure, patterns of brain activity and anatomy as assessed by brain imaging in MRI/DTI (diffusion tensor imaging) and PET scans. The AP Core accomplishes its goals through SubCores focused on (1) behavior, (2) metabolism, and (3) brain imaging. By combining existing capabilities and highly experienced faculty we have established an animal phenotyping facility uniquely suited to plan, perform and evaluate coordinated behavioral, metabolic, and functional neuroimaging assessments in developing and adult rodents. Through close collaborative efforts with the Neural Cell Engineering and Imaging (NCEI) Core and the Human Clinical Phenotyping (HCP) Core, as well as the Neurogenomics (NGEN) Core, the consequences of defined genetic, environmental and/or physiological alterations are thoroughly characterized to determine their impact in the context of measures most relevant and translatable to the human disease phenotype. The AP Core also makes critical contributions to Aim 2 of the RFK IDDRC signature research project focused on links between mutations in KDM5C and IDD. In addition to the wide range of expertise of its leadership and the resources they bring to this effort, the AP Core also emphasizes the importance of integration across measurement and analytical capabilities, i.e., it facilitates a combination of methodological approaches such as pursuit of brain imaging simultaneously with behavioral studies. We also emphasize, when possible, phenotyping techniques that are most comparable to those used in children with IDD. In pursuit of these scientific objectives, the AP Core leadership, in concert with ADM Core oversight, also carefully monitors IDDRC investigator Core access and user satisfaction, cost effectiveness and quality control.