Darleen A. Sandoval, Ph.D. - US grants

DESCRIPTION (provided by applicant): Intensive glucose treatment (maintaining plasma glucose at normal levels) in type 1 DM patients can delay the onset and slow the progression of microvascular complications. Unfortunately, intensive therapy is associated with an ~ threefold increase in severe hypoglycemia. Exercise, which improves glycemic control, insulin sensitivity, and quality of life, is an important clinical adjunct to diabetes treatment. Unfortunately, exercise also increases the incidence of hypoglycemia, but the mechanisms for this are not clearly understood. Thus, the aims of this proposal are to determine in type 1 DM: 1) whether the metabolic consequences following antecedent hypoglycemia or prior exercise will significantly alter the ability of an individual to defend against same day hypoglycemia and 2) whether following antecedent hypoglycemia, exercise in the presence of modest hyperinsulinemia will lead to substantial deficits in metabolic counterregulatory mechanisms. We will use glucose clamping and exercise and to assess neuroendocrine (plasma glucagon, catecholamines, growth hormone, cortisol, ACTH, and pancreatic polypeptide), in vivo metabolism (glucose and glycerol turnover, substrate levels, and substrate oxidation via indirect calorimetry), and muscle sympathetic nerve activity to thoroughly study these specific aims. The studies will allow us to elucidate in-vivo mechanisms responsible for counterregulatory failure during exercise related hypoglycemia in type 1 DM and to identify new treatment strategies to prevent this problem.

[unreadable] DESCRIPTION (provided by applicant): [unreadable] Obesity and type 2 diabetes mellitus are national and worldwide epidemics. The physiological mechanisms that lead from obesity to abnormal glucose homeostasis and eventual type 2 diabetes mellitus remain elusive. Interestingly, accumulating evidence suggests that neurons within the hypothalamus are sensitive to peripheral nutrients and hormones such as insulin and leptin to regulate peripheral glucose homeostasis. These data point to the brain as an important link between obesity and type 2 diabetes mellitus. [unreadable] A wide range of evidence points to an important role for glucagon-like peptide-1 (GLP-1) in the control of glucose levels in the blood and this has resulted in the recent approval of long-acting GLP-1 analogs for the treatment of type 2 diabetes mellitus. GLP-1 is made in the intestine and the conventional model hypothesizes that it acts on peripheral receptors to increase insulin secretion. However, GLP-1 is also made in the brain and GLP-1 receptors are found in several key regions of the brain that have been linked to the control of peripheral glucose levels. In fact, preliminary data conducted by Dr. Sandoval suggest that GLP-1, is a central nervous system signal that regulates peripheral glucose levels. Therefore, this proposal is aimed at understanding the mechanisms that link the brain GLP-1 system to peripheral glucose homeostasis. Specifically, the goals of this proposal are: to determine which populations of neuronal GLP-1 receptors mediate the effect of GLP-1 on peripheral glucose levels; the degree to which changing glucose levels are due to central GLP-1 effects on the liver or pancreas; and the key intracellular mediators of central GLP-1's action on peripheral glucose levels. Finally, we will also determine whether the glucose impairments produced by diet-induced obesity are associated with impairments of central nervous system GLP-1 function. The long-term goals of this research are to elucidate the specific cellular and neuronal mechanisms that link obesity and type 2 diabetes mellitus. Further, the current proposal will provide Dr. Sandoval an ideal opportunity to add a number of basic research skills to her already well-developed human research expertise. This will allow her to pursue an independently funded career of hypothesis-driven translational research to elucidate the etiology and develop better treatment strategies for obesity and type 2 diabetes mellitus. [unreadable] [unreadable]

DESCRIPTION (provided by applicant): Obesity and type 2 diabetes mellitus (T2DM) are worldwide epidemics and the mechanisms that lead from obesity to abnormal glucose homeostasis remain elusive. Accumulating evidence suggests that the neurons within the hypothalamus that sense peripheral nutrients and hormones to regulate food intake also regulate glucose homeostasis. This raises the possibility that CNS mechanisms could link obesity and T2DM. Glucagon-like peptide-1 (GLP-1), secreted by the intestine, is a potent stimulus for insulin secretion and is essential for normal glucose homeostasis. Several long-acting GLP-1 analogs have recently been developed for the treatment of T2DM. These new GLP-1 based drugs are presumed to work directly on the pancreatic beta- cell to promote insulin release. However, GLP-1 is also made in the brain, with receptors in several key regions implicated in the control of food intake and glucose homeostasis. Our preliminary data indicate that GLP-1 signaling within the hypothalamus regulates peripheral glucose levels and that the ability of CNS GLP-1 to regulate both glucose levels and food intake is dependent on glucose availability. In the beta-cell, the ability of GLP-1 to induce insulin secretion is dependent on elevated ambient glucose concentrations, a key control for homeostatic regulation. Glucokinase (GK) has been proposed to function as a glucose sensor, and we propose that the ability of central GLP-1 to regulate energy and glucose homeostasis is dependent upon glucose availability and that this in turn is regulated via GK. Specifically, the goals of this proposal are to determine which populations of GLP-1r within the CNS are linked to GK function and glucose sensing in regulation of both food intake and glucose homeostasis. The long term goals of this proposal are to elucidate specific cellular and neuronal mechanisms that link obesity to type 2 diabetes mellitus.

Project Summary: A novel paracrine role for GLP-1 in the islet The preproglucagon gene (Gcg), expressed in the intestine, the pancreas, and a small cluster of neurons in the hindbrain, encodes multiple peptides in a tissue-specific manner. One of these peptides, glucagon like peptide-1 (GLP-1) increases following meals, functions to stimulate insulin secretion, and is essential for normal glucose tolerance. The dogma is that intestinally-derived GLP-1 acts as a hormone binding to pancreatic GLP-1 receptors (GLP-1r) to stimulate insulin secretion. However, in both human and rodent models, there is emerging in vitro and pathophysiological evidence to support the production of GLP-1 in the endocrine pancreas. Our preliminary data reveal in vivo evidence that not only does a pancreatic source of GLP-1 exist, but that this pool of active peptide plays a necessary physiological role in normal glucose tolerance. These data represent a paradigm shift in our understanding of the GLP-1 system. In this model, the acute insulinotropic effects of endogenous GLP-1 are paracrine rather than endocrine, derived from islet ?- cells and acting on ?-cell GLP-1r to stimulate insulin secretion. This model would address many of the questions faced when arguing that GLP-1 has classical endocrine action on the islets. Limited by rapid intravascular metabolism, the plasma concentrations of GLP-1 are relatively low and are only modestly elevated during meal ingestion. Interestingly, there are multiple experimental models available that manipulate plasma and/or pancreatic GLP-1 and even more interesting is that two, in particular, represent extremes in ?- cell function. Both streptozotocin-induced diabetes and bariatric surgery raise plasma and/or pancreatic GLP-1 with opposite effects on islet function. In this proposal, we will use our unique genetic models combined with pharmacological and surgical interventions in order to advance the understanding of the in vivo role of pancreatic GLP-1 and in the process will help elucidate many controversies surrounding this source of GLP-1. We propose to do this in 2 specific aims: Specific Aim 1 is focused on understanding the physiological function of pancreatic GLP-1 and will test the hypothesis that pancreatic GLP-1 stimulates insulin secretion and production through paracrine mechanisms. Specific Aim 2 is focused on the pharmacological function of pancreatic GLP-1 and will test the hypothesis that the contribution of pancreatic GLP-1 to ?-cell function increases in bariatric surgery and streptozotocin-induced diabetes.

Project Summary/Abstract - Animal Studies Core As it has for over a decade, the fee-for-service MDRC Animal Studies Core (ASC) (previously, the Animal Phenotyping Core- APC) provides state-of-the art equipment, services, training, and consultation regarding the detailed metabolic phenotyping of mouse and rat models of metabolic disease. T o address the previously unmet needs of MDRC members , the MDRC has invested in new technology and established a host of new services over the past five years . The ASC will continue to support these technologies and services, providing access to crucial equipment, expertise and training to empower specialized studies of rodent models of diabetes and related diseases. The ASC consists of four labs: 1) The Rat Metabolic Phenotyping Lab: includes the assessment of glucose homeostasis, whole animal metabolic assessment, body composition, and other specialized metabolic assessments in rats. 2) The Optogenetics and Behavioral Phenotyping Lab: provides training and access to optogenetic equipment to examine physiologic and behavioral responses to neural circuit manipulation, as well as with equipment to measure relevant behaviors, such as homeostatic and non-homeostatic feeding, activity, reward, and other behaviors that impact and/or are regulated by metabolic parameters in rodents. 3) The Continuous Glucose Monitoring Lab: provides continuous assessment of blood glucose concentrations in conscious, unrestrained rodents by radiotelemetry. This technology minimizes the stress of handling rodents and permits a detailed analysis of glucose fluctuations within normal feeding patterns and across extended time-frames. 4) The Islet Lab: provides islet isolation from mice and rats and ex vivo studies (including perifusion) of islets and other endocrine tissues. The ASC directly supports the goals of the MDRC. The services that the ASC provide are unique and are an important means to study rodent models of diabetes and related diseases, without which crucial aspects of diabetes-related research could not be accomplished. This core provides the necessary i nfrastructure to perform advanced, standardized, metabolic phenotyping of animal models of diabetes and related disorders that arise from genetic, pharmacologic, dietary, or other perturbations. The centralized equipment and services expedites research for many MDRC investigators in a cost-effective manner and provides access to complex metabolic techniques that they may not have otherwise.

Neural mediators of the metabolic effects of vertical sleeve gastrectomy Abstract Decisions about what and how much to eat are regulated by a complex communication network between the CNS and gut and involve a variety of hormonal, metabolite, and neuronal feedback systems. Bariatric surgery, arguably the most effective treatment for obesity and its complications, alters every aspect of these feedback systems and results in substantial weight loss and metabolic improvements. We have demonstrated that nutrient-induced neuronal activation (FOS) is greater within a specific subset of neurons (calcitonin receptor; CALCR) within the nucleus of the solitary tract (NTS), a CNS region that is critical for integrating peripheral signals and initiating changes in feeding behavior, after a particular bariatric surgery, vertical sleeve gastrectomy (VSG). VSG, a procedure where 80% of the stomach along the greater curvature is removed, generates several potential chemo- and mechano-sensing signals that these neurons respond to including the levels of nutrients themselves, greater gastric pressure, or the several-fold increase in many postprandial gut- secreted peptides. The overall aim of this proposal is to determine the identity and function of these activated NTS neurons and the mechanism(s) by which these neuronal populations are activated. In projects 1&2 of this program project, we have generated preliminary data demonstrating that distinct populations of neurons within the NTS (LEPRb, CALCR, and CCK) and PBN (GLP-1R and CGRP) are critical for regulation of feeding and responses to toxins as measured by conditioned taste aversion. Given that our data also demonstrates that CALCR within the NTS are specifically activated by VSG, our over-arching hypothesis is that obesity impairs, and bariatric surgery ?fixes? these circuits to reduce feeding and induce weight loss. To test this hypothesis, we will use genetic and chemogenetic strategies in combination with electrophysiology (using the neural physiology core, NPC, Goforth) to determine the relevant circuits within the NTS (Aim 1) responsible for changes in feeding behavior with obesity and after bariatric surgery, and in Aim 2, we will define the mechanisms that underlie the surgery-induced NTS activation.

Abstract Bariatric surgery is increasingly recognized as a potent tool for the treatment of type 2 diabetes (T2D), yielding not only weight loss but also rapid improvements in glycemia allowing discontinuation of diabetes-related medication within days after surgery1. However, along with this metabolic success comes an increased incidence of severe hypoglycemia (termed post-bariatric hypoglycemia; PBH) for a subset of patients. Severe hypoglycemia causes not only distressing adrenergic and cholinergic symptoms but also neuroglycopenia and hypoglycemia unawareness, impairing safety and increasing risk for disability, syncope, arrhythmias, seizures, coma, and death. Although initially thought to occur in <1% of patients and isolated to roux-en-Y gastric bypass (RYGB), latest estimates suggest that it occurs in ~30% of patients, after both RYGB and vertical sleeve gastrectomy (SG), with similar presentation and spectrum of severity. Increased postprandial incretin and insulin secretion, increased insulin sensitivity, and altered bile acid metabolism have all been implicated in the pathogenesis of PBH. Recent work also suggests that RYGB reduces counterregulatory responses to hypoglycemia. Together, these data suggest that, like the mechanisms that underlie the metabolic success of surgery, the mechanism(s) that underlie PBH are multi-factorial. However, an important feature of PBH is the timing of symptoms. While increases in incretins and improvements in glucose homeostasis occur almost immediately, the onset of PBH typically occurs years postoperatively. Thus, there is more to PBH than increased incretin and consequently insulin responses. The goal of this proposal is to utilize a combination of preclinical and clinical studies to identify physiological and molecular mechanisms that underlie PBH, to determine whether these changes also contribute to surgery-induced improvements in glucose homeostasis, and to define potential therapeutic interventions for PBH. We will determine if decreases in the counterregulatory hormonal responses to hypoglycemia precede bariatric surgery-induced improvements in peripheral insulin sensitivity in rodents (Aim 1). We will test the hypothesis that counterregulatory hormonal responses to hypoglycemia are impaired in post-bariatric patients with or without PBH (Aim 2). Finally, we see that FGF19 is increased in patients with PBH and we will utilize both clinical and preclinical strategies to answer the critical questions of what drives this increase in FGF19 (Aim 3) and whether it is required for the development of PBH (Aim 4).

Abstract/Summary ? P/F Program The primary goal of the MNORC Pilot and Feasibility Grants Program (P&F) is to promote research on the biological and behavioral determinants of obesity, the effects of altered composition and quantity of nutrients to alter metabolism and modulate disease risk, and to develop interventions to reduce obesity and its disease sequelae using basic, clinical, or population approaches. To achieve this goal, we provide opportunities for both new and established University of Michigan investigators to generate sufficient preliminary information for a successful application for major research funding from NIH or other national granting agencies. The $100K/year for the MNORC P&F grants program is largely funded from cost- sharing provided by several Schools within the University of Michigan, and is currently in the midst of its second cycle of funding. Two to three standard grants have been given out each year, and despite that the program has only completed two funding cycles, this investment has resulted in a considerable return of published papers, funded NIH grants, and collaborations with other MNORC members. In addition to the P&F grants currently provided, the P&F program will expand over the next cycle to include shared grants funded with the Michigan Diabetes Research Center and the Taubman Center for common research interests.