2010 — 2011 |
De Kloet, Annette Diane |
F31Activity Code Description: To provide predoctoral individuals with supervised research training in specified health and health-related areas leading toward the research degree (e.g., Ph.D.). |
The Role of the Renin Angiotensin System in the Cns Regulation of Energy Balance @ University of Cincinnati
DESCRIPTION (provided by applicant): Millions of people suffer from obesity and the concomitant susceptibility to cardiovascular disease. This project is based on the observation that a key feature of obesity is hyperactivity of the renin-angiotensin- system (RAS). Angiotensin-ll (ANGII), the effector peptide of the RAS, exerts peripheral effects that promote increased energy storage and elevate blood pressure. Although the peripheral actions of the RAS have been studied extensively, the role of ANGII in the neural regulation of energy balance is unclear. Unlike what occurs in the periphery, ANGII promotes negative energy balance in the central nervous system (CNS). Captopril, a drug that blocks the formation of ANGII (i.e., an angiotensin converting enzyme [ACE] inhibitor, resulting in increased plasma ANGI), reduces blood pressure and favors weight loss. However, because captopril does not enter the brain to reduce CNS ACE activity, the increased ANGI is converted locally to ANGII by brain-generated ACE. Increased CNS ANGII results in decreased body weight that is partially explained by alterations in food intake. In contrast, administration of captopril directly into the CNS inhibits brain-generated ACE and reduces CNS ANGII, resulting in increased food intake and implying that the CNS RAS normally influences body weight regulation. However, the role of altered energy expenditure and the underlying mechanism(s) of captopril-induced negative energy balance have not been unequivocally discerned. The proposed experiments will utilize rats to test the overall hypothesis that ANGII plays a key role in the neural control of energy balance by acting to promote negative energy balance via angiotensin receptor type-1 (ATI) in the paraventricular nucleus of the hypothalamus. First, we will test the hypothesis that systemic ACE inhibition (via captopril) results in decreased food intake, increased energy expenditure and altered cardiovascular tone, in part, by increasing central ANGII. Second, we will use a lentiviral vector containing ATI antisense oligonucleotides in order to downregulate AT1 expression in the hypothalamic paraventricular nucleus and thereby test the hypothesis that this ATI population is necessary for maintaining basal energy balance and for captopril-induced negative energy balance. Body weight, body composition, food intake, energy expenditure (indirect calorimetry), cardiovascular function (telemetry), plasma norepinephrine and the expression of corticotrophin-releasing hormone in the paraventricular nucleus of the hypothalamus will be assessed. The significance of the proposed research is that it may lead to development of novel therapeutics to treat or prevent obesity while also preventing concomitant hypertension, diabetes and stroke.
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2012 — 2014 |
De Kloet, Annette Diane |
F32Activity Code Description: To provide postdoctoral research training to individuals to broaden their scientific background and extend their potential for research in specified health-related areas. |
Angiotensin-Ii, Hypothalamic Inflammation and Neurogenic Hypertension
DESCRIPTION (provided by applicant): Hypertension is an epidemic health concern and a major risk factor for cardiovascular disease, the leading cause of death in the U.S. Many forms of hypertension are of neurogenic origin; however, the neural mechanism(s) underlying the development and progression of neurogenic hypertension are incompletely understood. Angiotensin-II (Ang-II), the effector peptide of the renin-angiotensin system, is a potent mediator of cardiovascular function that has pleiotropic actions within the brain. Ang-II is known to induce inflammation via the activation of the angiotensin type 1a receptor (AT1a) in a number of peripheral tissues and in the brain and this is thought to contribute to its hypertensive actions. Consistent with this, hypertension, in addition to being accompanied by enhanced renin-angiotensin system activity is also associated with a mild inflammatory state. One critical site of Ang-II actions within the brain is the paraventricular nucleus of the hypothalamus (PVN), which densely expresses AT1a and integrates signals to and from brain regions critical for the regulation of cardiovascular function and sympathetic nerve activity. This proposal investigates the role of the PVN AT1a in the inflammatory and sympathoexcitatory actions of elevated Ang-II. The proposed experiments will test the overall hypothesis that Ang-II acts at the PVN AT1a receptor to enhance inflammation and microglial activation and that this is an important mechanism for Ang-II-induced hypertension and augmented sympathetic outflow. In the first Specific Aim, experiments will utilize the Cre/lox system in mice to test the specific hypothesis that PVN AT1a are necessary for Ang-II-induced increases in blood pressure and sympathetic nervous system activity. Angiotensin-II-induced cardiovascular dysfunction (telemetric blood pressure assessment) and neuronal activation (c-Fos immunohistochemistry) of cardiovascular control centers of the brain will be assessed in mice that lack AT1a in the PVN and controls. In the second Specific Aim, a combination of the Cre/lox system and pharmacological approaches in mice will be used to determine the role of interactions between PVN AT1a, transforming growth factor beta and inflammation in the regulation of cardiovascular function during elevated Ang-II. Specifically, the necessity of PVN AT1a and transforming growth factor beta signaling for the inflammatory and hypertensive consequences of elevated Ang-II will be assessed. Important endpoints for Aim 2 will include the assessment of proinflammatory cytokines and microglial activation within the PVN and other cardiovascular control centers, as well as the telemetric assessment of cardiovascular function. The proposed research is significant because uncovering the mechanisms of Ang-II regulation of inflammation and cardiovascular function may lead to new strategies for the treatment and prevention of neurogenic hypertension. PUBLIC HEALTH RELEVANCE: Although there are a number of treatment strategies for high blood pressure, less than 50% of hypertensive patients have their condition under control. The objective of the proposed research is to understand mechanisms contributing to hypertension by investigating interactions between angiotensin-II's inflammatory actions in the brain and high blood pressure. This is of particular relevance to public health because uncovering the mechanisms of Ang-II regulation of inflammation and cardiovascular function may lead to new strategies for the treatment and prevention of hypertension.
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0.964 |
2015 — 2018 |
De Kloet, Annette Diane |
K99Activity Code Description: To support the initial phase of a Career/Research Transition award program that provides 1-2 years of mentored support for highly motivated, advanced postdoctoral research scientists. R00Activity Code Description: To support the second phase of a Career/Research Transition award program that provides 1 -3 years of independent research support (R00) contingent on securing an independent research position. Award recipients will be expected to compete successfully for independent R01 support from the NIH during the R00 research transition award period. |
Novel Role of Central At2r in Blood Pressure Regulation
DESCRIPTION (provided by applicant): Project Summary/Abstract Hypertension is a widespread health problem and a major risk factor for cardiovascular disease, the leading cause of death in the USA. Of particular concern is drug-resistant hypertension, which is accompanied by enhanced sympathetic nervous system activity, indicating that the increased blood pressure arises from neurogenic origins. Nearly one-third of hypertensive patients fall into this category for which there are no effective medications and determining strategies to treat or prevent neurogenic hypertension has great significance for public health. My overall career goal is to become an independent academic scientist that studies impairments in neural circuits that elicit neurogenic hypertension, as well as the development of therapeutics that alleviate these impairments. The expectation is that my research will contribute substantively to the understanding of the causes of neurogenic hypertension and to the development of therapeutics used to treat it. The activities proposed in this application are designed to facilitate reaching tis goal and will investigate a novel therapeutic target for neurogenic hypertension - angiotensin type-2 receptors (AT2R) expressed within the brain. The experiments test the overall hypothesis that activation of AT2R on neurons that project to the paraventricular nucleus of the hypothalamus (PVN; a brain region important for controlling sympathetic outflow and blood pressure) negatively-regulate blood pressure, potentially making activation of AT2R a suitable target for antihypertensive medications. My graduate training used laboratory rodents to examine how angiotensin-II, a peptide heavily implicated in the development of hypertension and cardiovascular disease, influenced the neural control of body weight and glucose metabolism. This line of research introduced me to the central pathways that were sensitive to angiotensin-II, which piqued my interest in neurogenic hypertension. Consequently, I chose the laboratory of Dr. Colin Sumners at the University of Florida to conduct my postdoctoral training. Dr. Sumners is a leading expert in the field of neurogenic hypertension and his laboratory is part of the Hypertension Center at UF, which is comprised of core facilities and nearly 50 faculty members dedicated to studying high blood pressure. This training environment contributed to my successful post-doctoral NRSA proposal that afforded competence with the assessment of cardiovascular function in rodents, and perhaps more importantly, found that experimentally-induced hypertension elicited changes within the electrophysiological properties of neurons controlling blood pressure that ultimately increased their excitation. Taking these results into account, I determined that effective therapeutics should decrease or reverse this increased excitation; however, I also determined that additional training in conceptual and technical approaches aimed at understanding the electrophysiological properties of neurons was imperative to developing this line of research. Accordingly, the primary objectives of the K99-phase are to answer some fundamental questions regarding the structure and function of specific AT2R that are positioned to decrease sympathetic outflow and blood pressure, while providing additional training for in vitro patch-clamp electrophysiology. It is anticipated that determining the therapeutic utility of AT2R for neurogenic hypertension and expertise in subcellular neural electrophysiology can be complemented by professional development activities to launch my independent research career. In the first Aim, experiments will combine genetic and neuroanatomical techniques to test the specific hypothesis that AT2R-expressing neurons that make contacts onto preautonomic neurons within the paraventricular nucleus of the hypothalamus express the inhibitory neurotransmitter (GABA), thereby positioning them to decrease blood pressure and autonomic function. In Aim 2, experiments will use patch- clamp electrophysiological techniques to test the specific hypothesis that activation of AT2R on GABA neurons that project to the PVN will facilitate inhibitory (i.e., GABAergic) neurotransmission and that this will lead to reduced activity of PVN preautonomic neurons. These experiments will not only determine precisely how activation of AT2R impact activity within a neuronal network, but they will also serve as a training vehicle for me to learn patch-clamp electrophysiology, a technique that is essential to understanding how subcellular changes in a discrete population of neurons can impact whole animal physiology. Importantly, my background, combined with expertise in electrophysiology will make my research program unique, as it will allow for the study of precisely how angiotensin-II acting through AT2R influences the excitability of specific neurons that control cardiovascular function. Aim 3 will be contained within the R00-phase and will partner my past training with my newly-acquired skills to determine the role of AT2R in blood pressure regulation basally and during neurogenic hypertension. Using the Cre/lox system and pharmacological approaches, I will selectively activate or inhibit AT2R on neurons that project to the PVN and test the specific hypothesis that these AT2R negatively regulate blood pressure and sympathetic nervous system outflow. Collectively, the proposed studies are significant because they may uncover a novel therapeutic target for treatment of neurogenic hypertension while preparing me to establish an independent research program that addresses a problem with high
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0.964 |
2019 — 2021 |
De Kloet, Annette Diane |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Interrogating Distinct Angiotensin Type-1 and Type-2 Receptor Containing Brain Circuits to Understand and Alleviate Hypertension
Project Summary Hypertension is a widespread health problem and a major risk factor for cardiovascular disease. Nearly one- third of hypertensive patients suffer from drug-resistant hypertension; a condition associated with activation of brain angiotensin receptors, enhanced sympathetic nervous system activity and elevated levels of circulating vasopressin. The proposed experiments aim to identify neurons within the brain whose excitation or inhibition is coupled to the pathophysiology underlying resistant hypertension. Our preliminary studies using mice with Cre recombinase or green fluorescent protein directed by the genes for the angiotensin type 1a or type 2 receptors (AT1R or AT2R) have provided intriguing insight. We have discovered that neurons in the organum vasculosum of the lamina terminalis (OVLT) and median preoptic nucleus (MnPO) express AT1R or AT2R and send dense excitatory projections to the paraventricular nucleus of the hypothalamus (PVN) and peri-PVN area, respectively. Fascinatingly, optogenetic excitation of such AT1R neurons elicits robust (>40 mmHg) and sustained increases in blood pressure, suggestive of sympathoexcitation and augmented vasopressin secretion. Within the MnPO/OVLT the vast majority of AT2R(s) are NOT expressed on neurons that also synthesize AT1R, but rather, are a separate population of neurons whose excitation may oppose the onset of hypertension. Consistent with this interpretation, we recently discovered that pharmacological activation of AT2R facilitates GABAergic mediated inhibition of vasopressin neurons and reduces systemic vasopressin and blood pressure. We have developed the overall hypothesis that neurons within the MnPO/OVLT that express AT1R or AT2R project to the PVN and peri-PVN area to coordinate sympathetic outflow and vasopressin secretion, and that the relative activities of these neurons predict resistance or susceptibility to hypertension. To address this hypothesis, the proposed studies combine Cre-LoxP technology, in vitro/in vivo optogenetics and classical systems physiology with a mouse model of hypertension. Aim 1 will use optogenetics to probe the connection between neurons in the MnPO/OVLT that express AT1R or AT2R and neurons in the PVN that express vasopressin. Sufficiency and/or necessity of these connections will be determined for the increased sympathetic nervous system activity and vasopressin secretion that promote neurogenic hypertension. Then, Aim 2 will use the Cre-loxP system to delete AT1R and/or AT2R within the MnPO/OVLT and determine whether AT1R/AT2R signaling within these brain nuclei contribute to the etiology of hypertension. Collectively, these experiments will determine whether excitability and/or AT1R/AT2R signaling within this neural circuit can be altered to prevent high blood pressure. These studies have the potential to uncover, at a detailed and mechanistic level, the neural circuits that are compromised during hypertension and, in the long-term, may inform a therapeutic strategy that optimally targets excitation of AT1R- and AT2R-containing neurons to relieve hypertension.
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0.964 |