Neurohumoral regulation of PVAT

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Project Summary – Project II Cardiovascular disease is the leading cause of death worldwide and encompasses multiple disorders including atherosclerosis, hypertension, stroke, and heart failure. There is a direct positive relationship between blood pressure and cardiovascular disease risk in humans, but the factors that contribute to abnormal blood pressure regulation in disease are incompletely understood. Perivascular adipose tissue (PVAT) has recently emerged as a key regulator of vascular tone and blood pressure. PVAT surrounds the majority of blood vessels and releases anticontractile factors. The production of anticontractile mediators is driven by the sympathetic neurotransmitter norepinephrine (NE), but the source of NE that drives this effect remains unknown. The overall goal of this proposal is to understand the mechanisms that mediate neurotransmission in PVAT in health and disease. Potential mechanisms that drive adipocyte activity include local production of mediators by non-neuronal cells, neural innervation, or circulating neurotransmitters. This proposal tests the central hypothesis that autocrine signaling mediated by the uptake and release of NE by adipocytes is the primary regulator of PVAT adipocyte activity, while neural innervation and circulating factors play minor roles. Further, we propose that the presynaptic functions of PVAT adipocytes are altered in conditions that produce vascular disease. This hypothesis will be tested in two specific aims that will study mesenteric and aortic PVAT in male and female animals during health and disease. Experiments will utilize targeted genetic tracing and tissue clearing to identify physical interactions between nerves and adipocytes, genetically encoded calcium indicators to study adipocyte activation, chemogenetics to test the functional relevance of adipocyte activation, fluorescent false neurotransmitters to study the dynamics of NE uptake and release in adipocytes, and RNA sequencing to understand how specific mechanisms and pathways are regulated during disease. Specific Aim 1 will test the hypothesis that the stimulation of PVAT adipocytes by NE involves autocrine signaling mediated by adipocytes rather direct neural innervation or circulating neurotransmitters. Experiments in this aim include anatomical studies in genetic reporter lines, calcium imaging with genetically encoded indicators, chemogenetics in functional assays of vascular tone, and measures of fluorescent false neurotransmitter uptake and release. Specific Aim 2 will test the hypothesis that presynaptic functions of PVAT adipocytes are impaired in conditions that produce vascular disease. This aim will use the high fat diet model of vascular dysfunction and will study candidate mechanisms with RNA sequencing. Effects on adipocyte neurotransmitter uptake, release, and responsiveness will be studied using fluorescent false neurotransmitters and genetically encoded calcium indicators. The results of this study will identify novel mechanisms that regulate the functions of PVAT. New insight into mechanisms that drive the release of anticontractile factors will facilitate the development of therapies for vascular dysfunction in common disorders, such as hypertension.