Creation of Optical Biosensor Mice for Longitudinal Studies of Vascular Function

用于血管功能纵向研究的光学生物传感器小鼠的创建

基本信息

批准号：
9242698
负责人：
MEGAN A RIZZO
金额：
$ 38.59万
依托单位：
UNIVERSITY OF MARYLAND BALTIMORE
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-04-01 至 2020-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9242698
关键词：
Adrenergic Receptor Angiotensin II Angiotensin II Receptor Animals Arteries Autonomic ganglion Biosensor Blood Pressure Blood Vessels Blood flow Calibration Calmodulin Cardiovascular system Cells Characteristics Chronic Color Conscious Coupled DNA cassette Data Development Diet Disease Ear Endocrine Endothelin Endothelin A Receptor Enzymes Event Fluorescence Resonance Energy Transfer G-Protein-Coupled Receptors Goals Hexamethonium Hypertension Image Implant Individual Injection of therapeutic agent Knowledge Lead Ligands Longitudinal Studies Measurement Measures Methods Modeling Molecular Monomeric GTP-Binding Proteins Mus Muscle Cells Muscle Contraction Myosin ATPase Myosin Light Chain Kinase Myosin Regulatory Light Chains Nerve Neurons Nonmuscle Myosin Type IIA Operative Surgical Procedures Optics Pathway interactions Pharmacology Phosphorylation Physiological Pressure Transducers Principal Investigator Procedures Process Reagent Recovery Regulation Reporting Research Rho-associated kinase Role Rosa Signal Transduction Smooth Muscle Smooth Muscle Myocytes Smooth Muscle Myosins Sodium Chloride Sphingosine-1-Phosphate Receptor Tamoxifen Techniques Telemetry Testing Time Tissues Transgenic Mice Transgenic Organisms Vascular Smooth Muscle Vascular remodeling Vascular resistance arteriole base constriction experimental study fluorescence imaging genetic regulatory protein human disease in vivo insight intravital imaging molecular imaging myosin phosphatase novel novel strategies public health relevance quantitative imaging salt sensitive salt sensitive hypertension sensor sphingosine 1-phosphate

项目摘要

DESCRIPTION (provided by applicant): Hypertension involves elevated vascular resistance and only exists in a living animal, where the physiologic regulators of vascular tone (i.e. neuronal activity, blood flow, and endocrine factors) are intact. Therefore, hypertension research will greatly benefit from the in vivo exploration of the molecular regulators of arterial smooth muscle cell contraction. Our overall goal is to elucidate mechanisms of increased vascular resistance, for the first time, in conscious animals, during experimental salt-dependent hypertension. The use of conscious animals (as opposed to anesthetized) is key to understanding the putative role of sympathetic nerve activity (SNA), increasingly recognized as a key mechanism of salt-dependent hypertension. This overall goal is to be achieved through the use of non-invasive, fluorescence imaging of molecular signaling in arterioles of conscious optical biosensor mice. Given that phosphorylation of myosin regulatory light chains is the critical determinant of smooth muscle contraction, Specific Aims 1 and 2 are to develop optical biosensor mice that express novel, genetically-encoded activity biosensors for the key molecular regulators of smooth muscle myosin phosphorylation; a) myosin light chain kinase, (MLCK), b) myosin light chain phosphatase, MLCP), and c) a key upstream regulator of MLCP, the small GTPase, RhoA. In Aim 3, we will use these optical biosensor mice to determine, in vivo, 1) the regulation of MLCK, MLCP and RhoA by certain vascular G-protein coupled receptors (GPCR) putatively involved in hypertension, including adrenoceptors (α1-AR), Angiotensin II receptors (AT1-R), endothelin-1 receptors, (ETA), and sphingosine-1-phosphate receptors (S1P1). The regulation of MLCK, MLCP and RhoA by SNA will be determined through the use of complete autonomic ganglionic blockade (hexamethonium) in conscious mice. Mice will be implanted with telemetric arterial blood pressure transducers to allow continuous measurement of arterial blood pressure during imaging (and all other times). In Specific Aim 4, the time course of the activation levels of MLCK, MLCP and RhoA will be measured in ear arterioles of conscious individual mice (i.e. a `longitudinal' study) during 14 days of Angiotensin II/salt hypertension. Mice are infused chronically with Angiotensin II and fed a high-salt (NaCl) diet. Increased vascular resistance in this model of salt-dependent hypertension is believed to involve heightened SNA (`sympathoexcitation) emanating from salt-sensitive CNS cardiovascular control regions, mandating use of conscious mice. These Specific Aims will be performed in this multi-PI project under the direction of two Principal Investigators with the necessary expertise to generate the proposed sensors (Rizzo) and perform the physiologic experiments (Wier). In summary, we expect to achieve dynamic imaging of myosin phosphorylation regulatory processes during the development of salt-sensitive hypertension in a living animal for the first time. These studies will reveal new insights on the molecular basis of increased vascular resistance in hypertension, as it actually occurs in living animals.

描述（由申请人证明）：血管抑制在活性动物中，即神经元活动，血液流量和内分泌因子是完整的在有意识的动物中，在实验性盐 - 盐盐中，抗药性是与盐的盐分相反的在摩西肌肉收缩的施工小鼠的动脉中，特定的目标是开发光学生物传感器的小鼠，这些小鼠为平滑肌迈罗迈罗迈罗甲磷酸化的关键分子调节剂MLCP的关键分子调节剂，MLCP的ream ream ream ream ream ream ream ream ream ，我们将使用这些光学生物传感器小鼠来确定具有高血压，包括肾上腺受体（α1-AR），血管紧张素II受体（AT1-R），entother-1受体，（ETA）和鞘氨酸-1-磷酸受体（ S1P1）。。 II/盐高血压是II，并喂养高盐（NaCl）饮食。这些特定的目标将在此多PI WO主要研究人员中具有必要的专业知识，以产生支撑性的传感器（Rizzo）（Rizzo）并进行生理实验（否）。这些研究首次对活动物的盐敏感。