Investigations of cAMP-dependent brain-barrier permeability in choroid plexus

脉络丛 cAMP 依赖性脑屏障通透性的研究

基本信息

批准号：
10753098
负责人：
Dario Xavier Figueroa Velez
金额：
$ 7.41万
依托单位：
BOSTON CHILDREN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-01 至 2025-08-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10753098
关键词：
Adult Agonist Apical Biochemical Biological Assay Biology Blood Blood - brain barrier anatomy Blood Circulation Blood-Borne Pathogens Boston Brain Brain Diseases Brain Hypoxia-Ischemia Calcium Cell Nucleus Cells Central Nervous System Cerebral Ventricles Cerebrospinal Fluid Choroid Plexus Epithelium Collaborations Coupled Cyclic AMP Cyclic AMP Receptors Cyclic AMP-Dependent Protein Kinases Data Development Dextrans Disease Electron Microscopy Endothelial Cells Endothelium Environment Epilepsy Epithelial Cells Exposure to Fellowship Fluorescent Dyes G-Protein-Coupled Receptors Glutamates Goals Health Image Imaging Techniques Imaging technology In Vitro Individual Intravenous Investigation Laboratories Ligands Link Maps Measures Mediating Mental Depression Mental disorders Mentors Methods Molecular Molecular Weight Mus Neuroglia Neurologic Neurotransmitters Pediatric Hospitals Peripheral Peripheral Nervous System Permeability Play Population Predisposition Preparation Process Production Property Protocols documentation Publications Regulation Reporter Research Resolution Role Schizophrenia Signal Transduction Source Stains Structure of choroid plexus Surface Techniques Testing Therapeutic Intervention Tight Junctions Tissues Training Traumatic Brain Injury Vascular Endothelial Cell Vascularization Visualization apical membrane awake bench to bedside blood cerebrospinal fluid barrier brain tissue career cell type experimental study imaging modality improved in vivo in vivo imaging in vivo optical imaging in vivo two-photon imaging intravenous injection lateral ventricle metabotropic glutamate receptor 8 nervous system disorder neuroprotection novel strategies receptor selective expression sensor tool two-photon

项目摘要

PROJECT SUMMARY The choroid plexus (ChP) comprises a network of cells that form a critical brain barrier that can mediate secondary damage in certain brain disorders and trauma. The Lehtinen lab has developed a suite of tools to study the ChP across development ex vivo and in vivo. This project applies imaging technology to study blood- CSF barrier permeability regulation at the cellular level. Our overarching hypothesis is that the intracellular messenger cAMP regulates endothelial tight junctions between ChP epithelial cells and thereby blood-CSF barrier integrity as it does in the nearby blood-brain barrier (BBB). The main effectors of cAMP, PKA and Epac, regulate endothelial tight junction redistribution and barrier permeability. Gi/o-linked G-Protein Coupled Receptors (GPCRs) are strong upstream regulators of cAMP. The Lehtinen laboratory's single nucleus sequencing data suggest that ChP epithelial cells selectively and developmentally express mGlur8, which via Gαi, activation in other brain tissues inhibits cAMP production. To elucidate mechanisms of neurotransmitter alteration of the blood-CSF barrier we will study the cellular mechanisms of neurotransmitter-activated GPCR- cAMP signaling in ChP epithelial cells. The Lehtinen lab, in collaboration with co-sponsor Mark Andermann's lab, recently established a protocol for ex vivo and in vivo imaging of ChP structure and function based on using fluorescent reporters of calcium activity. I will apply these techniques to reveal how GPCRs modulate cAMP levels using fluorescent cAMP indicators in ChP explants (Aim 1). In Aim 2 we will use an in vivo preparation to map the populations of receptors that are accessible to central vs peripheral ligands. Those located on the apical membrane are in contact with the CSF and those on the basal surface are exposed to the blood. With functional assays including peripheral delivery of low molecular weight fluorescent dyes, we will assess the effects of central (CSF) vs peripheral (intravenous) delivered ligands such as mGluR8 agonists on cAMP and blood-CSF barrier permeability. Together these studies will reveal mechanisms how neurotransmitters, specifically glutamate, may contribute to blood-CSF barrier integrity in health and disease. The research and training proposed will take place at Boston Children's Hospital, a world-renowned pediatric hospital that offers an exceptional research environment and countless opportunities to carry research from bench to bedside. Importantly, the research proposed will take place under the guidance of Dr. Maria Lehtinen, an expert in the field of choroid plexus and CSF biology. In addition, Dr. Mark Andermann (co-mentor) is a leader in in vivo optical imaging techniques. The results from this proposal will result in first-authored publications and a wealth of preliminary data for a competitive K99/R00 application. This fellowship will provide the candidate with the opportunity to begin training in choroid plexus biology and make significant scientific contributions while also helping launch an independent academic career.

项目摘要脉络丛（CHP）包括一个形成临界脑屏障的细胞网络，可以介导某些脑部疾病和创伤的次要损害。 Lehtinen Lab开发了一套工具研究跨体内和体内发育的CHP。该项目应用成像技术研究血液 - CSF屏障渗透率调节在细胞水平上。我们的总体假设是细胞内 Messenger Camp调节CHP上皮细胞之间的内皮紧密连接，从而使血液CSF 在附近的血脑屏障（BBB）中，屏障完整性如它所做的。营地，PKA和EPAC的主要影响，调节内皮紧密连接重新分布和屏障渗透性。 GI/O连接的G蛋白耦合受体（GPCR）是CAMP的强大上游调节器。 Lehtinen实验室的单核测序数据表明CHP上皮细胞有选择地和开发表达MGLUR8，通过 GαI，其他脑组织中的激活抑制了cAMP的产生。阐明神经递质的机制血液-CSF屏障的改变我们将研究神经递质激活GPCR-的细胞机制 CHP上皮细胞中的cAMP信号传导。 Lehtinen Lab与共同赞助商Mark Andermann合作实验室，最近建立了基于体内的体内和体内成像的协议，基于CHP结构和功能使用钙活性的荧光记者。我将应用这些技术来揭示GPCR如何调节使用CHP外植体中的荧光营地指标（AIM 1）的cAMP水平。在AIM 2中，我们将使用一个体内准备绘制中央与外围配体可访问的接收器种群。那些位于顶部膜上的CSF和基本表面上的膜接触血。通过功能分析，包括低分子量荧光染料的外围递送，我们将评估中央（CSF）与外围（静脉内）递送的配体（例如MGLUR8激动剂）对营地和血液-CSF屏障渗透性。这些研究将共同揭示机制神经递质，特别是谷氨酸，可能有助于健康和疾病中的血液-CSF屏障完整性。提议的研究和培训将在波士顿儿童医院举行，这是一个世界知名的儿科提供了卓越的研究环境和无数机会进行研究的医院长凳到床边。重要的是，提出的研究将在Maria Lehtinen博士的指导下进行。脉络丛和CSF生物学领域的专家。此外，马克·安德曼（Mark Andermann）博士（Co-Insor）是体内光学成像技术的领导者。该提案的结果将导致首次实现竞争性K99/R00应用程序的出版物和大量的初步数据。这个奖学金将提供有机会开始接受脉络丛生物学培训并进行大量科学的候选人贡献同时也有助于开展独立的学术职业。