Organellar Na+/H+ Exchangers and Intracellular pH Regulation in Schizophrenia Brain

精神分裂症脑细胞器 Na /H 交换器和细胞内 pH 调节

基本信息

批准号：
10559700
负责人：
Brandon Scott Pruett
金额：
$ 19.14万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-02-01 至 2026-12-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10559700
关键词：
AMPA Receptors Acidity Acute Affect Astrocytes Autopsy Biology Brain Calibration Cell Culture Techniques Cell membrane Cells Closure by clamp Co-Immunoprecipitations Confocal Microscopy Development Diagnosis Disease Doctor of Philosophy Endosomes Energy Metabolism Enrollment Enzymes Excision Fluorescence Funding Future Glycolysis Goals Health Hypoxia Individual Knowledge Learning Measures Mediating Mentorship Metabolic Methods Modeling Molecular Neurons Neurotransmitter Receptor Organelles PHluorin Patients Peripheral Pharmaceutical Preparations Positioning Attribute Post-Translational Protein Processing Pre-Clinical Model Prefrontal Cortex Proteins Psychiatry Psychopathology Regulation Reporting Research Personnel Research Project Grants Residencies Resolution Role Scaffolding Protein Schizophrenia Sodium-Hydrogen Antiporter Stress Subcellular Fractions Synapses Techniques Testing Time Tissues Training Transfection Universities Visualization Work brain tissue cancer cell career cell type cost economic cost experience experimental study first episode schizophrenia high throughput screening induced pluripotent stem cell mortality multimodality neuroimaging new therapeutic target novel novel therapeutics professor programs protein distribution protein transport ratiometric receptor response severe mental illness skill acquisition stem cell differentiation stressor trafficking

项目摘要

Candidate: I am an Assistant Professor in UAB’s Department of Psychiatry with a background in molecular methods used in preclinical models and the role of intracellular pH dysregulation in neurodevelopmental illnesses. Additionally, I have expertise in diagnosis and treatment of psychopathology having received my MD- PhD from LSU Health Shreveport and completed a Psyhciatry Residency at Brown University. Career Goals and Development: I hope to gain expertise in assessing schizophrenia (SZ)-associated molecular disruptions in postmortem brain, in generating SZ patient-derived induced pluripotent stem cells (iPSCs) and differentiating them into disease relevant cell types, and in measuring cellular trafficking and luminal pH through the use of fluorescently-tagged protein constructs. By acquiring these skills and completing the studies laid out in this proposal, I will be well positioned and competitive for independent funding. Research Project: Deficits in protein post-translational modifications (PTM) and trafficking are reported in schizophrenia (SZ) brain, but the underlying cause is unknown. The function of organelles involved in PTM and trafficking is greatly impacted by pH disruptions, and Na+/H+ Exchangers (NHE) 6-9 are major regulators of organelle pH. In cancer cells, hypoxia causes altered energy metabolism and redistribution of NHE6 from endosomes to the plasma membrane. Similar metabolic alterations are reported in SZ brain suggesting that NHE6-9 intracellular distribution may also be affected, which could contribute to disrupted protein PTM and trafficking. So far, I have found that NHE7/8 expression is decreased in SZ dorsolateral prefrontal cortex (DLPFC) while NHE6/9 is unchanged. Still, NHE6/9 show increased expression in a tissue fraction enriched for synapses suggesting altered distribution of these proteins. Here, I propose to more extensively determine the expression and distribution of NHE6-9 first in SZ postmortem DLPFC and then in excitatory cortical neurons and astrocytes differentiated from patient-derived iPSCs. I will also determine how the introduction and removal of an acute stressor (hypoxia) affects the distribution of these proteins in these cells. Finally, I will transfect cells with fluorescently-tagged protein constructs to measure NHE6-9 and neurotransmitter receptor trafficking as well as organelle pH in live cells. These studies could help identify novel treatment targets for SZ and lead to high throughput assays to identify drugs that reverse SZ-associated molecular disruptions. Mentorship: The primary mentorship team for this proposal consists of Dr. James Meador-Woodruff, a world renowned expert in molecular disruptions in schizophrenia brain and analysis of postmortem brain tissue, Dr. Marek Napierala, an expert on molecular mechanisms of repeat expansion disorders and of modeling these illnesses using iPSCs differentiated into a variety of cell types including cortical neurons, and Dr. Vladimir Parpura, an expert in glial biology and visualization of vesicular trafficking in live cells through the use of fluorescently-tagged protein constructs.

候选人：我是UAB精神病学系的助理教授，具有分子的背景临床前模型中使用的方法以及细胞内pH失调在神经发育中的作用疾病。此外，我在诊断和治疗心理病理学方面具有专业知识，因为我接受了我的MD- 来自LSU Health Shreveport的博士学位，并在布朗大学完成了一家院子里的住院医师。职业目标和发展：我希望获得评估精神分裂症（SZ）相关的专业知识在产生SZ患者衍生的多能干细胞时，死后大脑中的分子破坏（IPSC）并将它们区分为相关的细胞类型，并在测量细胞运输和通过使用荧光标记的蛋白质构建体的腔pH。通过获得这些技能并完成在这项提案中阐述的研究中，我将在独立资金方面拥有良好的位置和竞争力。研究项目：蛋白质后翻译后修饰（PTM）和贩运的缺陷在精神分裂症（SZ）大脑，但根本原因尚不清楚。涉及PTM和细胞器的功能贩运受到pH中断的影响很大，NA+/H+交换器（NHE）6-9是主要的监管机构细胞器ph。在癌细胞中，缺氧会改变能量代谢和NHE6的重新分布质膜内体。 SZ大脑中有类似的代谢改变表明 NHE6-9细胞内分布也可能受到影响，这可能导致蛋白质PTM的破坏和贩运。到目前为止，我发现SZ背侧前额叶皮层的NHE7/8表达降低（DLPFC）而NHE6/9不变。尽管如此，NHE6/9仍显示出富含组织的组织分数的表达突触表明这些蛋白质分布改变。在这里，我建议更广泛地确定 NHE6-9在SZ后DLPFC中首先表达和分布，然后在兴奋性皮质神经元中表达和分布和星形胶质细胞与患者衍生的IPSC不同。我还将确定介绍和去除急性应激源（缺氧）会影响这些细胞中这些蛋白质的分布。最后，我会的用荧光标记的蛋白质构建体转染细胞，以测量NHE6-9和神经递质接收器活细胞中的运输和细胞器pH。这些研究可以帮助确定SZ的新型治疗靶并导致高吞吐量的大师识别逆转SZ相关的分子破坏的药物。指导：该提案的主要精力团队由詹姆斯·米多尔（James Meador-Woodruff）博士组成精神分裂症大脑中著名的分子破坏专家和死后脑组织的分析，博士 Marek Napierala，重复扩张障碍分子机制的专家，并建模这些使用IPSC的疾病分为各种细胞类型，包括皮质神经元和弗拉基米尔博士 Parpura，神经胶质生物学专家和通过使用的活细胞中囊泡运输的可视化荧光标记的蛋白质构建体。