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

• 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
• 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

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，具有分子生物学背景。 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，我在精神病理学的诊断和治疗方面有专业知识，我已经收到了我的MD- 路易斯安那州立大学健康什里夫波特博士学位，并在布朗大学完成了精神病学住院医师。 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患者来源的诱导多能干细胞中，死后脑中的分子破坏 （iPSC）并将其分化为疾病相关细胞类型，以及测量细胞运输和 通过使用荧光标记的蛋白质构建体测定管腔pH。通过掌握这些技能并完成 在这项建议中列出的研究，我将处于有利地位，并有竞争力的独立资金。 研究项目：蛋白质翻译后修饰（PTM）和运输的缺陷报告在 精神分裂症（SZ）的大脑，但根本原因是未知的。细胞器参与PTM的功能， pH值的破坏极大地影响了运输，Na+/H+交换剂（NHE）6-9是运输的主要调节剂。 在癌细胞中，缺氧导致能量代谢改变和NHE 6从细胞器中重新分布。 内体到质膜。在SZ脑中报告了类似的代谢改变，表明 NHE 6 -9细胞内分布也可能受到影响，这可能有助于破坏蛋白PTM和 贩卖人口到目前为止，我发现NHE 7/8在SZ背外侧前额叶皮层的表达减少， （DLPFC），而NHE 6/9不变。尽管如此，NHE 6/9在富含NHE 6/9的组织部分中显示出增加的表达。 突触表明这些蛋白质的分布发生了变化。在这里，我建议更广泛地确定 NHE 6 -9在SZ死后DLPFC和兴奋性皮层神经元中的表达和分布 和从患者来源的iPSC分化的星形胶质细胞。我还将确定如何介绍和 急性应激源（缺氧）的去除影响这些蛋白质在这些细胞中的分布。最后要 用荧光标记的蛋白质构建体检测细胞以测量NHE 6 -9和神经递质受体 运输以及活细胞中的细胞器pH。这些研究可以帮助确定SZ的新治疗靶点 并导致高通量测定以鉴定逆转SZ相关分子破坏的药物。 导师：本提案的主要导师团队由James Meador-Woodruff博士组成， 著名的精神分裂症大脑分子破坏和死后脑组织分析专家， Marek Napierala是重复扩展障碍分子机制和建模的专家， 利用iPSC分化成包括皮层神经元在内的多种细胞类型的疾病，弗拉基米尔博士 Parpura是神经胶质生物学和通过使用 荧光标记的蛋白质构建体。