Novel optogenetic tool for noninvasive neuronal inhibition

用于非侵入性神经元抑制的新型光遗传学工具

基本信息

批准号：
10353090
负责人：
Mark Gomelsky
金额：
$ 18.06万
依托单位：
UNIVERSITY OF WYOMING
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：
2022-09-01 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10353090
关键词：
Acids Adenylate Cyclase Amino Acids Anxiety Biochemical Biological Assay Biomedical Engineering Brain Brain region Carbon Dioxide Carboxy-Lyases Cell Therapy Cyclic AMP Cyclic GMP Deinococcus radiodurans Dependovirus Dopamine Engineering Enzymes Epilepsy Future GAD67 enzyme Glutamate Decarboxylase Goals Guanylate Cyclase Histamine Hour Hyperactivity In Vitro Interneurons Length Light Location Mammals Membrane Mental disorders Methodology Modeling Molecular Conformation Monitor Mus Mutagenesis Neurons Neurosciences Neurotransmitters Performance Pharmaceutical Preparations Photoreceptors Protein Engineering Protein Tyrosine Kinase Proteins Research Saccharomyces cerevisiae Schizophrenia Seizures Serotonin Signal Transduction Site Slice Somatosensory Cortex Source Surface Synapses Synaptic Vesicles Testing The Sun Time Variant Virus Visible Radiation Work Yeasts autism spectrum disorder brain tissue cranium design design-build-test designer receptors exclusively activated by designer drugs enzyme activity gamma-Aminobutyric Acid gene therapy high risk high throughput screening in vivo innovation monomer nervous system disorder neuronal excitability novel optogenetics phosphoric diester hydrolase preservation promoter prototype quantum side effect spatiotemporal synthetic biology tool vector

项目摘要

PROJECT SUMMARY/ ABSTRACT This exploratory bioengineering project is aimed at designing, building and testing a light-activated glutamate decarboxylase (GAD) as a novel optogenetic tool for noninvasive inhibition of neuronal activity. GAD produces gamma-aminoburyric acid, GABA, a central neurotransmitter involved in reducing neuronal excitability. Enzy- matic activity of the engineered enzyme will be controlled by light in the near-infrared window (NIRW) of the spectrum (670-900 nm) that penetrates through the skull and brain tissue much better than visible light. We expect the NIRW light-activated GAD (NIRW-GAD) expressed in specific brain regions to be activated via ex- tracorporeal light (e.g., transcranially). NIRW-GAD will represent a unique optogenetic research tool for nonin- vasive, spatiotemporally controlled long-term inhibition of neuronal activity in deep or surface brain regions. In the future, it may be developed into a gene therapy for neurological and psychiatric disorders involving hyper- active brain, such as epilepsy, schizophrenia, anxiety and autism spectrum disorder. The NIRW-GAD protein will be engineered using the homodimeric bacteriophytochrome engineering approach, where bacteriophyto- chromes are a class of photoreceptors that sense light within the NIRW spectrum. In Aim 1, the NIRW-GAD prototypes will be designed and screened for light-inducible GAD activity in yeast. The dynamic range and back- ground activity of the enzyme will be optimized using mutagenesis. In Aim 2, we will examine the NIRW-GAD expression and the effects of NIRW treatment in the somatosensory cortex neurons of mice. We will also quantify the effect of NIRW-GAD on GABAergic synapses and GABA quantal content in cortical neurons in vitro. Upon completion of this project, we expect to have engineered and tested a unique optogenetic tool for noninvasive control of the major inhibitory neurotransmitter in silencing of neuronal activity in mammals. 0

项目概要/摘要这个探索性生物工程项目旨在设计、构建和测试光激活谷氨酸脱羧酶（GAD）作为一种新型光遗传学工具，用于非侵入性抑制神经元活动。 GAD 生产 γ-氨基丁酸，GABA，一种参与降低神经元兴奋性的中枢神经递质。酶- 工程酶的活性将由近红外窗口（NIRW）中的光控制光谱（670-900 nm）比可见光更好地穿透颅骨和脑组织。我们期望在特定大脑区域表达的 NIRW 光激活 GAD (NIRW-GAD) 通过 ex- 体外光（例如，经颅光）。 NIRW-GAD 将代表一种独特的光遗传学研究工具，适用于非对深部或表层大脑区域的神经元活动进行广泛的、时空控制的长期抑制。在未来，它可能会发展成为一种基因疗法，用于治疗涉及过度兴奋的神经和精神疾病。大脑活跃，如癫痫、精神分裂症、焦虑症和自闭症谱系障碍。 NIRW-GAD 蛋白将使用同型二聚体细菌光敏色素工程方法进行工程设计，其中细菌光敏色素铬是一类感光器，可以感知 NIRW 光谱内的光。在目标 1 中，NIRW-GAD 将设计和筛选酵母中光诱导 GAD 活性的原型。动态范围和反向酶的基本活性将通过诱变进行优化。在目标 2 中，我们将研究 NIRW-GAD 小鼠体感皮层神经元的表达和 NIRW 治疗的影响。我们也会量化 NIRW-GAD 对体外皮质神经元 GABA 能突触和 GABA 量子含量的影响。之上该项目完成后，我们希望设计并测试一种独特的非侵入性光遗传学工具控制主要抑制性神经递质沉默哺乳动物的神经元活动。 0