| Transfection and Activation of CofActor, a Light and Stress Gated Optogenetic Tool, in Primary Hippocampal Neuron Cultures
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Author:
Date:
2021-04-20
[Abstract] Proteins involved in neurodegeneration can be coupled with optogenetic reagents to create rapid and sensitive reporters to provide insight into the biochemical processes that mediate the progression of neurodegenerative disorders, including Alzheimer’s Disease (AD). We have recently developed a novel optically-responsive tool (the ‘CofActor’ system) that couples cofilin and actin (key players in early stage cytoskeletal abnormalities associated with neurodegenerative disorders) with light-gated optogenetic proteins to provide spatial and temporal resolution of oxidative and ...
[摘要] [摘要]参与神经变性蛋白质可具有耦合光遗传学试剂来创建快速且灵敏的记者到provid Ë洞察介导的神经变性疾病，包括进展的生化过程阿尔茨海默氏病（AD）。我们最近开发了一种新型光学-响应工具（“辅”系统）夫妇COF伊林和行动中使用（与神经退行性疾病相关的早期阶段，细胞骨架异常关键球员）光门控光遗传学 蛋白质提供时空分辨率的氧化和高能应激依赖的生化事件。与目前可用的基于小分子的生物传感器来监测细胞氧化还原环境的变化相比，CofActor是一种光激活的，遗传编码的氧化还原传感器，可以通过精确的空间和时间控制来激活。在这里，我们描述了从新生小鼠制备的解离海马神经元培养物中CofActor系统的表达和激活的协议。将培养物转染用大号ipofectamine上的第五天体外（DIV5），然后暴露于细胞应激诱导刺激，导致的肌动蛋白的形成丝切蛋白可使用活细胞成像技术可以观察到杆。本文所述的方案可用于研究暴露于神经退行性刺激（例如毒性Aβ42低聚物）的活神经元中与压力相关的细胞骨架失调。此外，从AD的转基因小鼠模型和/或与KO相关的小鼠KO小鼠分离的神经元中传感器的表达可以促进我们对与神经变性相关的早期细胞骨架功能障碍的分子基础的理解。
[背景]神经变性疾病的生化标志（神经原纤维，团块和缠结，提高活性氧物质（ROS） ...
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| Clonal Culture of Mouse Liver Progenitor Cells
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Author:
Date:
2015-10-20
[Abstract] Liver stem/progenitor cells (LPCs) are defined as bipotential cells differentiating into both hepatocytes and cholangiocytes. For analyzing their differentiation potential, clonal culture has been used for LPCs isolated by a cell sorter. In addition, we can use the culture to assess functions of target genes on differentiation potential of LPCs. This protocol describes the process of cell isolation and colony assay to examine proliferative and differentiation potential of LPCs.
[摘要] 肝干/祖细胞（LPC）被定义为分化成肝细胞和胆管细胞的双能细胞。 为了分析其分化潜力，克隆培养已经用于通过细胞分选仪分离的LPC。 此外，我们可以使用文化来评估目标基因的功能分化潜力的LPCs。 该协议描述细胞分离和集落分析的过程，以检查增殖和分化潜力的LPCs。
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| Neuron Culture from Mouse Superior Cervical Ganglion
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Author:
Date:
2014-01-20
[Abstract] The rodent superior cervical ganglion (SCG) is a useful and readily accessible source of neurons for studying the mechanisms of sympathetic nervous system (SNS) development and growth in vitro. The sympathetic nervous system (SNS) of early postnatal animals undergoes a great deal of remodeling and development; thus, neurons taken from mice at this age are primed to re-grow and establish synaptic connections after in situ removal. The stereotypic location and size of the SCG make it ideal for rapid isolation and dissociation. The protocol described here details the ...
[摘要] 啮齿动物上级子宫颈神经节（SCG）是一种有用且易于获取的神经元来源，用于研究交感神经系统（SNS）体外发育和生长的机制。 早期出生后动物的交感神经系统（SNS）经历了大量的重塑和发育; 因此，在这个年龄的小鼠中取出的神经元被引发再生长并在原位移除后建立突触连接。 SCG的刻板位置和尺寸使其成为快速隔离和解离的理想选择。 这里描述的方案详述了SCG神经元的解剖，培养和分化的要求。 该方案适用于从晚期胚胎妊娠至大约出生后第3天培养神经元。下面讨论的培养技术利用玻璃盖玻片进行固定细胞的显微镜检查。
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