MEASURING OXYGEN CONSUMPTION OF A SINGLE CELL USING A SELF-REFERENCING ELECTRODE

使用自参考电极测量单个细胞的耗氧量

• 批准号: 7953826
• 负责人: PETER JS SMITH
• 金额: $ 3.36万
• 依托单位: MARINE BIOLOGICAL LABORATORY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-12-01 至 2009-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7953826
• 关键词: Aging; Cells; Computer Retrieval of Information on Scientific Projects Database; Consumption; Data; Degenerative Disorder; Development; Electrodes; Funding; Glutamates; Grant; Institution; Measurement; Measures; Membrane Potentials; Metabolic; Metabolism; Neurons; Neurotransmitters; Oxidative Phosphorylation; Oxygen Consumption; Research; Research Personnel; Resources; Rest; Retina; Retinal; Signal Transduction; Source; Techniques; United States National Institutes of Health; analog; cost; horizontal cell; information processing; kainate; response; visual process; visual processing; voltage clamp

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. This study seeks to establish the metabolic cost to an excitable cell associated with neurotransmitter-induced depolarization. Measurements of oxygen consumption from single cells were made using a modulation electrochemical technique. The cell chosen for these studies was the retinal horizontal cell, an important player in the processing of visual signals in the outer retina. Enzymatically isolated cells have a resting potential of approximately -70mV and depolarize in response to the excitatory neurotransmitter glutamate and its analogue, kainate. The depolarization induced by these agents mimics the tonic depolarization of horizontal cells normally observed in the intact retina under dark-adapted conditions. Our technique allows a direct comparison between the metabolic state measured at rest and following tonic depolarization. We found that the average resting O2 consumption for horizontal cells was 0.024 ¿mol.cm-2.sec-1. Application of FCCP (10¿M), an uncoupler of oxidative phosphorylation, elevated oxygen consumption to 0.035 ¿mol.cm2.sec-1; this elevation was followed by a sharp decline to below basal levels after 10min. 100¿M kainate elevated oxygen consumption from a basal average of 0.03 ¿mol.cm-2.sec-1 to 0.07¿mol.cm-2.sec-1. Further, we were able to follow changes in O2 consumption in response to voltage clamp at 0 and -70mV. This data is one of the few examples where the metabolic cost of information processing has been measured from a single identifiable cell. The development of this approach has great promise for furthering our understanding of neuronal metabolism, degenerative diseases and aging.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以出现在其他 CRISP 条目中 列出的机构是。 对于中心来说，它不一定是研究者的机构。 本研究旨在确定与神经递质诱导的去极化相关的可兴奋细胞的代谢成本，使用调制电化学技术测量单个细胞的耗氧量，为这些研究选择的细胞是视网膜水平细胞，它是其中的重要参与者。外视网膜中视觉信号的处理通过酶分离的细胞具有大约-70mV的静息电位，并响应兴奋性神经递质谷氨酸及其类似物而去极化，这些试剂诱导的去极化模拟了在暗适应条件下正常观察到的完整视网膜中的水平细胞的强直去极化，我们发现在休息时和强直去极化技术后测量的代谢状态之间的直接比较。水平细胞的静息 O2 消耗量为 0.024 ¿ mol.cm-2.sec-1。FCCP (10¿M)（一种氧化磷酸化解偶联剂）的应用，将耗氧量提高至 0.035 ¿ mol.cm2.sec-1；100 分钟后急剧下降至基础水平以下。红藻氨酸 M 使基础平均值的耗氧量提高 0.03 ¿ mol.cm-2.sec-1 至 0.07¿此外，我们能够跟踪电压钳位在 0 和 -70mV 时 O2 消耗的变化。 该数据是从单个可识别细胞测量信息处理的代谢成本的少数例子之一。这种方法的发展对于进一步加深我们对神经代谢、退行性疾病和衰老的理解具有很大的希望。