Department of Biomedical Engineering, University of California Irvine, Irvine, CA, USA.
Laboratory of Fluorescence Dynamics, Department of Biomedical Engineering, University of California Irvine, Irvine, CA, USA.
Geroscience. 2019 Feb;41(1):51-67. doi: 10.1007/s11357-019-00052-8. Epub 2019 Feb 6.
Nicotinamide adenine dinucleotide (reduced form: NADH) serves as a vital redox-energy currency for reduction-oxidation homeostasis and fulfilling energetic demands. While NADH exists as free and bound forms, only free NADH is utilized for complex I to power oxidative phosphorylation, especially important in neurons. Here, we studied how much free NADH remains available for energy production in mitochondria of old living neurons. We hypothesize that free NADH in neurons from old mice is lower than the levels in young mice and even lower in neurons from the 3xTg-AD Alzheimer's disease (AD) mouse model. To assess free NADH, we used lifetime imaging of NADH autofluorescence with 2-photon excitation to be able to resolve the pool of NADH in mitochondria, cytoplasm, and nuclei. Primary neurons from old mice were characterized by a lower free/bound NADH ratio than young neurons from both non-transgenic (NTg) and more so in 3xTg-AD mice. Mitochondrial compartments maintained 26 to 41% more reducing NADH redox state than cytoplasm for each age, genotype, and sex. Aging diminished the mitochondrial free NADH concentration in NTg neurons by 43% and in 3xTg-AD by 50%. The lower free NADH with age suggests a decline in capacity to regenerate free NADH for energetic supply to power oxidative phosphorylation which further worsens in AD. Applying this non-invasive approach, we showed the most explicit measures yet of bioenergetic deficits in free NADH with aging at the subcellular level in live neurons from in-bred mice and an AD model.
烟酰胺腺嘌呤二核苷酸（还原形式：NADH）作为氧化还原平衡和满足能量需求的重要氧化还原能量货币。虽然 NADH 以游离和结合形式存在，但只有游离的 NADH 可用于 I 复合物为氧化磷酸化提供动力，这在神经元中尤为重要。在这里，我们研究了衰老的活神经元线粒体中可用作能量产生的游离 NADH 有多少。我们假设老年小鼠神经元中的游离 NADH 低于年轻小鼠的水平，甚至低于阿尔茨海默病（AD）3xTg-AD 小鼠模型的神经元中的水平。为了评估游离 NADH，我们使用 2 光子激发的 NADH 自发荧光的终生成像来分辨线粒体、细胞质和细胞核中 NADH 的池。与非转基因（NTg）小鼠相比，老年小鼠的原代神经元具有更低的游离/结合 NADH 比值，而 3xTg-AD 小鼠的比值则更低。对于每个年龄、基因型和性别，线粒体区室比细胞质保持 26%至 41%更高的还原态 NADH 氧化还原状态。衰老使 NTg 神经元中的游离 NADH 浓度降低了 43%，3xTg-AD 神经元降低了 50%。随着年龄的增长，游离 NADH 减少表明再生游离 NADH 的能力下降，无法为氧化磷酸化提供能量，而 AD 则使这种情况进一步恶化。通过应用这种非侵入性方法，我们在近交系小鼠和 AD 模型的活神经元中，在亚细胞水平上显示了迄今为止最明确的衰老时游离 NADH 生物能量缺陷的措施。
