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首页> 外文期刊>Journal of Neuroscience Methods >Quantifying mitochondrial volume density in phrenic motor neurons
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Quantifying mitochondrial volume density in phrenic motor neurons

机译：量化膈电动机神经元的线粒体体积密度

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摘要

Background: Previous assessments of mitochondrial volume density within motor neurons used electron microscopy (EM) to image mitochondria. However, adequate identification and sampling of motor neurons within a particular motor neuron pool is largely precluded using EM. Here, we present an alternative method for determining mitochondrial volume density in identified motor neurons within the phrenic motor neuron (PhMN) pool, with greatly increased sampling. New method: This novel method for assessing mitochondrial volume density in PhMNs uses a combination of intrapleural injection of Alexa 488-conjugated cholera toxin B (CTB) to retrogradely label PhMNs, followed by intrathecal application of MitoTracker Red to label mitochondria. This technique was validated by comparison to 3D EM determination of mitochondrial volume density as a ?gold standard?. Results: A mean mitochondrial volume density of -11 % was observed across PhMNs using the new MitoTracker Red method. This compared favourably with mitochondrial volume density (-11 %) measurements using EM. Comparison with existing method: The range, mean and variance of mitochondrial volume density estimates in PhMNs were not different between EM and fluorescent imaging techniques. Conclusions: Fluorescent imaging may be used to estimate mitochondrial volume density in a large sample of motor neurons, with results similar to EM, although EM did distinguish finer mitochondrion morphology compared to MitoTracker fluorescence. Compared to EM methods, the assessment of a larger sample size and unambiguous identification of motor neurons belonging to a specific motor neuron pool represent major advantages over previous methods.
机译：背景：以前对运动神经元内线粒体体积密度的评估使用电子显微镜（EM）对线粒体进行成像。然而，使用EM在很大程度上无法对特定运动神经元池中的运动神经元进行充分的识别和采样。在这里，我们提出了一种替代方法，用于测定膈运动神经元（PhMN）池中已识别运动神经元的线粒体体积密度，大大增加了采样。新方法：这种评估PhMNs线粒体体积密度的新方法结合胸腔内注射Alexa 488结合霍乱毒素B（CTB）逆行标记PhMNs，然后鞘内应用MitoTracker Red标记线粒体。该技术通过与3D EM测定线粒体体积密度（作为一种？金本位？。结果：采用新的有丝分裂追踪红色法，在PHMN中观察到平均线粒体体积密度为-11%。与现有方法相比：在韧皮部线粒体体积密度估计值的范围、平均值和方差在EM和荧光成像技术之间没有差异。结论：荧光成像可用于估计大样本运动神经元中的线粒体体积密度，其结果与EM相似，但EM确实比MitoTracker荧光识别出更精细的线粒体形态。与EM方法相比，与以前的方法相比，评估更大的样本量和明确识别属于特定运动神经元库的运动神经元具有重大优势。

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