Determination of true Glia-Neuron Ratios: Validation of the Isotropic Fractionato

真实胶质细胞-神经元比率的测定：各向同性分数的验证

基本信息

批准号：
8725753
负责人：
CHRISTOPHER S VON BARTHELD
金额：
$ 7.1万
依托单位：
UNIVERSITY OF NEVADA RENO
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-01 至 2017-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8725753
关键词：
Animals Antibodies Astrocytes Autistic Disorder Basic Science Belief Biological Models Bipolar Depression Bipolar Disorder Brain Brain region Calibration Cell Count Cell Density Cell Nucleus Cells Chickens Clinical Research Conflict (Psychology)Confusion Corpus Callosum DNA Data Disease Disputes Endothelial Cells Frequencies Gold Health Histology Human Journals Knowledge Label Language Lead Measures Mental Depression Mental disorders Methods Microglia Neuroglia Neurons Neurosciences Oligodendroglia Optic Nerve Optics Paper Patients Ploidies Publishing Recovery Reporting Sampling Schizophrenia Schools Science Series Solutions Structure Techniques Textbooks Ursidae Family Validation Work base cell type density design evidence base frontal lobe gray matter interest morphometry nervous system disorder nonhuman primate novel novel strategies particle public health relevance tool white matter

项目摘要

DESCRIPTION (provided by applicant): Major neurological and psychiatric diseases have been reported to be associated with significantly changed glia densities, but considerable confusion surrounds the question of the true numbers and ratios of neurons and glial cells in the brains of humans and animals. Traditionally and per "textbook knowledge," it was thought that glial cells outnumber neurons by a ratio of between 10:1 and 50:1. Recently introduced new counting techniques have challenged this belief and postulate a ratio of approximately 1:1, reducing the number of glial cells in the human brain from previous estimates by as much as 4.9 trillion (from 5 trillion to a mere 100 billion). The new, much lower estimates of glial cells are primarily based on a novel methodological approach, the isotropic fractionator (IF). However, this technique has not been calibrated or validated. It is therefore uncertain whether it produces true numbers or may possibly be biased towards larger particles (neuronal nuclei) and may underestimate smaller ones (glial nuclei). The fractionator technique homogenizes whole brains (or dissected parts thereof), then takes samples of the cell nuclei in solution, and uses antibody labeling to distinguish neuronal nuclei from other nuclei such as glia in sample solutions. While conceptually elegant, it remains to be calibrated against known numbers and ratios of neurons and glial cells. Therefore, we propose a series of calibrations that will unambiguously determine the presence or absence of any biases in the isotropic fractionator (IF) technique. We will probe a variety of human and non-human primate brain regions (primarily white matter tracts) to determine whether the fractionator accurately estimates numbers of glial cells. The same brain regions will be examined by measuring DNA content (which can directly reflect cell numbers), and by histology, using systematic random sampling to independently estimate glial numbers and ratios. Preliminary data indicate that the histological approach is most suitable to provide a gold standard for comparison. Modern stereological sampling techniques will identify the ratios of cell types at the ultrastructural level. This work will clarify true neuron-glia ratios in the bain and other parts of the CNS. Validation of the new fractionator method will be crucial for understanding neurological and psychiatric diseases with reported imbalances of glia cell densities, such as schizophrenia, autism, bipolar disease, and depression.

描述（由申请人提供）：据报道，主要的神经和精神疾病与神经胶质细胞密度的显着变化有关，但围绕人类和动物大脑中神经元和神经胶质细胞的真实数量和比例的问题存在相当大的混乱。传统上，根据“教科书知识”，人们认为神经胶质细胞的数量多于神经元，其比例在 10:1 到 50:1 之间。最近推出的新计数技术挑战了这一信念，并假设比例约为 1:1，将人脑中神经胶质细胞的数量比之前的估计减少了多达 4.9 万亿个（从 5 万亿减少到仅仅 1000 亿）。对神经胶质细胞的新的、低得多的估计主要基于一种新的方法，即各向同性分馏器 (IF)。然而，该技术尚未经过校准或验证。因此，不确定它是否产生真实的数字，或者可能偏向较大的颗粒（神经元核）并可能低估较小的颗粒（神经胶质细胞核）。分级分离器技术将整个大脑（或其解剖部分）均质化，然后采集溶液中的细胞核样本，并使用抗体标记将神经元核与样本溶液中的其他核（例如神经胶质细胞）区分开。虽然概念上很优雅，但它仍然需要根据已知的神经元和神经胶质细胞的数量和比例进行校准。因此，我们提出了一系列校准，以明确确定各向同性分馏器 (IF) 技术中是否存在任何偏差。我们将探测各种人类和非人类灵长类动物的大脑区域（主要是白质束），以确定分馏器是否准确估计神经胶质细胞的数量。通过测量 DNA 含量（可以直接反映细胞数量）和组织学，使用系统随机抽样独立估计神经胶质细胞数量和比率，对相同的大脑区域进行检查。初步数据表明组织学方法最适合提供比较的金标准。现代体视取样技术将在超微结构水平上识别细胞类型的比例。这项工作将阐明贝恩和中枢神经系统其他部分的真实神经元-胶质细胞比率。新分馏器方法的验证对于了解神经和精神疾病（据报道神经胶质细胞密度不平衡）至关重要，例如精神分裂症、自闭症、双相情感障碍和抑郁症。