Department of Pharmacology and Cancer Biology, Duke University, Durham, NC, United States; Integrated Toxicology and Environmental Health Program, Duke University, Durham, NC, United States.
Integrated Toxicology and Environmental Health Program, Duke University, Durham, NC, United States; Nicholas School of the Environment, Duke University, Durham, NC, United States.
Neurotoxicology. 2019 Sep;74:209-220. doi: 10.1016/j.neuro.2019.07.002. Epub 2019 Jul 16.
In order to develop a better understanding of the role environmental toxicants may play in the onset and progression of neurodegenerative diseases, it has become increasingly important to optimize sensitive methods for quickly screening toxicants to determine their ability to disrupt neuronal function. The nematode Caenorhabditis elegans can help with this effort. This species has an integrated nervous system producing behavioral function, provides easy access for molecular studies, has a rapid lifespan, and is an inexpensive model. This study focuses on methods of measuring neurodegeneration involving the dopaminergic system and the identification of compounds with actions that disrupt dopamine function in the model organism C. elegans. Several dopamine-mediated locomotory behaviors, Area Exploration, Body Bends, and Reversals, as well as Swimming-Induced Paralysis and Learned 2-Nonanone Avoidance, were compared to determine the best behavioral method for screening purposes. These behavioral endpoints were also compared to morphological scoring of neurodegeneration in the dopamine neurons. We found that in adult worms, Area Exploration is more advantageous than the other behavioral methods for identifying DA-deficient locomotion and is comparable to neuromorphological scoring outputs. For larval stage worms, locomotion was an unreliable endpoint, and neuronal scoring appeared to be the best method. We compared the wild-type N2 strain to the commonly used dat-1p::GFP reporter strains BY200 and BZ555, and we further characterized the dopamine-deficient strains, cat-2 e1112 and cat-2 n4547. In contrast to published results, we found that the cat-2 strains slowed on food almost as much as N2s. Both showed decreased levels of cat-2 mRNA and DA content, rather than none, with cat-2 e1112 having the greatest reduction in DA content in comparison to N2. Finally, we compared and contrasted strengths, limitations, cost, and equipment needs for all primary methods for analysis of the dopamine system in C. elegans.
为了更好地了解环境毒物在神经退行性疾病的发生和进展中可能发挥的作用，优化快速筛选毒物的敏感方法以确定其破坏神经元功能的能力变得越来越重要。秀丽隐杆线虫可以为此做出贡献。这种物种具有完整的神经系统，产生行为功能，便于进行分子研究，寿命短，而且是一种廉价的模型。本研究重点介绍了测量涉及多巴胺能系统的神经退行性变的方法，并鉴定了在模型生物秀丽隐杆线虫中具有破坏多巴胺功能的化合物。几种多巴胺介导的运动行为，包括区域探索、身体弯曲和反转，以及游泳诱导的麻痹和习得的 2-壬酮回避，被比较以确定用于筛选目的的最佳行为方法。这些行为终点也与多巴胺能神经元的神经形态评分进行了比较。我们发现，在成年蠕虫中，区域探索比其他行为方法更有利于识别 DA 缺乏运动，并且与神经形态评分输出相当。对于幼虫阶段的蠕虫，运动是一个不可靠的终点，神经元评分似乎是最好的方法。我们将野生型 N2 菌株与常用的 dat-1p::GFP 报告菌株 BY200 和 BZ555 进行了比较，并且进一步对多巴胺缺乏菌株 cat-2 e1112 和 cat-2 n4547 进行了特征描述。与已发表的结果相反，我们发现 cat-2 菌株在食物上的速度几乎与 N2 一样慢。两者都显示出 cat-2 mRNA 和 DA 含量的降低，而不是没有，与 N2 相比，cat-2 e1112 的 DA 含量降低最大。最后，我们比较和对比了所有分析秀丽隐杆线虫中多巴胺系统的主要方法的优缺点、成本和设备需求。
