The nematode Caenorhabditis elegans has served as one of the primary model organisms in neuroscience. As C. elegans research became more specific, so have the biological tools for manipulating C. elegans improved and matured. Additionally, in some avenues of research, technologies have been developed to manipulate the animals in very efficient and quantitative ways. However, the field of dynamic studies has remained without significant technological support. Dynamic studies focus on processes occurring over time and span a range of time-scales of i) minutes to hours requiring continuous imaging for accurate observation, ii) hours to days requiring periodic imaging of the same animal, and iii) days to weeks requiring daily monitoring. Because of a lack of suitable tools and technologies to perform these studies, researchers have to either apply standard biological methods with limited ability to observe processes dynamically or simply cannot perform such studies with the desired set of experimental conditions.
To address this problem, a comprehensive microfluidic toolkit for dynamic studies has been created. The first element is a novel method for reversible and repeatable immobilization at benign conditions in tandem with a microfluidic system for isolated culture of C. elegans with integrated temperature control. The second element is a system for efficient handling of C. elegans embryos in a high-throughput and scalable fashion for chemical and thermal embryonic stimulation with subsequent study of development. The third component is a system capable of selective immobilization of animals’ bodies, while simultaneously facilitating feeding and normal physiological function for live imaging. The last component is capable of culturing animals over their life-span with efficient animal handling, environmental control (temperature and dietary conditions), and high data content experimentation.
As a whole, the work in this thesis enables dynamic studies over the whole range of time scales applicable to C. elegans research. These types of studies were previously very difficult or near impossible to perform practically. Now, instead of building population composites to understand the dynamics of a process, risking affecting physiology via the experiment itself, or dealing with extremely labor intensive physical handling of animals, a toolkit for efficient handling of C. elegans facilitating dynamic and direct observation of individual animals is available. The biological applications range from dynamically studying lipid droplet morphology or studying synaptic vesicle transport, through observing the dynamics of synaptic re-arrangement during development or the effect of cancer drugs on development, to performing high-content life-span experiments able to ascertain the relationship between aging and behavior. Additionally, many of the principles in these designs can be expanded to accommodate research on other model organisms, such as other nematode species, zebra fish embryos, or cells and embryoid bodies.
| Identifer | oai:union.ndltd.org:GATECH/oai:smartech.gatech.edu:1853/49115 |
| Date | 20 September 2013 |
| Creators | Krajniak, Jan |
| Contributors | Lu, Hang |
| Publisher | Georgia Institute of Technology |
| Source Sets | Georgia Tech Electronic Thesis and Dissertation Archive |
| Language | en_US |
| Detected Language | English |
| Type | Dissertation |
| Format | application/pdf |
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