Propagation of mechanical stimuli to the touch receptor neurons of Caenorhabditis elegans

The sense of touch is crucial for the survival and thriving of humans and other animals. Unfortunately, touch sensation can be degraded by cancer treatment, aging, and diseases such as diabetes. Compared to vision, smell, and taste, we know little about the process of transducing a mechanical stimulus into a signal in a neuron. Part of the reason we know less about mechanical senses is the lack of tools for studying the complex composite of touch neurons and skin that humans and other mammals use to sense touch. To address this problem, researchers use model organisms such as Caenorhabditis elegans, which is a small roundworm with a compact nervous system. C. elegans has six gentle touch receptor neurons (TRNs) that develop in stereotypical locations due to the deterministic cellular development of these worms. Combined with a well-developed genetic toolbox, this makes C. elegans ideal for studying the molecular mechanisms of touch sensation, but introduces the challenges of manipulating and applying controlled forces to an animal that is roughly one millimeter long and 50 micrometers in diameter. This thesis presents my doctoral research which endeavors to contribute to research on C. elegans touch sensation by characterizing existing tools, developing new tools, and using these tools to generate new insights into the process of transducing an external mechanical stimulus into the activation of a C. elegans TRN. I begin with an overview of our current knowledge of C. elegans touch sensation and the tools used to obtain that knowledge. Then, I present our research on the forces applied during the most common touch assay for C. elegans, the eyebrow hair touch test. By measuring the forces applied by volunteers wielding an eyebrow hair tool, we showed that all experimenters applied forces that are high enough to saturate the probability of a behavioral response. This indicates that, despite variability across experimenters and within the stimuli applied by a single experimenter, the eyebrow hair touch assay is able to reliably detect severe touch defects. The latter half of my thesis focuses on studying mechanosensation using microfluidic technology. I present a review of microfluidics for studying mechanobiology of various model organisms, and then discuss the development and application of a microfluidic device for applying controlled stimuli to the C. elegans TRNs while performing high-resolution imaging. This device uses pneumatic actuators to apply controlled stimuli to worms with TRNs expressing a fluorescent protein. In one set of experiments, this fluorescent protein is the Calcium sensor GCaMP6s, which changes its intensity when the concentration of Calcium in the TRN increases, indicating activation of the neuron. These experiments showed that the device could activate TRNs using dynamic stimuli targeted to the neuron of interest. Finally, I present experiments that use a fluorescent protein in the mitochondria of the TRN to measure the mechanical deformation of the TRN upon mechanical stimulation. This set of experiments showed that local indentation leads to local strain in C. elegans TRNs, and that local strain is not dependent on some of the extracellular matrix proteins associated with touch sensation. The contributions described in this thesis have helped advance our collective understanding of touch sensation, and will hopefully inspire future discoveries that slow or prevent degradation of touch sensation in humans.

Keywords: Mechanical Engineering.
Advisor: Chaudhuri Ovijit 
Goodman Miriam Beth 
Pruitt Beth