(473e) Microfluidic Systems for High-Throughput Functional Imaging of Mechanosensing Neurons in Caenorhabditis Elegans

Mechanosensation is an important modality to sense the environment in most animals, but the exact mechanism of how information is sensed and transduced is not well understood. Caenorhabditis elegans, a soil nematode and an important model organism for neuroscience, is a good model for studying mechanosensory biology. C. elegans has gentle and harsh touch neurons, which mediate behavior responses. Conventional methods for studying mechanosensation uses painstaking laser ablation and behavior assay by eyelash touches or glued worm mechanically stimulated by stylus. The limitations of these methods include relatively low throughput and difficulty to control the exact stimulus repeated and robustly on large number of worms, so studying functions of genes or screening drugs is impractical.

To address these technical limitations, we designed a microfluidic device for trapping and delivering mechanical stimuli to single worms while monitoring their neural activity. We used this mechanosensing device to monitor genetically encoded calcium indicator GCaMP6 changes in touch receptor neurons. Reponses to anterior touch require ALML, ALMR, and AVM, while responses to posterior touch require PLML and PLMR. We show the ability of the device to deliver mechanical stimulus in a graded manner. The higher pressure generated reliable increases in the fluorescence ratio averaged over the cell body. It is indicating our controlled stimuli correspond to gentle touch typically delivered in behavioral tests by eyelash stroke. We also show that the controlled mechanical stimuli can be delivered to ALM, AVM, and PVM neurons, and calcium changes of these neurons can be recorded. To assess the effect of mechanical stimulation other than cell body, we also record along ALM processes. Calcium transients show smaller peak and faster decays. Upon repeated stimulation, many sensory systems show habituation. Further, we demonstrate how the C. elegans gentle touch system habituates upon repeated stimulation.

We have shown in this work that the microfluidic device is flexible in stimulating different parts of the C. elegans and allowing for function calcium imaging. The assay is semi-quantitative in that the intensity and duration of the stimuli can be easily controlled. We envision this device as a general mechanosensing manipulation platform for neuronal imaging that will contribute to the neuronal functional map of the C. elegans nervous system, as well as serving as a vehicle for discovering genes affecting the function of neuronal functions and therapeutics for neuronal diseases.