An automated microfluidic platform for calcium imaging of chemosensory neurons in Caenorhabditis elegans

Abstract

Functional fluorescence imaging methods are widely used to study cellular physiology. When applied to small organisms, these methods suffer from low-throughput due to the laborious immobilization/stimulus delivery procedure that is typically involved during imaging. Here, we describe the development of an automated microfluidic-based platform for performing automated neuronal functional (calcium) imaging in the roundworm Caenorhabditis elegans. The platform, capable of processing tens to hundreds of worms per hour, immobilizes individual worms, delivers a chemical odor to their nose and collects calcium imaging data from single neurons without any manual intervention. We used the developed platform to obtain a large number of calcium responses from worms of different ages (212 worms were imaged in total). The calcium imaging data revealed significant difference in the responses from young and old worms, indicating that neural functionality is age-dependent. We believe that such a technology will be an essential tool for obtaining repeatable and accurate functional imaging data from a large population of worms, in order to minimize stochastic biological noise and identify statistically significant trends.

Analysis of Caenorhabditis elegans in microfluidic devices

“…Combined with fluorescence imaging of calcium transmitters and neurons, behavior mechanism could be explored in the neuronal level [60]. Based on the established microfluidic devices by several researchers, accurate stimuli could be delivered to individual worms flexibly and responses of worms behavior and neuron could be monitored [33,35,39,52,[61][62][63][64].…”

“…Chung and others [29] used the cooperation of a cooler and micro-valves to restrict the movement of C. elegans, and exploited software to realize automatic images identification and [52]; (b) a microfluidic platform enabled stable worm immobilizaton for axotomy and observation of neural regeneration process [27]. cell ablation.…”

“…Also, Chalasani et al [35] used a similar design to characterize the role of C. elegans olfactory neurons (AWC) and interneurons (AIB and AIY) in information processing of food-and odour-evoked behaviors. Chokshi et al [52] utilized an automated microfluidic-based e-chip (Figure 6(a), consisted of a worm trap, a set of ITO electrodes, an array of PDMS micropillars and a flush channel) in which single worm was immobilized in a tapered channel with its nose exposed to a flowing stream. With this device, the electrically induced ASH neuronal response was observed to be dependent on the magnitude, polarity, and spatial location of the electrical stimulus as well as on the age of the worm.…”

“…These studies are extremely useful for chemical screening and evaluation of molecular mechanisms. In microfluidic platform, precise chemical stimuli could be delivered to individual worms by flexible spatial and temporal control and the corresponding responses of worms could be monitored in real-time [34,[37][38][39][52][53][54]. Gray et al [37] established a hybridized PDMS-Agar chip ( Figure 5(b)), and generated a stable concentration gradient of oxygen in the chamber to study the behavioral responses of C. elegans to oxygen, and found the worms exhibited a strong behavioral preference for 5%-12% oxygen, avoiding higher and lower oxygen levels.…”

A perspective on optical developments in microfluidic platforms for Caenorhabditis elegans research

“…Chronis et al also demonstrated the ability to deliver the odors with precise temporal control while monitoring the neuronal activity, which is difficult with conventional imaging techniques. Upgraded versions of this design can now be used for the automation of the process from worm loading to image acquisition increasing the throughput up to 30 worms/h, 26 and the ability of delivering multiple chemical stimuli in specific temporal patterns. 27 These chips have had a great impact on calcium imaging experiments particularly in the dissection of the olfactory behavior circuitry.…”