Parallel-Channel Electrotaxis and Neuron Screening of Caenorhabditis elegans

Abstract

In this paper, we report a novel microfluidic method to conduct a Caenorhabditis elegans electrotaxis movement assay and neuronal imaging on up to 16 worms in parallel. C. elegans is a model organism for neurodegenerative disease and movement disorders such as Parkinson’s disease (PD), and for screening chemicals that alleviate protein aggregation, neuronal death, and movement impairment in PD. Electrotaxis of C. elegans in microfluidic channels has led to the development of neurobehavioral screening platforms, but enhancing the throughput of the electrotactic behavioral assay has remained a challenge. Our device consisted of a hierarchy of tree-like channels for worm loading into 16 parallel electrotaxis screening channels with equivalent electric fields. Tapered channels at the ends of electrotaxis channels were used for worm immobilization and fluorescent imaging of neurons. Parallel electrotaxis of worms was first validated against established single-worm electrotaxis phenotypes. Then, mutant screening was demonstrated using the NL5901 strain, carrying human α-synuclein in the muscle cells, by showing the associated electrotaxis defects in the average speed, body bend frequency (BBF), and electrotaxis time index (ETI). Moreover, chemical screening of a PD worm model was shown by exposing the BZ555 strain, expressing green fluorescence protein (GFP) in the dopaminergic neurons (DNs), to 6-hydroxydopamine neurotoxin. The neurotoxin-treated worms exhibited a reduction in electrotaxis swimming speed, BBF, ETI, and DNs fluorescence intensity. We envision our technique to be used widely in C. elegans-based movement disorder assays to accelerate behavioral and cellular phenotypic investigations.

Microfluidic Electric Egg-Laying Assay and Application toIn-vivoToxicity Screening of Microplastics usingC. elegans

“…Prior to EF stimulation, a 60 s acclimation period was used during which the flow was stopped by maintaining the inlet and outlet tubes at the same height level. Our previous experiments showed that the anode-facing worms could deposit significantly more eggs 27,35 , highlighting that the worm loading orientation with respect to the EF direction could affect our results. To maintain an equal exposure condition among randomly oriented worms in our device (as shown in Figure 2A), we adopted a new EF stimulation technique in which a series of 5s anodal and 5s cathodal pulses, separated by 25s acclimation periods, was applied for 10 min (Figure 2B).…”

“…This prevented the use of unidirectional pulses because approximately half of the worms would have been exposed to cathodal pulses, leading to no egg-laying response and waste of animals. Similar longitudinal orientation in the multi-worm device may have been achieved by preconditioning the worms with a longitudinal stimulus (e.g., controlled EF 37 or flow 38 ), but this would have added complexity to our procedures.…”

“…The code was also used to control the sourcemeter and apply the desired EF pulses while changing the EF polarity when needed. According to our single channel-based egg-laying experiments, an EF of 6 V/cm was needed to maximize egg-laying in C. elegans 22,35 The required voltage to obtain this EF in the new microfluidic device was estimated to be 34 V using a simple COMSOL Multiphysics simulation (Figure 1D), as described in the Supplementary file Section S1.…”

Editorial for the Topic on Micromachining for Advanced Biological Imaging

“…Today, various types of micro- and nano-structures play important roles in advanced bioimaging applications. Accordingly, in this Editorial, we highlight reviews and original research articles that focus on the use of micromachining for advanced biological imaging, including in micro-optics [ 1 , 2 , 3 ], nanopatterning [ 4 , 5 ], and optical imaging with microfluidics [ 6 , 7 , 8 ].…”

“…This is a simple and inexpensive method that can be performed on a conventional fluorescence microscope that uncouples the observation and selection of droplets. Youssef et al report the development of an easy-to-use microfluidic electrotaxis-based chip that can be used to investigate the behavior and neuron degeneration of 16 worms in parallel [ 8 ]. They show its applicability for genetic, chemical, and neuronal screening after validating it against a single-worm electrotaxis assay.…”