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A microfluidic device for whole-animal drug screening using electrophysiological measures in the nematode C. elegans.

ABSTRACT: This paper describes the fabrication and use of a microfluidic device for performing whole-animal chemical screens using non-invasive electrophysiological readouts of neuromuscular function in the nematode worm, C. elegans. The device consists of an array of microchannels to which electrodes are attached to form recording modules capable of detecting the electrical activity of the pharynx, a heart-like neuromuscular organ involved in feeding. The array is coupled to a tree-like arrangement of distribution channels that automatically delivers one nematode to each recording module. The same channels are then used to perfuse the recording modules with test solutions while recording the electropharyngeogram (EPG) from each worm with sufficient sensitivity to detect each pharyngeal contraction. The device accurately reported the acute effects of known anthelmintics (anti-nematode drugs) and also correctly distinguished a specific drug-resistant mutant strain of C. elegans from wild type. The approach described here is readily adaptable to parasitic species for the identification of novel anthelmintics. It is also applicable in toxicology and drug discovery programs for human metabolic and degenerative diseases for which C. elegans is used as a model.

SUBMITTER: Lockery SR

PROVIDER: S-EPMC3372093 | biostudies-literature | 2012 Jun

REPOSITORIES: biostudies-literature

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A microfluidic device for whole-animal drug screening using electrophysiological measures in the nematode C. elegans.

Lockery Shawn R SR Hulme S Elizabeth SE Roberts William M WM Robinson Kristin J KJ Laromaine Anna A Lindsay Theodore H TH Whitesides George M GM Weeks Janis C JC

Lab on a chip 20120515 12

This paper describes the fabrication and use of a microfluidic device for performing whole-animal chemical screens using non-invasive electrophysiological readouts of neuromuscular function in the nematode worm, C. elegans. The device consists of an array of microchannels to which electrodes are attached to form recording modules capable of detecting the electrical activity of the pharynx, a heart-like neuromuscular organ involved in feeding. The array is coupled to a tree-like arrangement of di ...[more]

PMID: 22588281

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Project description:Many anthelmintic drugs used to treat parasitic nematode infections target proteins that regulate electrical activity of neurons and muscles: ion channels (ICs) and neurotransmitter receptors (NTRs). Perturbation of IC/NTR function disrupts worm behavior and can lead to paralysis, starvation, immune attack and expulsion. Limitations of current anthelmintics include a limited spectrum of activity across species and the threat of drug resistance, highlighting the need for new drugs for human and veterinary medicine. Although ICs/NTRs are valuable anthelmintic targets, electrophysiological recordings are not commonly included in drug development pipelines. We designed a medium-throughput platform for recording electropharyngeograms (EPGs)-the electrical signals emitted by muscles and neurons of the pharynx during pharyngeal pumping (feeding)-in Caenorhabditis elegans and parasitic nematodes. The current study in C. elegans expands previous work in several ways. Detecting anthelmintic bioactivity in drugs, compounds or natural products requires robust, sustained pharyngeal pumping under baseline conditions. We generated concentration-response curves for stimulating pumping by perfusing 8-channel microfluidic devices (chips) with the neuromodulator serotonin, or with E. coli bacteria (C. elegans' food in the laboratory). Worm orientation in the chip (head-first vs. tail-first) affected the response to E. coli but not to serotonin. Using a panel of anthelmintics-ivermectin, levamisole and piperazine-targeting different ICs/NTRs, we determined the effects of concentration and treatment duration on EPG activity, and successfully distinguished control (N2) and drug-resistant worms (avr-14; avr-15; glc-1, unc-38 and unc-49). EPG recordings detected anthelmintic activity of drugs that target ICs/NTRs located in the pharynx as well as at extra-pharyngeal sites. A bus-8 mutant with enhanced permeability was more sensitive than controls to drug treatment. These results provide a useful framework for investigators who would like to more easily incorporate electrophysiology as a routine component of their anthelmintic research workflow.

Project description:This paper describes a new microfluidic platform for screening drugs and their dose response on the locomotion behavior of free living nematodes and parasitic nematodes. The system offers a higher sensitivity drug screening chip which employs a combination of existing and newly developed methods. Real-time observation of the entire drug application process (i.e. the innate pre-exposure locomotion, the transient response during drug exposure and the time-resolved, post-exposure behavior) at a single worm resolution is made possible. The chip enables the monitoring of four nematode parameters (number of worms responsive, number of worms leaving the drug well, average worm velocity and time until unresponsiveness). Each parameter generates an inherently different dose response; allowing for a higher resolution when screening for resistance. We expect this worm chip could be used as a robust cross species, cross drug platform. Existing nematode motility and migration assays do not offer this level of sophistication. The device comprises two principal components: behavioral microchannels to study nematode motility and a drug well for administering the dose and observing drug effects as a function of exposure time. The drug screening experiment can be described by three main steps: (i) 'pre-exposure study'- worms are inserted into the behavioral channels and their locomotion is characterized, (ii) 'dose exposure'- worms are guided from the behavioral microchannels into the drug well and held for a predefined time, during which time their transient response to the dose is characterized and (iii) 'post-exposure study'- worms are guided back into the behavioral microchannels where their locomotion (i.e. their time-resolved response to the dose) is characterized and compared to pre-exposure motility. The direction of nematodes' movement is reliably controlled by the application of an electric field within a defined range. Control experiments (e.g. in the absence of any drug) confirm that the applied electric fields do not affect the worms' motility or viability. We demonstrate the workability of the microfluidic platform on free living Caenorhabditis elegans (wild-type N2 and levamisole resistant ZZ15 lev-8) and parasitic Oesophagotomum dentatum (levamisole-sensitive, SENS and levamisole-resistant, LEVR) using levamisole (a well-studied anthelmintic) as the test drug. The proposed scheme of drug screening on a microfluidic device is expected to significantly improve the resolution, sensitivity and data throughput of in vivo testing, while offering new details on the transient and time-resolved exposure effects of new and existing anthelmintics.

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A microfluidic device for whole-animal drug screening using electrophysiological measures in the nematode C. elegans.

Publications

A microfluidic device for whole-animal drug screening using electrophysiological measures in the nematode C. elegans.

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