title: Microfluidic platform for electrophysiological recordings from host-stage hookworm and Ascaris suum larvae: A new tool for anthelmintic research.
abstract: The screening of candidate compounds and natural products for anthelmintic activity is important for discovering new drugs against human and animal parasites. We previously validated in Caenorhabditis elegans a microfluidic device ('chip') that records non-invasively the tiny electrophysiological signals generated by rhythmic contraction (pumping) of the worm's pharynx. These electropharyngeograms (EPGs) are recorded simultaneously from multiple worms per chip, providing a medium-throughput readout of muscular and neural activity that is especially useful for compounds targeting neurotransmitter receptors and ion channels. Microfluidic technologies have transformed C. elegans research and the goal of the current study was to validate hookworm and Ascaris suum host-stage larvae in the microfluidic EPG platform. Ancylostoma ceylanicum and A. caninum infective L3s (iL3s) that had been activated in vitro generally produced erratic EPG activity under the conditions tested. In contrast, A. ceylanicum L4s recovered from hamsters exhibited robust, sustained EPG activity, consisting of three waveforms: (1) conventional pumps as seen in other nematodes; (2) rapid voltage deflections, associated with irregular contractions of the esophagus and openings of the esophogeal-intestinal valve (termed a 'flutter'); and (3) hybrid waveforms, which we classified as pumps. For data analysis, pumps and flutters were combined and termed EPG 'events.' EPG waveform identification and analysis were performed semi-automatically using custom-designed software. The neuromodulator serotonin (5-hydroxytryptamine; 5HT) increased EPG event frequency in A. ceylanicum L4s at an optimal concentration of 0.5 mM. The anthelmintic drug ivermectin (IVM) inhibited EPG activity in a concentration-dependent manner. EPGs from A. suum L3s recovered from pig lungs exhibited robust pharyngeal pumping in 1 mM 5HT, which was inhibited by IVM. These experiments validate the use of A. ceylanicum L4s and A. suum L3s with the microfluidic EPG platform, providing a new tool for screening anthelmintic candidates or investigating parasitic nematode feeding behavior.
authors.affiliation: Institute of Neuroscience and African Studies Program, University of Oregon, 1254 University of Oregon, Eugene, OR 97403-1254, USA. Electronic address: jweeks@uoregon.edu.
authors.forename: Janis C
authors.initials: JC
authors.lastname: Weeks
authors.affiliation: Institute of Neuroscience, University of Oregon, 1254 University of Oregon, Eugene, OR 97403-1254, USA. Electronic address: billr@uoregon.edu.
authors.forename: William M
authors.initials: WM
authors.lastname: Roberts
authors.affiliation: Institute of Neuroscience, University of Oregon, 1254 University of Oregon, Eugene, OR 97403-1254, USA. Electronic address: kristinr@uoneuro.uoregon.edu.
authors.forename: Kristin J
authors.initials: KJ
authors.lastname: Robinson
authors.affiliation: Department of Microbiology, Immunology, and Tropical Medicine, School of Medicine and Health Sciences, The George Washington University, Washington, DC 20037, USA. Electronic address: keaney.melissa@gmail.com.
authors.forename: Melissa
authors.initials: M
authors.lastname: Keaney
authors.affiliation: Center for Discovery and Innovation in Parasitic Diseases, Dept. of Pathology and Laboratory Medicine, UC, San Francisco, USA. Electronic address: jon.vermeire@gmail.com.
authors.forename: Jon J
authors.initials: JJ
authors.lastname: Vermeire
authors.affiliation: US Department of Agriculture, Agricultural Research Service, Beltsville Human Nutrition Research Center, Diet, Genomic and Immunology Laboratory, Beltsville, MD, USA. Electronic address: joe.urban@ars.usda.gov.
authors.forename: Joseph F
authors.initials: JF
authors.lastname: Urban
authors.affiliation: Institute of Neuroscience, University of Oregon, 1254 University of Oregon, Eugene, OR 97403-1254, USA. Electronic address: shawn@uoregon.edu.
authors.forename: Shawn R
authors.initials: SR
authors.lastname: Lockery
authors.affiliation: Department of Microbiology, Immunology, and Tropical Medicine, School of Medicine and Health Sciences, The George Washington University, Washington, DC 20037, USA. Electronic address: jhawdon@email.gwu.edu.
authors.forename: John M
authors.initials: JM
authors.lastname: Hawdon
mesh_terms: D000722:Ancylostoma / Q000187:drug effects / Q000502:physiology*; D000818:Animals; D000871:Anthelmintics / Q000494:pharmacology*; D017165:Ascaris suum / Q000187:drug effects / Q000502:physiology*; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D055724:Electrophysiological Phenomena / Q000187:drug effects*; D007814:Larva / Q000187:drug effects / Q000502:physiology; D044085:Microfluidics / Q000379:methods*; D010274:Parasitology / Q000379:methods
chemical_list: D000871:Anthelmintics
keywords: Ancylostoma; Anthelmintic; Ascaris; Electropharyngeogram; Electrophysiology; Hookworm; Ivermectin; Microfluidic; ScreenChip; Serotonin