title: Embryonic stem cell-derived motoneurons provide a highly sensitive cell culture model for botulinum neurotoxin studies, with implications for high-throughput drug discovery.
abstract: Botulinum neurotoxins (BoNTs) inhibit cholinergic synaptic transmission by specifically cleaving proteins that are crucial for neurotransmitter exocytosis. Due to the lethality of these toxins, there are elevated concerns regarding their possible use as bioterrorism agents. Moreover, their widespread use for cosmetic purposes, and as medical treatments, has increased the potential risk of accidental overdosing and environmental exposure. Hence, there is an urgent need to develop novel modalities to counter BoNT intoxication. Mammalian motoneurons are the main target of BoNTs; however, due to the difficulty and poor efficiency of the procedures required to isolate the cells, they are not suitable for high-throughput drug screening assays. Here, we explored the suitability of embryonic stem (ES) cell-derived motoneurons as a renewable, reproducible, and physiologically relevant system for BoNT studies. We found that the sensitivity of ES-derived motoneurons to BoNT/A intoxication is comparable to that of primary mouse spinal motoneurons. Additionally, we demonstrated that several BoNT/A inhibitors protected SNAP-25, the BoNT/A substrate, in the ES-derived motoneuron system. Furthermore, this system is compatible with immunofluorescence-based high-throughput studies. These data suggest that ES-derived motoneurons provide a highly sensitive system that is amenable to large-scale screenings to rapidly identify and evaluate the biological efficacies of novel therapeutics.
authors.affiliation: Department of Target Discovery and Experimental Microbiology, US Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Frederick, MD 21702-5011, USA.
authors.forename: Erkan
authors.initials: E
authors.lastname: Kiris
authors.forename: Jonathan E
authors.initials: JE
authors.lastname: Nuss
authors.forename: James C
authors.initials: JC
authors.lastname: Burnett
authors.forename: Krishna P
authors.initials: KP
authors.lastname: Kota
authors.forename: Dawn C
authors.initials: DC
authors.lastname: Koh
authors.forename: Laura M
authors.initials: LM
authors.lastname: Wanner
authors.forename: Edna
authors.initials: E
authors.lastname: Torres-Melendez
authors.forename: Rick
authors.initials: R
authors.lastname: Gussio
authors.forename: Lino
authors.initials: L
authors.lastname: Tessarollo
authors.forename: Sina
authors.initials: S
authors.lastname: Bavari
mesh_terms: D000818:Animals; D001904:Botulinum Antitoxin / Q000494:pharmacology*; D001905:Botulinum Toxins / Q000037:antagonists & inhibitors* / Q000633:toxicity; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D055808:Drug Discovery; D004353:Drug Evaluation, Preclinical / Q000295:instrumentation / Q000379:methods*; D053595:Embryonic Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D057166:High-Throughput Screening Assays / Q000295:instrumentation / Q000379:methods*; D051379:Mice; D008810:Mice, Inbred C57BL; D008954:Models, Biological; D009046:Motor Neurons / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D050825:Synaptosomal-Associated Protein 25 / Q000378:metabolism
chemical_list: D001904:Botulinum Antitoxin; C494209:Snap25 protein, mouse; D050825:Synaptosomal-Associated Protein 25; D001905:Botulinum Toxins