A primary culture system for functional analysis of C. elegans neurons and muscle cells.
C. elegans has provided important insights into neuromuscular system function and development. However, the animal's small size limits access to individual neurons and muscle cells for physiological, biochemical, and molecular study. We describe here primary culture methods that allow C. elegans embryonic cells to differentiate into neurons and muscle cells in vitro. Morphological, electrophysiological, and GFP reporter studies demonstrate that the differentiation and functional properties of cultured cells are similar to those observed in vivo. Enriched populations of cells expressing specific GFP reporters can be generated by fluorescence-activated cell sorting. Addition of double-stranded RNA to the culture medium induces dramatic knockdown of targeted gene expression. Primary nematode cell culture provides a new foundation for a wide variety of experimental opportunities heretofore unavailable in the field.
Christensen
Michael
M
Department of Anesthesiology, Vanderbilt University Medical Center, Nashville, TN 37232, USA.
Estevez
Ana
A
Yin
Xiaoyan
X
Fox
Rebecca
R
Morrison
Rebecca
R
McDonnell
Maureen
M
Gleason
Christina
C
Miller
David M
DM
Strange
Kevin
K
D000818:Animals; D017173:Caenorhabditis elegans / Q000166:cytology* / Q000196:embryology / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D005434:Flow Cytometry; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects / Q000502:physiology; D017930:Genes, Reporter / Q000502:physiology; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents / Q000378:metabolism; D007473:Ion Channels / Q000187:drug effects / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009046:Motor Neurons / Q000166:cytology / Q000378:metabolism; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism; D009132:Muscles / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D009420:Nervous System / Q000166:cytology* / Q000378:metabolism; D009474:Neurons / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism
D007202:Indicators and Reagents; D007473:Ion Channels; D008164:Luminescent Proteins; D009124:Muscle Proteins; D009419:Nerve Tissue Proteins; D049452:Green Fluorescent Proteins