Document Detail


Single-cell electroporation of adult sensory neurons for gene screening with RNA interference mechanism.
MedLine Citation:
PMID:  18314198     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
RNA interference appears as a technique of choice to identify gene candidate or to evaluate gene function. To date, a main problem is to achieve high transfection efficiencies on native cells such as adult neurons. In addition, transfection on organ or mass culture does not allow to approach the cellular diversity. Dorsal root ganglia are composed with several cell types to convey somato-sensory sensations. Single-cell electroporation is the most recent method of transfection that allows the introduction into cells, not only dyes or drugs, but also large molecules such plasmid DNA expression constructs. In the present study, the application of the RNA interference technique with the use of single-cell electroporation was evaluated in primary culture of adult sensory neurons. With the use of fluorescent dextran as a co-transfectant, we first determined the non-specific siRNA concentration leading to cell death. Efficacy of siRNA at the non-toxic concentration was demonstrated at the protein level by extinction of GFP fluorescence in actin-GFP neurons and by the inhibition of the intracellular Cl- concentration increase following activation of the membrane co-transporter Na+-K+-2Cl- in regenerating axotomized sensory neurons. Altogether, these data show that delivery of siRNAs by single-cell electroporation leads to the induction of functional RNA interference.
Authors:
Mathieu Boudes; Simon Pieraut; Jean Valmier; Patrick Carroll; Frédérique Scamps
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Publication Detail:
Type:  Journal Article     Date:  2008-01-31
Journal Detail:
Title:  Journal of neuroscience methods     Volume:  170     ISSN:  0165-0270     ISO Abbreviation:  J. Neurosci. Methods     Publication Date:  2008 May 
Date Detail:
Created Date:  2008-04-18     Completed Date:  2008-07-18     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  7905558     Medline TA:  J Neurosci Methods     Country:  Netherlands    
Other Details:
Languages:  eng     Pagination:  204-11     Citation Subset:  IM    
Affiliation:
Inserm U583, Univ Montpellier II, Montpellier F-34000 France.
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MeSH Terms
Descriptor/Qualifier:
Animals
Cell Survival
Cells, Cultured
Chlorides / metabolism
Electrophysiology
Electroporation / methods*
Female
Ganglia, Spinal / cytology
Green Fluorescent Proteins / genetics
Image Processing, Computer-Assisted
Mice
Neurons, Afferent / physiology*
RNA, Small Interfering / diagnostic use*
Transfection
Chemical
Reg. No./Substance:
0/Chlorides; 0/RNA, Small Interfering; 147336-22-9/Green Fluorescent Proteins

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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