Document Detail

Detecting ATP release by a biosensor method.
MedLine Citation:
PMID:  15536175     Owner:  NLM     Status:  MEDLINE    
Cells release adenosine 5'-triphosphate (ATP) into the extracellular space in response to various stimuli. This released ATP plays an important physiological role in cell-to-cell signal transduction. The bulk ATP concentration can be detected using a conventional luciferin-luciferase assay. However, the ATP concentration in the vicinity of the cell surface is often different from the bulk concentration because of its rapid degradation by ecto-ATPases and because of delayed diffusion due to unstirred layer effects. Here, we describe a simple biosensor method to measure the local ATP concentration on the cell surface in real time. The method is based on the ATP-dependent opening of ligand-gated cation channels of purinergic P2X receptors expressed in undifferentiated pheochromocytoma (PC12) cells or in human embryonic kidney 293 (HEK293) cells stably transfected with recombinant P2X2 purinergic receptors. Under the whole-cell configuration of patch-clamp, a sensor PC12 cell or HEK293 is positioned within the proximity of a target cell, and the P2X-mediated currents induced by ATP released from a given site on the target cell surface is measured. The ATP release is quantified by a calibration procedure utilizing local puff applications of ATP at preset concentrations.
Seiji Hayashi; Akihiro Hazama; Amal K Dutta; Ravshan Z Sabirov; Yasunobu Okada
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Publication Detail:
Type:  Journal Article     Date:  2004-11-09
Journal Detail:
Title:  Science's STKE : signal transduction knowledge environment     Volume:  2004     ISSN:  1525-8882     ISO Abbreviation:  Sci. STKE     Publication Date:  2004 Nov 
Date Detail:
Created Date:  2004-11-10     Completed Date:  2005-03-17     Revised Date:  2010-01-14    
Medline Journal Info:
Nlm Unique ID:  100964423     Medline TA:  Sci STKE     Country:  United States    
Other Details:
Languages:  eng     Pagination:  pl14     Citation Subset:  IM    
Department of Cell Physiology, National Institute for Physiological Sciences, Okazaki 444-8585, Japan.
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MeSH Terms
Adenosine Triphosphate / metabolism*
Adrenal Gland Neoplasms / genetics,  metabolism,  pathology
Biosensing Techniques / methods*,  standards
Calibration / standards
Cell Line
Cell Line, Tumor
Kidney / chemistry,  embryology,  metabolism
Pheochromocytoma / genetics,  metabolism,  pathology
Receptors, Purinergic P2 / genetics
Transfection / methods
Reg. No./Substance:
0/Receptors, Purinergic P2; 0/purinergic P2X2 receptor; 56-65-5/Adenosine Triphosphate

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