Functional expression of Na-Ca exchanger clones measured with the fluorescent Ca(2+)-indicating dye fluo-3.
The process of Ca2+ homeostasis is of prime importance to all cells because of the ubiquitous role of cytoplasmic Ca2+ as an intracellular messenger and the cytotoxicity of sustained elevated cytosolic Ca2+ concentrations. Two classes of plasma membrane proteins are responsible for maintaining cytosolic free Ca2+ in the submicromolar range against a very large electrochemical Ca2+ gradient across the plasma membrane, the ATP-driven Ca2+ pump and Na-Ca exchangers. Two types of Na-Ca exchangers are known, the 3Na:1Ca exchangers found in heart, brain, kidney, and most other tissues and the 4Na:1Ca+ 1K exchanger found in retinal rod and cone photoreceptors. Functional expression of Na-Ca(/K) exchangers is most often measured as 45Ca uptake in Na(+)-loaded cells or as Na-Ca exchange currents with the giant excised patch technique. In this study, two functional assays used to detect expression of the bovine heart Na-Ca exchanger in CHO cells are described. Both assays are based on measurements of cytosolic free Ca2+ with the fluorescent Ca(2+)-indicating dye fluo-3 and should be equally applicable in the study of functional expression of both Na-Ca and Na-Ca/K exchanger clones.
Schnetkamp
P P
PP
Department of Medical Biochemistry, Faculty of Medicine, University of Calgary, AB, Canada.
D000814:Aniline Compounds; D000818:Animals; D016466:CHO Cells; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D002417:Cattle; D006224:Cricetinae; D005456:Fluorescent Dyes; D006706:Homeostasis; D007476:Ionophores / Q000494:pharmacology; D009206:Myocardium / Q000737:chemistry; D012964:Sodium / Q000378:metabolism; D019831:Sodium-Calcium Exchanger; D014162:Transfection; D014966:Xanthenes
D000814:Aniline Compounds; D002352:Carrier Proteins; D005456:Fluorescent Dyes; D007476:Ionophores; D019831:Sodium-Calcium Exchanger; D014966:Xanthenes; C059715:Fluo-3; D000001:Calcimycin; D012964:Sodium; D002118:Calcium