Design of an indicator of intracellular free Na+ concentration using 19F-NMR.
The development is described of an Na+ chelator with appropriate properties for an indicator of intracellular free Na+ concentration ([Na+]i). The new indicator, FCryp-1, is a tribenzo derivative of the parent (2:2:1) cryptand structure, incorporating the same F-substituted dibenzo 19F-NMR reporter group as the free [Ca2+] indicator, 5FBAPTA (Smith, G.A., Hesketh, T.R., Metcalfe, J.C., Feeney, J. and Morris, P.G. (1983) Proc. Natl. Acad. Sci., USA 80, 7178-7182). FCryp-1 has appropriate affinity for Na+ (KNa = 10(1.3) M-1) and selectivity over other intracellular cations (KK; KCa; K Mg less than 10(-1) M(-1)) for a [Na]i indicator. There is an 19F-NMR chemical shift of 2.00 ppm between free FCryp-1 and the Na-FCryp-1 complex which provides a direct read out of free [Na+]. FCryp-1 carries four carboxylate groups to confer aqueous solubility which can be esterified with acetoxymethyl groups to render the indicator membrane permeant. Experiments on pig lymphocytes loaded with FCryp-1 gave an indicated [Na+]i of 13.8 +/- 1.8 mM (n = 4). The FCryp-1 structure can also be readily modified to provide fluorescent [Na+]i indicators.
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002614:Chelating Agents / Q000138:chemical synthesis*; D005453:Fluorescence; D008214:Lymphocytes / Q000032:analysis; D008274:Magnesium / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D011188:Potassium / Q000378:metabolism; D012964:Sodium / Q000032:analysis*; D013329:Structure-Activity Relationship; D013552:Swine
D002614:Chelating Agents; D012964:Sodium; D008274:Magnesium; D011188:Potassium; D002118:Calcium