SMALL CHANGES OF CYTOSOLIC SODIUM IN RAT VENTRICULAR MYOCYTES MEASURED WITH SBFI IN EMISSION RATIO MODE

The spectral properties of SBFI (sodium-binding benzofurzan isophthala te) were re-examined to arrive at a more specific and sensitive method to measure small changes of intracellular sodium ([Na+](i)) particula rly at low concentration. Relative to spectra of SBFI in protein-and c ell-free solution, binding of SBFI to intracellular proteins caused a shift of excitation and emission spectra, and increased quantum effici ency. Excitation of SBFI at 340 nm caused an exclusively sodium-depend ent fluorescence from 400-420nm, and hardly any change of fluorescence above 530 nm upon replacing sodium by potassium. Due to these spectra l and quantum efficiency changes, SBFI excitated at 340 nm can be used in a dual emission ratio mode to measure [Na+](i). In dual emission r atio mode (410 and 590 nm, respectively), the fluorescence ratio incre ased by a factor of 13 upon replacing sodium for potassium, The appare nt equilibrium constant measured in single isolated rat ventricular my ocytes was 22.5 +/- 0.3 mmol/l. Control [Na+](i) was 9.6 +/- 0.4 mmol/ l. After abrupt reduction of extracellular sodium from 156 to 29 or 11 mmol/l, [Na+](i) decreased mono-exponentially to 2.5 +/- 0.3 and 1.9 +/- 0.3 mmol/l, respectively, with a rate constant of about 0.02/s. We conclude that SBFI used in dual emission mode provides a more sensiti ve and more specific method to measure small changes of [Na+]( )i in s ingle myocytes down to cytosolic sodium concentration as low as about 1 mmol/l. (C) 1997 Academic Press Limited.