NMR measurement of intracellular water volume in rat kidney proximal tubules

Abstract

Knowledge of cell water volume is essential for the measurement of concentrations of intracellular ions and metabolites in kidney proximal tubules. We have developed a method which utilizes 35Cl-NMR as a measure of extracellular volume and 2H-NMR, in combination with a membrane-impermeable shift-reagent [Dy-DTPA] 2−, as a measure of the ratio of intra- and extracellular water volumes. Measurement of extracellular volume by 35Cl-NMR is possible, since the resonance of intracellular 35Cl is too broad to be detectable in kidney cells. The 2H-NMR measurement exploits the fact that only extracellular water is in direct contact with [Dy-DTPA] 2−. However, rapid exchange of water across the cell membrane results in only a single 2H 2O resonance at a chemical shift which is a weighted average of the shifted extra- and unshifted intracellular water resonances. Expression of the extracellular volume as a fraction of the total volume by f Cl and as a fraction of the total water-volume by f D, permits the calculation of the fractional cell-water content f w = [( 1 f D ) − 1] [( 1 f C1 ) − 1] . This approach was applied to proximal tubular suspensions prepared from the rat kidney. The water content was found to be 76.9±1.8% ( n = 6) at 37°C. Increasing extracellular osmolality from 295 to 390 mOsm/kg H 2O, by addition of mannitol, decreased the water content by 21%. Our results are in good agreement with those obtained by the gravimetric method.