COULD CYTOPLASMIC CONCENTRATION GRADIENTS FOR SODIUM AND ATP EXIST ININTACT RENAL-CELLS

In renal cells, the Na+ pump maintains a transmembrane concentration g radient for sodium ensuring the net reabsorption of sodium with or wit hout cotransported species. This process requires a significant fracti on of the ATP turnover of proximal tubules and thick ascending limbs. To understand the potential regulatory influences of Na+ and ATP on th e activity of the Na+ pump in these nephron segments, the apparent kin etics of the membrane-bound Na+ -K+ ATPase and of the cellular Na+ pum p were studied in different preparations of dog proximal tubules and t hick ascending limbs (tubular suspensions, tissue homogenates, and bas olateral membrane vesicles) obtained from dog kidney cortex and red me dulla. Two determinant kinetic parameters, i.e., the apparent Michaeli s constant (K-m) and the saturating concentrations for sodium and ATP, were compared with the intracellular concentrations of Na+ and ATP me asured under physiological conditions. In both types of tubules, the a pparent K-m value for Na+ (5-15 mM) is set well below the measured mea n intracellular concentration of sodium (50-60 mM), suggesting that th e Na+ pump should be saturated by sodium ions under normal conditions. Nevertheless, a modest increment of the Na concentration in the vicin ity of the pump, obtained by equilibrating the intra- and extra-cellul ar sodium concentrations at various extracellular [Na+] with nystatin, increases the activity of the Na+ pump in intact cortical tubules and thick ascending limbs, even when the extracellular [Na+] is set at th e estimated intracellular [Na+], demonstrating that the pump is not sa turated by sodium in situ. Similarly, the kinetics of the renal Na+ pu mp as a function of the ATP concentration suggested that the pump shou ld be saturated by ATP in physiological conditions, since in both tiss ues the cellular ATP level (3-6 mM) is higher than the concentration r equired to achieve saturation of this activity (<2.5 mM). However, in renal cortical tubules, the steady-state intracellular [Na+] is affect ed by modest changes of ATP concentration, suggesting that the Na+ pum p is not functionally saturated by ATP. Our data suggest that concentr ation gradients for Na+ and ATP may exist in the cytosol of renal cell s. These gradients would be related to the polarity of sodium transpor t and of the ATP-consuming and ATP-regenerating processes in intact ce lls. According to this scheme, the intracellular concentration of both Na+ and ATP would be lower in the vicinity of the Na+ pump, the site of active extrusion of sodium and ATP utilization, than the mean cellu lar concentrations measured using classical techniques. Such an organi zation would amplify the signals given to the cell membranes by change s in ATP concentration.