CHARACTERIZATION OF THE ERYTHROCYTE SODIUM-LITHIUM COUNTERTRANSPORTER- LIMITATIONS AND ASSUMPTIONS OF TRADITIONAL AND KINETIC METHODOLOGIES

The present work examined the key elements featuring in the various me thods used to characterize the erythrocyte sodium-lithium countertrans port. Effects of medium composition on lithium efflux were investigate d in 20 subjects. Mean lithium efflux (mmol Li/l RBC.h) into a 150 mM sodium medium was significantly higher than efflux into a revised sodi um-rich medium (149 mM) containing 1 mM Mg (0.335 +/- 0.100 vs. 0.298 +/- 0.085 respectively; P < 0.03). Mean lithium efflux into sodium-fre e media where sodium had been entirely replaced by magnesium, was sign ificantly lower than efflux into a choline-based medium containing onl y 1 mM magnesium (0.088 +/- 0.027 vs. 0.109 +/- 0.034 respectively; P = 0.03). Sodium-lithium countertransport activity and the transporter' s kinetic profile were measured simultaneously in 35 subjects using tr aditional choline-based and kinetic methodologies. There was a signifi cant correlation between countertransport activity and maximal rate of turnover (V-max) (r = 0.62; P < 0.001); V-max values were consistentl y greater than their corresponding countertransport activities (P < 0. 001). On subdividing the subject group into tertiles based on the Mich aelis-Menten constant (k(m)) values (mM), <75, 75 - 150 and >150, the slopes of the regression lines for each group diminished progressively (0.64, 0.49 and 0.23 respectively), correlations within each group re mained significant (P < 0.001, P < 0.001 and P < 0.02). No sig nifican t correlation was found between k, values and countertransport activit y (r = 0.035; P = NS). Increasing the number of points representing so dium concentrations within the range 0-150 mM, improved the confidence in the emerging estimates of V-max and k(m) obtained by linear transf ormation. Comparison of kinetic data derived using four different anal ytical methods (two linear transformations, a nonlinear regression and a statistical method), showed no significant differences between the estimates yielded for either V-max (P = 0.88, NS) Or k(m) (P = 0.92, N S). This study has highlighted the critical roles of assay conditions and derivation techniques used when measuring sodium-lithium countertr ansport, emphasizing the need for standardization of the methodology.