Urea activation of K-Cl transport in human erythrocytes

This report prompted us to examine the effect of urea on K-Cl cotransport in human erythrocytes. In human erythrocytes, urea activated K-Cl cotransport reversibly and in a concentration-dependent manner. Pretreatment with okadaic acid abolished the urea activation of transport, suggesting that exposure to urea resulted in net dephosphorylation of the transporter or a key regulator and that the action of urea was exerted proximal to the phosphorylation-dephosphorylation step. At a concentration of 200 mM, urea activated K-Cl cotransport without any delay, even in the absence of cell swelling. However, with increasing urea concentrations, an appreciable increase in lag time was observed before the final steady-state flux was reached, suggesting that urea inhibits a regulatory kinase. The latter conclusion was also supported by the finding that, at any given urea concentration, the lag time for activation was greater than the lag time for deactivation. Mg depletion activated cotransport, and urea had no additional stimulatory effect in Mg-depleted cells. In urea-pretreated cells, swelling further activated cotransport, but without any measurable delay, in contrast to a time lag of 8 min when control cells (not exposed to urea) were swollen. The latter finding suggests that urea promotes the conversion of transporters from the resting to the partially activated state. These findings raise the possibility that high concentrations of urea in the renal medulla may play a role in the decrease in cell volume that occurs during the maturation of reticulocytes and young erythrocytes, both in normal subjects and in subjects with hemoglobinopathies.