Urea transport by cotransporters

The rabbit Na+-glucose cotransporter (rbSGLT1) was expressed in Xenopus laevis oocytes and urea transport in rbSGLT1 and non-injected (control) oocytes was studied using [14C]urea as a tracer. The level of rbSGLT1 expression in these batches of oocytes was monitored by measuring the uptake of alpha-methyl-D-[14C]glucopyranoside ([14C]alphaMDG). In rbSGLT1-expressing oocytes, there was a 4-fold increase in urea transport in the absence of sugar relative to that in control oocytes. Urea uptake was not Na+ dependent and was linear with both time of incubation (5-120 min) and increasing urea concentration (50 microM to 100 mM) in the bathing medium. rbSGLT1 urea uptake was blocked by the rbSGLT1-specific inhibitor phlorizin (Ki 1 microM) in 100 mM NaCl buffer, but was not affected in 100 mM choline chloride buffer. Phloretin inhibited rbSGLT1 urea uptake with a low affinity (Ki > 1 mM) in the presence and absence of Na+. The uptake of 55 m[mu]M urea through rbSGLT1 was not blocked by 100 mM urea analogues including thiourea, 1,3-dimethyl urea, 1,1-dimethyl urea and acetamide. The activation energies (Ea) of urea transport for control and rbSGLT1-expressing oocytes were 14+/-3 and 6+/-1 kcal mol(-1), respectively. The low Ea for urea transport through rbSGLT1 is comparable to the Ea of passive water transport through rbSGLT1. Urea transport through rbSGLT1 was further increased when the cotransporter was activated by the addition of sugar to the external medium. The rate of sugar-dependent urea uptake was directly proportional to the rate of Na+-glucose-H2O cotransport such that the amount of urea transport was approximately proportional to the molar concentration ratio of urea to H2O (55 microM/55 M). The low affinity Na+-glucose (pSGLT3), the Na+-iodide (rNIS) and the Na+-(Cl-)-GABA (hGAT1) cotransporters expressed in oocytes demonstrated similar urea transport properties. These observations suggest that cotransporters behave as urea channels in the absence of substrates. Furthermore, under substrate-transporting conditions, the same cotransporters serve as urea cotransporters. This could account for urea transport in cells that appear not to have urea uniporters or channels.