Passive water and ion transport by cotransporters

1. The rabbit Na+-glucose (SGLT1) and the human Na+-Cl--GABA (GAT1) cotransporters were expressed in Xenopus laevis oocytes, and passive Na+ and water transport were studied using electrical and optical techniques. Passive water permeabilities (L-p) of the cotransporters were deter mined from the changes in oocyte volume in response to osmotic gradients. The specific SGLT1 and GAT1 L-p values were obtained by measuring L-p in the presence and absence of blockers (phlorizin and SKF89976A). In the presence of the blockers, the L-p values of oocytes expressing SGLT1 and GAT1 were indistinguishable from the L-p of control oocytes, 2. Passive Na+ transport (Na+ leak) was obtained from the blocker-sensitive Naf currents in the absence of substrates (glucose and GABA). Passive Na+ and water transport through SGLT1 were blocked by phlorizin with the same sensitivity (inhibitory constant (K-i), 3-5 mu M). When Na+ was replaced with Li+, phlorizin also inhibited Li+ and water transport, but with a lower affinity (K-i, 100 mu M). When Na+ was replaced by choline, which is not transported, the SGLT1 L-p was indistinguishable from that in Na+ or Li+, but in this case water transport was less sensitive to phlorizin. 3. The activation energies (E-a) for passive Na+ and water transport through SGLT1 were 21 and 5 kcal mol(-1), respectively. The high E-a for Na+ transport is comparable to that of Na+-glucose cotransport and indicates that the process is dependent on conformational changes of the protein, while the low E-a for water transport is similar to that of water channels (aquaporins). 4. GAT1 also behaved as an SKF89976A-sensitive water channel. We did not observe passive Na+ transport through GAT1. 5. We conclude that passive water and Na+ transport through cotransporters depend on different mechanisms: Na+ transport occurs by a saturable uniport mechanism, and water permeation is through a low conductance water channel. In the case of SGLT1, we suggest that both the water channel and water cotransport could contribute to isotonic fluid transport across the intestinal brush border membrane.