Effects of urea on K-Cl cotransport in sheep red blood cells: evidence for two signals of swelling.

The activation proceeds with a delay, like activation by swelling. Swelling of cells in urea activates K uptake further, but with no delay. Inactivation after removal of urea also proceeds without delay. With cotransport partially activated by reducing intracellular Mg concentration ([Mg]i) or with staurosporine, urea did not activate cotransport further. However, swelling activated cotransport further in these two types of cells. In terms of the three-state process for swelling-activation of K-Cl cotransport (P. B. Dunham, J. Klimczak, and P. J. Logue, J. Gen. Physiol. 101: 733-765, 1993), these results indicate that urea activates the first conversion, A-->B, and does so by inhibiting the reverse reaction promoted by a kinase, just as reducing [Mg]i does. Stimulation of cotransport by urea is nearly completely reversed by shrinkage, whereas activation by reducing [Mg]i is reversed only approximately 35%. Therefore urea inhibits the kinase indirectly, like swelling, by reducing macromolecular crowding of cytoplasmic proteins (A. P. Minton, G. C. Coleclasure, and J. C. Parker. Proc. Natl. Acad. Sci. USA 89: 10504-10506, 1992). Since swelling activates cotransport in two ways, one mimicked by urea and one not, there must be two signals of swelling, one a reduction of macromolecular crowding and the other probably a mechanical signal.