Two Operational Modes of K-Cl Cotransport in Low K+ Sheep Red Blood Cells

The present study further characterizes the two operational modes of K-Cl cotransport in low K+ sheep red blood cells previously described and proposed by us: (i) a membrane-bound cotransport entity devoid of modulation by putative kinases/phosphatases, and (ii) an ATP- and Mg-modulated activity. We now report that quinine, spermine, and 300 µM Cai2+ inhibit (40-100%) K-Cl efflux in ATP- and Mgi2+-depleted cells and hence interact directly with the cotransporter moiety. The sensitivity of K-Cl cotransport to calyculin, however, was modulated by the ATP content of the cells: after depletion of both ATP and Mgi2+ the flux was insensitive to the inhibitor, whereas at about 120 µmol ATP/L cells calyculine inhibited K-Cl efflux by 95-100% in Mgi2+-free cells. The K-Cl cotransporter itself, free of modulation by kinases/phosphatases, ‘senses’ changes in cell volume perhaps at the membrane/cytoskeleton level. Cellular ATP then switches the co-transporter’s operational mode so that under physiological conditions the rate of K-Cl efflux is under the control of a putative kinases/phosphatases pathway.