Outward sodium and potassium cotransport in human red cells

This paper reports some kinetic properties of Na-K cotransport in human red cells. All fluxes were measured in the presence of 10(-4) M ouabain. We measured Na and K efflux from cells loaded by the PCMBS method to contain different concentrations of these ions into a medium that contained neither Na nor K (MgCl2-sucrose substitution) in the absence and presence of furosemide. Furosemide inhibited 30-60% of the total efflux depending on the internal ion concentration and the individual subject. We took the furosemide-sensitive fluxes to be a measure of Na-K cotransport. The ratio of Na to K cotransport was 1 over the entire range of internal Na and K concentrations studied. When Na was substituted for K as the only internal cation, cotransport was maximally activated when the Na and K concentrations were between 20 and 90 mmol/liter cells. The concentration of internal Na required to produce half-maximal cotransport was about 13 +/- 4 mmol/liter cells (n = 4), while the comparable concentration of K was somewhat lower. The activation curve was definitely sigmoid in character, suggesting that at least two Na ions are involved in the transport process. The maximum of Na-K cotransport was about 0.5 +/- 0.15 mmol/liter cells x hr (n = 5); it had a flat maximum in the medium at about pH 7.0, decreasing in both the acid and alkaline sides. Furosemide-resistant effluxes were found to be linear functions of internal Na and K concentrations and to yield rate coefficients of 0.019 +/- 0.002 hr-1 and 0.014 +/- 0.002 hr-1 (n=7), respectively. These values are of the same order of magnitude expected of ions moving across phospholipid bilayers.