Characterization of the Na -dependent Mg transport in sheep ruminal epithelial cells

Schweigel, Monika, Hi-Sung Park, Benjamin Etschmann, and Holger Martens. Characterization of the Na -dependent Mg transport in sheep ruminal epithelial cells. Am J Physiol Gastrointest Liver Physiol 290: G56–G65, 2006. First published August 18, 2005; doi:10.1152/ajpgi.00014.2005.—This study examines the routes by which Mg leaves cultured ovine ruminal epithelial cells (REC). Mg -loaded (6 mM) REC were incubated in completely Mg -free solutions with varying Na concentrations, and the Mg extrusion rate was calculated from the increase of the Mg concentration in the incubation medium determined with the aid of the fluorescent probe mag-fura 2 (Na salt). In other experiments, REC were also studied for the intracellular free Mg concentration ([Mg ]i; using magfura 2), the intracellular Na concentration (using Na -binding benzofuran isophthalate), the intracellular cAMP concentration ([cAMP]i; using an enzyme-linked immunoassay), and Na /Mg exchanger existence [using a monoclonal antibody (mAb) raised against the porcine red blood cell Na /Mg exchanger]. Mg -loaded REC show a Mg efflux that was strictly dependent on extracellular Na . The Mg extrusion rate increased from 0.018 0.009 in a Na -free medium to 0.73 0.3 mM l cells 1 min 1 in a 145 mM Na medium and relates to extracellular Na concentration ([Na ]e) according to a typical saturation kinetic (Km value for [Na ]e 24 mM; maximal velocity 11 mM l cells 1 min ). Mg efflux was reduced by imipramine (48%) and increased after application of dibutyryl-cAMP (55%) or PGE2 (17%). These effects are completely abolished in Na -free media. Furthermore, an elevation of [cAMP]i led to an [Mg ]i decrease that amounted to 375 105 M. The anti-Na /Mg exchanger mAb inhibits Mg extrusion; moreover, it detects a specific 70-kDa immunoreactive band in protein lysates of ovine REC. The data clearly demonstrate that a Na /Mg exchanger is existent in the cell membrane of REC. The transport protein is the main pathway (97%) for Mg extrusion and can be assumed to play a considerable role in the process of Mg absorption as well as the maintenance of the cellular Mg homeodynamics.