Intracellular Mg2+ and magnesium depletion in isolated renal thick ascending limb cells.

Abstract

Magnesium reabsorption and regulation within the kidney oc- cur principally within the cortical thick ascending limb (cTAL) cells of the loop of Henle. Fluorometry with the dye, mag-fura- 2, was used to characterize intracellular Mg2" concentration (IMg2j11) in single cTAL cells. Primary cell cultures were pre- pared from porcine kidneys using a double antibody technique (goat anti-human Tamm-Horsfall and rabbit anti-goat IgG an- tibodies). Basal [Ig2+"] was 0.520.02 mM, which was -2% of the total cellular Mg. Cells cultured (16 h) in high magne- sium media (5 mM) maintained basal [Mg2j1i, 0.480.02, in the normal range. However, cells cultured in nominally magnesium-free media possessed [Mg2J1j, 0.270.01 mM, which was associated with a significant increase in net Mg transport, (con- trol, 0.190.03 and low Mg, 0.350.01 nmol. mg-' pro- tein min-') as assessed by 'Mg uptake. Mg2'-depleted cells were subsequently placed in high Mg solution (5 mM) and the Mg2" refill rate was assessed by fluorescence. [Mg2"1 returned to normal basal levels, 0.530.03 mM, with a refill rate of 25737 nM/s. Mg2" entry was not changed by 5.0 mM Ca2" or 2 mM Sr2+, Cd2+, Co2+, nor Ba2+ but was inhibited by Mn2+ = La3+ .,Gd3+ Ni2+ , Zn2+ Be2+ at 2 mM. Intracellular Ca2`and "Ca uptake was not altered by Mg depletion or Mg2+ refill, indicating that the entry is relatively specific to Mg2e.