Mg2+ transport in sheep rumen epithelium: evidence for an electrodiffusive uptake mechanism.

The potential difference (PD)-dependent component of transcellular Mg2+uptake in sheep rumen epithelium was studied. Unidirectional28Mg2+fluxes were measured at various transepithelial PD values, and the unidirectional mucosal-to-serosal28Mg2+flux ([Formula: see text]) was correlated with the PD across the apical membrane (PDa) determined by mucosal impalement with microelectrodes. PDa was found to be -54 ± 5 mV, and [Formula: see text] was 65.9 ± 13.8 nmol ⋅ cm-2 ⋅ h-1under short-circuit conditions. Hyperpolarization of the ruminal epithelium (blood-side positive) depolarized PDa and, most noticeably, decreased [Formula: see text]. Further experiments were performed with cultured ruminal epithelial cells (REC). With the aid of the fluorescence probe mag-fura 2, we measured the intracellular free Mg2+ concentration ([Mg2+]i) of isolated REC under basal conditions at various extracellular Mg2+ concentrations ([Mg2+]e) and after alterations of the transmembrane voltage. Basal [Mg2+]iwas 0.54 ± 0.08 mM. REC suspended in media with [Mg2+]ebetween 0.5 and 7.5 mM showed an increase in [Mg2+]ithat was dependent on [Mg2+]eand that exhibited a saturable component (Michaelis-Menten constant = 1.2 mM; maximum [Mg2+]i= 1.26 mM). Membrane depolarization with high extracellular K+ (40, 80, or 140 mM K+) and the K+ channel blocker quinidine (50 and 100 μM ) resulted in a decrease in [Mg2+]i. On the other hand, hyperpolarization created by K+ diffusion (intracellular K+ concentration > extracellular K+ concentration) in the presence of valinomycin induced a 15% increase in [Mg2+]i. None of the manipulations had any effect on intracellular Ca2+ concentration and intracellular pH. The results support the assumption that the membrane potential acts as a principal driving force for Mg2+ entry in REC and suggest that the pathway for this electrodiffusive Mg2+ uptake across the luminal membrane is a channel or a carrier.