Kinetic properties and Na+ dependence of rheogenic Na(+)‐HCO3‐ co‐transport in frog retinal pigment epithelium.

Publisher Website
Google Scholar
Abstract
1. Na(+)‐HCO3‐ co‐transport across the retinal membrane of the frog retinal pigment epithelium was studied by means of double‐barrelled pH‐selective microelectrodes. Transient changes in the intracellular pH were monitored in response to abrupt changes in the Na+ concentration on the retinal side of the epithelium. 2. The experiments were performed as follows. The Na(+)‐HCO3‐ co‐transport was inhibited by perfusing the retinal side of the epithelium with a Na(+)‐free solution. The co‐transport was then stimulated by changing the perfusate from the Na(+)‐free solution to a solution which contained from 5 to 110 mM‐Na+. The resulting inward Na(+)‐HCO3‐ co‐transport produced an intracellular alkalinization, the initial rate of which was used to calculate the initial rate of Na(+)‐HCO3‐ co‐transport, JHCO3‐. 3. The Na+ dependence of the Na(+)‐HCO3‐ co‐transport was studied at two different values of extracellular pH (7.40 and 7.10), at constant extracellular HCO3‐ concentration (27.5 mM) and at two different extracellular HCO3‐ concentrations (27.5 mM and 55 mM) at constant extracellular pH (7.40). In these experiments, the calculated values of JHCO3‐ followed single Michaelis‐Menten kinetics with respect to the extracellular Na+ concentration. 4. The data are consistent with a model in which the co‐transporter has a single binding site for the Na+ ion with an apparent affinity constant (apparent Km) of 37 mM. The apparent affinity constant for Na+ was independent of the extracellular concentration of CO3(2‐) in the range of 16‐65 microM, and of the extracellular HCO3‐ concentration in the range 27.5‐55 mM. 5. The NaCO3‐ ion‐pair hypothesis, in which sodium binds to the co‐transporter and is translocated across the cell membrane as the NaCO3‐ ion pair, was analysed. For stoichiometries 1:2 and 1:3 of the Na(+)‐HCO3‐ co‐transport, the NaCO3‐ ion‐pair hypothesis was found incompatible with the data. 6. The intracellular buffer capacity as measured by the CO2 method was 15 mM.

This publication has 21 references indexed in Scilit: