Lactate transport in freshly isolated human fetal retinal pigment epithelium

To study transport mechanisms for small monocarboxylic acids in the apical and basolateral membranes of freshly isolated, human fetal retinal pigment epithelium.

The epithelium was mounted in a small Ussing chamber that allowed separate perfusion of both the apical and basal compartments and simultaneous measurements of intracellular pH, transepithelial potential, and tissue resistance. Intracellular pH was measured using a pH-sensitive dye, 2',7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein.

When 10-100 mM lactate or pyruvate was added to the apical bath the cells acidified by 0.10-0.25 pH units. There were no differences between the initial rates of intracellular acidification produced by L-lactate and D-lactate. These rates could be described as Michaelis-Menten functions of the concentrations of lactate and pyruvate. The Km values were 42 +/- 12 mM for L-lactate and 34 +/- 8 mM for pyruvate. The rates of acidification caused by 50 mM L-lactate were reversibly reduced by 44% or 35% after apical administration of probenecid (2 mM) or alpha-cyano-4-hydroxycinnamate (2 mM), and irreversibly reduced by 78% after apical administration of the sulfhydryl-reagent mersalyl acid (2 mM). The intracellular acidifications caused by apical pyruvate (50 mM) were completely and reversibly inhibited by 50 mM apical L-lactate. Addition of 50 to 100 mM lactate to the basal bath caused intracellular alkalinizations, which could be inhibited by Na+ removal in the basal bath or by 2 mM alpha-cyano-4-hydroxycinnamate in the apical bath.