LACTATE TRANSPORT IN FRESHLY ISOLATED HUMAN FETAL RETINAL-PIGMENT EPITHELIUM

Purpose. To study transport mechanisms for small monocarboxylic acids in the apical and basolateral membranes of freshly isolated, human fet al retinal pigment epithelium. Methods. The epithelium was mounted in a small Ussing chamber that allowed separate perfusion of both the api cal and basal compartments and simultaneous measurements of intracellu lar pH, transepithelial potential, and tissue resistance. Intracellula r pH was measured using a pH-sensitive dye, 2',7'-bis(2-carboxyethyl)- 5,6-carboxyfluorescein. Results. When 10-100 mM lactate or pyruvate wa s 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 a cidification produced by L-lactate and D-lactate. These rates could be described as Michaelis-Menten functions of the concentrations of lact ate and pyruvate. The K-m 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 administra tion of probenecid (2 mM) or alpha-cyano-4-hydroxycinnamate (2 mM), an d irreversibly reduced by 78% after apical administration of the sulfh ydryl-reagent mersalyl acid (2 mM). The intracellular acidifications c aused by apical pyruvate (50 mM) were completely and reversibly inhibi ted by 50 mM apical L-lactate. Addition of 50 to 100 mM lactate to the basal bath caused intracellular alkalinizations, which could be inhib ited by Na+ removal in the basal bath or by 2 mM alpha-cyano-4-hydroxy cinnamate in the apical bath. Conclusion. These results suggest the pr esence of a proton-lactate cotransport system in the apical membrane a nd a Nac-dependent transport mechanism for the lactate anion in the ba solateral membrane.