title: Ascorbate transport in cultured cat retinal pigment epithelial cells.
abstract: Transport of ascorbate by primary cultures of cat retinal pigment epithelial cells (RPE) was studied. Confluent primary cultures were incubated with 10-500 microM L-[carboxyl-14C] ascorbic acid in balanced salt solution (BSS) at 37 degrees C for 1 to 40 min. The uptake of radioactive ascorbate followed saturation kinetics with a Km of 42 microM and Vmax of 117 pmol min-1 microgram-1 DNA. Cells incubated with 10 microM radioactive ascorbate for 40 min showed a ratio of intracellular to extracellular radioactive ascorbate of greater than 40. The transport of ascorbate was sodium- and energy-dependent. Replacement of 150 mM NaCl in BSS with 150 mM LiCl reduced ascorbate uptake significantly. Ouabain, 2,4-dinitrophenol, alpha-D-glucose, 3-O-methyl-D-glucose, and the ascorbate analogues, D-isoascorbate and dehydroascorbate, each inhibited ascorbate uptake into RPE cells. The efflux of radioactivity into the incubation media was slow when cells were preloaded with either 50- or 500 microM radioactive ascorbate, but increased when cells preloaded with 50 microM ascorbate were incubated in the presence of excess non-radioactive ascorbate. These studies demonstrated that a sodium-dependent carrier system is involved in transport of ascorbate in primary cultures of cat RPE.
mesh_terms: D019325:3-O-Methylglucose; D000818:Animals; D001205:Ascorbic Acid / Q000378:metabolism*; D001693:Biological Transport, Active / Q000187:drug effects; D002250:Carbon Radioisotopes; D002415:Cats; D002478:Cells, Cultured; D004140:Dinitrophenols / Q000494:pharmacology; D005947:Glucose / Q000494:pharmacology; D007700:Kinetics; D008757:Methylglucosides / Q000494:pharmacology; D010042:Ouabain / Q000494:pharmacology; D010857:Pigment Epithelium of Eye / Q000187:drug effects / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism; D013997:Time Factors
chemical_list: D002250:Carbon Radioisotopes; D004140:Dinitrophenols; D008757:Methylglucosides; D019325:3-O-Methylglucose; D010042:Ouabain; D012964:Sodium; D005947:Glucose; D001205:Ascorbic Acid