L-DOPA transport properties in an immortalised cell line of rat capillary cerebral endothelial cells, RBE 4

Abstract

The present study aimed to determine the kinetics of l-3,4-dihydroxyphenylalanine ( l-DOPA) uptake in an immortalised cell line of rat capillary cerebral endothelial cells (clones RBE 4 and RBE 4B), to define the type of inhibition produced by l-5-hydroxytryptophan ( l-5-HTP), 2-aminobicyclo(2,2,1)-heptane-2-carboxylic acid (BHC) and N-(methylamino)-isobutyric acid (MeAlB) and its sodium dependence. Non-linear analysis of the saturation curves for l-DOPA and l-5-HTP revealed in RBE 4 cells K m values (in μM) of 72 and 102 and in RBE 4B cells K m values (in μM) of 60 and 118, respectively. IC 50 values for l-5-HTP (RBE 4, 1026 μM; RBE 4B, 831 μM) obtained in the presence of a nearly saturating (250 μM) concentration of l-DOPA were almost 5-fold those obtained when non-saturating (25 μM) concentrations of l-DOPA were used. IC 50 values for BHC obtained in the presence of a nearly saturating (250 μM) concentration of l-DOPA were also 6- to 5-fold those obtained when non-saturating (25 μM) concentrations of l-DOPA were used. MeAlB (up to 2.5 mM) was found not to interfere with the uptake of l-DOPA. In RBE 4 cells, V max values for l-DOPA uptake were identical in the absence and the presence of 150 μM l-5-HTP or 150 μM BHC, but K m values (μM) were significantly greater ( P<0.05) when l-DOPA uptake was studied in the presence of l-5-HTP or BHC. Similar findings were observed when RBE 4B cells were used. Uptake of (250 μM) l-DOPA in the absence of sodium in the incubation medium was similar to that observed in the presence of increasing concentrations of sodium (20 to 140 mM). It is concluded that RBE 4 and RBE 4B cells are endowed with the L-type amino acid transporter through which l-DOPA and l-5-HTP can be taken up, and suggested that this immortalised cell line of rat capillary cerebral endothelium might constitute an interesting in vitro model for the study of BBB mechanisms, namely those concerning solute and nutrient transfer across the brain capillary endothelium.