Glucose transporter asymmetries in the bovine blood-brain barrier

The transport of glucose across the mammalian blood-brain barrier is mediat ed by the GLUT1 glucose transporter, which is concentrated in the endotheli al cells of the cerebral microvessels, Several studies supported an asymmet ric distribution of GLUT1 protein between the luminal and abluminal membran es (1:4) with a significant proportion of intracellular transporters. In th is study we investigated the activity and concentration of GLUT1 in isolate d luminal and abluminal membrane fractions of bovine brain endothelial cell s, Glucose transport activity and glucose transporter concentration, as det ermined by cytochalasin B binding, were 2-fold greater in the luminal than in the abluminal membranes. In contrast, Western blot analysis using a rabb it polyclonal antibody raised against the C-terminal 20 amino acids of GLUT 1 indicated a 1:5 luminal:abluminal distribution. Western blot analysis wit h antibodies raised against either the intracellular loop of GLUT1 or the p urified erythrocyte protein exhibited luminal:abluminal ratios of 1:1, A si milar ratio was observed when the luminal and abluminal fractions were expo sed to the 2-N-4[H-3](1-azi-2,2,2,-trifluoroethyl)benzoxyl-1,3-bis-(d-manno s-4-yloxyl)-2-propylamine ([H-3]ATB-BMPA) photoaffinity label. These observ ations suggest that either an additional glucose transporter isoform is pre sent in the luminal membrane of the bovine blood-brain barrier or the C-ter minal epitope of GLUT1 is "masked" in the luminal membrane but not in the a bluminal membranes.