Abstract:The transport of glucose across the blood-brain barrier (BBB) is mediated by the high molecular mass (55-kDa) isoform of the GLUT1 glucose transporter protein. In this study we have utilized the tritiated, impermeant photolabel 2-N-[4-(1-azi-2,2,2-trifluoroethyl)[2-3 H]propyl]-1,3-bis(D-mannose-4-yloxy)-2-propylamine to develop a technique to specifically measure the concentration of GLUT1 glucose transporters on the luminal surface of the endothelial cells of the BBB. We have combined this methodology with measurements of BBB glucose transport and immunoblot analysis of isolated brain microvessels for labeled luminal GLUT1 and total GLUT1 to reevaluate the effects of chronic hypoglycemia and diabetic hyperglycemia on transendothelial glucose transport in the rat. Hypoglycemia was induced with continuous-release insulin pellets (6 U/day) for a 12-to 14-day duration; diabetes was induced by streptozotocin (65 mg/kg i.p.) for a 14-to 21-day duration. Hypoglycemia resulted in 25-45% increases in regional BBB permeability-surface area (PA) values for D-[ 14 C]glucose uptake, when measured at identical glucose concentration using the in situ brain perfusion technique. Similarily, there was a 23 Ϯ 4% increase in total GLUT1/mg of microvessel protein and a 52 Ϯ 13% increase in luminal GLUT1 in hypoglycemic animals, suggesting that both increased GLUT1 synthesis and a redistribution to favor luminal transporters account for the enhanced uptake. A corresponding (twofold) increase in cortical GLUT1 mRNA was observed by in situ hybridization. In contrast, no significant changes were observed in regional brain glucose uptake PA, total microvessel 55-kDa GLUT1, or luminal GLUT1 concentrations in hyperglycemic rats. There was, however, a 30 -40% increase in total cortical GLUT1 mRNA expression, with a 96% increase in the microvessels. Neither condition altered the levels of GLUT3 mRNA or protein expression. These results show that hypoglycemia, but not hyperglycemia, alters glucose transport activity at the BBB and that these changes in transport activity result from both an overall increase in total BBB GLUT1 and an increased transporter concentration at the luminal surface.

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The pharmacokinetics, cerebrovascular permeability, and tissue distribution of the neurotoxic plasticizer N-butylbenzenesulfonamide (NBBS) were determined in rats. A stable isotope-labeled form ([(13)C(6)]NBBS) was used to circumvent ubiquitous contamination that was evident whenever the native form was measured. Plasticizer decline in plasma, following an iv dose of 1 mg/kg, was described by a triexponential decay function. NBBS was cleared from plasma at a rate of 25 ml/min/kg, and 24 h after administration, plasma concentrations represented 0.04% of the administered dose. These data suggest rapid elimination and uptake into tissue; however, NBBS was not accumulated by any of the tissues studied (i.e., liver, kidney, muscle, adipose tissue, and brain). Given the critical interest in NBBS neurotoxicity, the brain uptake of [(13)C(6)]NBBS was further explored in experiments using the in situ brain perfusion technique. During perfusion with protein-free saline for 15-30 s, the single-pass brain extraction for free [(13)C(6)]NBBS was very high (73-100%) with a unidirectional blood-brain barrier transfer constant (K(in)) of > 0.08 ml/s/g. No significant differences were found in [(13)C(6)]NBBS content among the measured brain regions. Plasma protein binding (70%) only slightly lowered the single-pass brain extraction to 48%. In summary, the results demonstrate that NBBS distributes rapidly to tissues, including brain. Though highly lipophilic with a Log octanol/water partition coefficient of 2.17 +/- 0.09, brain:blood ratios (2:1) for NBBS were consistent throughout the experimental duration, with little indication of accumulation.