A complete understanding of the molecular events that account for the transcellular transport of glucose from plasma to the interstitial fluid of brain will include identification of the transporters, their sites of localization, and the mechanisms that regulate their activities. In this competing continuation proposal, the applicant outlines such an investigation of the brain's glucose transporters at the mRNA and protein levels. He will synthesize 12-25 amino acid polypeptides corresponding to the known sequence of the C-terminal and of GLUT3 (a transporter isoform found in brain) and to the deduced sequences of any previous unknown transporters (identified by PCR amplification of a 35-base consensus transporter cDNA sequence derived from a canine brain mRNA preparation). These synthesized polypeptides and identified cDNAs will be used to prepare polyclonal antibodies and labeled oligonucleotides for light and electron microscopic immunocytochemistry/ELISA and in situ hybridization/Northern blots, respectively These reagents will allow (a) determination of the localization, abundance, and subcellular distribution of transporters in hippocampal endothelial, neuronal, and glial cells; and (b) study of the regulation of the transporters by ischemia in the bilateral carotid arterial occlusion model (gerbil) and by hypoglycemia in the insulin osmotic pump model (rat). Finally, the induction of GLUT1 (a transporter isoform found in brain endothelial cells) will be evaluated in vitro by subjecting cultured canine brain endothelial cells to a combination of extracellular matrix proteins and/or astrocyte-conditioned Matrigel. Active extracts will be fractionated by size and chromatographic properties and characterized. It is hoped that research describing the molecular events of blood-brain glucose transport will be valuable in the understanding of a wide range of neurological disorders.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS027229-05
Application #
3413465
Study Section
Neurology A Study Section (NEUA)
Project Start
1989-04-01
Project End
1995-03-31
Budget Start
1993-04-01
Budget End
1994-03-31
Support Year
5
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of Minnesota Twin Cities
Department
Type
Schools of Medicine
DUNS #
168559177
City
Minneapolis
State
MN
Country
United States
Zip Code
55455
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Gerhart, D Z; Enerson, B E; Zhdankina, O Y et al. (1997) Expression of monocarboxylate transporter MCT1 by brain endothelium and glia in adult and suckling rats. Am J Physiol 273:E207-13
Leino, R L; Gerhart, D Z; van Bueren, A M et al. (1997) Ultrastructural localization of GLUT 1 and GLUT 3 glucose transporters in rat brain. J Neurosci Res 49:617-26
Gronlund, K M; Gerhart, D Z; Leino, R L et al. (1996) Chronic seizures increase glucose transporter abundance in rat brain. J Neuropathol Exp Neurol 55:832-40
Borson, N D; Salo, W L; Drewes, L R (1996) Canine brain glucose transporter 3: gene sequence, phylogenetic comparisons and analysis of functional sites. Gene 168:251-6
Gerhart, D Z; Leino, R L; Borson, N D et al. (1995) Localization of glucose transporter GLUT 3 in brain: comparison of rodent and dog using species-specific carboxyl-terminal antisera. Neuroscience 66:237-46
Gerhart, D Z; Leino, R L; Taylor, W E et al. (1994) GLUT1 and GLUT3 gene expression in gerbil brain following brief ischemia: an in situ hybridization study. Brain Res Mol Brain Res 25:313-22
Borson, N D; Salo, W L; Drewes, L R (1993) Application of novel PCR strategies to amplify and sequence glucose transporters in canine brain. Adv Exp Med Biol 331:19-24
Hemmila, J M; Drewes, L R (1993) Glucose transporter (GLUT1) expression by canine brain microvessel endothelial cells in culture: an immunocytochemical study. Adv Exp Med Biol 331:13-8

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