Thiamine deficiency results in downregulation of the GLAST glutamate transporter in cultured astrocytes

Alan S. Hazell, Pierre Pannunzio, K. V.Rama Rao, David V. Pow, Andrea Rambaldi

Research output: Contribution to journalArticlepeer-review

41 Scopus citations

Abstract

Pyrithiamine-induced thiamine deficiency (TD) is a well-established model of Wernicke's encephalopathy in which a glutamate-mediated excitotoxic mechanism may play an important role in determining selective vulnerability. In order to examine this possibility, cultured astrocytes were exposed to TD and effects on glutamate transport and metabolic function were studied. TD led to decreases in cellular levels of thiamine and thiamine diphosphate (TDP) after 24 h of treatment and decreased activities of the TDP-dependent enzymes α-ketoglutarate dehydrogenase and transketolase after 4 and 7 days, respectively. TD treatment for 10 days led to a reversible decrease in the uptake of [3H]-D-aspartate, a nonmetabolizable analogue of glutamate. Kinetic analysis revealed that the uptake inhibition was caused by a 47% decrease in the Vmax for uptake of [3H]-D-aspartate, with no change in the Km value. Immunoblotting showed that this decrease in uptake was due to an 81% downregulation of the astrocyte-specific GLAST glutamate transporter. Loss of uptake activity and GLAST protein were blocked by treatment with the protein kinase C inhibitor H7, while exposure to DCG IV, a group II metabotropic glutamate receptor (mGluR) agonist, resulted in improvement of [3H]-D-aspartate uptake and a partial reversal of transporter downregulation. These results are consistent with our recent in vivo findings of a loss of astrocytic glutamate transporters in TD and provide evidence that TD conditions may increase phosphorylation of GLAST, contributing to its downregulation. In addition, manipulation of group II mGluR activity may provide an important strategy in the treatment of this disorder.

Original languageEnglish (US)
Pages (from-to)175-184
Number of pages10
JournalGLIA
Volume43
Issue number2
DOIs
StatePublished - Aug 1 2003
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Neurology
  • Cellular and Molecular Neuroscience

Keywords

  • Cell culture
  • Excitotoxicity
  • Glutamate transporter
  • Metabotropic receptor
  • Pyrithiamine

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