No abstract

Glutamate is involved in gene expression regulation in neurons and glial cells through the activation of a diverse array of signaling cascades. In Bergmann glia, Ca2+ -permeable alpha-hydroxy-5-methyl-4-isoazole-propionic acid (AMPA) receptors become tyrosine phosphorylated after ligand binding and by these means form multiprotein signaling complexes. Of the various proteins that associate to these receptors, the phosphatidylinositol 3-kinase (PI-3K) deserves special attention since D3-phosphorylated phosphoinositides are docking molecules for signaling proteins with a pleckstrin homology domain. In order to characterize the role of PI-3K in AMPA receptors signaling, in the present report we analyze the involvement of the serine/threonine protein kinase B in this process. Our results demonstrate an augmentation in protein kinase B phosphorylation and activity after glutamate exposure. Interestingly, the effect is independent of Ca2+ influx, but sensitive to Src blockers. Our present findings broaden our current knowledge of glial glutamate receptors signaling and their involvement glutamatergic neurotransmission.

Appropriate removal of L: -glutamate from the synaptic cleft is important for prevention of the excitotoxic effects of this neurotransmitter. The Na+-dependent glutamate/aspartate transporter GLAST is regulated in the short term, by a transporter-dependent decrease in uptake activity while in the long term, a receptor's-dependent decrease in GLAST protein levels leads to a severe reduction in glutamate uptake. The promoter region of the mouse glast gene harbors an Activator Protein-1 site (AP-1). To gain insight into the molecular mechanisms triggered by Glu-receptors activation involved in GLAST regulation, we took advantage of the neonatal mouse cerebellar prisms model. We characterized the glutamate uptake activity; the glutamate-dependent effect on GLAST protein levels and over the interaction of nuclear proteins with a mouse glast promoter AP-1 probe. A time and dose dependent decrease in transporter activity matching with a decrease in GLAST levels was recorded upon glutamate treatment. Moreover, a significant increase in glast AP-1 DNA binding was found. Pharmacological experiments established that both effects are mediated through alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptors, favoring the notion of the critical involvement of glutamate in the regulation of its binding partners: receptors and transporters.

The regulation of the Na+-dependent glutamate/aspartate transporter system GLAST expressed in rat and mouse cerebellar and cortical astrocytic cultures was examined. Pretreatment of the cerebellar cells with L-glutamate and 12-O-tetradecanoyl-phorbol-13-acetate (TPA), a known Ca2+/diacylglicerol-dependent protein kinase (PKC) activator, produced a decrease in [3H]-D-aspartate uptake. This reduction was dose- and time-dependent and sensitive to PKC inhibitors. Furthermore, the L-glutamate-dependent [3H]-D-aspartate uptake decrease is a non-receptor dependent process, because neither of the agonists or antagonists were effective in mimicking or reverting the effect. Interestingly, transportable substrates could reproduce the L-glutamate effect. In sharp contrast, in cortical astrocytes, both L-glutamate and TPA pre-exposure result in an augmentation of the [3H]-D-aspartate uptake. These findings suggest that the Na+-dependent glutamate uptake GLAST undergoes a region-specific regulation.