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Modulation of human glutamate transporter activity by phorbol ester
1Department of Pharmacology, Texas Tech University Health Sciences Center, Lubbock 79430-0001, USA.
Journal of Neurochemistry
|March 7, 1998
Summary
This study identifies a specific glutamate transporter in Y-79 cells, showing protein kinase C activation inhibits glutamate uptake by reducing transporter affinity. This highlights a potential mechanism for regulating synaptic glutamate levels.
Area of Science:
- Neuroscience
- Molecular Biology
- Cell Biology
Background:
- Synaptic glutamate transmission relies on efficient glutamate removal by high-affinity transporters.
- Five subtypes of Na+-dependent mammalian glutamate transporters are known.
Purpose of the Study:
- To characterize the glutamate uptake system in Y-79 human retinoblastoma cells.
- To investigate the role of protein kinase C (PKC) in modulating glutamate transporter activity.
Main Methods:
- Pharmacological characterization of glutamate uptake kinetics in Y-79 cells.
- Assessing the effect of PKC activator (phorbol 12-myristate 13-acetate) on glutamate uptake.
- Western blot analysis to confirm transporter subtype.
Main Results:
- Y-79 cells exhibit a single Na+-dependent glutamate uptake system with a Km of 1.7 microM, sensitive to dihydrokainate and DL-threo-beta-hydroxyaspartate.
- PKC activation by phorbol 12-myristate 13-acetate inhibited glutamate uptake by decreasing transporter affinity (fivefold) without altering Vmax.
- This inhibition was reversed by staurosporine and did not affect glycine uptake.
Conclusions:
- The identified transporter is the dihydrokainate-sensitive EAAT2/GLT-1 subtype.
- PKC modulation of glutamate transporter activity offers a mechanism for regulating extracellular glutamate levels and synaptic responses.