Ascorbic acid-dependent GLUT3 inhibition is a critical step for switching neuronal metabolism.

Intracellular ascorbic acid is able to modulate neuronal glucose utilization between resting and activity periods. We have previously demonstrated that intracellular ascorbic acid inhibits deoxyglucose transport in primary cultures of cortical and hippocampal neurons and in HEK293 cells. The same effect was not seen in astrocytes. Since this observation was valid only for cells expressing glucose transporter 3 (GLUT3), we evaluated the importance of this transporter on the inhibitory effect of ascorbic acid on glucose transport. Intracellular ascorbic acid was able to inhibit (3)H-deoxyglucose transport only in astrocytes expressing GLUT3-EGFP. In C6 glioma cells and primary cultures of cortical neurons, which natively express GLUT3, the same inhibitory effect on (3)H-deoxyglucose transport and fluorescent hexose 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxyglucose (2-NBDG) was observed. Finally, knocking down the native expression of GLUT3 in primary cultured neurons and C6 cells using shRNA was sufficient to abolish the ascorbic acid-dependent inhibitory effect on uptake of glucose analogs. Uptake assays using real-time confocal microscopy demonstrated that ascorbic acid effect abrogation on 2-NBDG uptake in cultured neurons. Therefore, ascorbic acid would seem to function as a metabolic switch inhibiting glucose transport in neurons under glutamatergic synaptic activity through direct or indirect inhibition of GLUT3.