Substrate regulation of ascorbate transport activity in astrocytes

Astrocytes possess a concentrative l-ascorbate (vitamin C) uptake mechanism involving a Na⁺-dependent l-ascorbate transporter located in the plasma membrane. The present experiments examined the effects of deprivation and supplementation of extracellular l-ascorbate on the activity of this transport system. Initial rates of l-ascorbate uptake were measured by incubating primary cultures of rat astrocytes with l-[¹⁴C]ascorbate for 1 min at 37°C. We observed that the apparent maximal rate of uptake (V_max) increased rapidly (<1 h) when cultured cells were deprived of l-ascorbate. In contrast, there was no change in the apparent affinity of the transport system for l-[¹⁴C]ascorbate. The increase in V_max was reversed by addition of l-ascorbate, but not D-isoascorbate, to the medium. The effects of external ascorbate on ascorbate transport activity were specific in that preincubation of cultures with l-ascorbate did not affect uptake of 2-deoxy-D-[³H(G)]glucose. We conclude that the astroglial ascorbate transport system is modulated by changes in substrate availability. Regulation of transport activity may play a role in intracellular ascorbate homeostasis by compensating for regional differences and temporal fluctuations in external ascorbate levels.