ASCORBATE TRANSPORT AND INTRACELLULAR CONCENTRATION IN CEREBRAL ASTROCYTES

Regulation of the initial rate of uptake and steady-state concentratio n of ascorbate (reduced vitamin C) was investigated in rat cerebral as trocytes. Although these cells did not synthesize vitamin C, they accu mulated millimolar concentrations of ascorbate when incubated with med ium containing the vitamin at a level (200 mu M) typical of brain extr acellular fluid. Initial rate of [C-14]- ascorbate uptake and intracel lular ascorbate concentration were dependent on extracellular Na+ and sensitive to the anion transport inhibitor sulfinpyrazone. Comparison of the efflux profiles of ascorbate and 2',7'-bis(carboxyethyl)-5 (or -6)-carboxyfluorescein from astrocytes permeabilized with digitonin lo calized most intracellular ascorbate to the cytosol. Pretreatment of a strocytes with dibutyryl cyclic AMP (dBcAMP) doubled their initial rat e of sulfinpyrazone-sensitive [C-14]ascorbate uptake compared with cel ls treated with either n-butyric acid or vehicIe. dBcAMP also increase d steady-state intracellular ascorbate concentration by 39%. The relat ively small size of the change in astrocytic ascorbate concentration w as explained by the finding that dBcAMP increased the rate of efflux o f the vitamin from ascorbate-loaded cells. These results indicate that uptake and efflux pathways are stimulated by cyclic AMP-dependent mec hanisms and that they regulate the cytosolic concentration of ascorbat e in astrocytes.