Retinoic acid treatment of fibroblasts causes a rapid decrease in [3H]inositol uptake

NIH 3T3 fibroblasts treated with all-trans-retinoic acid (RA) showed a dramatic decrease in the uptake of [3H]inositol compared to solvent-treated controls. The onset of RA-induced inhibition of [3H]inositol uptake was rapid with a 10-15% decrease occurring after 2-3 h of RA exposure and 60-70% reduction after 16 h of RA treatment. A progressive dose-dependent decrease in inositol uptake was found as the concentration of RA increased from 10(-8) to 10(-5) M and the effect was fully reversible within 48 h after RA removal. The Vmax and Kt for the controls were 10 nmol/2.5 x 10(6) cells/2 h and 51 microM; and for RA-treated cells the values were 4 nmol/2.5 x 10(6) cells/2 h and 52 microM. The decreased [3H]inositol uptake was not due to a change in the affinity (Kt) of the transporter for the inositol but to a decrease in the Vmax. The maximal effect on inositol uptake was dependent on RA treatment of the cells after they reached saturation density or if made quiescent by serum starvation. RA was the most active of the different retinoids examined in the order RA greater than 13-cis-RA = retinyl acetate greater than all-trans-retinol greater than 5,6-dihydroxyretinoic acid methyl ester greater than N-4-hydroxyphenyl retinamide. In contrast to this effect on inositol, the uptake of fucose, mannose, galactose, and glucose was either not affected or enhanced (for mannose and fucose) by RA treatment. RA inhibition of inositol uptake was also observed in 3T3-Swiss and Balb/3T3 cells but not in two virally transformed 3T3 cell lines. Phlorizin, amiloride, and monensin inhibited inositol uptake by 66, 74, and 58%, respectively, and this inhibition was additive when the cells were treated with RA as well as these inhibitors. A decreased incorporation of [3H]inositol into polyphosphoinositides was also observed in RA-treated cells but not to the same extent as for [3H]inositol uptake. In conclusion, RA treatment of 3T3 fibroblasts decreases the uptake of [3H]inositol by up to 70% within 8 to 10 h at near physiological concentrations in a reversible and specific manner.