In a recent study we have demonstrated that induced changes in several parameters of one-carbon metabolism in livers of rats fed either a choline deficient diet or treated with low doses of methotrexate may be reversed through restoration of an adequate choline diet and discontinuing of methotrexate (MTX) administration. The present study was aimed at comparing these two treatments for their effect on brain: total folate was 0.69+/-0.35 nmol/g in the control group and was significantly lower in the other two groups (0.25+/-0.06 in the MTX and 0.45+/-0.24 nmol/g in the choline deficient groups, p< 0.01). Both treatments were associated with elongations of the glutamate chains of the folate molecules: the proportion of hepta-and octaglutamyl folates increased from 16% to 31% in the MTX treated and to 33% in the choline deficient group, accompanied with concomitant decreases in the concentrations of tetra-and pentaglutamyl folates. MTX could not be detected in brain tissue in contrast to previous observations of significant amounts found in liver. DNA was significantly (p< 0.05) undermethylated in both treated groups compared to controls. Results are discussed on the basis of the possibility that choline deficiency and MTX treatment appear to impair the capacity of tissues, either a peripheral one as liver or a central one as brain, to incorporate folate.