Effect of purine starvation on uptake of Ado and NT1 expression in wild type L. major promastigotes. A. Parasites grown in M199 medium supplemented with 100 µM Xan (early log phase, 4 × 106 cells ml−1) were pelleted, washed, and resuspended (at 0.2–5 × 106 cell ml−1 with samples to be assayed earliest resuspended at higher densities than those assayed later) in medium containing 20% dialyzed fetal bovine serum and supplemented with either 100 µM Ado (squares, 100 µM Ado) or no purine (triangles, 0 µM Ado). Uptake of 1 µM [3H]Ado was measured in triplicate at various time points from 0 to 48 h (cells at ~5 × 106 cell ml−1 at time of uptake assay). The level of induction for adenosine transport at 48 h was ~60-fold for purine-starved compared to purine-replete cells. B. Parasites grown in purine deficient medium for 48 h (4 × 106 cell ml−1) were pelleted, washed, and resuspended as in part A. Uptake of 1 µM [3H]Ado was measured as in part A. The level of decrease for Ado transport at 48 h was ~47-fold for purine-replete versus purine-starved cells. C. Membrane fractions (20 µg) from parasites cultured for 24 h in medium containing 0 or 100 µM Ado were analyzed by immunoblotting. D. Increase in rate of uptake of Ado, Ino, and Ade for purine-starved versus purine-replete cells. Uptake of [3H]Ado, [3H]Ino, and [3H]Ade was measured for parasites cultured for 24 h in the presence of 0 or 100 µM Ado. Values plotted on the y-axis represent the relative uptake (fold increase) of each purine in purine-starved compared to purine-replete parasites.