Fig. 3.

Transport of [³H]-uridine by L. mexicana promastigotes. (A) Transport of 0.25 μM [³H]-uridine was linear (r² = 0.992) over 120 s with a rate of 0.0043 ± 0.0002 pmol(10⁷ cells⁾⁻¹s⁻¹ (●). In the presence of 5 mM uridine (□) transport was not significantly different from zero (P = 0.43). (B) Transport of 0.25 μM [³H]-uridine was dose-dependently inhibited by unlabeled uridine (●), resulting in an apparently mono-phasic sigmoid curve that could be converted to a Michaelis-Menten saturation plot (inset) to determine K_m and V_max values. Transport was inhibited by 48% by up to 2.5 mM uracil (□), and ∼100% by adenosine (▲). (C) Transport of 0.25 μM [³H]-uridine was determined in the presence (●) or absence (□,▲) of 1 mM unlabeled uracil in order to inhibit the uracil-sensitive component of uridine uptake. Inhibitors shown are uracil (▲) and thymidine (□,●). The dotted line indicates the level of uridine uptake in the presence of 1 mM uracil with zero thymidine added. (D) Transport of 0.25 μM [³H]-uridine in the presence of 1 mM uracil was dose-dependently inhibited by adenosine (■), uridine (□) and cytidine (●). The uridine inhibition data was converted to a Michaelis-Menten saturation plot (inset). The level of [³H]-uridine in the absence of uracil or other inhibitor is also indicated (▲). Unit for transport was pmol(10⁷ cells)⁻¹ for frame A and pmol(10⁷ cells)⁻¹s⁻¹ in frames B-D; symbols represent the average of triplicate determinations in a single representative experiment, and error bars represent SEM.