Effect of tryptophan on the early tri-iodothyronine uptake in mouse thymocytes

Objective: We have studied the effect of tryptophan on cellular [I-125]tri-iodothyronine (T3) uptake by mouse thymocytes. Materials and methods: Mouse thymocytes (20 x 10(6) cells/ml) were suspended in Krebs-Ringer solution buffered by Tris-HCl and incubation (23 degrees C at pH 7.45 +/- 0.6), in the presence or absence of 1 mM tryptophan, was started by adding 25 pM [I-125]T3. At th, end of incubation, samples were cooled in ice, centrifuged over a 30% sucrose cushion and the cell-associated radioactivity was measured in the pellet. Results: Tryptophan reduced both the total and the saturable fraction of [I-125]T3 uptake by 44% (P = 0.0009) and 60% (P = 0.0006) respectively, following 1 min of incubation. This effect was specific and dose-dependent, being maximal at 5 mM concentration (-82%). In contrast, the preexposure of cells to tryptophan for up to 2 h had no effect on the subsequent uptake of [I-125]T3, in the absence of tryptophan. The effect of D-tryptophan on saturable T3 uptake was not different from that obtained using the L-stereoisomer, Tryptophan reduced the V-max of the initial rate of saturable [I-125]T3 uptake by two-thirds without affecting the apparent K-m (2.2 nM) of the process, thus indicating the non-competitive nature of the inhibition. In sodium-free medium the saturable [I-125]T3 uptake was reduced by 43%. The inhibitory effect of tryptophan on [I-125]T3 uptake was exerted in both the presence and the absence of sodium. In fact, the inhibitory effect of tryptophan on T3 transport was greater and significantly different (P = 0.0046) from that obtained by sodium depletion alone. Conclusions: Tryptophan interferes with both the sodium-dependent and -independent components of [I-125]T3 uptake by a dose-dependent, non-competitive mechanism which operates in cis-modality at the plasma membrane level of mouse thymocytes.