UPTAKE OF THYROXINE IN CULTURED ANTERIOR-PITUITARY-CELLS OF EUTHYROID RATS

The uptake of [I-125]T-4 was investigated in cultured anterior pituitary cells isolated from adult fed Wister rats and cultured for 3 days in medium containing 10% fetal calf serum. Experiments were performed with [I-125]T-4 (10(5) to 2 x 10(6) cpm; 0.35-7 nM) in medium containing 0.5% or 0.1% BSA. The uptake of [I-125]T-4 increased with time and showed equilibrium after around 1 h of incubation. The presence of 10 mu M unlabeled T-4 during incubation decreased the uptake of [I-125]T-4 by 65-70% at all time intervals. After 24 h of incubation, 1.5% iodide and 3.2% conjugates were detected in the medium, whereas around 20% of cellular radioactivity represented [I-125]T-3. The 15-min uptake of [I-125]T-4 was significantly reduced by simultaneous incubation with 100 nM T-4 (by 24%; P < 0.05), 100 nM T-3 (by 38%; P < 0.001), or 10 mu M rT(3) (by 32%; P < 0.001), whereas 10 mu M tetraiodothyroacetic acid (Tetrac) had no effect. Furthermore, preincubation (30 min) and incubation (15 min) with 10 mu M monodansylcadaverine, oligomycin, or monensin reduced the uptake of [I-125]T-4 by 30%, 50%, and 40%, respectively (all P < 0.001). Substitution of Na+ in the buffer by K+ diminished the uptake of [I-125]T-4 by 39% (P < 0.005); 2 mM phenylalanine, tyrosine, or tryptophan reduced [I-125]T-4 uptake by 18% (P < 0.05), 18% (P = NS), and 33% (P < 0.005), respectively. Our data suggest that the pituitary contains a specific carrier-mediated energy-requiring mechanism for [I-125]T-4 uptake that is partly dependent on the Na+ gradient. In addition, part of [I-125]T-4 uptake in the pituitary might occur through an amino acid transport system. When expressed per pM of free hormone, the 15-min uptake of [(I-125]T-4 was approximately as high as that of [I-125]T-3. Because the reduction of [I-125]T-4 uptake by T-4, T-3, monodansylcadaverine, oligomycin, and monensin was roughly the same as the previously reported reduction of [I-125]T-3 uptake by the same compounds, it is further suggested that T-4 and T-3 Share a common carrier in cultured anterior pituitary cells.