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A cloned rat thyroid cell line (FRTL-5) was examined for both insulin binding and responsiveness. The characteristics of insulin binding to thyroid cells were similar to those observed in typical insulin target cells. The 125I-insulin binding was time and temperature dependent and Scatchard analysis suggested the presence of two major binding sites with high and low affinity constant (Kd = 1.4 X 10(-10) mol/L and 1.5 X 10(-9) mol/L, respectively). 125I-insulin was also internalized and degraded in a temperature-dependent manner. IGF1 was weakly effective in completing 125I-insulin binding to FRTL-5 cells (57% inhibition at 333 nmol/L), whereas noninsulin-related peptide hormones were ineffective. Exposure of FRTL-5 cells to insulin stimulated both methyl-aminoisobutyric acid (M-AIB) and 2-deoxy-D-glucose (2DG) transport. These effects were evident at 10(-9) mol/L and maximal at 10(-7) mol/L insulin. Maximal stimulation was three- to four-fold over basal value for both M-AIB and 2DG transport. These data suggest that insulin specifically binds to FRTL-5 cells and regulates different biological functions of this thyroid cell line.

Nicotinamide (50mM) prevented insulin-mediated down-regulation of insulin receptors in IM-9 lymphoblastoid cells. Half-maximum effectiveness was between 10 and 33mM. Nicotinamide did not influence insulin binding to the cells, cell viability, insulin degradation or protein synthesis. A variety of inhibitors of ADP-ribosylation reactions besides nicotinamide, most of them pyridine analogues, similarly prevented insulin-induced receptor loss. Spermine decreased the number of insulin receptors in IM-9 cells, but this effect was not inhibited by nicotinamide.

A new sensitive radioimmunoassay method for measuring reverse triiodothyronine (rT3) concentrations in dried blood samples, designed to screen newborn infants for congenital hypothyroidism, has been developed. Paper strips are impregnated with cord blood and dried. Duplicate 5-mm diameter discs are punched from the paper strips and added directly to the radioimmunoassay reaction mixture. After incubation, bound and free hormone are separated by dextran-coated charcoal. The disc remains in the solution throughout the procedure and the assay can be completed within 24 hr. Recovery of rT3 is greater than 95% and coefficients of variation are 9.4% (intraassay) and 12.2% (interassay) at an rT3 concentration of 220 ng/dl. At very low rT3 concentrations (25 ng/dl), coefficients of variation are 14.2% (intraassay) and 18.7% (interassay). The method readily detects 12.5 ng/dl of rT3. With this paper disc method, rT3 was measured in 38 newborns and compared with serum rT3 measured in the same subjects by a standard radioimmunoassay method. The correlation between rT3 values measured in dried blood disc and in serum was very high (r = 0.918). The rT3 in dried blood discs from the cord blood of 745 normal newborns was 228.9 +/- 76.0 ng/dl (mean +/- SD). In contrast, two infants with proven congenital hypothyroidism had rT3 values of 35 and 75 ng/dl, respectively. This study indicates that rT3 can be easily measured in dried blood discs and suggests that the described method may be a useful screening procedure in a program for the detection of neonatal hypothyroidism.

Glucose transport by FRTL-5 cells, a rat thyroid cell line, was found to be TSH dependent. The effect of TSH on the uptake of 2-deoxy-D-glucose, a nonmetabolizable glucose analogue, was prompt, being 200% over basal value after 10 min and maximal after 12 h (600-700% increase). The TSH effect was dose dependent, with half-maximum stimulation at 10 microU TSH/ml, and maximum stimulation at 1 mU TSH/ml. TSH enhanced also the uptake of 3-O-methyl-D-glucose by FRTL-5 cells. The TSH activation of glucose transport had the following characteristics: it was mimicked by (Bu)2-cAMP (1 mM) and by agents that increase cAMP levels in thyroid cells, such as forskolin (10 microM) and cholera toxin (50 micrograms/ml); it involved the facilitated glucose transport system in that it was inhibited in a dose-related manner by both cytochalasin B and phloretin; it showed a glucose stereochemical sensitivity, being affected by D-glucose and 3-O-methyl-glucose, and not by L-glucose; it was characterized by an increase in the maximum velocity (Vmax) of glucose uptake (from 15.3 to 66.0 fmol/min X micrograms DNA) without change in the Michaelis-Menten constant (Km) (5.3 mM); the effect on the Vmax was due to an increase in the number of surface glucose transporters as indicated by the enhancement of the D-glucose-sensitive fraction of [3H]cytochalasin B binding sites that in thyroid plasma membranes of cells exposed to TSH for 2 and 8 h, increased from 5.0 (basal value) to 10.4 and 23.1 pmol/mg protein, respectively. These data indicate that in FRTL-5 cells TSH stimulates the glucose transport system by an enhancement of the number of functional glucose transporters in the thyroid plasma membrane.

Studies were conducted to examine the mechanism by which 'escape' from TSH desensitization of the cyclic AMP response to TSH (Endocrinology 109, 1156, 1981) occurs in confluent cultured thyroid cells. At confluence, cell replication and DNA synthesis are suppressed. An attempt was therefore made to reproduce escape in sparse thyroid cell monolayers using inhibitors of DNA synthesis. The concurrent presence of TSH and mitomycin C (5 micrograms/ml) did not influence the induction of desensitization to TSH after 6 h of stimulation, but cAMP levels then rebounded by 24 h; that is, mitomycin C reproduced escape in sparse cells. Hydroxyurea (10 mM) did not reproduce escape in sparse cells. Adenylate cyclase activity was unaltered in plasma membranes prepared from sparse thyroid cells treated with mitomycin C for 24 h. These data suggest that 'escape' from TSH desensitization is related to events occurring during the cell cycle associated with DNA synthesis, and is caused by an alteration in adenylate cyclase substrate or co-factor availability rather than in enzyme activity itself.

We describe the effect of administration of repeated doses of sodium ipodate in a newborn infant with hyperthyroidism due to transient Graves' disease. Pretreatment (day 3) serum T4 and T3 concentrations were 49 micrograms/dl and 590 ng/dl, respectively. With 24 h after the first dose of ipodate, serum T3 fell by 40%, and it subsequently ranged from 209-278 ng/dl throughout the 39-day ipodate treatment period. Serum T4 also decreased after ipodate administration to 69% and 41% of the pretreatment value after 72 h and 7 days of treatment, respectively; values thereafter during treatment ranged from 19-22 micrograms/dl. These plateau values are in the upper range of normal for the neonatal period. Rapid clinical improvement occurred as the hyperiodothyroninemia abated. Serum rT3 concentrations increased from 468-672 ng/dl to greater than 1400 ng/dl 24 h after each ipodate dose. Thyroid-stimulating immunoglobulin was present in maternal and cord sera, and the half-life of serum thyroid-stimulating immunoglobulin in the infant was approximately 12 days. Antithyroglobulin and antimicrosomal antibodies were present in the infant at 10 days of age, and the titers decreased progressively thereafter; the half-life for the antimicrosomal antibody titer was 3 weeks. The data suggest that sodium ipodate can be useful for treatment of neonatal hyperthyroidism due to Graves' disease.