Paul A. Rue

Reduced nuclear triiodothyronine receptors in starvation-induced hypothyroidism

Abstract Starvation of male rats during 48 hours causes a marked reduction of serum thyroxine, serum triiodothyronine, and liver nuclear triiodothyronine content. Liver nuclear receptors capacity for binding of triiodothyronine was reduced, in contrast to thyro-previc hypothyroidism in which binding capacity is normal. DNA-dependent RNA polymerase activities were reduced. In contrast to typical hypothyroidism, serum thyrotropin was low. This form of pituitary non-responsive hypothyroidism may represent a selective response to caloric and/or amino acid deprivation.

T3 receptor occupancy and metabolic responses

The relationship between receptor occupancy and metabolic response was examined in normal rats, hypothyroid rats, and hypothyroid rats treated with various doses of T3 for one week. Serum and nuclear T3 levels were restored to normal by 0.3 — 1 μg T3/day, and increased progressively with higher dosage. Weight gain and liver weight were restored to normal by 0.3 — 3 μg T3/day and were lower with maximal doses of T3. Total nuclear polymerase activity, polymerase I activity, and mRNA content were restored to normal with 0.1 — 1 μg/day and polymerase activity decreased at highest T3 doses. α-Glycerophosphate dehydrogenase activity rose linearly with T3 dose up to the level of 15 /μg T3/day. The response curves for weight gain, liver weight, polymerase activity, and mRNA content indicate a correlation between nuclear receptor occupancy and response within the physiologic dose range, and then a decrement in these parameters as nuclear receptors are saturated, producing a condition that presumably represents hyperthyroidism. The response curve for α-glycerophosphate dehydrogenase activity showed essential linearity until near complete saturation of the receptors was reached. α-Glycerophosphate dehydrogenase activity may represent an effect on a different group of receptors or a different steady state mechanism in comparison to other metabolic parameters measured.

Alterations in hepatic protein kinase activity induced by triiodothyronine

Quantitative and qualitative alterations in protein kinase (PK) activity in preparations of rat liver from hypothyroid (H) and T3-treated hypothyroid animals (T) were analyzed in comparison to epzyme from normal (N) animals. Qualitative variations in type of protein kinase were assessed by chromatography of cytosol preparations on DE-52 cellulose columns. Cytosol kinases resolved into a small fraction I containing a catalytic subunit, fraction II containing a type 1 holoenzyme, fraction III containing a cyclic-AMP independent PK, and fraction IV containing a type 2 holoenzyme. A cytosolic kinase active with casein as substrate was also identified. In H the type 1 holoenzyme was reduced in comparison to N. There were no other qualitative changes. Nuclear PKs were extracted with solvents containing 0.3 M KCl. Qualitative changes were evaluated by chromatography on phosphocellulose columns, but available methodology did not give reproducible evidence of changes in individual PKs. H had significantly more cytosolic PK active with protamine substrate than did normal animals, and by administration of T3, the H level was reduced progressively over 48 h to the N level. The activation ratio of total PK in cytosol was higher in H, and was also reduced to the N level by T3 administration. This suggests a higher steady state level of cyclic-AMP may be present in H rat liver cytosol. Cytosolic protein kinase reactive with casein as substrate increased gradually over 48 h after T3 administration from the H to the higher N level. It was significantly elevated to 110% of the H value by 48 h after T3 administration. The behavior of nuclear PK was entirely different. Nuclear PK reactive with protamine as substrate was increased within 1–1/2 h after T3 administration, reaching a peak of 130% of the control value at 5 h and returning to the normal level by48h. In contrast, nuclear PK reactive with casein as substrate also increased by 1–1/2 h after T3 administration and remained elevated at 110–115% of the control value throughout 48 h after T3 administration. The early changes in nuclear PK activity were prevented by administration of cycloheximide or or-amanitin. The observed changes in cytosolic PK, including the increment induced by T3 in type 1 enzyme, reduction in the activation ratio, reduction in kinase active with protamine, and increase in the amount of enzyme reactive with casein, occurring at 24–48 h, presumably are part of the generalized metabolic response to T3. The early quantitative increment in nuclear PK reactive with protamine and with casein could be associated with the early changes in phosphorylation of nuclear proteins previously identified. Inhibition of the changes by cycloheximide and α-amanitin suggest that the changes are secondary to neosynthesis of PK, which is then translocated to nucleus.