Relation of plasma thyroxine binding to thyroidal activity and determination of thyroxine binding proteins in a turtle, Pseudemys scripta

Abstract

The ability of plasma to bind thyroxine (T 4) was examined in the turtle, Pseudemys scripta, in relation to variations in thyroidal state associated with age, sex, environment, and surgical and chemical manipulations. Relative plasma binding activity was assessed by use of binding to [ 125I]T 4 on minicolumns of Sephadex G-25 (fine). Hypothyroidism induced by surgical thyroidectomy (Tx) or goitrogen (Methimazole) treatment resulted in a marked depression of plasma binding (50- to 100-fold) in juveniles, and T 4 treatment restored binding after 4–6 weeks in long-term Tx animals and increased levels in intact animals. Among intact turtles or those made slightly hypothyroid by partial thyroidectomy, binding was consistently correlated with plasma T 4. For example, juvenile turtles kept under continuous light and constant temperature (28°) for 4.5 months showed a pronounced depression of plasma T 4 (2.6 ± 1.1 ng/ml) and binding capacity compared to animals raised under variable conditions (T 4 = 69.6 ± 22 ng/ml) for the last 2 months. Plasma T 3 was < 1 ng/ml in all cases. Binding levels in adult turtles were similar to juveniles, but females had significantly higher binding levels than males which paralleled differences in their plasma T 4 (137 ± 17.4 vs 83.9 ± 13.8 ng/ml). These variations in binding were independent of total plasma protein and albumin. Plasma T 4 binding measured on Sephadex G-25 was reversible and reduced by addition of exogenous T 4. The affinity for T 3 was 10- to 100-fold less than for T 4. When plasma preincubated with [ 125I]T 4 was electrophoresed on polyacrylamide slab gels (7% nonreducing) only a small percentage of radiolabel was associated with albumin and the majority with a slower migrating protein(s). Addition of unlabeled T 4 displaced binding from the slower migrating region to the albumin and dye front (unbound). In contrast, plasma from Tx turtles showed only minimal binding and radiolabel was associated primarily with the albumin fraction. Elution of proteins from gels confirmed that only the slower migrating components bound T 4 when tested on Sephadex G-25, and Tx animals lacked this binding component. The protein(s) responsible for most of the T 4 binding appears to exist in low concentration. Limited comparative studies with human blood showed a similar binding activity on Sephadex G-25, but electrophoretic mobilities of binding proteins were distinct from those in the turtle. Evidence suggests that this binding protein is not prealbumin. Thus, contrary to prevailing views for ectotherms, the levels of total plasma T 4 in this turtle are higher than those typical of mammals and the turtle may possess a unique T 4 binding protein that is functionally analogous to thyroxine binding globulins in mammals.