In Vitro Demonstration of Putative Nuclear 3,5,3'-Triiodothyronine Receptors in Isolated Liver Nuclei of Singi Fish, Heteropneustes Fossilis (Bloch)

Summary

Putative thyroid hormone (TH) receptors have been demonstrated in the isolated liver nuclei of Singi fish, Heteropneustes fossilis (Bloch), and their binding characteristics have been examined. Nuclear T₃ saturation analyses were carried out in vitro at 27 °C in a sucrose-Tris-HCl buffer (pH 7.5) containing calcium (2 mM), magnesium (3 mM) and 2-mercaptoethanol (5 mM). After incubation the bound and free hormones were separated by centrifugation and the nuclei were treated with Triton X-100 (final concentration 0.25%) to reduce the non-specific binding. The binding was saturable and reached equilibrium by 20 minutes of incubation and was also stable for 2 hours. The binding was reversible and the rate of dissociation was more or less equal to the rate of association. The binding was linearly increased with the increased concentrations of the DNA (nuclei). Scatchard analyses of the equilibrium binding data revealed that only one class of binding sites for T₃ did exist in the hepatic nuclei of Singi fish. The affinity of these sites or the mean dissociation constant (Kd=0.20±0.07 × 10^-10 M) and the mean maximum binding capacity (MBC=0.17±0.04 pmol/mg DNA) were in reasonable agreement with the values reported for other teleost fishes.

Key words

Thyroid Hormone Receptor - Heteropneustes fossilis Bloch - Fish Liver