The Appearance,Distribution,and Longevity of Receptor-[ 125 I]T 3 Complexes within the Nuclei of Isolated Rat Hepatocytes

The nuclei of isolated rat hepatocytes were separable into three receptor compartments based upon their differential salt extractabilities: nucleoplasmic receptors (NP) extractable with 0.15 M KC1, high-salt extractable receptors (HSE) extractable with 0.4 M KC1, and salt-resistant receptors (SR) extractable with 0.4 M KC1/5 mM dithiothreitol. The receptor distribution among the three compartments was approximately NP, 45 %; HSE, 30 %; SR, 25 %. The mean percent occupancy with endogenous T₃ of the SR receptors (86%) was higher than the occupancies of the NP receptors (68%) and the HSE receptors (63%). When hepatocytes were pulsed with 3 nM [¹²⁵I]T₃ at 37°C for brief intervals, receptor-[¹²⁵I]T₃ complexes were detectable in all three nuclear compartments within 15 sec. With increasing pulse intervals up to 120 sec, the receptor content of each nuclear compartment increased progressively and without evidence of preferential accumulation in any of the three compartments. To determine the life span and intercompartmental "migration" pattern of nuclear receptors, hepatocytes were pulsed with 3 nM [¹²⁵I]T₃ at 37°C for 2.5 min or 5 min, followed by a chase with a 500-fold excess of nonlabeled T₃. The population of receptor-[¹²⁵I]T₃ complexes generated during the pulse was serially recovered at increasing intervals after the chase. The complexes of each compartment dissociated with a half-life of approximately 3 min and manifested no predilection to accumulate in any of the compartments. Exposure of isolated hepatocytes to 3 nM T₃ for 5 min or 10 min at 37°C induced no change in the gross intercompartmental distribution of receptors compared to control hepatocytes incubated without T₃. These studies support the concept that extracellular T₃ is rapidly taken up by the hepatocyte and enters the nucleus well within 1 min. Within the nucleus, the T₃ has immediate access to the unoccupied receptors of each compartment. The receptor-T₃ complexes formed therefrom exist for only a few minutes (t _½ ≃ 3 min), during which time there is no net shift of the complexes from one compartment to another. Exposure to T₃ does not appear to drive or influence the gross intercompartmental receptor distribution.