Modulation of Adenylate Cyclase/Cyclic AMP Response by Thyrotropin and Prostaglandin E2 in Cultured Thyroid Cells. 2. Positive Regulation

Abstract

Two different independent processes are operating in cultured thyroid cells to regulate adenylate cyclase/cyclic AMP responsiveness to thyroid stimulators (thyrotropin and prostaglandin Ez) :firstly, refractoriness or negative regulation [preceding paper], which is specific for each thyroid stimulator, is not mediated by cyclic AMP and is not accompanied by alteration of adenylate cyclase activity; secondly, positive regulation which is characterized by an augmentation of the cyclic AMP response stimulated by thyrotropin and prostaglandin E2. This process is not specific for each thyroid stimulator and is a state of increased susceptibility of cyclic AMP synthesis to stimulation, accompanied by increased activity of the catalytic subunit of adenylate cyclase. Positive regulation is apparently mediated by increased intracellular cyclic AMP levels. It is a time-dependent and dose-dependent process. Very low concentrations ( 5 -50 pU/ml) of thyrotropin augmented cyclic AMP synthesis stimulated by thyrotropin and prostaglandin E2 whereas higher concentrations (above 0.1 mU/ml) augmented prostaglandin El stimulation but induced refractoriness to thyrotropin. Prostaglandin E2 (0.1 to 10 pM) augmented thyrotropin stimulation and dibutyryl adenosine 3' : 5'-monophosphate (0.3 to 2 mM) augmented thyrotropin and prostaglandin Ez stimulation. Positive regulation is a slow process which develops within days and increases up to day 5 in culture. Experiments using inhibitors suggested that protein synthesis is required for the full expression of the increase in adenylate cyclase activity induced by the studied thyroid stimulators.We have recently shown that the cyclic AMP response to thyrotropin of thyroid cells in culture was under a dual regulation by thyrotropin : chronic exposure of the cells to physiological concentrations of the hormone augmented the subsequent cyclic AMP response to thyrotropin (positive regulation), whereas moderate to high concentrations of thyrotropin induced refractoriness to further thyrotropin stimulation [l].The conditions for the development of refractoriness to thyrotropin and prostaglandin Ez have been described in a preceding paper [2]. In this report, we analyze the mechanism of the positive regulation of the acute cyclic AMP response to thyrotropin and prostaglandin E2 as influenced by chronic treatment of cultured thyroid cells with these stimulators. MATERIALS AND METHODSThe methods and the materials used have been described in the preceding paper [ 2 ] . Cells cultured in the presence of thyrotropin (10 mU/ml), prostaglandin EZ (1 pM) or dibutyryl cyclic AMP (0.4 mM) will be referred to further as thyrotropin-treated, prostaglandin E2-treated or BtzcAMP-treated cells, respectively. Cells cultured in the absence of these compounds will be referred to as control cells.Cyclic AMP synthesis using prelabeling technique with 14C-labeled adenine [3] was measured as reported previously [4] with minor modifications. Duration of preincubation with [14C]adenine was 40 rnin. To preserve cell viab...

The Functional Expression of Recombinant Human Thyrotropin Receptors in Nonthyroidal Eukaryotic Cells Provides Evidence that Homologous Desensitization to Thyrotropin Stimulation Requires a Cell-Specific Factor*

“…4) containing NaCl instead of sucrose, the number of TSH binding sites was reduced by approximately 10-fold, without any alteration in the affinity of TSH binding (data not shown). Previous studies with FRTL5 rat (14) and pig (18) thyroid cells indicate that prolonged (16-24 h) exposure of these cells to TSH reduces the number of TSH receptors available for TSH binding. This TSH receptor down-regulation cannot be the basis for TSH desensitization in FRTL5 thyroid cells because the latter is evident much earlier, within 2 h of TSH stimulation (12).…”

Iodide transport and breast cancer

“…In the thyroid, the expression of NIS is principally regulated by the thyroid-stimulating hormone (TSH) (Kogai et al 1997). When TSH binds to the TSH receptor (TSHR), adenylate cyclase is activated, leading to increased intracellular cAMP levels (Takasu et al 1978), which further activates the transcription factors, cAMP response element-binding protein (CREB) and Pax8 (Poleev et al 1997). The NIS upstream enhancer (NUE), which is fundamental in the initiation of NIS transcription, contains binding sites for both Pax8 and CREB, which stimulate NIS transcription upon transcription factor binding (Ohno et al 1999).…”

Tonic effect of endogenous TSH on the in vitro thyroid cAMP response to TSH

“…The fourth possibility seems also to be unlikely, since phos¬ phodiesterase activity has been reported to increase by chronic stimulation with endogenous TSH (Bastomsky et al 1971). On the other hand, Takasu et al (1978) reported that adenylate cyclase activity in cultured porcine thyroid cells was decreased when cultured without TSH as compared to when cultured in 5-50 µ /ml of TSH.…”