Studies of the mechanism by which 3,5,3'- triiodothyronine stimulates 2-deoxyglucose uptake in rat thymocytes in vitro. Role of calcium and adenosine 3':5'-monophosphate.

Abstract

A B S T R A C T The present experiments were designed to explore the mechanism whereby 3,5,3'-triiodothyronine (T3) stimulates the uptake of 2-deoxy-D-glucose (2-DG) in rat thymocytes in vitro. Addition of T3 evoked a transient, dose-related increase in cellular cyclic (c) AMP concentrations, evident within 5 min, followed soon by an increase in 2-DG uptake. The effects of T3 on both cAMP concentration and 2-DG uptake were dependent upon the presence of extracellular calcium. Epinephrine also induced a sequential increase in thymocyte cAMP concentration and 2-DG uptake. These responses were more prompt than those to T3, but were calcium independent. As with their combined effects on 2-DG uptake, T3 and epinephrine produced synergistic or additive effects on cellular cAMP concentration. Dibutyryl cAMP also stimulated 2-DG uptake, an effect that was more prompt than that of epinephrine, and, like that of epinephrine, was calcium independent.Prior or simultaneous addition of L-alprenolol (10 AM), which, we have previously shown, blocks the effect of both T3 and epinephrine on 2-DG uptake, also blocked the increase in thymocyte cAMP concentration induced by these agents. In contrast, Lalprenolol failed to block the increase in 2-DG uptake produced by dibutyryl cAMP. On the basis of these observations we suggest that T3 increases 2-DG uptake in the rat thymocyte by increasing the cellular concentration of cAMP, which then acts to enhance sugar transport. The increase in 2-DG uptake induced by epinephrine is also mediated by an increase in cAMP concentration. The greater

Specific Binding Sites for Triiodothyronine in the Plasma Membrane of Rat Thymocytes

“…First, the apparent dissociation constants of the T3 binding sites found were in the range of concentrations at which T3 alone was found to be effective in increasing both cellular cyclic AMP concentration and 2-DG uptake in intact thymocytes (1,3). These concentrations are substantially higher than the assumed free T3 concentration in vivo (5-10 pM).…”

“…However, in the case of these T3 binding sites, there is no evidence that links them to a physiological response. Our interest in determining whether analogous T3-binding sites are present in the plasma membrane of the rat thymocyte stemmed from our previous observations in intact thymocytes, in which substantial evidence was obtained that the prompt increase in 2-DG uptake induced by T3 in vitro, as well as its synergistic interaction with epinephrine, is indicated at a plasma membrane level (1)(2)(3)(4)(5).…”

“…Several lines of evidence indicate that T3 induces its effect by acting at the level of the plasma membrane. Thus, the effect is rapid, is independent of new protein synthesis, and is mediated via an increase in cellular cyclic (c)AMP concentration, as a consequence of an increase in the activity of adenylate cyclase (2)(3)(4), an enzyme that resides within the plasma membrane. Furthermore, epinephrine, which is known to initiate its action upon binding to its plasma membrane receptors and thence activating adenylate cyclase, produces with T3 a synergistic increase in the 2-DG uptake (5 At the end of the incubation period, triplicate 250-Ml aliquots of reaction mixture were quickly removed and applied onto 0.3-um Millipore filters (Millipore Corp., Bedford, MA) under vacuum.…”

In vivo stimulation of sugar uptake in rat thymocytes. An extranuclear action of 3,5,3'-triiodothyronine.

“…3H content of the samples was measured in a liquid scintillation counter. 4 In Cell viability. Cell viability was assessed by means ofthe trypan blue technique (1), in which a viable cell is one that excludes the dye.…”

“…We have previously demonstrated that 3,5,3'-triiodothyronine (T3)' increases the uptake ofthe glucose analogue 2-deoxyglucose (2-DG) in isolated rat thymocytes in vitro (1)(2)(3)(4)(5). This effect of T3 is evident within 20 min after the addition of the hormones; it is independent of new protein synthesis, is calcium-dependent, and is mediated by an increase in cellular cyclic AMP (cAMP) concentration secondary to an increase in adenylate cyclase activity.…”