Stimulation of Adenylate Cyclase Activity in Rat Thymocytes in Vitro by 3,5,3′-Triiodothyronine*

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Abstract
In view of the previous demonstration that T3 [triiodothyronine] promptly increases the cAMP concentration in freshly isolated rat thymocytes in vitro, the effects of T3 on adenylate cyclase activity in a crude thymocyte plasma membrane preparation were studied. In common with adenylate cyclase in other tissues, the enzyme in rat thymocytes was activated by NaF, GTP, 5''-guanylylimidodiphosphate and .beta.-adrenergic agonists and was inhibited by high concentrations of Ca. In the presence of 1 .mu.M M Ca+2, T3 induced a time-dependent increase in adenylate cyclase activity that was statistically significant between 1 and 2 min and maximum between 2 and 5 min after hormone addition. As judged from observations made at 5 min, the effect of T3 was dose dependent over the range 1 nM to 1 .mu.M. The stimulatory effect of T3 was Ca dependent, since it was abolished by EGTA [ethylene glycol bis(.beta. aminoethyl ether) N, N, N'', N''-tetraacetic acid] at a concentration (0.5 .mu.M) that did not alter basal enzyme activity; the effect of T3 in the presence of EGTA was restored by the addition of either 0.1 or 1 mM Ca+2. As judged from the lack of hydrolysis of added cAMP, phosphodiesterase activity in the assay mixture was nil in both the presence and absence of T3. Both epinephrine and the specific b-adrenergic agonist isoproterenol, but not the .alpha.-agonist phenylephrine, increased adenylate cyclase activity, and their effects appeared to be additive to that of T3. The .beta.-adrenergic antagonist L -alprenolol, in doses that did not influence basal adenylate cyclase activity, produced a dose-related inhibition of the stimultory effect of T3 and of the effects of epinephrine and isoproterenol as well. Neither D- alprenolol nor the .alpha.-antagonist phentolamine had any effect. Various thyronine analogs displayed a rank order of potency in stimulating adenylate cyclase activity very similar to their relative potencies in increasing cAMP concentration in the intact thymocyte. Apparently, T3 stimulates adenylate cyclase activity in rat thymocyte plasma membrane preparations. With respect to Ca dependence, inhibition by alprenolol and response to thyronine analogs, this effect has properties similar to those of the increase in cellular cAMP concentration induced by T3 in the intact thymocyte. Evidently, the effect of T3 to increase 2-deoxyglucose uptake by the rat thymocyte in vitro, a response consequent to an increase in thymocyte cAMP concentration, derives from a stimulatory effect of the hormone on adenylate cyclase itself.