Half lobes of mouse thyroid gland were incubated in vitro with TSH. They released cyclic AMP (cAMP) into the medium in amounts depending on the concentration of TSH. The release of cAMP was greatest during the first hour of incubation then it occurred at a lower rate. With an incubation time of 45 min the medium cAMP levels ranged from 43.0 +/- 11.9 pmole per mg tissue protein for controls to 296.5 +/- 29.2 pmole per mg tissue protein with 5 mU of TSH in the medium. The tissue cAMP level reached a maximum after 15--30 min of incubation with TSH, then it gradually decreased towards control level during 4 h of incubation. With 25 min of incubation the tissue cAMP level was 28.5 +/- 8.8 pmole per mg tissue protein for controls compared to 194.3 +/- 27.0 pmole per mg tissue protein with 5 mU TSH in the incubation medium. The release of thyroxine was of the same order during the later part of the 4 h incubation period compared to the first one. The results illustrate the quantitative importance of cAMP release, and the fact that in the later part of the incubaion period the cell content of cAMP was low while the release of thyroxine remained high.