Thyrotropin, through a cAMP-dependent pathway, stimulates function, differentiation, and proliferation of dog and human thyroid cells. Our previous findings suggested that, in addition to PKA activation, another cAMP-dependent mechanism is involved in TSH action. In this work, we assess whether the newly identified cAMP-Epac-Rap1 cascade is involved in TSH-cAMP-mediated effects in dog thyroid cells. We first demonstrate that TSH and forskolin strongly activate Rap1 in a PKA-independent manner. However, activation of Rap1 is not specific for TSH or cAMP. Indeed, carbachol, TPA, insulin, or EGF, which activate different cAMP-independent cascades, all independently activate Rap1. Rap1 is therefore a common step in all these cascades which exert various effects on proliferation, differentiation, and function of thyroid cells. Moreover, the microinjection of the Rap1 protein alone or in combination with the catalytic C subunit of PKA fails to induce proliferation or expression of thyroglobulin.