Resistance to thyroid hormone (RTH) syndrome is an inherited inability to respond appropriately to T3 hormone. In generalized RTH, the T3 response of both the pituitary and periphery is disrupted. In pituitary (or central) RTH, the ability of the pituitary to sense (and down-regulate) elevated T3 is selectively impaired, whereas the periphery remains relatively T3 responsive, resulting in peripheral thyrotoxicity. Both forms of disease are linked to mutations in thyroid hormone receptor (TR)-beta. TRbeta is expressed by alternate mRNA splicing as two isoforms: TRbeta2, found primarily in the pituitary/hypothalamus, and TRbeta1, expressed broadly in many tissues. We report here that the wild-type TRbeta2 isoform displays an enhanced T3 response relative to the TRbeta1 isoform. Mutations associated with generalized RTH (P453S, G345S) impair both TRbeta2 and TRbeta1 function proportionally, whereas mutations associated with pituitary-specific RTH (R338L, R338W, R429Q) disproportionately disrupt TRbeta2 function. We propose that in the normal organism, and in generalized RTH, TRbeta2 in the pituitary can sense rising T3 levels in advance of TRbeta1 in the periphery, preventing thyrotoxicity. In contrast, the TRbeta mutations associated with pituitary RTH disproportionately disrupt the pituitary's ability to sense and suppress elevated T3 levels in advance of the periphery, producing symptoms of thyrotoxicity.