The effect of hypothyroidism on gastrointestinal beta(1)- and beta(3)-adrenoceptor function and expression was examined in rat ileal smooth muscle preparations. (-)-Isoprenaline and the selective beta(3) agonist disodium (R,R)-5-[2-[[2-3-chlorophenyl)-2-hydroxyethyl]-amino]propyl]-1,3-benzodioxole-2,2-dicarboxylate (CL 316234) relaxed both control and hypothyroid tissues in a dose-dependent manner. Responses to isoprenaline were reduced in tissues from hypothyroid rats, as was the shift produced with the beta(3)-adrenoceptor antagonist, 3-(2-ethylphenoxy)-1-[(1S)-1,2,3,4-tetrahydronaphth-1-ylamino]-(2S)-2-propanol oxalate (SR 59230A). No change was seen in responses to CL 316243. Experiments with a selective beta(1)-adrenoceptor antagonist produced results suggesting that isoprenaline did not act at this receptor. Messenger RNA levels for both beta(1)- and beta(2)-adrenoceptors were not affected by hypothyroidism. These results show that, unlike in adipose tissues, ileal beta(1)- and beta(3)-adrenoceptors are not directly regulated by thyroid hormone and that beta(3)-adrenoceptor coupling to the relaxation response is reduced in a rat model of hypothyroidism.