Long-acting thyroid stimulator (LATS) increased glucose oxidation and (32)P incorporation into phospholipids in in vitro experiments with dog thyroid slices. The time course of the response was different from that obtained with thyroid-stimulating hormone (TSH), but was very similar to the delayed effect observed in vivo. During a 45 min incubation, TSH, but not LATS increased glucose oxidation, whereas in longer experiments up to 6 hr, both substances augmented (14)CO(2) production. Amounts of pooled human gamma globulin equivalent to LATS were inactive. Although TSH stimulated (32)P incorporation into phospholipids during a 2 hr incubation, LATS was ineffective. In longer incubations, from 4(1/2) to 8 hr, LATS did increase (32)P incorporation. The stimulatory effect of LATS was not abolished by anti-TSH antibody capable of neutralizing human TSH. Effects of LATS were also obtained with beef and pig thyroid slices. In addition to stimulation of glucose oxidation in dog thyroid slices, LATS occasionally also stimulated glucose oxidation in dog spleen and liver slices. Despite a 54-fold increase in LATS concentration, a satisfactory dose-response curve could not be demonstrated when (14)CO(2) production was measured.