Liver and kidney mitochondrial [alpha]-glycerophosphate dehydrogenase (GPD) and liver malic enzyme responses to graded doses of thyroxine (T4) were measured in adrenalectomized (adx.), hypophysectomized (hypox.), gonadectomized and alloxan-diabetic rats. Although the malic enzyme was sometimes affected by other factors, the GPD enzyme response to T4 was highly specific, and it appeared to measure the peripheral activity of T4 better than the metabolic rate (MR). For example, estrogen suppressed the MR response to T4 in intact and gonadectomized male and female rats, and cortisone and growth hormone stimulated the MR response to T4 in hypox. animals. Since none of these hormones had any affect on the GPD response to T4, their influence on the MR was not mediated by changes in the peripheral activity of T4. Testosterone, on the other hand, may have altered the peripheral activity of T4 because it partially suppressed both the MR and GPD enzyme responses to T4. Both the MR and GPD enzyme levels were elevated in diabetic control animals, but the liver malic enzyme activity was below normal. Whereas diabetes did not affect the GPD and MR responses to T4, it did suppress the liver malic enzyme response.