In view of the importance of the caliotropic hormones to calcium homeostasis and bone strength we have studied the regulation of the expression of the calcitonin gene in vivo in the rat. Normal rats were injected with 1,25-dihydroxyvitamin D3 (12.5-200 pmol/100g body weight) and calcitonin mRNA levels measured. 1,25(OH)2D3 led to a marked decrease in calcitonin mRNA levels, which reached 4% of basal at 24 h. Nuclear transcript experiments showed that the effect was transcriptional. After large changes in serum calcium for periods from 6 h to 3 weeks there were no differences in calcitonin mRNA levels. Estrogens given to ovariectomized rats led to increases in calcitonin mRNA. The estrogen receptor was present in the thyroid C-cells. Therefore the calcitonin gene is regulated by vitamin D and estrogens, but not by calcium.