An extracellular Ca(2+)-sensing mechanism consisting of a G protein-coupled receptor linked to phosphoinositide turnover and inhibition of PTH secretion, has recently been identified in bovine parathyroid cells. In C cells, voltage-dependent L-type calcium channels are thought to be involved in calcium-sensing mechanisms, but evidence exists for additional calcium-sensing mechanisms, such as via a calcium-sensing receptor (CaSR). Using the human medullary C cell carcinoma cell line TT, which lacks L-type calcium channels, we found that Ca2+ or cations specific for the CaSR lead to the release of calcium ions from intracellular stores and to an increase in calcitonin secretion. By molecular cloning we isolated the complete protein-coding complementary DNA of a CaSR from human TT cells, which are derived from a human medullary thyroid carcinoma. The CaSR is derived from the same CaSR gene expressed in the parathyroid gland. In addition, TT cells contain an alternative receptor form of CaSR, CaSRb. These findings provide strong evidence for the presence of a functional CaSR in the human C cell line TT. This receptor contributes not only to the inhibition of PTH secretion in the parathyroid, but also to the stimulation of calcitonin secretion in C cells.