The differential regulation of neurotensin (NT), calcitonin (CT), and CT gene-related peptide (CGRP) production was studied in the clonal, rat C cell-derived, 44-2C cell line. Two experimental paradigms were used: cells were incubated with maximally effective concentrations of calcium (4.0 mM); alternatively, cells were treated with the synthetic glucocorticoid, dexamethasone (DEX). The specificity of the DEX-mediated response was assessed by using the synthetic antiglucocorticoid, RU-486. Calcium was not mitogenic in 44-2C cells and did not affect cell growth. Calcium increased the secretion and cellular accumulation of NT. In contrast, calcium treatment decreased CT content and release while it diminished the levels of CT- and CGRP-specific messenger RNA (mRNA) levels. DEX (10(-8) M) inhibited cell proliferation. NT content and secretion increased after DEX treatment, and this was potentiated by the addition of calcium. DEX-treated cells showed diminished CT content and secretion. The levels of CT- and CGRP-specific mRNA were significantly reduced in DEX-treated cultures. RU-486 antagonized the action of DEX and blocked DEX-inhibited cell proliferation. Inhibition of CT secretion by DEX was blocked by RU-486; CT- and CGRP-specific mRNA levels were increased in response to treatment with equimolar or 100-fold excess concentrations of RU-486. We conclude that NT secretion as well as CT/CGRP expression and release can be differentially regulated in the 44-2C cell line.