Long term culture of primary rat pituitary adrenocorticotropin/endorphin-producing cells in serum-free medium.

Abstract

Insulin, transferrin, and serum albumin were found to be essential additives for maintenance of primary rat pituitary ACTH/endorphin cells in complete serum-free defined medium (CSFM). Primary anterior pituitary cultures maintained in CSFM exhibited a 3- to 5-fold increase in cell content of ACTH/endorphin-related peptide during the first 2 weeks in culture, and this level remained stable for at least the next week. Immunocytochemical and morphometric studies indicated that the number of corticotropes increased only slightly, so that hormone content per corticotrope increased. Anterior pituitary cultures maintained in CSFM exhibited a basal secretory rate of 0.3-0.4% of cell hormone content/h throughout the 3-week period in culture, and total hormone production increased 6-fold. Primary anterior pituitary cultures maintained chronically in CSFM containing 10 nM CRF demonstrated a 15- to 20-fold increase in total ACTH/endorphin production over 3 weeks in culture. Chronic treatment with CRF brought about a sustained 6-fold increase in secretory rate (2.5% of cell hormone content/h), and corticotrope content of hormone was diminished 3-fold. Corticotropes maintained chronically in CSFM containing CRF did not increase in number and exhibited a rim of immunocytochemically identifiable hormone around the cell periphery. Anterior pituitary cultures maintained chronically in CSFM containing 100 nM dexamethasone (DEX) exhibited decreased cell hormone content and an unaltered secretory rate. In the DEX-treated cultures the number of immunocytochemically identifiable corticotropes declined, as did the staining intensity per corticotrope. Primary cultures of rat intermediate pituitary cells maintained in CSFM exhibited a 1.5- to 2-fold increase in hormone content after 1-2 weeks in culture, maintained a constant basal secretory rate of 0.4-0.5% cell content/h, and were not responsive to chronic treatment with CRF or DEX.