Cytochemical characterization of anterior pituitary target cells for the neuropeptide, pituitary adenylate cyclase activating polypeptide (PACAP), using biotinylated ligands

Abstract

Two novel peptides, named PACAP (pituitary adenylate cyclase activating polypeptide) containing 38 (PACAP38) and 27 residues (PACAP27) were recently isolated from ovine hypothalami. In order to investigate the pituitary cell type(s) that bear a receptor for PACAP, PACAP38 was biotinylated and used for cytochemical examination of binding. The cells were also identified by immunocytochemical methods using the antisera against each of the rat anterior pituitary hormones or an antiserum against S-100 protein, a marker for pituitary folliculo-stellate (FS) cells. Biotinylated PACAP38 (biot-PACAP) exhibited adenylate cyclase stimulating activity (ACSA) comparable to PACAP38 in rat pituitary cell cultures, and displaced the bound 125I-PACAP27 to the rat pituitary membrane preparation to the same extent as PACAP38. After 2–4 days of culture, dispersed rat pituitary cells were incubated with varying concentrations of biot-PACAP at room temperature or 4°C. The bound biot-PACAP38 was visualized by avidin-biotin-peroxidase complex (ABC) method with nickel intensification. Biot-PACAP-positive and pituitary hormone or S-100-positive cells were counted. More than 90% of S-100-positive cells bound biot-PACAP38. A considerable number of GH and PRL cells and a lesser number of ACTH cells also bound biot-PACAP38, whereas only a few identified LH, FSH, or TSH cells bound biot-PACAP38. These results suggest that FS cells are a major target cell type for PACAP. A recent study from our laboratory demonstrated that PACAP stimulated the release of interleukin (IL)-6 in rat pituitary cell cultures. FS cells are known to produce IL-6.