Receptors for thyrotropin-releasing hormone on rat lactotropes and thyrotropes.
Primary cultures of rat pituitary cells were stained with an antibody to the native thyrotropin-releasing hormone (TRH) receptor and with a bioactive, fluorescent analogue of TRH, Rhod-TRH. Rhod-TRH specifically stained 86% of lactotropes and 21% of nonlactotropes from primary pituitary cell cultures. Lactotropes and thyrotropes accounted for 90% of cells that stained with Rhod-TRH, but there were occasional lactotropes and thyrotropes that did not show detectable staining with antireceptor antibodies or with Rhod-TRH. The intensity of staining was generally higher in the GH3 line of tumor cells than in normal pituicytes, and 100% of the tumor cells stained with Rhod-TRH. To determine whether the TRH receptor undergoes ligand-directed endocytosis in normal cells, TRH receptor immunocytochemistry was performed before and after TRH binding. TRH receptors were localized on the surface of cells prior to TRH exposure, and Rhod-TRH fluorescence was confined to the plasma membrane when TRH binding was performed at 0 degrees C, where endocytosis is blocked. When cells were incubated with TRH at 37 degrees C, receptors were found in intracellular vesicles in both lactotropes and thyrotropes, and Rhod-TRH was rapidly internalized into endosomes at elevated temperatures. Internalization of Rhod-TRH was inhibited by hypertonic sucrose, indicating that it occurs through clathrin-coated pits. These findings show that some of the heterogeneity in the secretory and calcium responses of pituicytes to TRH occurs at the level of the TRH receptor.
Yu
R
R
Department of Pharmacology and Physiology, Cancer Center, University of Rochester School of Medicine and Dentistry, New York, USA.
Ashworth
R
R
Hinkle
P M
PM
D000595:Amino Acid Sequence; D000818:Animals; D002478:Cells, Cultured; D005260:Female; D005455:Fluorescent Antibody Technique; D005456:Fluorescent Dyes; D008297:Male; D008969:Molecular Sequence Data; D010903:Pituitary Gland, Anterior / Q000737:chemistry* / Q000378:metabolism*; D010911:Pituitary Neoplasms / Q000737:chemistry; D011388:Prolactin / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D018025:Receptors, Thyrotropin-Releasing Hormone / Q000032:analysis* / Q000378:metabolism; D012235:Rhodamines; D013972:Thyrotropin / Q000378:metabolism*; D013973:Thyrotropin-Releasing Hormone / Q000494:pharmacology
D005456:Fluorescent Dyes; D018025:Receptors, Thyrotropin-Releasing Hormone; D012235:Rhodamines; D013973:Thyrotropin-Releasing Hormone; D011388:Prolactin; D013972:Thyrotropin