Effects of Thyrotropin-Releasing Hormone on Prolactin Compartments in Normal Rat Pituitary Cells in Primary Culture*

Abstract

PRL compartments have been studied in normal rat pituitary cells cultured for 6 days. The cells were pulse-labeled for 15 min with 35S-methionine and then chased for 24 h in the absence or presence of cycloheximide (3.6 X 10(-5) M). TRH (30 nM) was introduced into the medium either at the beginning or after increasing durations of chase. The findings were compared with those obtained with GH3B6 cells in similar experimental conditions. Despite the fact that normal PRL cells differ from GH3B6 cells by a large intracellular PRL store, several similarities were found between the two systems: newly synthesized PRL was rapidly and preferentially released in basal conditions, the pattern of the decay of the specific radioactivity of PRL released into the medium suggested the existence of at least two PRL pools with different half-lives: 2.5 h and 22 h, respectively, TRH induced the preferential release of stored PRL synthesized before the pulse, only 20% of the pulse-labeled PRL was released into the medium after 24 h of chase. However, normal PRL cells differed in several respects from GH3B6 cells: the turnover time of the two PRL pools is 8 times greater in normal PRL cells, an asynchrony in the time of appearance of labeled PRL in the medium was observed, suggesting a functional heterogeneity of these cells, at the end of the chase, 40% of the pulse-labeled PRL was lost in the case of normal cells, but not of GH3B6 cells, and this was prevented by cycloheximide, polyacrylamide gel electrophoresis analysis of this labeled immunoprecipitated intracellular material revealed the existence, in addition to the mol wt of 23,000 PRL and the large PRL-like forms (mol wt, 45,000 and 50,000), as observed with GH3B6 cells, of smaller proteins (mol wts, 39,000, 36,000, 20,000, 18,000, 15,000), which might represent degradation products.

Axons Containing a Prolactin-Like Peptide Project into the Perivascular Layer of the Median Eminence: An Immunocytochemical Light and Electron Microscope Study in Adult and Infant Rats

“…I). In good agreement with data obtained in GH.iBi, cell lines [16] and in pituitary cells in primary culture [17], immunostaining was found here to be essentially associated with a protein band corresponding exactly to the prolactin reference (mo lecular weight 23 kilodaltons; kD), and with two minor bands of higher molecular weight (45 and 70 kD) which may possibly repre sent the dimeric and polymeric forms of prolactin previously re ported [20]. On the other hand, no immunostaining could be delec ted in protein bands lighter than 23 kD.…”

Modulation of thyrotrophin release from an intracellular pool by pre-exposure to thyrotrophin-releasing hormone and dibutyryl cyclic AMP

“…This suggests that depletion of an intracellular pool of TSH occurs, with a predominant effect on basal and TRH-induced release. Preferential release of hormones from specific intracellular pools has been postulated in the case of prolactin and growth hormone, where data from differential isotopie labelling experiments have suggested preferential release of newly formed peptide in some circumstances (Swearingen, 1971;Stachura, 1982;Morin, Rosenbaum & Tixier-Vidal, 1984). The hypothesis of an intracellular pool of TSH can be only a functional description based on the present data, but it seems likely that there might be topographic localiza¬ tion of TSH within the cell, possibly into receptorassociated compartments.…”

Secretion of Renin–Angiotensin System (RAS) Components by Normal and Tumoral Lactotropes: A Comparative Study Using Reverse Hemolytic Plaque Assay (RHPA) and Immunoelectron Microscopy

“…GH3B6 cells contained only few small secretory granules even when cultured in a medium supplemented with EGF and hormones (see Materials and Methods) ( Figure 5). Such a small storage compartment has been linked to a high basal release of their major hormone product, prolactin, and to a more rapid intracellular transport of the newly synthesized hormone (Morin et al 1984;Tougard and Tixier-Vidal 1994). This feature of GH3B6 cells may explain the high percentage of cells releasing RAS precursors.…”