Gonadotropes in a novel rat pituitary tumor cell line,RC-4B/C. Establishment and partial characterization of the cell line

Summary An epithelial cell line (RC-4B/C) was established from a pituitary adenoma obtained from a 3-yr-old (ACI/fMai × F344/fMai)F1 male rat. Before Year 5 in vitro, RC-4B/C cells could not be viably recovered from cryogenic storage. Recovery of viable cells from cryogenic storage in Year 5 was associated with a more transformed phenotype, including the appearance of endogenous C-type rat retroviral particles. The ultrastructural appearance of the cells was similar to that of differentiated anterior pituitary cell; the cultured cells contained numerous, electron dense, secretory granules, Golgi complexes, and extended arrays of rough endoplasmic reticulum. Immunocytochemical study showed that all cell types present in the rat anterior pituitary gland were present in the cell line. The percentage of luteinizing hormone beta (LHβ) cells in the cell line was higher (19.9%) and that of growth hormone cells was lower (12.2%) than in normal male rat pituitary, whereas the cell line contained a comparable percentage of follicle stimulating hormone beta (FSHβ), prolactin (PRL), ACTH, and thyrotropin beta cells. Radioimmunoassay data demonstrated the PRL content of the cells was comparable to that of normal male rat pituitary gland, whereas the content of LH and FSH was 70- and 800-fold lower, respectively. Assay of specific receptor sites for gonadotropin releasing hormone (GnRH) using Scatchard plots of the data established the RC-4B/C cells contained GnRH receptor sites of the same affinity as in the pituitary gland, but of twofold lower capacity. These data suggest the RC-4B/C cell line warrants further study as a model for the induction and maintenance of the gonadotropic function of the pituitary gland. An abstract of portions of these results was presented at the 8th International Congress of Endocrinology, Kyoto, Japan, 1988. This work was supported in part by grants DK-17631 (E.H.L.), CA-24145 (W.G.B.), CA-31102 (H.G.B.), AG-01753 (D.E.H.) and HD-1778 (M.T.D.) from the National Institutes of Health, Bethesda, MD, and by a grant from the Association pour la Recherche sur le Cancer, France (M.J.). The NIH is not responsible for the contents of this publication nor do the contents necessarily represent the official views of that agency. Jolanta Polkowska was a recipient of a Foundation Simone et Cino del Duca grant.     

A unique ball-shaped Golgi apparatus in the rat pituitary gonadotrope: its functional implications in relation to the arrangement of the microtubule network

In polarized exocrine cells, the Golgi apparatus is cup-shaped and its convex and concave surfaces are designated as cis and trans faces, functionally confronting the rough endoplasmic reticulum and the cell surface, respectively. To clarify the morphological characteristics of the Golgi apparatus in non-polarized endocrine cells, the investigators immunocytochemically examined its precise architecture in pituitary gonadotropes, especially in relation to the arrangement of the intracellular microtubule network. The Golgi apparatus in the gonadotropes was not cup-shaped but ball-shaped or spherical, and its outer and inner surfaces were the cis and trans faces, respectively. Centrioles were situated at the center of the Golgi apparatus, from which radiating microtubules isotropically extended to the cell periphery through the gaps in the spherical wall of the Golgi stack. The shape of the Golgi apparatus and the arrangement of microtubules demonstrated in the present study could explain the microtubule-dependent movements of tubulovesicular carriers and granules within the gonadotropes. Furthermore, the spherical shape of the Golgi apparatus possibly reflects the highly symmetrical arrangement of microtubule arrays, as well as the poor polarity in the cell surface of pituitary gonadotropes.     

Is cellular disruption the mechanism of release of basic fibroblast growth factor from anterior pituitary gonadotropes?

Adult female Fischer 344 (F344) and Sprague-Dawley (SD) rats, intact and ovariectomized (10-30d), have been used for immunolocalization of basic fibroblast growth factor (FGF). Tissues were selected from three specific sites (postero-lateral, lateral wing, and anterior wedge) of the periphery of the anterior pituitary (AP) carefully maintaining the association between the gland and the highly vascular meningeal connective tissue which envelopes it. In both rat strains, most of the periphery of the AP was characterized by intact parenchymal cells delimited from the meningeal connective tissue by an intact basal lamina. However, foci also were evident in which parenchymal cells projected directly into the connective tissue without a basal lamina intervening. These zones, designated the Non-Delimited Peripheral Parenchyma (NDPP), were present minimally in control rats, but were more numerous in ovariectomized rats. Profiles of focally disrupted gonadotropes were evident within the NDPP of 20-30d ovariectomized rats juxtaposed against intact, granulated parenchymal cells. Partially disrupted gonadotropes also were evident within the peripheral parenchyma within approximately 100 mu of the edge, and occasionally the disruptions resulted in an association of neighboring gonadotropes as a syncytium. FGF was localized only within the cytosol of gonadotropes, i.e., cells immunopositive for LH-beta and FSH-beta subunits. Gonadotropes nearer to the edges of the AP, especially the postero-lateral edge, were the most intensely stained. Electron microscopy and immunostaining for S-100 protein, a marker for folliculo-stellate cells (FSC), demonstrated that in intact and ovariectomized SD rats FSC were present in all peripheral zones of the AP, whereas portions of the postero-lateral periphery of the AP of intact and ovariectomized F344 rats often lacked FSC. We propose that FGF may be released from the cytosol of gonadotropes by a mechanism of cellular disruption. FGF released at peripheral sites of the AP would be well-positioned to stimulate angiogenesis from systemic blood vessels within the meninges. Since FSC are known phagocytes within the AP, their consistent presence in the periphery of the AP of SD rats may help regulate the effects of the released FGF, and by contrast, their absence in F344 rats may intensify or prolong the effects of released FGF. Such differences may underlie the higher incidence of pituitary tumors in F344 rats.     

Co-existence of gonadotrophins (FSH,LH) and thyrotrophin (TSH) in single anterior pituitary cells of the musk shrew,Suncus murinus

According to recent immunocytochemical studies of anterior pituitary cells, it is obvious that the one cell-one hormone theory must be modified. Many pituitary morphologists have demonstrated that there are some cells that contain two hormones. In this study, we demonstrate by means of immuno-electronmicroscopy the co-existence of gonadotrophins (FSH and LH) and thyrotrophin (TSH) in the same anterior pituitary cells of the musk shrew. These cells were remarkably altered in their ultrastructural features by either gonadectomy or thyroidectomy. Double labeling for gonadotrophins and thyrotrophin was present not only in the same cells but also in the same secretory granules. Our ability to demonstrate co-existence of gonadotrophins and thyrotrophin in the same cell may be due to our selection of fixative and embedding media for electron-microscopic immunocytochemistry. Our conclusion that gonadotrophins and thyrotrophin are produced in a single cell type of the anterior pituitary gland in the musk shrew, i.e., thyrogonadotrophs, suggests the need to consider a modification of the classic scheme for classification of anterior pituitary cells.     

Ovulation is triggered by a cyclical modulation of gonadotropes into a hyperexcitable state

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Etude du mechanisme de la regulation hormonale dans le placenta humain: Mise en evidence d'une activite adenyl cyclasique dans les homogenats de placenta a terme

Human full-term placentas possess adenyl cyclase activity which was assayed by the transformation of [α-³²P]ATP to cyclic 3′,5′-[³²P]AMP according to the method of Krishnaet al. (G. Krishna, B. Weiss and B. B. Brodie), J. Pharmacol. Exptl. Therap., 163 (1968) as modified by Rodbell (M. Rodbell, J. Biol. Chem., 242 (1967) 5744).