M. J. Klein

Expression of the α-subunit of glycoprotein hormones in the pars tuberalis-specific glandular cells in rat, mouse and guinea-pig

Abstract.The nature of the hormone(s) secreted by the pars tuberalis (PT) is still unknown. This pituitary lobe is mainly formed by specific glandular cells that differ in their ultrastructural features from the other adenohypophysial cell types. Data from the literature indicate the presence of thyroid-stimulating hormone immunoreactivity in the PT-specific cells of the rat and the Djungarian hamster but not of other species, including the mouse and guinea-pig. The PT also encloses variable numbers of pars distalis cells, essentially gonadotrophs that are mainly dispersed in its caudal area. We studied the expression of the glycoprotein hormone α-subunit in the PT of the rat, mouse and guinea-pig by in situ hybridization and immunocytochemistry. In situ hybridization, using an oligonucleotide probe complementary to rat cDNA sequence 196–237 revealed the expression of the α-subunit gene throughout the PT of the rat and the mouse; in the guinea-pig, the probe labelled no pituitary cells. Light- and electron-microscopic immunocytochemistry demonstrated α-subunit immunoreactivity in the secretory granules of the PT-specific cells in the three species examined. These cells did not react with a specific antibody against the β-subunit of luteinizing hormone, an antibody that labelled scattered gonadotrophs. The present data suggest that hormone(s) produced by the PT-specific glandular cells are, at least partly, related to glycoprotein hormones.

Immunocytochemical evidence for intragranular processing of pro-opiomelanocortin in the melanotropic cells of the rabbit

SummaryThe immunogold technique, employing antisera with clear-cut specificities, was used to localise different processing stages of pro-opiomelanocortin (POMC) in rabbit melanotropic cells. While the antiserum against γ3-MSH labelled all the secretory granules including intrasaccular condensations in the Golgi apparatus, antisera against α-MSH only labelled extra-Golgi secretory vesicles (SV). All extra-Golgi SV were likewise labelled with the three antisera against α-MSH used, despite their different specificities for the desacetylated, N-acetylated or C-amidated forms of the peptide. The antibody against β-endorphin also labelled the extra-Golgi SV, while only some SV were labelled with the antibody against γ-endorphin. These results correlate with biochemical data in favour of mainly — if not exclusively — intragranular processing of POMC. Except for γ3-MSH, the cleavage of which could coincide with Golgi packaging of secretory material, other post-translational modifications of the precursor seem to occur when SV are discharged outside the Golgi area. The cleavage of γ-endorphin appears to be a later step in POMC processing, occurring in some mature SV.

Oxytocin-immunoreactive nerve fibers in the pars intermedia of the pituitary in the rabbit and hare

SummaryThe pars intermedia of the pituitary in the rabbit and hare is abundantly innervated by axons reacting selectively with antibodies against oxytocin. These axons contain dense secretory vesicles about 140 nm in diameter, i.e., smaller than those in the neurosecretory axons of the neural lobe. No fiber elements staining for other peptides (vasopressin, somatostatin, substance P) were observed in the pars intermedia, except rare leu-enkephalin axons restricted to the rostral zone of the gland. Dopaminergic innervation appears to be completely absent from the intermediate lobe. This was shown by the lack of reaction with an antibody against tyrosine-hydroxylase, which did reveal a well-developed tubero-infundibular system of nerve fibers. Axons reacting with an antibody against serotonin were irregularly distributed in the pars intermedia.In the absence of dopaminergic axons, the extensive oxytocin-like innervation may play a major role in regulating the melanotrophic cell activity in the Leporidae.

Corticotrophic cells in the rostral zone of the pars intermedia and in the adjacent neurohypophysis of the rat and mouse

SummaryIn the mouse, the rostral zone of the pars intermedia is almost exclusively composed of typical corticotrophic cells. They are located around and even within the neural stalk, at the level of transition between stalk and neural lobe. In the rat, the corticotrophic cells of the rostral zone are found in scattered islets among the MSH producing cells, and also in the neural lobe. In both the rat and mouse, these cells are in direct contact with various types of nerve terminals. Synaptoid contacts with aminergic and neurosecretory nerve fibers are observed. Furthermore they are also closely related to the hypophysial portal vessels. Following adrenalectomy, the cells located in the neurohypophysis always react more intensely than tose in the rostral zone. The functional significance of these corticotrophic cells which are subject to both humoral and neural regulation remains as yet hypothetical. Their participation in neurogenic stress response seems probable.

Effects of experimental hypo-or hypernatremia on the fine structure of the pars intermedia of the murine pituitary

SummaryQuantified ultrastructural observations of the pars intermedia (PI) of the murine hypophysis enable evaluation and kinetic study of relatively fine secretory changes in the gland. Changes in volume of rER and newly formed dense secretory granules (Golgi granules) appear to best translate functional variations in the PI, as shown by the morphological effects of drugs affecting the dopaminergic control of the gland. Our morphometric results show that the PI is stimulated, but only briefly (no longer than 8–12 days), by both salt-loading and Na deprivation. However, the PI displays different secretory patterns in salt-loaded and Na-deprived mice; moreover, bromocriptine, which abolishes PI stimulation in Na-deprived mice, has only a slight inhibitory effect in salt-treated animals. Thus, it appears that the stimulation of the PI under both experimental conditions is triggered by different mechanisms. These results underline the plurifactorial control of the PI and show that the gland may have complex effects on hydromineral regulation.

Differentiation of the melanotrophic cells of rat pituitary primordium in organotypic culture in defined medium

SummaryOrganotypic cultures, in defined medium, of pituitary primordia obtained from 15-day-old rat fetuses were performed in order to study the in vitro differentiation of melanotrophic cells. The morphological and ultrastructural features of the transplants resembled those of the gland developing in vivo. In situ hybridization on semi-thin sections, using a 35S-labelled oligonucleotide probe, revealed pro-opiomelanocortin-mRNA-containing cells on the first day of culture in the anterior lobe and after 2–3 days in the intermediate lobe. Immunoperoxidase labelling of adjacent sections showed that the same cells reacted with antibodies against α-melanocyte-stimulating hormone (αMSH), γ3 and adrenocorticotropic hormone in both lobes. The pro-opiomelanocortin-mRNA-containing cells formed progressively conspicuous areas in the intermediate lobe, which was almost uniformly labelled after 6 days. In the anterior lobe, these cells remained scattered in small cell groups, and colloidal gold immunolabelling showed the progressive disappearance of αMSH labelling from the secretory vesicles in cells exhibiting morphological features of adult corticotrophic cells. Both the αMSH content of the explants and αMSH release into the culture medium increased with time. Treatment with the dopamine agonist bromocriptine induced a strong dose-dependent decrease in αMSH secretion, which was significant after 3 days in culture, indicating that dopamine D2 receptors are able to regulate hormonal release of melanotrophic cells at early stages. This system constitutes a suitable model for further studies of factors controlling cell differentiation and cellular interactions involved in histogenesis.

Ontogenesis of the rabbit pituitary intermediate lobe

SummaryThe development of the intermediate lobe of the rabbit pituitary was investigated by light and electron microscopy and by using immunocytological techniques. The first immunoreactive melanotrophic cells were detected at the fetal day 17 in the dorsal zone of Rathkes pouch epithelium facing the neural lobe; this coincided ultrastructurally with the appearance in this area of a few cells exhibiting secretory vesicles and granular condensations in the Golgi saccules. The differentiation of the gland probably required an infundibular inductive effect. Secretory cells increased in number following a dorsoventral gradient during the next fetal and neonatal stages until postnatal day 20, the stage at which the intermediate lobe exhibited its definitive organization. The gland innervation occurred during the first days after birth. The advent of these oxytocin- and neurophysin-immunoreactive fibres coincided with an obvious stimulation of the synthetic activity of the melanotrophic cells. The possible neurotrophic effect of these cells on their innervating system remains to be established.