Mitotic activity of prolactin cells in the pituitary glands of male and female rats of different ages

Summary The ultrastructural localization of the glycoprotein D2 in rat adrenal gland was investigated using immunohistochemical methods, and D2 localization in cultures of adult bovine chromaffin cells was studied by immunofluorescence. D2 was found to be situated on nerve fibers passing through the adrenal cortex and in the medulla zone, and also on the surface of all chromaffin cells. In addition, it was strongly expressed on the surface of glial (Schwann) cells. Cortical cells were unreactive to the antiserum. In cultures, all adrenalin and noradrenalin [dopamine--hydroxylase (DBH)-positive] cells were surface labelled for D2. A less frequent second cell type was recognized in vitro which was DBH negative but D2 positive. Such cells were presumed to be Schwann cells. These data are discussed in terms of the developmental origin of the cells and with regard to the putative functional rôle of D2 in cell adhesion phenomena.     

Isocitrate dehydrogenase activity and its regulation by estradiol in tissues of rats of various ages

The activity and hormonal regulation of NAD- and NADP-linked isocitrate dehydrogenase (EC.1.1.1.41 and EC.1.1.1.42, respectively) in the brain, liver and kidney cortex of female rats of various ages was investigated. The activity of NAD-ICDH of brain was greater than extramitochondrial (-c) or intramitochondrial (-m) NADP-ICDH. In contrast, liver c-NADP-ICDH was much higher than NAD- or m-NADP-ICDH, whereas in kidney cortex the activity of m-NADP-ICDH is dominant over both NAD- and c-NADP-ICDH in all the age group of rats studied. The activity of the NAD-ICDH of brain and all the enzymes of liver and kidney cortex increases until adulthood (33-weeks) and decreases thereafter in old rats (85-weeks). In brain c-NADP-ICDH was much higher in immature (6-weeks) rats and decreases with increasing age of the animal, whereas m-NADP-ICDH showed no significant change with the age of the rats. Bilateral ovariectomy decreases the level of all the three forms of enzyme in all the tissues of 6-, 13- and 33-week rats but failed to show any significant effect in 85-week old rats. Exogenous administration of estradiol induces all the three forms of enzyme in all the tissues of ovariectomized rats. The degree of response is tissue- and age-specific.     

Circadian rhythm of the mitotic activity of pulmonary interalveolar septum cells in rats of different ages]

The daily rhythm of mitotic activity in the lungs of the 20-day-dd embryo coincides with the rhythm of the adult organism. The mitotic activity of the 1-, 3- and 10-day-old animals was the maximum in the evening and the minimum-in the morning hours. A definitive rhythm of cells division (with the maximal mitotic activity in the morning and the minimal-in the evening) is established beginning from the 17th day of the postnatal development. The average mitotic activity is very high in the embryos, but it falls immediately after birth. It rises on the 3rd day, and begins to decrease again from the 7th day after birth.     

DNA synthesis in lactotrophs of primary rat anterior pituitary cell cultures. Effects of thyroliberin, bromocriptine and somatostatin

Prolactin immunostaining in combination with thymidine autoradiography was used to characterize changes in the DNA-synthesizing activity of lactotrophs in primary monolayer cultures of the rat anterior pituitary gland treated for 3 days with thyroliberin (TRH), somatostatin (SRIF) and bromocriptine (CB 154). The number of lactotrophs labelled with 3H-thymidine within the total pool of labeled pituitary cells was used to estimate DNA synthesis in prolactin-producing cells. TRH (10 ng/ml) stimulated DNA synthesis in the whole population of cultured cells but not in lactotrophs. TRH only weakly counteracted the noticeable inhibitory effect of CB 154 (0.75 microM/l) on thymidine incorporation into lactotrophs. SRIF (20 ng/ml) inhibited DNA synthesis in lactotrophs to a lesser extent than CB 154. The combination of methods used in this paper may be useful for studying the selective effects of regulators on the proliferative activity of various pituitary cell types in vivo and in culture.     

Secretory plasticity of pituitary cells: a mechanism of hormonal regulation

Pituitary somatotropes and melanotropes have enabled us to investigate the molecular basis and functional dynamics underlying secretory plasticity, an ability of endocrine cells to adapt their activity to the changing physiologic requirements, which generates discrete cell subpopulations within each cell hormonal type. Porcine somatotropes comprise two morphologically distinct subpopulations of low- (LD) and high-density (HD) cells, separable by Percoll gradient, that respond differently to hypothalamic regulators. In LD somatotropes, somatostatin (SRIF) inhibits growth hormone (GH)-releasing hormone (GHRH)-induced GH secretion. Conversely, SRIF alone stimulates GH release from HD somatotropes. These disparate SRIF actions entail a molecular signaling heterogeneity, in that SRIF increases cAMP levels in HD but not in LD cells as a requisite to stimulate GH release. GHRH-stimulated GH release also involves differential signaling in LD and HD cells: although it acts primarily through the cAMP/extracellular Ca2+ route in both somatotrope subsets, full response of LD somatotropes also requires the inositol phosphate/intracellular Ca2+ pathway. Amphibian melanotropes, which regulate skin adaptation to background color by secreting POMC-derived alpha-melanocyte-stimulating hormone (alphaMSH), also comprise two subpopulations with divergent secretory phenotypes. LD melanotropes show high biosynthetic and secretory activities and high responsiveness to multiple hypothalamic factors. Conversely, HD melanotropes constitute a hormone-storage subset poorly responsive to regulatory inputs. Interestingly, in black-adapted animals most melanotropes acquire the highly-secretory LD phenotype, whereas white-background adaptation, which requires less alphaMSH, converts melanotropes to the storage HD phenotype. These same interconversions can be reproduced in vitro using appropriate hypothalamic factors, thus revealing the pivotal role of the hypothalamus in regulating the functional dynamics of the secretory plasticity. Furthermore, this regulation likely involves a precise control of the secretory pathway, as suggested by the differential distribution in LD and HD melanotropes of key components of the intracellular transport, processing, and storage of secretory proteins. Hence, molecular signaling heterogeneity and unique secretory pathway components seem to relevantly contribute to the control of secretory plasticity, thereby enabling endocrine cells to finely adjust their dynamic response to the specific hormonal requirements.     

Thyrotropin regulation of plasminogen activator activity in primary cultures of ovine thyroid cells

In primary cultures of ovine thyroid cells, a high level of plasminogen activator (PA) activity was detected in the culture media. This level is much higher than in primary cultures of Sertoli cells, granulosa cells, and pituitary cells. PA activity increased with time in culture and was regulated by TSH and insulin. Activity gel analysis of the culture media revealed a major band of 43,000 daltons and a minor one of 70,000 daltons, suggesting the presence of both of the urokinase-type and the tissue-type PA in the media.     

Altered hypothalamic monoamine metabolism and pituitary prolactin regulation in female spontaneously hypertensive rats

Prolactin (PRL) secretion after aromatic amino acid decarboxylase inhibition with NSD-1015 was significantly elevated in female spontaneously hypertensive rats (SHR) as compared to normotensive (WKY) controls. Although basal PRL levels tended to be elevated in SHR rats, the differences were not significant. In vitro PRL secretion was also significantly elevated in the SHR rats as compared to the WKY rats, but the SHR rats were more responsive to the inhibitory effects of dopamine (DA). Despite changes in pituitary PRL secretion and DA response, there was no apparent difference in tubero-infundibular DA activity between the two rat strains. Hypothalamic serotonin levels were elevated in SHR rats, but metabolism did not appear to be significantly changed based on measurements of 5-hydroxytryptophan accumulation after NSD-1015 treatment.     

Effect of dexamethasone on the release of pituitary hormones in monolayer cultures of rat pituitary cells

The effect of dexamethasone on the release of ACTH, GH, PRL, LH and TSH was studied in monolayer cultures of rat pituitary cells in 4-hour incubation. With or without the addition of rat hypothalamic extract, the release of GH was significantly inhibited by dexamethasone at concentrations higher than 10(-9) M, although less remarkably than that of ACTH. Intracellular ACTH and GH were unchanged. PRL, LH and TSH were not affected. These results indicate that dexamethasone, when exerted for 4 hours, suppressed the release of GH as well as ACTH, at least in part, at the pituitary level.     

Rapid down regulation of pyroglutamyl peptidase II activity by arachidonic acid in primary cultures of adenohypophyseal cells

Thyrotropin releasing hormone (TRH; pglu-his-proNH2) is inactivated, in the extracellular space, by pyroglutamyl aminopeptidase II (PPII), a narrow specificity ectopeptidase. In adenohypophysis, multiple hormones regulate PPII surface activity. The intracellular pathways of regulation are still poorly understood. Since some of the neurohormones which regulate PPII activity, including TRH and dopamine, transduce in part their effect through modulation of arachidonic acid (AA) mobilization, we have tested its role in regulation of PPII activity in primary cultures of rat adenohypophyseal cells. Melittin concentrations from 0.25 to 1 ug/ml induced a rapid decrease of PPII activity; 0.5 ug/ml caused a maximum effect (38-45% inhibition) at 20-30 min. AA (0.5 or 5 uM) also inhibited PPII activity (42-72%, maximum at 20 min); AA effect was reversible, with values approaching control at 1 h. The inhibitory effect of AA was blocked by lipoxygenase (10 uM nordihidroguaiaretic acid) but not ciclooxygenase inhibitors (10 uM indomethacin) suggesting the involvement of the lipoxygenase pathway. These data show that production of arachidonic acid by adenohypophyseal cells can rapidly but transiently down regulate surface PPII activity. This is the first evidence that AA mobilization can regulate the activity of an ectopeptidase.     

Somatotrophs and lactotrophs in the anterior pituitary of fetal and neonatal rats

Summary The ultrastructure of immunoreactive somatotrophs and lactotrophs in pituitaries of fetal rats at 19, 20 and 21 days of gestation and on the day of birth was studied. Somatotrophs, first detectable at 19 days of gestation, undergo only minor modifications before reaching the structure described for adults. In particular there is an increase in the endoplasmic reticulum and Golgi apparatus. Lactotrophs, first identifiable in newborn rats, are very different in ultrastructure from adult cells, because the secretory granules are generally small, but variable in shape and size, and the Golgi complex is prominent.     

The regulation of saccharase and phosphofructokinase activity in the small intestine of chicks of different ages]

Studies have been made on the onset and regulation of the activity of sucrase and phosphofructokinase from different parts of the small intestine of chicks after hatching up to the 30th day of postnatal life. It was shown that within this period the activity of both enzymes undergoes synchronous changes. Carbohydrate loading increases the activity of sucrase only in 30-day chicks, its effect being similar to that of a stimulator from the duodenum. Changes in the activity of phosphofructokinase exhibit a different pattern being significantly dependent on the region of the small intestine.     

Stimulation of arachidonic acid metabolism in primary cultures of osteoblast-like cells by hormones and drugs

The effects of parathyroid hormone (PTH), dihydroxycholecalciferol (1,25-(OH)₂ D₃), thrombin, epidermal growth factor (EGF) and 12-o-tetradecanoylphorbol-13-acetate (PMA) on the biosynthesis and release of arachidonic acid metabolites were studied in primary cultures of osteoblast-like cells isolated from 18-day-old chick embryo calvaria. Cells were labelled with (¹⁴C)-arachidonic acid for 30 h. The radioactive eicosanoids were extracted from the cell culture media after a further 30 h stimulation period and analysed on a PRP-1 column by HPLC. The radioactive products were characterized by co-elution of (³H) standard prostanoids. Osteoblasts showed a basal release of the prostanoids 6-keto-PGF_1α, TXB₂, PGF_2α, PGE₂, PGD₂ and PGB₂, the latter being the most abundant one. Indomethacin (10⁻⁵ M) effectively inhibited the basal release, but not that of an as yet unidentified compound. The release of prostanoids was stimulated by PTH (2 U/ml), thrombin (0.4 NIH/ml), EGF (50 ng/ml) and PMA (25 ng/ml), the latter being by far the most potent one. 1,25-(OH)₂D₃ was found to slightly inhibit the prostanoid release. These results indicate: (1) primary cultures of osteoblasts synthesize several prostaglandins, thromboxane B₂ and one unidentified product. (2) the action on bone of PTH and the various drugs tested may be, at least partly, mediated by an increased prostaglandin production by osteoblasts. Clearly this does not apply to 1,25-(OH)₂D₃.     

Stimulation of arachidonic acid metabolism in primary cultures of osteoblast-like cells by hormones and drugs

The effects of parathyroid hormone (PTH), dihydroxycholecalciferol (1,25-(OH)2 D3), thrombin, epidermal growth factor (EGF) and 12-o-tetradecanoylphorbol-13-acetate (PMA) on the biosynthesis and release of arachidonic acid metabolites were studied in primary cultures of osteoblast-like cells isolated from 18-day-old chick embryo calvaria. Cells were labelled with (14C)-arachidonic acid for 30 h. The radioactive eicosanoids were extracted from the cell culture media after a further 30 h stimulation period and analysed on a PRP-1 column by HPLC. The radioactive products were characterized by co-elution of (3H) standard prostanoids. Osteoblasts showed a basal release of the prostanoids 6-keto-PGF1 alpha, TXB2, PGF2 alpha, PGE2, PGD2 and PGB2, the latter being the most abundant one. Indomethacin (10(-5) M) effectively inhibited the basal release, but not that of an as yet unidentified compound. The release of prostanoids was stimulated by PTH (2 U/ml), thrombin (0.4 NIH/ml), EGF (50 ng/ml) and PMA (25 ng/ml), the latter being by far the most potent one. 1,25-(OH)2D3 was found to slightly inhibit the prostanoid release. These results indicate: (1) primary cultures of osteoblasts synthesize several prostaglandins, thromboxane B2 and one unidentified product. (2) the action on bone of PTH and the various drugs tested may be, at least partly, mediated by an increased prostaglandin production by osteoblasts. Clearly this does not apply to 1,25-(OH)2D3.     

Thyroid hormones influencing renal electrolyte excretion in saline loaded rats of different ages

Thyroid hormones are involved in the postnatal development of kidney functions. The influence of pretreatment with thyroid hormones on renal electrolyte excretion was measured in rats of different ages. Pretreatment with triiodothyronine or tetraiodothyronine (10 micrograms/100 g b.wt. i.p. for 3 days, once a day) cannot alter the low sodium excretion in young rats receiving a saline load. In 33-, 55- and 105-day-old rats the pretreatment with thyroid hormones is followed by a preferential increase of renal sodium excretion. Both thyroid hormones are effective.     

Participation of hypothalamic adrenergic and cholinergic systems in the regulation of pituitary adrenocorticotropic function]

Experiments were conducted on male rats; a study was made of the activity of pituitary-adrenal cortex system under conditions of electrolyte destruction of various hypothalamic areas. Pharmacological analysis pointed to the irregular distribution in the hypothalamus of the cholinergic and adrenergic systems controlling the secretion of ACTH-glucocorticoids. As suggested, the cholinergic systems are situated in the mammillary region, alpha-adrenergic ones--in the anterior or the posterior regions of the hypothalamus, and beta-adrenergic receptors--in the ventromedial nuclei of the hypothalamus. The latter possibly play an inhibitory role.