Extensive ultrastructural changes in rat mammotrophs following administration of the dopamine agonist ergocristine-reflecting inhibition of prolactin release

Summary We have demonstrated an extensive reorganization of organ elles in mammotrophs immediately following administration of ergocristine (a dopamine agonist) to estradiol-primed male rats. Our ultrastructural findings are consistent with our previous results that ergocristine can block prolactin release without any noticeable latent period. Following three-week priming of male rats with estradiol implants, ergocristine was administered by a bolus injection through an indwelling cannula. Within two min of its administration, ergocristine induced dramatic changes in the ultrastructure of mammotrophs, i.e., (1) increased numbers of secretory granules, (2) peripheral relocation of rough endoplasmic reticulum which tends to sequester secretory granules, (3) change in location of nucleus and (4) increased numbers of intracellular bodies associated with secretory granules. We suggest that the extensive ultrastructural changes that occurred in such a short period following ergocristine administration may be indications of specific factors associated with blockage of hormone release.     

Secretory granules of prolactin cells of neonatally thyroidectomized female rats

In the anterior pituitary glands of neonatally thyroidectomized female rats sacrificed at 30 days of age, the prolactin granules were small and spherical in shape. The administration of thyroxine to neonatally thyroidectomized rats produced an obvious increase in the number and size of secretory granules in prolactin cells; comparatively large, pleomorphic secretory granules were frequently observed in these cells. These enlarged and pleomorphic granules closely resembled those observed in the prolactin cells of sham-operated control rats. These results may indicate that thyroxine stimulates the basic metabolism or cellular function of prolactin cells of neonatally thyroidectomized rats and leads to the formation of prolactin granules that are similar to those of sham-operated control rats.     

Calcium concretions in the pineal gland of aged rats: an ultrastructural and microanalytical study of their biogenesis

The genesis of calcium concretions in aged rats was studied by means of transmission and scanning electron microscopy. The potassium pyroantimonate method, combined with X-ray microanalysis, allowed us to study the distribution of cations and calcium. Notable accumulations of calcium (associated with phosphorus) were localized in vesicles, vacuoles, lipid droplets, lipopigments, and mitochondria of dark pinealocytes. The results obtained in the present investigation suggest that these organelles are involved in the genesis of the concretions. The presence of sulfur indicates the existence of an organic matrix. We propose that genesis takes place in dark pinealocytes, which contain more calcium than light pinealocytes. Mineralization foci are some-times associated with cellular debris and enlarge by further apposition of material. Two types of concretions, as determined by electron microscopy and confirmed by electron diffraction, could be observed: the amorphous type with concentric layers and the crystalline type with needle-shaped crystals. Once formed, the concretions reach the extracellular space and the cell breaks down. Possible extracellular calcification is suggested in the extracellular calcium-rich floculent material. The mineralization process is interpreted as being an age-related phenomenon and mainly a consequence of the degeneration of pinealocytes.     

Subcellular distribution of laminin and prolactin in stimulated and blocked prolactin cells in the pituitary of lactating rats

Summary Laminin (LAM), a glycoprotein component of basement membranes, has been previously detected within several subcellular compartments of prolactin (PRL) cells in the pituitary gland. The present work was aimed at comparing the subcellular localization of PRL, a specific secretory product, with that of LAM, in relation to the secretory activity of PRL cells. LAM and PRL were located in parallel, by ultrastructural immunocytochemistry, in PRL cells of lactating female Wistar rats, either stimulated by suckling, or blocked by weaning, or reactivated by suckle following short-term weaning. Variations in physiological conditions were correlated with a redistribution of PRL immunoreactivity within morphologically modified compartments. The Golgi apparatus became hypertrophied, and PRL impressively accumulated within saccules of the Golgi stacks of blocked cells. On the contrary, no apparent changes occurred in LAM distribution, at least at the Golgi level. Only a slight increase of LAM immunoreactivity was observed in rough endoplasmic reticulum after a long weaning period. PRL could be detected in most of the secretory granules and particularly in forming elements, whereas LAM was observable at the peripheral edge of some mature granules. Such a labeling was not markedly influenced by the physiological state. The prominent structures, indicative of crinophagic activity, characteristic of blocked cells, contained masses of dense material, which were always immunopositive with antibodies to PRL, but never to LAM. These observations could suggest that, in PRL cells, intracellular transport and exportation of LAM are controlled by mechanisms independent from those involved in the regulation of PRL secretion.     

Immuno-electron-microscopic study of the prolactin cells in the pituitary gland of male Wistar rats during aging

Summary Prolactin cells were identified by means of immunocytochemistry with protein-A gold as a marker on ultrathin sections of the pituitary gland of young (3–4 months), middle-aged (16–19 months), and aged (26–30 months) male Wistar rats. Point-counting volumetry revealed that the prolactin (PRL) cell-volume density in middle-aged rats was significantly increased in comparison to the volume densities in young and aged rats. Within the PRL-cell population, four types of PRL cells were distinguished on the basis of the shape and size of their secretory granules. During aging, dramatic changes occurred in the relative volumes of the four cell types. The volume percentage of cells with round granules (type I, granule diameter 150–250 nm, and type IIA, granule diameter 250–350 nm) increased from ±30% in young rats to ±90% in old rats. The volume percentage of cells with round and polymorphic granules (type IIB; granule diameter 350–400 nm and type III; granule diameter 500–600 nm) decreased from ±70% in young rats to ±7% in old rats. Age-related changes in serum PRL levels were not found. It is concluded that although during the life span of the male Wistar rat considerable changes in PRL-cell volume densities and in the ratios of PRL-cell types occur serum, PRL levels remain more or less constant.     

Effect of parachlorophenylalanine,a brain serotonin depletor,on the prolactin cells of the eel pituitary

Summary Parachlorophenylalanine (pCPA), an inhibitor of tryptophan hydroxylase depleting brain serotonin in higher vertebrates, was injected into freshwater eels. After 4 or 6 injections (200 mg/kg/day) or 10 injections (100 and 140 mg/kg/day) plasma electrolyte values were not modified. Prolactin (PRL) cells appear less active, with increased granulation after 6 and 10 injections. Their cell height (P < 0.01) and their nuclear area (P < 0.001) are reduced. As injections of 5-hydroxytryptophan stimulate PRL cells, these findings suggest that a serotoninergic system may participate in the regulation of PRL cell activity. Brain serotonin depletion probably decreases granule release in PRL cells, a result comparable to the lowering action of pCPA on the plasma PRL level in some mammals.     

Immunocytochemical identification of prolactin cells in the pituitary gland of tilapia larvae (Oreochromis mossambicus: Teleostei)

Summary Using an antiserum to highly purified chum salmon prolactin, prolactin cells were identified in the putative rostral pars distalis of newly hatched tilapia larvae (Oreochromis mossambicus) by the immunogold method for the electron microscope. In the putative rostral pars distalis, some cells had another kind of secretory granule which was much less numerous, much smaller in size, and without immunoreactivity to salmon prolactin antiserum. Controls incubated with salmon prolactin-preabsorbed antiserum or normal serum showed no immunoreactive cells, confirming the specificity of the antiserum. The possible role of prolactin in the osmoregulation of tilapia larvae is discussed.     

Modification of the hypothermic circadian cycles induced by DSIP and melatonin in pinealectomized and hypophysectomized rats

Both melatonin and DSIP (a nine amino acid peptide) effects have been previously shown to be (a) circadian rhythm related and (b) involved in inducing hypothermic effects in rats. In this study we report the hypothermia effects by each of these drugs alone and in combination when studied in normal (unoperated), pinealectomized, and hypophysectomized rats at various time points of the corresponding circadian cycle. A clear differential effect of drugs × time × preparation was found. While both DSIP and melatonin hypothermic effects were both circadian cycle dependent in intact rats the rhythmicity of melatonin hypothermic effect in pinealectomized rats, and DSIP hypothermic effect in hypophysectomized rats was missing. Although several hypotheses have been offered to account for the physiological mechanism(s) that govern the effects of the drugs, it is not yet possible to reliably relate the findings to existing neurochemical theory.     

Effect of an antagonistic analog of LH-RH on haloperidol-induced hyperprolactinemia in female rats

The effects of prolonged treatment with the antagonistic analog of LH-RH (N-Ac-D-p-Cl-Phe1,2, D-Trp3,D-Arg6,D-Ala10) LH-RH (ORG 30276) on the hyperprolactinemia induced by haloperidol were investigated in intact or ovariectomized female rats. Treatment with ORG 30276 for 20 days significantly reduced prolactin levels elevated by daily injections of haloperidol in intact as well as in ovariectomized rats. Administration of ORG 30276 also significantly decreased serum LH levels in both types of rats. It is concluded that the LH-RH antagonist ORG 30276 is able to counteract the hyperprolactinemic effect of haloperidol. This effect might be due to a blockade of the action of endogenous LH-RH on the gonadotrophs, which results in a suppressing of the paracrine action of these cells on the lactotroph.     

The effects of bromocriptine and prolactin on porphyrin biosynthesis and morphology in the female hamster Harderian gland

Porphyrin biosynthesis was examined in the Harderian gland of the female golden hamster by fluorometric assays of gland porphyrin content and by measuring the activity of a rate-limiting enzyme for haem biosynthesis, -aminolaevulinic acid synthase. Both porphyrin content and enzyme activity are high in normal female glands. Enzyme activity was lowered in females ovariectomised for 6 weeks, and both enzyme activity and porphyrin content were greatly lowered in ovariectomised females given the dopamine agonist bromocriptine; this suppression could be prevented by simultaneous prolactin administration. Bromocriptine (but not ovariectomy alone) also masculinised the morphology of the Harderian gland, resulting in the appearance of type II cells and polytubular complexes; again, the simultaneous administration of prolactin prevented masculinisation. The results support the hypothesis that while androgens have an inhibitory effect on porphyrin synthesis within this model, prolactin may have a major facilitatory role.Abbreviations ALA -aminolaevulinic acid - ALA-s -aminolaevulinate synthase     

Immunocytochemical and immuno-electron-microscopical study of growth hormone cells in male and female rats of various ages

Summary Growth hormone (GH) secretory cells were identified by immunogold cytochemistry, and were classified on the basis of the size of secretory granules. Type I cells contained large secretory granules (250\2-350 nm in diameter). Type II cells contained the large secretory granules and small secretory granules (100\2-150 nm in diameter). Type III cells contained the small secretory granules. The percentages of each GH cell type changed with aging in male and female rats of the Wistar/Tw strain. Type I cells predominated throughout development; the proportion of type I cell was highest at 6 months of age, and decreased thereafter. The proportion of type II and type III cells decreased from 1 month to 6 months of age, but then increased at 12 and 18 months of age. The pituitary content of GH was highest at 6 months of age, and decreased thereafter. Estrogen and androgen, which are known to affect GH secretion, caused changes in the proportion of each GH cell type. The results suggest that when GH secretion is more active the proportion of type I GH cell increased, and when GH secretion is less active the proportion of type II and type III cells increased. The type III GH cell may therefore be an immature type of GH cell, and the type I cell the mature type of GH cell. Type II cells may be intermediate between type I and III cells.     

Occurrence and regional distribution of calcareous concretions in the rat pineal gland

Summary Serial sections of 90 Sprague-Dawley rat brains with the pineal in situ were scanned to determine the occurrence and regional distribution of calcareous concretions within the pineal gland and its surrounding leptomeningeal tissue. In 90 % of the cases examined concretions were found in varying number and appearance, predominantly lying in the dorsal region of the pineal gland and in the distal portion of the pineal stalk.Discussing the hypothesis advanced by Lukaszyk and Reiter (1975) that the origin of pineal concretions may be related to a neurosecretory process involving a pineal carrier protein, called neuroepiphysin, it is thought that, in view of the intra- and extra-pineal occurrence of concretions, processes other than secretion should be considered. Since in the pineal organ lymphatics are lacking it may well be that, due to a reduced drainage of tissue fluid, the coagulation of intercellular organic debris mingled with minerals increases with age.Supported by a grant (Vo 135/4) of the Deutsche Forschungsgemeinschaft within the Schwerpunktprogramm Neuroendokrinologie     

Occurrence and regional distribution of striated muscle fibers in the rat pineal gland

Summary In a total of 96 rat pineals studied 31 were found to contain striated muscle fibers or their precursors. The muscle fibers were most frequently present in the stalk region and more frequently found in the left than in the right hemisphere. Size measurements revealed that the lengths of pineal muscle cell nuclei differ only slightly from those of the sphincter muscle of the iris. However, the yellowish appearance of pineal muscle cell nuclei under darkfield investigation, a phenomenon observed in all muscular tissues of mesenchymal origin and connective tissue cells, may support the hypothesis that pineal musculature is of mesenchymal rather than ectodermal origin.Supported by a grant (Vo 135/4) of the Deutsche Forschungsgemeinschaft within the Schwerpunktprogramm Neuroendokrinologie     

Effect of TRH (thyrotropin-releasing hormone) on the fine structure and replication of TSH and prolactin cells in the rat

Summary In intact male rats after TRH administration for 7 and 14 days, TSH cells showed similar morphological changes to those observed after thyroidectomy. These changes were paralleled by small numerical increases in TSH cell counts. After 34 days of TRH treatment, however, most of the TSH cells had a normal appearance and the number of TSH cells also had returned to normal. TRH treatment for 7, 14 and 34 days caused morphological changes in Prolactin cells similar to those obtained after a suckling stimulus. In the three groups these changes were also paralleled by small numerical increases in Prolactin cell counts. The cell replication after TRH for 7 and 14 days, as measured by incorporation of tritiated thymidine to obtain a labeling index, was slightly but significantly increased.This work was supported by grants MA-552 and MT-2701 from the Medical Research Council of Canada. The authors wish to thank Dr. D.A.J. Ives, Connaught Medical Research Laboratories, Toronto, for providing the TRH, and Mr. G. Penz for technical assistance.Fellow of the Medical Research Council of Canada.     

Tentative immunohistochemical demonstration of melatonin in the rat pineal gland

Summary In the present study an attempt was made to demonstrate melatonin in the rat pineal gland by means of immunohistochemistry. The anti-body used was raised against 5-methoxy-N-acetyltryptophan which is chemically similar to melatonin. Specific fluorescence was demonstrable only in pineals from rats killed during the night, when melatonin formation is high. It was restricted to parenchymal cells lying in a marginal zone of the organ. These results are discussed in relation to a subdivision of the pineal parenchyma into cortical and medullary areas.Supported by a grant of the Deutsche Forschungsgemeinschaft (VO 135/4) within the Schwerpunktprogramm Neuroendokrinologie     

Ultrastructural localization of prolactin,growth hormone and luteinizing hormone by immunocytochemical techniques in the bovine pituitary

Summary The technique of ultrastructural immunocytochemistry involving the unlabeled antibody and the soluble peroxidase-antiperoxidase complex was used to identify and describe the prolactin (P) cells, somatotropic (STH) cells and luteinizing hormone (LH) cells in the bovine anterior pituitary gland. This method was used to localize the three hormones at the electron microscopic level. Staining of varying intensity was found on the secretory granules and on the small granules and vesicles within the Golgi complex. No stain was found in nuclei, on mitochondria or in the endoplasmic reticulum.     

Effects of prolactin-releasing peptide on tuberoinfundibular dopaminergic neuronal activity and prolactin secretion in estrogen-treated female rats

Both systemic and central effects of a newly discovered prolactin (PRL)-releasing factor (PRF), prolactin-releasing peptide (PrRP), were determined in this study. Systemic injection of PrRP (1 and 10 microg/rat, i.v.) stimulated PRL secretion in ovariectomized, estrogen-treated rats similar to the effect of another PRF, thyrotropin-releasing hormone (TRH). Pretreatment with a dopamine D2 receptor antagonist, sulpiride (1 microg/rat, i.v.), potentiated the stimulatory effect of both PrRP and TRH on PRL secretion. Using the double-labeling immunohistochemical method, PrRP-immunoreactive terminals were found in close contact with tyrosine-hydroxylase-immunoreactive neurons in the hypothalamic arcuate nucleus. Central administration of PrRP (0.1-1,000 ng/rat, i.c.v.) stimulated tuberoinfundibular but not nigrostriatal dopaminergic neuronal activity in 15 min. Levels of 3,4-dihydroxyphenylacetic acid (DOPAC) in the median eminence and striatum were used as indices for tuberoinfundibular dopaminergic (TIDA) and nigrostriatal dopaminergic neuronal activities, respectively. The serum PRL level, however, was not significantly changed. Similar treatment with TRH (10 ng/rat, i.c.v.) stimulated and inhibited TIDA neuronal activity and serum PRL, respectively, at 30 min. In summary, PrRP may play a role in both the central and peripheral control of PRL secretion.     

Calcium content and concretions of pineal glands of young and old rats

Summary Calcium content and pineal concretions were studied in young (2–3 months) and old (28 months) Wistar rats. Samples, deep-frozen by liquid propane/isopentane and freeze-dried were analysed by means of X-ray microanalysis in a scanning electron microscope. Total semi-quantitative measurements revealed that pineals of old rats showed a marked increase of calcium compared with the pineals of young rats. It is thus suggested that a calcium-rich environment is responsible for the growth of pineal concretions, which only appear in old rats. Pineal calcifications in rats could thus be an indicator of aging and/or of a degenerating state.     

Changes in vasopressin-converting aminopeptidase activity in the rat pineal gland during summer: Relationship to vasopressin contents

Vasopressin (VP)-converting aminopeptidase (VP-AP) activity and VP contents were measured in single rat pineal glands during the summer of two successive years. The peptidase activity decreased significantly in August. The lowest activity (±SEM) of 0.18±0.02 pmol·hour⁻¹ was recorded on August 14, compared to the basal activity of 0.25±0.01 pmol·hour⁻¹ in July and September of 1986. The change with similar percentage occurred in the same period of 1987. The specific activity of the enzyme in the crude homogenate, 15,000 g pellet and supernatant fraction of rat pineal glands, exhibited the same pattern of variations. The decrease in peptidase activity coincided with the previously reported dramatic rise in pineal VP content in early August which was confirmed in this series of experiments. Another peptidase, the so-called γ-endorphin generating endopeptidase (γ-EGE) activity, and β-endorphin-related peptides in the pineal gland did not change in this period. The results show that the variations of pineal VP contents and VP-AP activity during summer are not general for other peptides and peptidases. The coincidence of opposite changes in VP content and VP-AP activity of the pineal gland may indicate a role of the peptidase activity to regulate the VP content.