Innervation of the mink pineal gland with neuropeptide Y (NPY)-containing nerve fibers

Summary An immunohistochemical investigation of the mink pineal gland was performed by use of antibodies raised in rabbits against neuropeptide Y (NPY) and Cys-NPY (32–36)-amide recognizing neuropeptide Y with an amidation at position 36 (NPYamide). NPY-immunoreactive nerve fibers were located predominantly in the rostral part of the pineal gland and in the pineal stalk. Immunoreactive nerve fibers were found throughout the pineal gland, but the number of fibers in the caudal part of the gland was low. The fibers were present both in the perivascular spaces and between the pinealocytes. Many NPY-immunoreactive fibers were also located in the posterior and habenular commissures; some of these fibers were connected with the fibers in the rostral part of the mink pineal gland, indicating that at least some of the NPY-immunoreactive nerve fibers are of central origin. The nerve fibers immunoreactive to amidated NPY were distributed in a similar manner. However, the number of fibers immunoreactive to NPYamide was lower than the number of fibers immunoreactive to NPY itself. After removal of the superior cervical ganglia bilaterally 22 days or 12 months before sacrifice, NPY-immunoreactive nerve fibers remained in the gland. This immunohistochemical study of the mink pineal gland therefore shows that the NPY/NPYamide-immunoreactive nerve fibers innervating the pineal gland in this spegcies are a component of the central innervation or originnate from extracerebral parasympathetic ganglia.     

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     

Immunohistochemical evidence for the presence of melatonin in the pineal gland,the retina and the Harderian gland

Summary The presence of melatonin is demonstrated in the pineal gland, the retina and the Harderian gland in some mammalian and non-mammalian vertebrates, using a specific fluorescence labelled antibody technique. Four different potent antibodies against melatonin have been used and compared. In the pineal gland of hamsters, mice, rats and snakes, specific fluorescence, mostly restricted to the cytoplasm of the cells, is detected in pinealocytes. Fluorescence is also detected in the pineal organ of fishes, tortoises and lizards, but it has not been possible, from cryostat sections of fresh tissue, to assert which kind of cell is reacting (photoreceptor cells or interstitial ependymal cells). In the retina, fluorescence is almost exclusively restricted to the outer nuclear layer. In the Harderian gland of mammals and reptiles, fluorescence is localized in the secretory cells of the alveoli and mostly restricted to the cytoplasm surrounding the nucleus. These results are discussed in relation to the concept of melatonin synthesis at extrapineal sites independent of pineal production.Parts of this work have been presented in the Xth Conference of Comparative Endocrinologists, Sorrento, May 20–25, 1979 (Vivien-Roels and Dubois 1980) and the VIth International Congress of Endocrinology, Melbourne, February 10–16, 1980 (Vivien-Roels et al. 1980)The author wishes to thank Professor Lutz Vollrath who has accepted her in his laboratory for a short period, Doctor George M. Bubenik for his suggestions and critical remarks, Dr. L.J. Grota for producing the melatonin diazobenzoic acid-BSA and Dr. Castro for preparing one of the melatonin derivates     

Ontogenesis of corticotropes and lactotropes in situ in the pituitary gland of the hamster

Summary The development of corticotropes and lactotropes was investigated in the golden Syrian hamster using an anti-porcine ACTH antiserum and a homologous antihamster PRL antiserum. Oval corticotropes were first visible in the ventral region of the pars distalis at 13 days of gestation. By the end of gestation, corticotropes were found throughout the pars distalis and in the pars intermedia. Corticotropes in the pars distalis of postnatal hamsters were either round or irregularly-shaped, often appearing in clusters. Throughout development, corticotropes often appeared to be surrounding other cells. Scarce, very small lactotropes were first observed in the pars distalis of hamsters on the first postnatal day. The number of these cells, which were either round or polyhedral, increased dramatically between 4 and 20 days of postnatal life. These observations indicate that the sequence of appearance of corticotropes and lactotropes in the hamster is similar to that in other species and that lactotropes are confined to the pars distalis of postnatal hamsters.     

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.     

Population genetic structure and distribution of introduced American mink (<Emphasis Type="Italic">Mustela vison</Emphasis>) in Spain,based on microsatellite variation

The population genetic structure of an invasive species in Spain, the American mink (Mustela vison), was investigated using microsatellite DNA markers. This semi-aquatic carnivore, originating from North America, was imported into Europe for fur farming since the beginning of the 20th century. Due to massive escapes, farm damages, deliberate releases and/or accidents, feral mink populations were established in the aquatic ecosystems of many European countries, including Spain. We genotyped 155 American mink originating from the Spanish regions Basque Country, Catalonia, Castilla-Leon, Aragon, Valencia and Galicia using 10 polymorphic microsatellite loci to highlight population genetic structure, distribution and dispersal. M. vison populations in Spain appear differentiated and not yet connected by gene flow. Bayesian clustering analyses and spatial analyses of molecular variance detected four inferred clusters, overall coinciding with the sampled geographical localities. Preliminary testing shows moderate to large estimated effective population sizes. Molecular analyses result useful to provide baseline data for further research on the evolution of invasive mink populations, as well as support local management strategies and indirectly benefit the conservation of threatened species in Spain, such as the endangered European mink (Mustela lutreola), and the polecat (Mustela putorius), which share the habitat with the American mink. This paper is dedicated to the memory of Xavier Domingo-Roura.     

Gonadotropin-releasing hormone neurons and pathways in the brain of the female mink (Mustela vison)

Summary The distribution of gonadotropin-releasing hormone-immunoreactive neurons and processes was mapped in the female mink brain using coronal, horizontal and sagittal sections. Perikarya were found along a ventral continuum including the olfactory tubercle, the diagonal band of Broca, the lateral septum, the preoptic and anterior hypothalamic area and the mediobasal hypothalamus; 80% of the perikarya were counted in the mediobasal hypothalamus. Fibres were mainly observed in the organum vasculosum of the lamina terminalis and the median eminence. A few processes terminated in the ependymal cells lining the third and lateral ventricles. The total number of immunoreactive perikarya was the highest in the brains of females sacrificed in July; it then significantly decreased until December. This variation is discussed in relation to the annual breeding cycle.     

Characterization of oligosaccharides in milk of a mink, Mustela vison

Carbohydrates were extracted from a sample of milk from a mink, Mustela vison (Family Mustelidae). Free neutral and acidic oligosaccharides were isolated from the carbohydrate fraction and their chemical structures were compared with those of white-nosed coati (Nasua narica, Procyonidae) and harbour seal (Phoca vitulina, Phocidae) that we had studied previously. The ratio of free lactose to milk oligosaccharides was similar to that in milk of the white-nosed coati; in both species, this ratio was much lower than that in the milk of most eutherians. The neutral oligosaccharides of mink milk had alpha(1-3)-linked Gal or alpha(1-2)-linked Fuc residues at their non-reducing ends, as in the neutral oligosaccharides of white-nosed coati milk. Some of the neutral and acidic oligosaccharides, determined here, had been found also in harbour seal milk, but the harbour seal oligosaccharides did not contain alpha(1-3)-linked Gal residues.     

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.     

Ligh- and electron-microscopic immunocytochemistry of somatotropes in the anterior pituitary gland of European ferret,Mustela putorius furo

Light-microscopic immunocytochemistry of ferret anterior pituitary revealed the localization of somatotropes in the pars distalis, but no immunoreactive cells were detected in the pars tuberalis. Ultrastructural studies by superimposition immunocytochemistry and immuno-electron microscopy, clucidated the morphological heterogeneity of these somatotropic cells. They were classified into 2 subtypes on the basis of size of the secretory granules. Type-I cells with small granules (mean diameter, 192 nm), were considered to be the immature somatotrop, while Type-II cells, with comparatively larger secretory granules (mean diameter, 257 nm), were considered to be the matured form of Type-I cells and the typical somatotropic cell-type, and were much more predominant than the Type-I cells. The fact that Type-II cells had a distinct Golgi zone and many mitochondria, while in Type-I cells the intracellular organelles were generally less developed, supports this suggestion. In addition to these two extreme subtypes, several intermediate forms were also encountered that may represent different transitional phases during the conversion of Type I to Type II. Protein A-gold immuno-electron microscopy illustrated the specific localization of growth hormone over the granules, with no labelling over any other cytoplasmic organelles of the 2 somatotrope subtypes.     

In vivo correlation between c-Fos expression and corticotroph stimulation by adrenocorticotrophic hormone secretagogues in rat anterior pituitary gland

In the anterior pituitary gland, c-Fos expression is evoked by various stimuli. However, whether c-Fos expression is directly related to the stimulation of anterior pituitary cells by hypothalamic secretagogues is unclear. To confirm whether the reception of hormone-releasing stimuli evokes c-Fos expression in anterior pituitary cells, we have examined c-Fos expression of anterior pituitary glands in rats administered with synthetic corticotrophin-releasing hormone (CRH) intravenously or subjected to restraint stress. Single intravenous administration of CRH increases the number of c-Fos-expressing cells, and this number does not change even if the dose is increased. Double-immunostaining has revealed that most of the c-Fos-expressing cells contain adrenocorticotrophic hormone (ACTH); corticotrophs that do not express c-Fos in response to CRH have also been found. However, restraint stress evokes c-Fos expression in most of the corticotrophs and in a partial population of lactotrophs. These results suggest that c-Fos expression increases in corticotrophs stimulated by ACTH secretagogues, including CRH. Furthermore, we have found restricted numbers of corticotrophs expressing c-Fos in response to CRH. Although the mechanism underlying the different responses to CRH is not apparent, c-Fos is probably a useful immunohistochemical marker for corticotrophs stimulated by ACTH secretagogues. This work was supported by the Jichi Medical University young investigator award.     

Ultrastructural studies on prolactin and growth hormone cells in Anguilla pituitaries in vitro

Summary Eel hemi-pituitaries were cultured in vitro on high or low sodium media which are known to affect differentially prolactin and growth hormone release. Ultrastructural examination of the prolactin cells after 24 h culture showed the Golgi bodies were markedly more abundant and widely distributed in hemi-pituitaries from the low sodium medium. Secretory granule release profiles and dense bodies were also more frequent, but the percentage of the cytoplasmic volume occupied by secretory granules was lower than on the high sodium medium. RER was only slightly modified. Significant differences were noted in the shape and processes of the non-granulated (stellate) cells of the RPD, but there were only slight differences in the ultrastructure of the somatotropes.     

Alterations in the lipid content of pituitary gland and serum prolactin and growth hormone in cadmium treated rats

The present study was undertaken to assess whether chronic exposition to cadmium (Cd, 0.133 mM per liter for 2 months) through drinking water may affect the lipid contents in the pituitary anterior lobe (PAL) of adult male Wistar rats. As compared to metal non-exposed controls, PALs exposed to cadmium showed an increase in total phospholipid contents, which was associated to an increase of the incorporation of [1–¹⁴C]-methyl choline into phosphatidylcholine and of [U–¹⁴C]-glucose into total phospholipids. The incorporation of [1–¹⁴C]-methyl choline into sphingomyelin was not changed. Incorporation of [1–¹⁴C]-acetate into total fatty acids also increased but incorporation of [1–¹⁴C]-acetate into cholesterol did not change. The activity of phospholipase D decreased both in PALs from Cd exposed rats and in PAL dispersed cells treated with Cd in the culture medium from Cd non-exposed rats. In PALS from Cd exposed rats, a decrease of serum prolactin and growth hormone concentrations was determined. The results shown that cadmium modifies the lipid contents of pituitary gland and directly or indirectly the levels of prolactin and growth hormone in serum.     

Plasma leptin and thyroxine of mink (Mustela vison) vary with gender, diet and subchronic exposure to PCBs

Female minks (Mustela vison) fed diets based on freshwater, marine or mixed fish were exposed to 1 mg of polychlorinated biphenyls (PCBs) a day for 21 weeks. The plasma leptin and thyroxine concentrations and the glucose-6-phosphatase and glycogen phophorylase activities in the liver were measured at the end of the experiment. The plasma thyroxine concentrations were significantly higher in the group exposed to PCBs. The mean plasma leptin concentration and glucose-6-phosphatase activity was the highest in the group that had the lowest body-mass index (BMI). The glycogen phophorylase activity was the highest in the freshwater fish-control group. The results suggest that the amount of fat in the body of the female minks is not the only determinant of the plasma leptin levels, but the leptin levels seem to rise with a lowered BMI unlike in rodents or humans. The positive correlation between the leptin levels and the glucose-6-phosphatase activity suggests increased gluconeogenesis with high leptin levels. Subchronic exposure to PCBs seems to have no effect on the plasma leptin levels or the glucose-6-phophatase activities, but it elevates significantly the plasma thyroxine levels with a mechanism that remains unknown.     

The effects of bromocriptine and prolactin on porphyrin biosynthesis in the Harderian gland of the male hamster,Mesocritecus auratus

Porphyrin biosynthesis was examined in the Harderian gland of the male golden hamster by fluorometric assays of gland porphyrin content and by measuring the activity of a rate-limiting enzyme for haem biosynthesis, 5-aminolaevulinic acid synthase. Both porphyrin content and enzyme activity are low in normal male glands but were greatly raised in males castrated for 6 weeks. However, porphyrin synthesis remained at basal levels in castrates given the dopamine agonist bromocriptine; this suppression could be reversed by simultaneous prolactin administration, and castrated males receiving prolactin alone exhibited very high enzyme activity and porphyrin content. Bromocriptine also prevents the morphological feminisation of the Harderian gland which would normally occur after castration; again, the simultaneous administration of prolactin permits feminisation to occur. The results support the hypothesis that, while androgens have an inhibitory effect on porphyrin synthesis within this model, other factors, including prolactin, are permissive.Abbreviations ALA 5-aminolaevulinic acid - ALA-s 5-aminolaevulinate synthase - GH growth hormone     

哺乳动物昼夜节律组构中的下丘脑视交叉上核和松果腺

哺乳动物下丘脑视交叉上核（SCN）是昼夜节律最主要的起搏器,控制着机体的生理和行为的节律。它具有自身内在的节律性,同时也受光照周期信号和一些内源性化学物质的调节。检查腺分泌裉黑素（MEL）受SCN的调控,MEL通过作用于SCN上高亲和性MEL受体,启动第二、第三信使系统,调整SCN的昼夜节律活动。这种调整具有时间敏感性。     

Studies on the growth and development in Spirogyra

The phototropic response of Spirogyra sp. filaments and its relation with the different phases of their diurnal movements were studied. The filaments rapidly responded to unilateral irradiation; curvature towards the light source began in their tip region but shifted down to more basal regions. However, this typical and steady phototropic curvature could be observed only in the GnSt phase of the diurnal movement. In the other phases competitive states between the phototropic movement and other kinds of movement were evident. Thus, from the results of our previous and present studies it is proposed that the diurnal movement of Spirogyra filaments is composed of various individual movements, including a phototropic one; among these movements there exists a certain balance determined by the culture conditions and the time of day, and the phototropic movement is relatively inferior to the other movements.Abbreviations Gn negative gravitropism - Gp positive gravitropism - Sh shrinking (spiralling) growth - St straight form of growth V=Tanaka et al. (1983)     

Characterization and localization of gonadotropin-releasing hormone in the brain and pituitary of the three-spined stickleback,Gasterosteus aculeatus

Radioimmunoassay (RIA) studies on highperformance liquid chromatography (HPLC) fractions of brain extracts of the three-spined stickleback, Gasterosteus aculeatus, provided evidence for at least two forms of gonadotropin-releasing hormone (GnRH). One form showed chromatographic and immunological properties similar to that of synthetic salmon GnRH (sGnRH). A second, unidentified form of GnRH eluted in the same position as chicken GnRH I (cGnRH-I); however, it did not cross-react in a cGnRH-I RIA. Furthermore, it cannot be excluded that chicken GnRH II (cGnRH-II) and maybe one other unidentified form are present in the stickleback. The distribution of GnRH in the brain of breeding adult male sticklebacks was studied by use of immunohistochemistry. Two antisera against sGnRH and antisera against mGnRH and cGnRH-II were applied on cryosections and visualized using the peroxidase-antiperoxidase method. Staining patterns were similar after incubations with all four antisera. Immunoreactive fibers were found in most parts of the brain. Three distinct groups of GnRH-immunoreactive perikarya were found in the nucleus olfactoretinalis, in the nucleus anterior periventricularis, and in the nucleus lateralis tuberis. Moreover, weakly stained cells occurred in a periventricular position in the midbrain. The proximal pars distalis of the pituitary, housing the gonadotropic cells, was richly innervated by GnRH-positive fibers. In the pars intermedia and in the rostral pars distalis, immunoreactive fibers were absent.