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  • 1
    Electronic Resource
    Electronic Resource
    Springer
    Histochemistry and cell biology 97 (1992), S. 133-139 
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Pituitaries of the African catfish, Clarias gariepinus, were prefixed in aldehyde fixatives, frozen in liquid propane and submitted to a cryosubstitution procedure. Ultrathin sections of the Lowicryl HM20-embedded tissue were treated with primary antisera raised in rabbits to gonadotropin releasing hormone (GnRH), vasopressin or gamma amino butyric acid (GABA) respectively. Binding of the primary antisera was visualized with goat anti-rabbit (GAR) labeled with gold. The general morphology of the tissue components in the cryosubstituted pituitaries matches with that obtained after routine embedding procedures. In addition, a strong labeling intensity of the neuropeptides/neurotransmitters investigated in the present study was demonstrated. Due to these qualities cryosubstitution provides optimal conditions for studying co-localization of neurosecretory products, using double-immunostaining procedures. In the pars distalis of the catfish pituitary several types of hypothalamus-derived nerve fibers are present between or synapting on the secretory cells. It is demonstrated that the two known catfish GnRHs are co-localized in the same nerve fiber and within these nerve fibers even co-exist in the same neurosecretory granules. GABA and vasopressin-immunolabeling each occurred in different nerve fibers. The present data demonstrate that cryosubstitution and low temperature-embedding results in an excellent morphological preservation compared to ultracryotomy and a better preserved immunoreactivity of small antigenic molecules in comparison to conventional fixation and embedding techniques.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary In the pituitary of the African catfish, Clarias gariepinus, calcium precipitates were ultrastructurally visualized with the oxalate-pyroantimonate procedure (OPP). The presence of calcium in these precipiates was validated with several methods, including “Electron Energy Loss Spectrometry” (EELS). In the OPP-treated tissue calcium precipitates were seen in a) non-secretory stellate cells and b) gonadotropic (GTH-) cells. In the latter the amount of precipitate is generally low, but stimulation of the gonadotropin release, either in vivo or in vitro, resulted in a considerable increase. This increase is discussed in relation to the role of calcium as second messenger in the GTH-cells. Ca2+-ATPase was exclusively represented in stellate cells and GTH-cells, its strongest activity associated with the plasma membrane and with the membranes of the endoplasmic reticulum. The localization of this enzyme is discussed in relation to its role in the regulation of the intracellular calcium concentration in the GTH-cells. The stellate cells are considered to be involved in the regulation of extracellular calcium concentrations in the pituitary.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1432-0878
    Keywords: Tyrosine hydroxylase ; Dopamine ; Serotonin ; Immunohistochemistry ; CNS amphibian ; Triturus alpestris (Urodela)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Mapping of monoaminergic systems in the brain of the newt Triturus alpestris was achieved with antisera against (1) thyrosine hydroxylase (TH), (2) formaldehyde-conjugated dopamine (DA), and (3) formaldehyde-conjugated serotonin (5-HT). In the telencephalon, the striatum was densely innervated by a large number of 5-HT-, DA-and TH-immunoreactive (IR) fibers; IR fibers were more scattered in the amygdala, the medial and lateral forebrain bundles, and the anterior commissure. In the anterior and medial diencephalon, TH-IR perikarya contacting the cerebrospinal fluid (CSF-C perikarya) were located in the preoptic recess organ (PRO), the organum vasculosum laminae terminalis and the suprachiasmatic nucleus. Numerous TH-IR perikarya, not contacting the CSF, were present in the posterior preoptic nucleus and the ventral thalamus. At this level, DA-IR CSF-C neurons were only located in the PRO. In the posterior diencephalon, large populations of 5-HT-IR and DA-IR CSF-C perikarya were found in the paraventricular organ (PVO) and the nucleus infundibularis dorsalis (NID); the dorsal part of the NID additionally presented TH-IR CSF-C perikarya. Most regions of the diencephalon showed an intense monoaminergic innervation. In addition, numerous TH-IR, DA-IR and 5-HT-IR fibers, orginating from the anterior and posterior hypothalamic nuclei, extended ventrally and reached the median eminence and the pars intermedia of the pituitary gland. In the midbrain, TH-IR perikarya were located dorsally in the pretectal area. Ventrally, a large group of TH-IR cell bodies and some weakly stained DA-IR and 5-HT-IR neurons were observed in the posterior tuberculum. No dopaminergic system equivalent to the substantia nigra was revealed. The possible significance of the differences in the distribution of TH-IR and DA-IR neurons is discussed, with special reference to the CSF-C neurons.
    Type of Medium: Electronic Resource
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  • 4
    ISSN: 1432-0878
    Keywords: Median eminence ; Catecholamines ; Neuropeptides ; Immunocytochemistry ; Double labeling ; Ultrastructure ; Triturus alpestris (Urodela)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Dopaminergic and peptidergic nerve fibers were simultaneously demonstrated with a double-labeling technique at the ultrastructural level. The first antibody, raised against tyrosine hydroxylase, was applied during the preembedding phase and visualized with the peroxidase method. The second antibody, raised against one of the peptides met-enkephalin, somatostatin or gonadotropin-releasing hormone (GnRH), was applied to the ultrathin sections and visualized with gold-labeled goat anti-rabbit IgG. The fibers of both categories were present in the zona externa of the median eminence, frequently contacting the basal lamina of the portal vessels. In addition, topographical relationships between different types of nerve fibers were observed in the perivascular areas, although there were no morphological signs of synaptic specializations. Using serial sections, it could be established that one GnRH-fiber contacted both a dopaminergic fiber and a fiber immunoreactive for met-enkephalin. The observations support earlier physiological data concerning the regulation of the hypothalamo-hypophyseal axis, with special emphasis on the release of neurohormones in the median eminence of the newt.
    Type of Medium: Electronic Resource
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  • 5
    ISSN: 1573-5168
    Keywords: catfish ; pituitary ; ontogeny ; gonadotrops ; ultrastructure ; stereological analysis ; immunocytochemistry ; GTH subunits ; plasma androgen
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Description / Table of Contents: Résumé A l'aide d'une étude d'immunocytochimie ultrastructurale, nous avons suivi chez le poisson chat africain mâle âgé de 2 à 6 mois le développement des cellules gonadotropes hypophysaires en relation avec celui des testicules. Pendant cette période, 5 prélèvements hypophysaires et testiculaires ont été effectués (groupe I–V) alors que les prélèvements sanguins n'ont pu être pratiqués que dans les groupes III–V. Le développement testiculaire a été divisé en 3 stades c'est à dire immature (spermatogonies seulement, groupe I), début de spermatogenèse (spermatogonies et spermatocytes, groupe II et III), spermatogenèse avancée (tous les stades de cellules germinales incluant les spermatozïdes, groupe IV et V). Les niveaux sanguins en 11-ketotestostèrone étaient bas excepté dans le dernier groupe. Des anticorps ont été fabriqués contre l'α,βGtH-II et les sousunités α et β de GtH-II de poisson chat. Dans la partie proximale distale de l'hypophyse du poisson immature, des cellules indifférenciées, des cellules somatotropes, des cellules putatives thyrotropes (pTSH) et gonadotropes (pGtH) ont été trouvées. Dans les deux derniers types cellulaires, des granules de sécrétion ont été marqués avec l'anti-αGtH mais pas avec l'antiβ-GtHII. Les cellules présumées à TSH et GtH ont été distinguées par la taille des granules de sécrétion. Pendant le début de la spermatogenèse, 2 classes de cellules gonadotropes putatives ont pu être distinguées. Un type a les mêmes caractéristiques immunocytochimiques et ultrastructurales que chez les immatures; les granules de sécrétion dans le second type cellulaire, qui était plus abondant, répondaient aussi immunopositivement à l'anti-βGtHII. Le volume moyen des granules de sécrétion de ces cellules à GtHII était 3 fois supérieur à celui des cellules putatives à GtH apparaissant tôt. De plus, les cellules à GtHII apparaissant tardivement contiennent de grandes inclusions, connues sous le nom de globules. Ces structures étaient marquées avec l'anti-αβGtHII et avec l'anti-βGtHII, mais pas avec l'anti-αGtH. Il a été supposé que les globules sont impliqués dans un stockage différentiel et/ou destruction des sous-unités de GthII. Pendant les stades plus avancés de la spermatogenèse les 2 typev gonadotropes pouvaient encore être distingués, mais les cellules putatives à GtH apparaissant tôt étaient rares. Les présentes observations permettent de supposer que les cellules à GtH apparaissant tôt peuvent être des cellules à GtHI. Cependant, la preuve définitive de leur identité dépend de la disponibilité d'anti-GtHI ou de cDNAs spécifiques de la GtHI.
    Notes: Abstract In an ultrastructural immunocytochemical study we investigated the development of the gonadotropic cells in the pituitary of two to six months old male African catfish in relation to testicular development. In this period, pituitary and testicular tissue samples were collected on five occasions (groups I–V). Blood samples could only be taken from the fish in groups III–V. The testicular development was divided in three stages i.e., immature (only spermatogonia, group I), early (spermatogonia and spermatocytes, groups II and III) and advanced (all germ cell stages including spermatozoa, groups IV and V) spermatogenesis. 11-Ketotestosterone blood levels were low, except for the last group. Antisera were raised against the complete catfish α,βGTH-II, as well as to the separate α- and β-subunits of catfish GTH-II. In the proximal pars distalis of immature fish, undifferentiated cells, somatotrops, putative thyrotrops (pTSH) and putative gonadotrops (pGTH) were found. In the two latter, secretory granules were labeled with anti-αGTH, but not with anti-βGTH-II. pTSH- and pGTH-cells were distinguished on the basis of the size of their secretory granules. During early spermatogenesis, two classes of putative gonadotrops could be distinguished. One type had the same immunocytochemical and ultrastructural characteristics as in immature fish; the secretory granules in the second cell type, which was more abundant, were also immunopositive for anti-βGTH-II. The mean volume of the secretory granules in these GTH-II cells was three times larger than that in the early appearing pGTH-cells. In addition, the later appearing GTH-II cells contained large inclusions, known as globules. These structures labeled with anti-αβGTH-II and with anti-βGTH-II, but not with anti-αGTH. It is assumed that the globules are involved in a differential storage and/or breakdown of the GTH-II subunits. During advanced spermatogenesis the two gonadotropic cell types could still be distinguished, but the early appearing pGTH-cell type was scarce. The present observations permit the conclusion that the early appearing cells may be GTH-I cells. However, definitive proof about their identity depends on the availability of antibodies or cDNA probes specific for GTH-I.
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