Ultrastructural characterization of the pars distalis of the Indian female Sheath-tailed bat, Taphozous longimanus (Hardwicke) / Caracterización ultraestructural de la pars distalis del murciélago cola de vaina hembra indio, Taphozous longimanus (Hardwicke)

The present ultrastructural observations demonstrate the presence of six cell types in the pars distalis of non-pregnant and pregnant bats of Taphozous longimanus. In the pars distalis of T. longimanus, STH cells are round to oval with eccentrically placed nucleus, numerous secretory granules and well developed Golgi indicate a cell under vigorous synthetic activity while those filled with secretory granules with reduced Golgi complex suggest reserve or storage state of cells. LTH cell is characterized by the large secretory granules, dilated endoplasmic reticulum and numerous mitochondria in the cytoplasm which indicate that these cells are hypertrophied and synthetically very active during pregnancy. ACTH cells are found either singly or in groups and are elongated or angular with long cytoplasmic processes. The size and peripheral arrangement of secretory granules are characteristic of ACTH cell. TSH cells are distributed mostly towards the periphery of the pars distalis of T. longimanus. They are elongated, polygonal or triangular in shape. The secretory granules are small, electron dense, 150-200 nm in diameter. The rough endoplasmic reticulum is very well developed. In FSH, the secretory granules are small (200 to 400 nm) and less in number and are distributed towards the periphery of the cell. FSH cells show well developed mitochondria, Golgi and rough endoplasmic reticulum indicating active state of FSH during estrus and pregnancy. The hypertrophy of FSH and LH cells during pregnancy is associated with filigreed cytoplasmic pattern giving a bizarre appearance. At late pregnancy, FSH and LH cells are highly active and synthesize large quantities of hormone as indicated by the development of cell organelles.

Las observaciones ultraestructurales actuales demuestran la presencia de seis tipos de células en la pars distalis de murciélagos Taphozous longimanus preñadas y no preñadas. En la pars distalis del T. longimanus, las células STH son redondas u ovaladas con un núcleo excéntrico, numerosos gránulos de secreción y un Golgi bien desarrollado que indican una célula en actividad de síntesis vigorosa, mientras que las llenas de gránulos de secreción con un complejo de Golgi reducido sugieren un estado celular de reserva o almacenamiento. Las células LTH se caracterizan por grandes gránulos de secreción, el retículo endoplásmico dilatado y numerosas mitocondrias en el citoplasma, indicando que estas células están hipertrofiadas y con una actividad sintética muy activa durante el embarazo. Células de ACTH se encuentran de forma individual o en grupos, son alargadas o angulares, con largos procesos citoplásmicos. El tamaño y la disposición periférica de los gránulos de secreción de ACTH son característicos de la célula. Células de TSH se distribuyen principalmente hacia la periferia de la pars distalis del T. longimanus. Ellos son alargadas, poligonales o de forma triangular. Los gránulos de secreción son pequeños, electrodensos, de 150-200 nm de diámetro. El retículo endoplasmático rugoso está muy bien desarrollado. En células FSH, los gránulos de secreción son pequeños (200 a 400 nm), menores en número y se distribuyen hacia la periferia de la célula. Células FSH muestran mitocondrias bien desarrolladas, Golgi y retículo endoplasmático rugoso que indica el estado activo de la FSH durante el estro y la preñez. La hipertrofia de las células de FSH y LH durante la preñez se asocia con un patrón citoplasmático filigrana dando una extraña apariencia. Al final de la preñez, las células de FSH y LH son muy activas y sintetizan grandes cantidades de hormonas, como producto del desarrollo de las organelos celulares.

Genistein affects the morphology of pituitary ACTH cells and decreases circulating levels of ACTH and corticosterone in middle-aged male rats

The soybean phytoestrogen, genistein, is increasingly consumed as an alternative therapeutic for age-related diseases, namely cardiovascular conditions, cancer and osteoporosis. However, despite the beneficial effects on health, concern has been raised that this isoflavone also acts as an endocrine-disrupting chemical. The aim of this study was to examine the effects of genistein on immunohistomorphometric features of pituitary adrenocorticotropic cells (ACTH) and blood concentrations of ACTH and corticosterone in orchidectomized middle-aged male rats. Sixteen-month-old Wistar rats were divided into sham-operated (SO), orchidectomized (Orx) and genistein-treated orchidectomized (Orx+G) groups. Genistein (30mg/kg/day) was administered subcutaneously for three weeks, while the control groups received the vehicle alone. ACTH cells were identified by the peroxidase-antiperoxidase (PAP) immunohistochemical procedure. Circulating concentrations of ACTH and corticosterone were measured by immunoassay. Orchidectomy reduced (p<0.05) the cell volume and the relative volume of ACTH cells in comparison to SO rats. Genistein treatment further decreased (p<0.05) these morphometric parameters and reduced (p<0.05) circulating ACTH and corticosterone concentrations by more than 20 percent in comparison to both Orx and SO rats. In conclusión, genistein modulated the immunohistomorphometric features of ACTH cells and decreased blood ACTH and corticosterone levels, which supports evidence that this isoflavone affects the hypothalamic-pituitary-adrenal axis and suppresses glucocorticoid hormone secretion.

Intracellular trafficking and metabolic turnover of yeast prepro-alpha-factor-SRIF precursors in GH3 cells

Chimeric genes coding for prepro region of yeast alpha-factor and anglerfish SRIF were expressed in rat GH3 cells to determine whether yeast signals could regulate hormone processing in mammalian cells. We report that nascent hybrid polypeptides were efficiently targeted to ER, where cleavage of signal peptides and core glycosylation occurred, and were localized mainly in Golgi. These data indicate that prepro region of yeast alpha-factor functions in sorting molecules to secretory pathway in mammalian cells. A hybrid construct with a mutated signal peptide underwent similar ER translocation, whereas such a mutation resulted in defective translocation in yeast (Cheong et al., 1997). This difference may be due to the differences in ER translocation between yeast and mammalian cells, i.e., posttranslational versus cotranslational translocation. Processing and secretion of metabolically labeled hybrid propeptides to mature SRIF peptides were assessed by HPLC. When pulse-labeled cells were chased for up to 2 h, intracellular propeptides disappeared with a half-life of approximately 25 min, showing that -68% of initially synthesized propeptides were secreted constitutively. About 22% of SRIF-related products were proteolytically processed to mature SRIF, of which 38.7% were stored intracellularly with a half-life of - 2 h. In addition, immunocytochemical localization showed that a small proportion of SRIF molecules accumulated in secretory vesicles. All these results suggest that yeast prepropeptide could direct hybrid precursors to translocate into ER lumen and transit through secretory pathway to the distal elements of Golgi compartment, but could process and target it less efficiently to downstream in rat endocrine cells.

Intracellular trafficking and metabolic turnover of yeast prepro-alpha-factor-SRIF precursors in GH3 cells

Chimeric genes coding for prepro region of yeast alpha-factor and anglerfish SRIF were expressed in rat GH3 cells to determine whether yeast signals could regulate hormone processing in mammalian cells. We report that nascent hybrid polypeptides were efficiently targeted to ER, where cleavage of signal peptides and core glycosylation occurred, and were localized mainly in Golgi. These data indicate that prepro region of yeast alpha-factor functions in sorting molecules to secretory pathway in mammalian cells. A hybrid construct with a mutated signal peptide underwent similar ER translocation, whereas such a mutation resulted in defective translocation in yeast (Cheong et al., 1997). This difference may be due to the differences in ER translocation between yeast and mammalian cells, i.e., posttranslational versus cotranslational translocation. Processing and secretion of metabolically labeled hybrid propeptides to mature SRIF peptides were assessed by HPLC. When pulse-labeled cells were chased for up to 2 h, intracellular propeptides disappeared with a half-life of approximately 25 min, showing that -68% of initially synthesized propeptides were secreted constitutively. About 22% of SRIF-related products were proteolytically processed to mature SRIF, of which 38.7% were stored intracellularly with a half-life of - 2 h. In addition, immunocytochemical localization showed that a small proportion of SRIF molecules accumulated in secretory vesicles. All these results suggest that yeast prepropeptide could direct hybrid precursors to translocate into ER lumen and transit through secretory pathway to the distal elements of Golgi compartment, but could process and target it less efficiently to downstream in rat endocrine cells.

Immunocytochemical localization of different cell types in the adenohypophysis of the cichlid fish Cichlasoma dimerus (Heckel, 1840)

The adenohypophysis of the cichlid fish Cichlasoma dimerus was studied using the avidin-biotin-peroxidase method with antisera raised against piscine pituitary hormones and heterologous antisera against mammalian pituitary hormones. Antiserum raised against rabbit ACTH recognized a group of cells bordering the neurohypophysis (NH) in the rostral pars distalis (RPD). Anti-chum salmon prolactin (PRL) identified a compact group of cells in the periphery of the RPD. Gonadotropin II (GTH II), thyrotropin (TSH) and growth hormone (GH)-ir cells were localized in the proximal pars distalis. Ir-GTH II cells were also observed in the dorsal area of the pars intermedia (PI). Ir-GTH I cells could not be identified using anti-chum salmon GTH I, this may be due either to a failure of the antisera to recognize the gonadotropin or to a low expression of the hormone in adults of this species. PAS positive cells from the PI bound specifically with three different antisera raised against somatolactin (SL) of four different fish species. These cells surrounded deep branches of the NH in the PI.

Immunohistochemical and ultrastructural age-related changes in rat lactotroph cells

Ageing produces alterations in some functions of the hypothalamo-pituitary axis leading to sexually dimorphic changes in the prolactin (PRL)-secreting cells. Since quantitative morphological data of these age-associated alterations are scarce, we carried out a morphometric immunohistochemical assessment as well as an ultrastructural study of the PRL cell population in male and female rats of different ages. Young (3-month-old), old (20-month-old), and senescent (31-month-old) Sprague-Dawley rats of both sexes were sacrificed by rapid decapitation, their pituitaries immediately dissected out and processed for both immunohistochemistry and electron microscopy. Analysis of different morphometric parameters revealed that the cell density (CD) and volume density (VD) significantly decreased with age in male rats. In females, while CD showed a significant age-related diminution when young rats were compared to old ones, this parameter increased in senescent animals. The VD presented higher values in senescent rats. When the data were compared between sexes, VD was found to be higher in females if old and senescent rats were considered. Finally, CD increased significantly in females when compared to males. The ultrastructure of the PRL cells from old and senescent animals of both sexes exhibited changes suggestive of an hyperstimulation state, with some prolactotrophs having the appearance of cells undergoing an involutive process. We conclude that ageing has a differential impact on the PRL cells of male and female rats with respect to the immunohistochemical and ultrastructural features of that cell population

Hormone uptake and morphological changes in cultured pituicytes exposed to oxytocin and vasopressin

Enzymatically dispersed cells, isolated from adult female rat neural lobes, were cultured for 7 days. Routine cultures showed pituicytes with compact, sometimes ovoid, cell bodies. The cytoplasmic processes of these cells exhibited several varicosities and made contact with neighboring cells forming networks. The cultured pituicytes were immunocytochemically characterized using antisera to glial fibrillary acidic protein and to S-100. Most pituicytes, when exposed during culture to oxytocin (OXY) and vasopressin (VP; 1 microM each), were devoid of their characteristic processes. Immunocytochemical staining for OXY or VP revealed that the pituicytes were capable of incorporating these hormones during culture. In cultures without added hormones, no significant staining reaction for OXY or VP could be detected. The lack of projections in pituicytes exposed to the hormones during culture is in agreement with the morphological changes observed by other authors in situ after acute hormone release. The uptake of OXY and VP may be indicative for a regulatory mechanism, by which the pituicytes control the amount of hormones present in the intercellular space

Efecto de secreciones de pars tuberalis de la adenohipofisis sobre la liberacion de prolactina desde pars distalis / Effect of adenohypophyseal pars tuberalis secretions on pars distalis prolactin liberation

El presente trabajo aporta evidencias que indican que factores producidos por la pars tuberalls (PT) afectan la liberación de prolactina desde la pars distalis (PD). Para determinar el efecto de las secreciones al medio de cultivo de células de la PT bovina sobre células dispersas de PD de rata, se trabajó con estas últimas tanto en incubaciones de 30 minutos, como en superfusiones continuas en columnas. Cuando se utilizó medio de cultivo de la totalidad de las células de la PT, se requirieron 9 mug de proteína total para la máxima estimulación de la liberación de prolactina. En cambio, cuando se empleó el correspondiente a las fracciones del 50-60 por ciento de un gradiente de Percoll, sólo se requirieron 4 mug de proteína total. Con una purificación parical en Sephadex G 50 de ese medio, se logró el mismo efecto com 80 ng de proteína total. El principio activo tendría un tamaño molecular superior a los 40 kDal. Los resultados obtenidos permiten proponer, al menos para las células prolactotropas, a la PD como el órgano efector de algunas secreciones de la PT.

Morphological and cytochemical characterization of cell types of the adenohypophysis of Manjuba, Anchoviella lepidentostole (Fowler, 1911) (Osteichthyes, Engraulidae)

A hipófise de Anchoviella lepidentostole apresenta-se dividida em neuro-hipófise e adeno-hipófise, sendo que a caracterizaçäo morfológica e citoquímica dos tipos celulares desta regiäo foi a proposta deste trabalho. A adeno-hipófise divide-se em pars intermedia (PI) e pars distalis (PD), sendo que esta última se divide em pars distalis rostralis (PDR) e pars distalis proximalis (PDP). As células da PDR organizam-se em folículos. No epitélio folicular foram caracterizados quatro tipos celulares: I-PDR (basófilo), II-PDR (positivo à hematoxilina-chumbo/HPb+), III-PDR (PAS+, AB pH2,5+ e AF+), e IV-PDR (acidófilas). A PDP possui dois tipos celulares: I-PDP (PAS+, AB pH2,5+ e AF+) e II-PDP (acidófilas). Na PI também foram caracterizados dois tipos celulares: I-PI (HPb+) e II-PI (cromófobo aos métodos empregados)

The adenohypophysis of the swamp eel, Synbranchus marmoratus, an immunocytochemical analysis

The adenohypophyseal cell types of the protogynous fish Synbranchus marmoratus were studied by histochemical and immunocytochemical staining with antisera raised against piscine and human pituitary hormones to ascertain their distribution. The prolactin (PRL) cells were distributed in the rostral pars distalis and showed specific binding to antisera to carp and chum salmon prolactin. No reaction was observed with antiserum to human prolactin. The corticotrops showed strong immunoreactivity with anti-human ACTH, these cells bordered the neurohypophysis and islets between PRL cells in the rostral pars distalis. Growth hormone (GH) cells were densely distributed and associated with the neurohypophysis only in pars distalis proximal. They reacted with antisera to piscine GH but not with antisera to human growth hormone. The thyrotrops were scattered in the proximal pars distalis and showed strong immunoreactivity to the human thyrotropin Beta subunit antiserum. Gonadotrops were located in the central area of the proximal pars distalis and in the external border of the pars intermedia. These cells were alcian blue and PAS positive, and reacted with anti-croaker GTH and anti-coho GTH I and GTH II. The PAS positive cells from the pars intermedia bound specifically to anti-chum somatolactin.

Obtención, purificación y caracterización de dos formas de hormona luteinizante de la adenohipófisis caprina (gLH) / Obtention, purification and characterization of two forms of caprine luteinizing hormone from adenohypophsis (gLH)

Se describe la obtención, purificación y caracterización de dos proteínas hipofisiarias de origen caprino con carga eléctrica diferente y con características de hormona luteinizante (LH). La fracción no retenida en la cromatografía de intercambio catiónico (CM-celulosa) designada CM-lab y la correspondiente al segundo pico de dicha cromatografía (CM-2ab), se repurificaron en una cromatografía de intercambio aniónico (DEAE-celulosa) en condiciones idénticas, para obtener LH-I (CM-lab-DEAE-la) con un rendimiento de 76 mg/kg de adenohipófisis, y a la LH-II (CM2ab-DEAE-lb) con 15 mg/kg. La diferencia de carga de cada proteína se determinó por su movilidad relativa (Rf) mediante una electroforesis en geles de poliacrilamida (PAGE) en condiciones nativas. La LH-I mostró un Rf de 0.09 (banda mayoritaria) y 0.415, mientras la LH-II presentó 2 bandas de tipo catiónico con un Rf de 0.14, 0.19 (banda mayoritaria). La determinación del peso molecular relativo de LH-I y LH-II, se llevo a cabo por medio de una electroforesis en geles de poliacilamida-dodecilsulfato de sodio (SDS-PAGE). En condiciones no reductoras (NR), ambas proteínas presentaron un peso molecular relativo de 36.5 kilodaltones (KDa) correspondiente a la forma monomérica, semejante a los estándares NIDDK-oLH-26 y USDA-bLH5, mientras que en presencia de 2ß-mercaptoetanol (condiciones reducidas), las proteínas presentaron un peso molecular de 22.4 y 20.2 KDa. El análisis por inmunotransferencia (Western-blot) en condiciones NR utilizando el anti-oLH (CSU-204), permitió identificar bandas inmunorreactivas de peso molecular de 50,43, 36.5 (mayoritaria), y 22.4 KDa, tanto en los estándares como en las fracciones obtenidas en este estudio. La potencia biológica realizada en un bioensayo in vivo 3 + 3 balanceado fue de 1.03 UI/mg para la LH-I. y de 0.65 UI/mg para la LH-II con intervalos de confianza de 95 por ciento. Su actividad inmunológica se determinó con un radioinmunoensayo (RIA) homólogo de LH caprina, a la dosis esperada del 50 por ciento (ED 50). Los resultados fueron de 1.7 ng/ml y 3.5 ng/ml para LH-I y LH-II respectivamante. Se concluye que esta técnica permite la obtención de dos proteínas con carga eléctrica diferente, con actividad biológica e inmunológica de LH, y con peso molecular idéntico

Effect of cimetidine on adenohypophysis of male albino mice under certain experimental condition.

Effect of 75 mg/kg of body weight of cimetidine administered intraperitoneally daily for 14 days to two groups of experimental animals (one of the experimental group was having intact testis and another group was bilaterally orchidectomized) was observed on cell population & cell volume of gonadotrophs and lactotrophs in pituitary gland, as it has not been studied earlier. In the experimental group of intact testis, there was significant reduction in the cell population of FSH cells in the cephalomedian area (P < 0.001) and in the lateral lobe (P < 0.01); the volume of both FSH and LH cells was also significantly reduced. In group 4 and group 5 there was significant increase in the population of lactotrophs and also in the volume of LH cells, FSH cells & lactotrophs. The change in the gonadotrophs in group 2 was due to increased production of testosterone from hypertrophied Leydig cells of testis rather then it's direct effect on adenohypophysis; in group 4 and group 5 the changes were due to lack of testosterone as in those cases bilaterally orchidectomy already done.

Regulacion de la secrecion de prolactina a nivel celular / Regulation of prolactin secretion at cellular level

En la presente revisión se recopilan y discuten dos décadas de investigación científica relacionada con los mecanismos involucrados en la regulación de la actividad secretora de las células prolactínicas en mamíferos. Los temas desarrollados abarcan algunos aspectos metodológicos (dispersión, cultivo y aislamiento de células anterohipofisarias) y los mecanismos de acción de las catecolaminas, los segundos mensajeros (calcio, AMP0-cíclico y los metabolitos del fosfatidil-inosito) y una serie de neuromoduladores y péptidos nuroendocrinos. Además se incluyó un capítulo sobre la morfología y bioquímica del gránulo secretorio de prolactina. En la mayoría de los casos los datos - a menudo dispersos o aislados - no permiten conclusiones definitivas y queda claro que, a pesar de algunos adelantos significativos, estamos aún lejos de comprender la complejidad de los mecanismos entrelazados y a veces superpuestos que intervienen en la regulación de la secreción de prolactina

Estudio inmunocitoquimico de las hormonas de la adenohipofisis humana: hormona del crecimiento / Immunocytochemical study of the human adenohypophysis hormones: growth hormone

Con el objeto de verificar el concepto de que la hormona del crecimiento esta contenida en poblaciones de celulas acidofilas, se realizo la deteccion inmunocitoquimica de la hormona en cortes histologicos de adenohipofisis humana fijados en formalina. Se utilizo la reaccion inmunocitoquimica de peroxidasa antiperoxidasa (PAP). se encontro que las celulas detectadas inmunocitoquimicamente como productoras de la hormona del crecimiento (GH), corresponden a las celulas acidofilas naranjas en la reaccion de tincion naranja G. delineadas clara y especificamente por el metodo PAP, estas celulas fueron observadas formando filas a lo largo de sinusoides. Se presenta la ultraestructura de estas celulas y se compara con las observaciones hechas por otros autores. Se describe ademas la utilidad del metodo para detectar la hormona de crecimiento en adenomas de la pituitaria y sus metastasis. Finalmente se discute el papel de la hormona en el crecimiento y se analizan las perspectivas de investigacion y la utilizacion practica de ella en el tratamiento de algunas entidades clinicas y en el incremento del crecimiento