Use of a pituitary cell dispersion method and primary culture system for the studies of gonadotropin-releasing hormone action in the goldfish, Carassius auratus I. Initial morphological, static, and cell column perifusion studies

Chang, John P.; Cook, Harry; Freedman, G L; Wiggs, A.J.; Somoza, Gustavo M.; Leeuw, R. de; Peter, Richard

doi:10.1016/0016-6480(90)90310-i

Cited by 166 publications

(129 citation statements)

References 33 publications

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“…Fish were killed by spinosectomy after anesthesia in 0.05% tricane methanesulphonate (Syndel, Vancouver, BC, Canada). Pituitaries were excised, diced into 0.6-mm fragments, and dispersed by using the trypsin/DNA II digestion method (38) with minor modifications (39). Pituitary cells were cultured in 24-well cluster plates at a density of 0.25 ϫ 10 6 cells per well.…”

Section: Methodsmentioning

confidence: 99%

Discovery of growth hormone-releasing hormones and receptors in nonmammalian vertebrates

Lee¹,

Siu²,

Tam³

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

111

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In mammals, growth hormone-releasing hormone (GHRH) is the most important neuroendocrine factor that stimulates the release of growth hormone (GH) from the anterior pituitary. In nonmammalian vertebrates, however, the previously named GHRH-like peptides were unable to demonstrate robust GH-releasing activities. In this article, we provide evidence that these GHRH-like peptides are homologues of mammalian PACAP-related peptides (PRP). Instead, GHRH peptides encoded in cDNAs isolated from goldfish, zebrafish, and African clawed frog were identified. Moreover, receptors specific for these GHRHs were characterized from goldfish and zebrafish. These GHRHs and GHRH receptors (GHRHRs) are phylogenetically and structurally more similar to their mammalian counterparts than the previously named GHRH-like peptides and GHRH-like receptors. Information regarding their chromosomal locations and organization of neighboring genes confirmed that they share the same origins as the mammalian genes. Functionally, the goldfish GHRH dose-dependently activates cAMP production in receptor-transfected CHO cells as well as GH release from goldfish pituitary cells. Tissue distribution studies showed that the goldfish GHRH is expressed almost exclusively in the brain, whereas the goldfish GHRH-R is actively expressed in brain and pituitary. Taken together, these results provide evidence for a previously uncharacterized GHRH-GHRH-R axis in nonmammalian vertebrates. Based on these data, a comprehensive evolutionary scheme for GHRH, PRP-PACAP, and PHI-VIP genes in relation to three rounds of genome duplication early on in vertebrate evolution is proposed. These GHRHs, also found in flounder, Fugu, medaka, stickleback, Tetraodon, and rainbow trout, provide research directions regarding the neuroendocrine control of growth in vertebrates. molecular evolution ͉ genome duplication ͉ pituitary adenylate cyclase-activating polypeptide

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Section: Methodsmentioning

confidence: 99%

Discovery of growth hormone-releasing hormones and receptors in nonmammalian vertebrates

Lee¹,

Siu²,

Tam³

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

111

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“…Animal use protocols were approved by the University of Alberta Animal Care Committee in accordance with national guidelines. Pituitaries from goldfish (Aquatic Imports, Calgary, AB, Canada) were dispersed using a trypsin-DNase protocol that has been described previously (8). Dispersed cells were cultured overnight on poly-L-lysine-coated coverslips (0.25 million cells/ coverslip, 1-ml medium) for imaging experiments (24), preswollen Cytodex beads (1.5 million/dish, 6-ml medium) for cell column perifusion studies (8), or Primaria surface-modified tissue culture plates (Becton-Dickinson, Franklin Lakes, NJ) for static incubation experiments designed to measure long-term hormone secretion/contents (24-well plates; 0.25 million cells/well; 1-ml medium) or GH mRNA levels (6-well plates; 2 million cells/well; 4-ml medium) in Medium 199 (M-199) with Earle's salts (GIBCO, Grand Island, NY) supplemented with 2.2 g/l NaHCO 3, 25 mM HEPES, 100,000 U/l penicillin, 100 mg/l streptomycin, and 1% horse serum (pH adjusted to 7.2 with NaOH) at 28°C, 5% CO 2, and in saturated humidity.…”

Section: Methodsmentioning

confidence: 99%

“…Prolonged cellular GH content and GH release were assayed in testing medium (GIBCO M-199 with Hanks' salts, supplemented with 2.2 g/l NaHCO 3, 25 mM HEPES, 100,000 U/l penicillin, 100 mg/l streptomycin, 20 mg/l phenol red, and 0.1% BSA, pH adjusted to 7.2 with NaOH) under static incubation conditions [same as culture conditions described above (8)]. After the incubation period, the medium was aspirated, and cells were lysed and snap frozen.…”

Section: Methodsmentioning

confidence: 99%

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Function-specific calcium stores selectively regulate growth hormone secretion, storage, and mRNA level

Johnson

Klausen

Habibi

et al. 2002

American Journal of Physiology-Endocrinology and Metabolism

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Ca+ stores may regulate multiple components of the secretory pathway. We examined the roles of biochemically independent intracellular Ca2+stores on acute and long-term growth hormone (GH) release, storage, and mRNA levels in goldfish somatotropes. Thapsigargin-evoked intracellular Ca2+ concentration ([Ca2+]i) signal amplitude was similar to the Ca2+-mobilizing agonist gonadotropin-releasing hormone, but thapsigargin (2 μM) did not acutely increase GH release, suggesting uncoupling between [Ca2+]i and exocytosis. However, 2 μM thapsigargin affected long-term secretory function. Thapsigargin-treated cells displayed a steady secretion of GH (2, 12, and 24 h), which decreased GH content (12 and 24 h), but not GH mRNA/production (24 h). In contrast to the results with thapsigargin, activating the ryanodine (Ry) receptor (RyR) with 1 nM Ry transiently increased GH release (2 h). Prolonged activation of RyR (24 h) reduced GH release, contents and apparent production, without changing GH mRNA levels. Inhibiting RyR with 10 μM Ry increased GH mRNA levels, production, and storage (2 h). Increasing [Ca2+]i independently of Ca2+stores with the use of 30 mM KCl decreased GH mRNA. Collectively, these results suggest that parts of the secretory pathway can be controlled independently by function-specific Ca2+ stores.

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“…Furthermore, recent studies in goldfish have shown that GnRH, until now considered to be strictly a gonadotropin releasing hormone, may also function as a GH releasing factor. Treatments with GnRH peptides are capable of increasing GH plasma levels and, in vitro, to stimulate GH release from pituitary fragments or dispersed pituitary cells Chang et al 1990;Flett et al 1992). Weil and collaborators found that, in trout, GnRH is also able to stimulate GH secretion by cultured pituitary cells obtained from males and females at different sexual stages (Blaise 1991).…”

Section: Gonadotropin Releasing Hormone (Gnrh)mentioning

confidence: 99%

Growth hormone (GH) and reproduction: a review

Gac

Blaise

Fostier

et al. 1993

Fish Physiol Biochem

171

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ResumeLes interactions entre les fonctions de croissance et de reproduction mises en evidence chez de nombreux vert6br6s, sont particulierement aigiies A certaines tapes du cycle vital des poissons. Nous d6crivons les interactions endocrines existant entre les axes somatrope et gonadotrope en insistant sur le role jou6 par l'hormone de croissance (GH). L'analyse compare de ces phenomenes chez les mammiferes, les poissons et les amphibiens permet de sugg6rer que la GH joue un rle sp6cifique dans la physiologie de la puberty, la gametog6nese et la fertility. Nous mettons en evidence l'apport original des tudes effectu6es sur le module poisson dans ce champ d'investigation. AbstractInteraction between growth and reproduction occurs in many vertebrates and is particularly obvious at certain stages of the life cycle in fish. Endocrine interactions between the gonadotropic axis and the somatotropic axis are described, the potential role of GH being emphasised. A comparative analysis of these phenomena in mammals, amphibians and fish, suggests a specific role of GH in the physiology of puberty, gametogenesis and fertility. It also shows the original contribution made by studies on the fish model in this field of investigations.

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Use of a pituitary cell dispersion method and primary culture system for the studies of gonadotropin-releasing hormone action in the goldfish, Carassius auratus I. Initial morphological, static, and cell column perifusion studies

Cited by 166 publications

References 33 publications

Discovery of growth hormone-releasing hormones and receptors in nonmammalian vertebrates

Discovery of growth hormone-releasing hormones and receptors in nonmammalian vertebrates

Function-specific calcium stores selectively regulate growth hormone secretion, storage, and mRNA level

Growth hormone (GH) and reproduction: a review

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