Summary
Organotypic cultures, in defined medium, of pituitary primordia obtained from 15-day-old rat fetuses were performed in order to study the in vitro differentiation of melanotrophic cells. The morphological and ultrastructural features of the transplants resembled those of the gland developing in vivo. In situ hybridization on semi-thin sections, using a 35S-labelled oligonucleotide probe, revealed pro-opiomelanocortin-mRNA-containing cells on the first day of culture in the anterior lobe and after 2–3 days in the intermediate lobe. Immunoperoxidase labelling of adjacent sections showed that the same cells reacted with antibodies against α-melanocyte-stimulating hormone (αMSH), γ3 and adrenocorticotropic hormone in both lobes. The pro-opiomelanocortin-mRNA-containing cells formed progressively conspicuous areas in the intermediate lobe, which was almost uniformly labelled after 6 days. In the anterior lobe, these cells remained scattered in small cell groups, and colloidal gold immunolabelling showed the progressive disappearance of αMSH labelling from the secretory vesicles in cells exhibiting morphological features of adult corticotrophic cells. Both the αMSH content of the explants and αMSH release into the culture medium increased with time. Treatment with the dopamine agonist bromocriptine induced a strong dose-dependent decrease in αMSH secretion, which was significant after 3 days in culture, indicating that dopamine D2 receptors are able to regulate hormonal release of melanotrophic cells at early stages. This system constitutes a suitable model for further studies of factors controlling cell differentiation and cellular interactions involved in histogenesis.
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References
Allaerts W, Carmeliet P, Denef C (1990) New perspectives in the function of pituitary folliculo-stellate cells. Mol Cell Endocrinol 71:73–81
Autelitano DJ, Lundblad JR, Blum M, Roberts JL (1989) Hormonal regulation of POMC gene expression. Annu Rev Physiol 51:715–726
Begeot M, Dupouy JP, Dubois MP, Dubois PM (1981) Immunocytological determination of gonadotropic and thyrotropic cells in fetal rat anterior pituitary during normal development and under experimental conditions. Neuroendocrinology 32:285–294
Begeot M, Dubois MP, Dubois PM (1982) Comparative study in vivo and in vitro of the differentiation of immunoreactive corticotropic cells in fetal rat anterior pituitary. Neuroendocrinology 35:255–264
Begeot M, Hemming FJ, Aubert ML, Dubois PM (1987) Différenciation des cellules antéhypophysaires en culture. Ann Endocrinol (Paris) 48:367–377
Bodner M, Castrillo JL, Theill LE, Deerinck T, Ellisman M, Karin M (1988) The pituitary-specific transcription factor GHF-1 is a homeobox-containing protein. Cell 55:505–518
Bottenstein JE, Sato GH (1979) Growth of a rat neuroblastoma cell-line in serum-free supplemented medium. Proc Natl Acad Sci USA 79:514–517
Buma P, Roubos EW, Buijs RM (1984) Ultrastructural demonstration of excytosis of neural, neuroendocrine and endocrine secretions with an in vitro tannic acid (TARI-) method. Histochemistry 80:247–256
Chatelain A, Dupouy JP (1981) Adrenocorticotrophic hormone in the anterior and neurointermediate lobes of the fetal rat pituitary gland. J Endocrinol 89:181–186
Chatelain A, Dupois MP, Dupouy JP (1976) Hypothalamus and cytodifferentiation of the foetal pituitary gland. Study in vivo. Cell Tissue Res 169:335–344
Chatelain A, Dupouy JP, Dubois MP (1979) Ontogenesis of cells producing polypeptide hormones (ACTH, MSH, LPH, GH, prolactin) in the fetal hypophysis of the rat: influence of the hypothalamus. Cell Tissue Res 196:409–427
Cote TE, Felder R, Kebabian JW, Sekura RD, Reisine T, Affolter HU (1986) D-2 dopamine receptor-mediated inhibition of proopiomelanocortin synthesis in rat intermediate lobe. J Biol Chem 261:4555–4561
Couly GF, Le Douarin NM (1985) Mapping of the early neural primordium in quail-chick chimeras. I. Developmental relationships between placodes, facial ectoderm, and prosencephalon. Dev Biol 110:422–439
Daikoku S, Chikamori M, Adachi T, Maki Y (1982) Effect of the basal diencephalon on the development of Rathke's pouch in rats: a study in combined organ cultures. Dev Biol 90:198–202
Daikoku S, Chikamori M, Adachi T, Okamura Y, Nishiyama T, Tsuruo Y (1983) Ontogenesis of hypothalamic immunoreactive ACTH cells in vivo and in vitro: role of Rathke's pouch. Dev Biol 97:81–88
Davis MD, Lichtensteiger W, Schlumpf M, Bruinink A (1984) Early postnatal development of pituitary intermediate lobe control in the rat by dopamine neurons. Neuroendocrinology 39:1–12
Dupouy JP (1980) Differentiation of MSH-, ACTH-, endorphin-, and LPH-containing cells in the hypophysis during embryonic and fetal development. Int Rev Cytol 68:197–249
Dupouy JP, Dubois MP (1975) Ontogenesis of the α-MSH, β-MSH and ACTH cells in the foetal hypophysis of the rat. Correlation with the growth of the adrenals and adrenocortical activity. Cell Tissue Res 161:373–384
Eberle AN (1988) The melanotropins. Chemistry, physiology and mechanisms of action. Karger, Basel
Ferrand R, Nanot J (1968) Differenciation de la poche de Rathke isolée de l'ébauche nerveuse de l'hypophyse et associée à un mésenchyme hétérologue chez la Souris. C R Soc Biol 162:983–986
Ferrara N, Schweigerer L, Neufeld G, Mitchell R, Gospodarowicz D (1987) Pituitary follicular cells produce basic fibroblast growth factor. Proc Natl Acad Sci USA 84:5773–5777
Follénius E, Van Dorsselaer A, Meunier A (1985) Separation and partial characterization by high-performance liquid chromatography and radioimmunoassay of different forms of melanocyte-stimulating hormone from fish (Cyprinidae) neurointermediate lobes. Gen Comp Endocrinol 57:198–207
Gash D, Ahmad N, Schechter J (1982) Comparison of gonadotroph, thyrotroph and mammotroph development in situ, in transplants and in organ culture. Neuroendocrinology 34:222–228
Gospodarowicz D, Abraham JA, Schilling J (1989) Isolation and characterization of a vascular endothelial cell mitogen produced by pituitary-derived folliculo-stellate cells. Proc Natl Acad Sci USA 86:7311–7315
Guitteny AF, Böttlen P, Bloch B (1988) Analysis of vasopressin gene expression by in situ hybridization and immunohistochemistry in semi-thin sections. J Histochem Cytochem 36:1373–1378
Hindelang C, Félix JM, Laurent FM, Klein MJ, Stoeckel ME (1990) Ontogenesis of proopiomelanocortin gene expression and regulation in the rat pituitary intermediate lobe. Mol Cell Endocrinol 70:225–235
Ingraham HA, Chen R, Mangalam HJ, Elsholtz HP, Flynn SE, Lin CR, Simmons DM, Swanson L, Rosenfeld MG (1988) A tissue-specific transcription factor containing a homeodomain specifies a pituitary phenotype. Cell 55:519–529
Keck PJ, Hauser SD, Krivi G, Sanzo K, Warren T, Feder J, Connolly DT (1989) Vascular permeability factor, an endothelial cell mitogen related to PDGF. Science 246:1309–1312
Khachaturian H, Alessi NE, Munfakh N, Watson SJ (1983) Ontogeny of opioid and related peptides in the rat CNS and pituitary: an immunocytochemical study. Life Sciences 33:61–64
Kurosumi K, Tosaka H, Ijima K (1989) The immature type of proopiomelanocortin cell of the rat anterior pituitary as observed by immunogold electron microscopy. Arch Histol Cytol 52:135–150
Leung DW, Cachianes G, Kuang WJ, Goeddel DV, Ferrara N (1989) Vascular endothelial growth factor is a secreted angiogenic mitogen. Science 246:1306–1309
Lowry OH, Rosenbrough NJ, Randall R (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275
Lutz-Bucher B, Schimchowitsch S, Félix JM, Stoeckel ME, Koch B (1989) Stimulation by atrial natriuretic factor of cyclic GMP production in cultured anterior and intermediate pituitary tissues: evidence of a major contribution of proliferating nonendocrine cells. Mol Cell Endocrinol 64:257–266
Nemeskéri A, Halász B (1989) Cultured fetal rat pituitaries kept in synthetic medium are able to initiate synthesis of trophic hormones. Cell Tissue Res 255:645–650
Nemeskéri A, Nemeth A, Sétáló G, Vigh S, Halász B (1976) Cell differentiation of the fetal rat anterior pituitary in vitro. Cell Tissue Res 170:263–273
Nemeskéri A, Sétáló G, Halász B (1988) Ontogenesis of the three parts of the fetal rat adenohypophysis. A detailed immunohistochemical analysis. Neuroendocrinology 48:534–543
Pintar JE, Lugo DI (1987) Proopiomelanocortin gene expression, prohormone processing, and secretion during rat pituitary development. Ann NY Acad Sci 512:318–327
Sales N, Martres MP, Bouthenet ML, Schwartz JC (1989) Ontogeny of dopaminergic D2 receptors in the rat nervous system: characterization and detailed autoradiographic mapping with [125I] iodosulpiride. Neuroscience 28:673–700
Schechter J, Gash D, Ahmad N (1985) Mesenchymal influences on the development of the adenohypophysis in the rat. Cell Tissue Res 241:67–76
Schimchowitsch S, Stoeckel ME, Klein MJ, Garaud JC, Schmitt G, Porte A (1983) Oxytocin-immunoreactive nerve fibers in the pars intermedia of the pituitary in the rabbit and hare. Cell Tissue Res 228:255–263
Schmitt G, Briaud B, Mialhe C, Stutinsky F (1979) Different effects of K+ and Ca++ on αMSH and ACTH release from superfused neurointermediate lobe of the rat hypophysis. Neuroendocrinology 28:289–348
Schwartzberg DG, Nakane PK (1982) Ontogenesis of adrenocorticotropin-related peptide determinants in the hypothalamus and pituitary gland of the rat. Endocrinology 110:855–864
Simmons DM, Voss JW, Ingraham HA, Holloway JM, Broide RS, Rosenfeld MG, Swanson LW (1990) Pituitary cell phenotypes involve cell-specific Pit-1 mRNA translation and synergistic interactions with other classes of transcription factors. Genes Dev 4:695–711
Smith AI, Funder JW, (1988) Proopiomelanocortin processing in the pituitary, central nervous system, and peripheral tissues. Endocrinol Rev 9:159–179
Stoeckel ME, Schimchowitsch S, Garaud JC, Schmitt G, Vaudry H, Klein MJ, Porte A (1985) Immunocytochemical evidence for intragranular processing of pro-opiomelanocortin in the melanotropic cells of the rabbit. Cell Tissue Res 242:365–370
Valentino KL, Crumrine DA, Reichardt LF (1985) Lowicryl K4M embedding of brain tissue for immunogold electron microscopy. J Histochem Cytochem 33:969–973
Vuillez P, Carbajo-Pérez S, Stoeckel ME (1987) Colocalization of GABA and tyrosine hydroxylase immunoreactivities in the axons innervating the neurointermediate lobe of the rat pituitary: an ultrastructural immunogold study. Neurosci Lett 79:53–58
Watanabe YG (1982) Effects of brain and mesenchyme upon the cytogenesis of rat adenohypophysis in vitro. I. Differentiation of adrenocorticotropes. Cell Tissue Res 227:257–266
Watanabe YG, Daikoku S (1979) An immunohistochemical study on the cytogenesis of adenohypophysial cells in fetal rats. Dev Biol 68:557–567
Watanabe YG, Matsumura H, Daikoku S (1973) Electron microscopic study of rat pituitary primordium in organ culture. Z Zellforsch 146:453–461
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Vuillez, P., René, F., Plante, M. et al. Differentiation of the melanotrophic cells of rat pituitary primordium in organotypic culture in defined medium. Cell Tissue Res 267, 169–183 (1992). https://doi.org/10.1007/BF00318702
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DOI: https://doi.org/10.1007/BF00318702