Summary
Calcium plays an important role in the release of insulin. When the GBHA [glyoxal bis (2-hydroxyanil)] reaction is employed, calcium can be clearly demonstrated by light microscope in pancreatic islets of mice. The specificity of this finding has now been proved by elemental X-ray analysis. Electron microscopically, certain cations can be visualized by a precipitation technique using potassium pyroantimonate as the precipitating agent. In the B cell of mice this technique reveals a characteristic precipitation pattern. Elemental X-ray analysis suggests that the precipitates contain high amounts of calcium. The pattern of the precipitates changes dependent on the functional state of the B cell. In normoglycemia the deposits are mainly associated with the granule membranes, the cell membranes and the cytoplasmic matrix. In hypoglycemia there is a shift of precipitates into the endoplasmic reticulum and the mitochondria, which are thought to be storage organelles for intracellular calcium. The deposits within the halos of the numerous secretory granules, are diminished. In hyperglycemia there is a marked ion translocation across the cell membrane to its inner surface and particularly into the halos of the secretory granules, while the deposit content of mitochondria and endoplasmic reticulum is decreased. Within the saccules of the secretory granules, the deposits sometimes seem to impregnate a filamentous network, which encloses the secretory granule and cannot be seen by conventional electron microscopical preparations. The morphological data suggest that emiocytosis of hormone granules is associated with a release of cellular calcium. The presented observations in treated and untreated animals extend and support the conceptions on the specific role of calcium within the insulin releasing mechanism of the B cell.
Zusammenfassung
Calcium spielt eine wichtige Rolle bei der Insulinsekretion. Lichtmikroskopisch läßt sich unter Anwendung der GBHA [Glyoxal bis (2-hydroxyanil)]-Methode Calcium eindeutig in den Pankreasinseln der Maus nachweisen. Die Spezifität dieses Befundes wurde durch die Röntgenelementaranalyse gesichert. Elektronenmikroskopisch können bestimmte Kationen durch eine Präzipitationstechnik mit Hilfe von Kaliumpyroantimonat sichtbar gemacht werden. In der B-Zelle der Maus ergibt diese Technik ein charakteristisches Verteilungsmuster von Präzipitaten. Die Röntgenelementaranalyse zeigt, daß die Präzipitate große Calciummengen enthalten. Das Verteilungsmuster der Niederschläge verändert sich in Abhängigkeit von dem funktionellen Zustand der B-Zelle. Bei Normoglykämie treten die Ausfällungen hauptsächlich in Verbindung mit den Membranen der Hormongranula, den Zellmembranen und der Matrix des Cytoplasmas auf. Bei Hypoglykämie zeigt sich eine Verschiebung der Präzipitate in das endoplasmatische Retikulum und in die Mitochondrien, die als Speicherorganellen für das intracelluläre Calcium angesehen werden. Die Ausfällungen innerhalb der Halos der zahlreichen Sekretgranula sind vermindert. Bei Hyperglykämie ergibt sich eine erhebliche Ionenverlagerung durch die Zellmembran zu deren innerer Oberfläche und besonders in die Halos der Sekretgranula, während die Präzipitatmengen in Mitochondrien und endoplasmatischem Retikulum vermindert sind. Innerhalb der Vesikel der Sekretgranula scheinen die Präzipitate manchmal ein filamentöses Netzwerk zu imprägnieren, welches das Sekretgranulum umhüllt und bei konventioneller elektronenmikroskopischer Präparation nicht sichtbar ist. Die morphologischen Befunde deuten darauf hin, daß die Emiocytose der Hormongranula mit einer Ausschleusung von cellulärem Calcium einhergeht. Die vorliegenden Beobachtungen an intakten Tieren erweitern und bestätigen die Konzeptionen über die spezifische Rolle des Calciums im Insulinsekretionsmechanismus der B-Zelle.
Similar content being viewed by others
References
Borle, A.B.: Calcium transport in kidney cells and its regulation. In: Cellular mechanisms for calcium transfer and homeostasis, ed. by G. Nichols, and R.H. Wasserman, p. 151–174. New York-London: Academic Press 1971
Curry, D.L., Bennett, L.L., Grodsky, G.M.: Requirement for calcium ion in insulin secretion by the perfused rat pancreas. Amer. J. Physiol. 214, 174–178 (1968)
Dean, P.M., Matthews, E.K.: Electrical activity in pancreatic islet cells: effect of ions. J. Physiol. (Lond.) 210, 265–275 (1970)
Fleckenstein, A.: Physiologie und Pharmakologie der transmembranären Natrium-, Kalium und Calcium-Bewegungen. Arzneimittel-Forsch. (Drug Res.) 22, 2019–2028 (1972)
Garfield, R.E., Henderson, R.M., Daniel, E.E.: Evaluation of the pyroantimonate technique for localization of tissue sodium. Tissue & Cell 4, 575–589 (1972)
Gomba, S., Szabo, J., Soltesz, B.M.: Electronhistochemical observations on the localization of sodium ions in glomerular basement membrane. Acta histochem. (Jena) 42, 367–372 (1972)
Grodsky, G.M., Bennett, L.L.: Cation requirements for insulin secretion in the isolated perfused pancreas. Diabetes 15, 910–913 (1966)
Hales, C.N., Milner, R.D.G.: The role of sodium and potassium in insulin secretion from rabbit pancreas. J. Physiol. (Lond.) 194, 725–743 (1968)
Hasselbach, W.: Die sarkoplasmatische Calciumpumpe. Arzneimittel-Forsch. (Drug Res.) 22, 2028–2036 (1972)
Heinze, E., Fussgänger, R., Teller, W.M.: Influence of calcium on insulin secretion in newborns. Pediat. Res. 7, 100–102 (1973)
Herman, L., Sato, T., Hales, C.N.: The electron microscopic localization of cations to pancreatic islets of Langerhans and their possible role in insulin secretion. J. Ultrastruct. Res. 42, 298–311 (1973)
Kashiwa, H.K.: Calcium in cells of fresh bone stained with glyoxal bis (2-hydroxyanil). Stain Technol. 41, 49–55 (1966)
Klöppel, G., Altenähr, E., Freytag, G.: Elektronenmikroskopische Untersuchungen zur experimentellen Insulitis nach Injektion von Anti-Insulin-Serum. Virchows Arch. Abt. A Path. Anat. 354, 324–335 (1971)
Klöppel, G., Schäfer, H.-J.: In preparation
Komnick, H., Komnick, U.: Elektronenmikroskopische Untersuchungen zur funktionellen Morphologie des Ionentransportes in der Salzdrüse von Larus argentatus. Z. Zellforsch. 60, 163–203 (1963)
Lacy, P.E., Howell, S.L., Young, D.A., Fink, C.J.: New hypothesis of insulin secretion. Nature (Lond.) 219, 1177–1179 (1968)
Legato, M.J., Langer, G.A.: The subcellular localization of calcium ion in mammalian myocardium. J. Cell Biol. 41, 401–423 (1969)
Malaisse, W.J.: Insulin secretion: multifactorial regulation for a single process of release. Diabetologia 9, 167–173 (1973)
Malaisse, W. J., Brisson, G. R., Baird, L.E.: The stimulus-secretion coupling of glucose-induced insulin release. X. Effect of glucose on 45 calcium efflux from perifused islets. Amer. J. Physiol., in press (1973)
Malaisse-Lagae, R., Malaisse, W.J.: Stimulus-secretion coupling of glucose-induced insulin release. III. Uptake of 45 calcium by isolated islets of Langerhans. Endocrinology 88, 72–80 (1971)
Matthews, J.L., Martin, J.H., Arsenis, C., Eisenstein, R., Kuettner, K.: The role of mitochondria in intracellular calcium regulation. In: Cellular mechanisms for calcium transfer and homeostasis, ed. by G. Nichols, and R.H. Wasserman, p. 239–255. New York-London: Academic Press 1971
Milner, R.D.G., Hales, C.N.: The role of calcium and magnesium in insulin secretion from rabbit pancreas studied in vitro. Diabetologia 3, 47–49 (1967)
Mizuhira, V.: Demonstration of the elemental distribution in biological tissues by means of the electron microscope and electron probe X-ray microanalyzer. Acta histochem. cytochem. 6, 44–52 (1973)
Rasmussen, H.: Cell communication, calcium ion, and cyclic adenosine monophosphate. Science 170, 404–412 (1970)
Rasmussen, H., Allen, J. E.: The calcium cyclic AMP relationship in cell activation. 9. Schwabinger Symposion der Forsch,gruppe Diabetes vom 21. 6. 73 in München
Sampson, H.W., Matthews, J.L., Martin, J.H., Kunin, A.S.: An electron microscopic localization of calcium in the small intestine of normal, rechitic and vitamin-D-treated rats. Calcif. Tiss. Res. 5, 305–316 (1970)
Schäfer, H.-J.: Ultrastructure and ion distribution of the intestinal cell during experimental vitamin-D deficiency rickets in rats. Virchows Arch. Abt. A Path. Anat. 359, 111–123 (1973)
Schäfer, H.-J., Klöppel, G.: Demonstration of calcium in pancreatic islets. Light microscopical observations in activated and inactivated B-cells of mice. Virchows Arch. Abt. A Path. Anat. in press
Schäfer, H.-J., Otto, H.F.: In preparation
Shiina, S., Mizuhira, V., Amakawa, T., Futaesaku, Y.: An analysis of the histochemical procedure for sodium ion detection. J. Histochem. Cytochem. 18, 644–649 (1970)
Shlatz, L., Marinetti, G.V.: Hormone-calcium interactions with the plasma membrane of rat liver cells. Science 176, 175–177 (1972)
Spicer, S.S., Swanson, A.A.: Elemental analysis of precipitate formed in nuclei by antimonate-osmium tetroxide fixation. J. Histochem. Cytochem. 20, 518–526 (1972)
Tandler, C.J., Kierszenbaum, A.L.: Inorganic cations in rat kidney. Localization with potassium pyroantimonate-perfusion fixation. J. Cell Biol. 50, 830–839 (1971)
Thureson-Klein, A., Klein, R.L.: Cation distribution and cardiac jelly in early embryonic heart: a histochemical and electron microscopic study. J. Molec. Cell. Cardiol. 2, 31–40 (1971)
Torack, R.M., Lavalle, M.: The specificity of the pyroantimonate technique to demonstrate sodium. J. Histochem. Cytochem. 18, 635–643 (1970)
Yarom, R., Meiri, U.: N lines in striated muscle: a site of intracellular Ca2+. Nature (Lond.) New Biol. 234, 254–256 (1971)
Author information
Authors and Affiliations
Additional information
Supported by SFB 34, Hamburg.
The authors wish to thank Miss Zeiger, Miss Fischer and Mrs. Baack for their skilful technical assistance. We are also indebted to Mr. and Mrs. Liebel in Kontron Application Laboratories, München, for effective cooperation in carrying out the X-ray analyses.
Rights and permissions
About this article
Cite this article
Schäfer, H.J., Klöppel, G. The significance of calcium in insulin secretion. Virchows Arch. A Path. Anat. and Histol. 362, 231–245 (1974). https://doi.org/10.1007/BF00432197
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00432197