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Pflügers Archiv

, Volume 411, Issue 5, pp 529–539 | Cite as

Aldosterone metabolism in rat renal tissue in vitro

Formation of lipid soluble metabolites
  • D. l'Allemand
  • H. Siebe
  • D. Tsiakiras
  • G. -A. Hoyer
  • P. Vecsei
  • K. Hierholzer
Transport Processes, Metabolism and Endocrinology; Kidney, Gastrointestinal Tract, and Exocrine Glands

Abstract

In the present study the formation of lipid soluble metabolites from3H-aldosterone was investigated in vitro in isolated kidneys and kidney and liver slices of Sprague Dawley rats. The steroids were separated by HPLC (forward and reversed phase systems) and detected on-line as UV- or3H-chromatograms. Apart from an unenzymatically formed substance, isoaldosterone, three less polar metabolites were traced (A1, A2, A3). The structure of the quantitatively most important metabolite (A1), was identified as 5α-dihydroaldosterone using a combination of techniques such as chromatographic comparison with reference steroids, antibody binding and mass spectrometry. Evidence for further conversion of DHaldo to 3α,5α-tetrahydroaldosterone was obtained in chromatographic and antibody binding studies. The formation of metabolites was not dependent on glomerular filtration. Furthermore it displayed regional heterogeneity with highest activity in the outer medulla. Finally it was observed that the in vitro metabolism of aldosterone was not saturable over a range of initial aldo concentration of 10−9 to 10−5 M.

Key words

Corticosteroid metabolism, renal Aldosterone Isolated rat kidney Kidney slices Liver slices HPLC 

Abbreviations

Aldo

aldosterone

B

corticosterone

CS

corticosteroids

HPLC

high pressure liquid chromatography

KH

Krebs-Henseleit

MCS

mineralocorticosteroid

MeOH

methanol

met

metabolite

MS

mass spectrometry

TH(DH)

tetrahydro-(dihydro)

tR

retention time

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References

  1. 1.
    l'Allemand D, Siebe H, Vecsei P, Hierholzer K (1985) Conversion of aldosterone (aldo) into less polar metabolites in rat renal tissue. Deutsche Gesellschaft für Physiologie, Berlin, Pflügers Arch, 403 (Suppl.):R18Google Scholar
  2. 2.
    Brien TG, Slater JDH (1967) Isomerization of radioactive aldosterone at C-17: enzymic purification of the 17α- and 17β-isomers. J Endocrinol 38:197–198Google Scholar
  3. 3.
    Botzen H (1987) Aldosteron- und Corticosteronmetabolismus in der Glandula mandibularis von Ratte, Schwein und Rind. Veterinärmedizinische Dissertation, Freie Universität Berlin (submitted)Google Scholar
  4. 4.
    Chao HP (1985) Vergleich des Corticosteronmetabolismus von Nieren- und Lebergewebe der Ratte in vitro. Medizinische Dissertation, Freie Universität BerlinGoogle Scholar
  5. 5.
    Conolly TM, Vecsei P, Haack D, Kohl KH, Abdelhamid S, Ammenti A (1978) Aldosterone diagnosis in hypertension: comparative evaluation of radioimmunoassays for urinary aldosterone and 18-OH-corticosterone. Klin Wochenschr 56 (Suppl. I):173–181Google Scholar
  6. 6.
    Crabbé J (1963) Sodium-retaining action of aldosterone. Presses Acad Eur, BruxellesGoogle Scholar
  7. 7.
    Deck KA, Siegenthaler W (1967) Further experimental evidence for the formation of the acid hydrolyzable conjugate of aldosterone by the kidney. Acta Endocrinol (Copenh) 55:637–647Google Scholar
  8. 8.
    Farman N, Bonvalet JP (1983) Aldosterone binding in isolated tubules. III. Autoradiography along the rat nephron. Am J Physiol 245:F606-F614Google Scholar
  9. 9.
    Farman N, Pradelles P, Bonvalet JP (1984) HPLC-analysis of bound aldosterone in kidney slices and isolated tubules. Int. Congr Nephrol, Renal Metabolism, Los Angeles, Abstr 367AGoogle Scholar
  10. 10.
    Farman N, Pradelles P, Bonvalet JP (1985) Binding of aldosterone metabolites in isolated tubular segments. Am J Physiol 249:F923-F932Google Scholar
  11. 11.
    Flood C, Layne DS, Ramcharan S, Rossipal E, Tait JF, Tait SAS (1961) An investigation of the urinary metabolites and secretion rates of aldosterone and cortisol in man and a description of methods for their measurement. Acta Endocrinol 36:237–264Google Scholar
  12. 12.
    Flood C, Pincus G, Tait JF, Tait SAS, Willoughby S (1967) A comparison of the metabolism of radioactive 17-isoaldosterone and aldosterone administered intravenously and orally to normal human subjects. J Clin Invest 46:717–727Google Scholar
  13. 13.
    Grogan WM, Fidelman ML, Newton DE, Duncan RL, Watlington CO (1985) A corticosterone metabolite produced by A6 (toad kidney) cells in culture: Identification and effects on Na+ transport. Endocrinology 116:1189–1194Google Scholar
  14. 14.
    Hierholzer K, Tsiakiras D, Schöneshöfer M, Siebe H, Weskamp P (1981) Renal handling of hormones. In: Greger R, Lang F, Silbernagel S (eds) Renal transport of organic substances. Springer, Berlin Heidelberg New York, pp 278–289Google Scholar
  15. 15.
    Hierholzer K, Lichtenstein I, Siebe H, Tsiakiras D, Witt I (1982) In vitro conversion of corticosterone (B) by rat renal tissue. Biochem of kidney functions. In: Morel F (ed) INSERM Symp. No. 21. Elsevier, Amsterdam, pp 233–240Google Scholar
  16. 16.
    Hierholzer K, Lichtenstein I, Siebe H, Tsiakiras D, Witt I (1982) Renal metabolism of corticosteroid hormones. Klin Wochenschr 60:1127–1135Google Scholar
  17. 17.
    Hierholzer K, Schöneshöfer M, Siebe H, Tsiakiras D, Weskamp P (1984) Corticosteroid metabolism in isolated rat kidney in vitro. I. Formation of lipid soluble metabolites from corticosterone in renal tissue. Pflügers Arch 400:363–371Google Scholar
  18. 18.
    Hoyer G-A, Tsiakiras D, Siebe H, Hierholzer K (1984) Corticosteroid metabolism in isolated rat kidney in vitro. III. Structure analysis of lipid soluble metabolites of corticosterone. Pflügers Arch 400:377–380Google Scholar
  19. 19.
    Kelly WG, Bandi L, Lieberman S (1962) Isolation and characterization of human urinary metabolites of aldosterone. III. Three isomeric tetrahydro metabolites. Biochemistry 1:792–803Google Scholar
  20. 20.
    Kelly WG, Bandi L, Lieberman S (1963a) Isolation and characterization of human urinary metabolites of aldosterone. IV. The synthesis and stereochemistry of two bicyclic acetal metabolites. Biochemistry 2:1243–1248Google Scholar
  21. 21.
    Kelly WG, Bandi L, Lieberman S (1963b) Isolation and characterization of human urinary metabolites of aldosterone. V. Dihydroaldosterone and 21-deoxy-tetrahydroaldosterone. Biochemistry 2:1249–1256Google Scholar
  22. 22.
    Kenyon CJ, Brem AS, McDermott MJ, Deconti GA, Latif SA, Morris DJ (1983) Antinatriuretic and kaliuretic activities of the reduced derivatives of aldosterone. Endocrinology 112:1852–1856Google Scholar
  23. 23.
    Kirk DN, Miller BW (1982) 18-substituted steroids. 9. Studies on the stability of aldosterone in dilute alkali. J Steroid Biochem 16:269–276Google Scholar
  24. 24.
    Kobayashi N, Schulz W, Hierholzer K (1987) Corticosteroid metabolism in rat kidney in vitro. IV. Subcellular sites of 11β-hydroxysteroid dehydrogenase activity. Pflügers Arch 408:46–63Google Scholar
  25. 25.
    Kohler H, Hesse RH, Pechet MM (1964) The metabolism of aldosterone. J Biol Chem 239:4117–4123Google Scholar
  26. 26.
    Luetscher JA, Hancock EW, Dowdy AJ, Nokes GW (1965) Conjugation of 1,2-3H-aldosterone in human liver and kidneys and renal extraction of aldosterone and labeled compounds from blood plasma. J Endocrinol 25:628–638Google Scholar
  27. 27.
    McCaa CS, Sulya LL (1966) Pathways of excretion of 1,2-3H-aldosterone radiometabolites in the rat. Endocrinology 79:815–818Google Scholar
  28. 28.
    McDermott M, Latif S, Morris DJ (1983) The metabolism of aldosterone in kidney. J Steroid Biochem 19:1205–1211Google Scholar
  29. 29.
    McDermott M, Freiberger M, Latif SA, Morris DJ (1985) The synthesis of reduced metabolites of aldosterone by subcellular fractions of rat kidney: effects of antimineralocorticoids. J Steroid Biochem 23:503–509Google Scholar
  30. 30.
    Möhring J, Möhring B, Siegenthaler W (1968a) Bildung und Ausscheidung von 18-Aldosteronglucuronid in den Nieren des Menschen. Z Gesamte Exp Med 146:336–345Google Scholar
  31. 31.
    Möhring J, Möhring B, Siegenthaler W (1968b) Die biologische Aktivität von Tetrahydroaldosteron, Tetrahydroaldosteronglucuronid und 18-Aldosteronglucuronid im Vergleich zu Aldosteron. Klin Wschr 46:22–24Google Scholar
  32. 32.
    Morris DJ (1981) The metabolism and mechanism of action of aldosterone. Endocr Rev 2:234–247Google Scholar
  33. 33.
    Morris DJ (1986) Further studies on aldosterone metabolism. Ann Clin Lab Science 16:94–102Google Scholar
  34. 34.
    Morris DJ, Tsai R (1981) Chromatographic separation of aldosterone and its metabolites. In: Giddings JC (ed). Advances in Chromatogr 19:261–285Google Scholar
  35. 35.
    Morris DJ, Berek JS, Davis RP (1973) Sex dependance of the metabolism of aldosterone in adrenalectomized and intact rats. Steroids 21:397–401Google Scholar
  36. 36.
    Morris DJ, McDermott MJ, Latif SA, Keating AW, Kenyon CJ (1981) The metabolism of aldosterone in target tissues. J Steroid Biochem 15:473–477Google Scholar
  37. 37.
    Nakane H, Nakane Y, Reach G, Corvol P, Menard J (1978) Aldosterone metabolism in isolated perfused rat kidney. Am J Physiol 234:E472-E479Google Scholar
  38. 38.
    Rossier BC, Wilce PA, Edelman IS (1974) Kinetics of RNA labeling in toad bladder epithelium: effects of aldosterone and related steroids. Proc Natl Acad Sci USA 71:3101–3105Google Scholar
  39. 39.
    Sandor T, Lanthier A (1962) The metabolism of aldosterone. II. Studies “in vitro” and “in vivo” in man. Acta Endocrinol (Copenh.) 39:87–102Google Scholar
  40. 40.
    Schulz W, Spieth A, Hierholzer K (1987) Renal handling of corticosteroids: localization of 11β-hydroxysteroid-dehydrogenase (11-HSD) within the nephron. Acta Endocrinol (Copenh) 114 (Suppl 283) 42 (Abstr)Google Scholar
  41. 41.
    Schurek HJ, Brecht JP, Lohfert H, Hierholzer K (1975) The basic requirements for the function of the isolated cell free perfused rat kidney. Pflügers Arch 354:349–365Google Scholar
  42. 42.
    Schöneshöfer, M, Fenner A (1981) A convenient and efficient method for the extraction and fractionation of steroid hormones from serum or urine. J Clin Chem Clin Biochem 19:71–74Google Scholar
  43. 43.
    Sekihara H, Island DP, Liddle GW (1978) New mineralocorticoids: 5α-dihydroaldosterone and 5α-dihydro-11-deoxycorticosterone. Endocrinology 103:1450–1452Google Scholar
  44. 44.
    Sharp GWG, Komuck CL, Leaf A (1966) Studies on the binding of aldosterone in the toad bladder. J Clin Invest 45:450–460Google Scholar
  45. 45.
    Siebe H, Tsiakiras D, Hierholzer K (1984) Corticosteroid metabolism in isolated rat kidney in vitro. II. Sex dependancy and formation of 11-dehydro-corticosterone. Pflügers Arch 400:372–376Google Scholar
  46. 46.
    Spieth A, Hierholzer K (1987) Longitudinal heterogeneity of renal corticosterone metabolism in mouse nephron. Abstr 64. Meeting Deutsche Physiologische Gesellschaft, Homburg. Pflügers Arch 408 (Suppl 1):R42Google Scholar
  47. 47.
    Stolte H, Brecht JP, Wiederholt M, Hierholzer K (1968) Einfluß von Adrenalektomie und Glucocorticoiden auf die Wasserpermeabilität corticaler Nephronabschnitte der Rattenniere. Pflügers Arch 299:99–127Google Scholar
  48. 48.
    Ulick S (1961) Stereospecificity in the metabolism of aldosterone in man. J Biol Chem 236:680–684Google Scholar
  49. 49.
    Ulick S, Lieberman S (1957) Evidence for the occurrence of a metabolite of aldosterone in urine. J Am Chem Soc 79:6567Google Scholar
  50. 50.
    Ulick S, Laragh JH, Lieberman S (1958) The isolation of a urinary metabolite of aldosterone and its use to measure the rate of secretion of aldosterone by the adrenal cortex of man. Trans Assoc Am Physicians 71:225–229Google Scholar
  51. 51.
    Ulick S, Kusch K, August JT (1961) Correction of the structure of a urinary metabolite of aldosterone. J Am Chem Soc 83:4482Google Scholar

Copyright information

© Springer-Verlag 1988

Authors and Affiliations

  • D. l'Allemand
    • 1
  • H. Siebe
    • 1
  • D. Tsiakiras
    • 1
  • G. -A. Hoyer
    • 2
  • P. Vecsei
    • 1
    • 3
  • K. Hierholzer
    • 1
  1. 1.Institut für Klinische PhysiologieKlinikum Steglitz der Freien Universität BerlinBerlin 45Germany
  2. 2.Forschungslaboratorien der Schering AGBerlin 65Germany
  3. 3.Pharmakologisches Institut der Universität HeidelbergHeidelbergGermany

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