Acta Neuropathologica

, Volume 65, Issue 2, pp 150–157

Limitations of tetrazolium salts in delineating infarcted brain

  • T. M. Liszczak
  • E. T. Hedley-Whyte
  • J. F. Adams
  • D. H. Han
  • V. S. Kolluri
  • F. X. Vacanti
  • R. C. Heros
  • N. T. Zervas
Original Works

Summary

Tetrazolium salts, histochemical indicators of mitochondrial respiratory enzymes, have been used by some pathologists to detect infarcts in myocardium. We explored the utility of this technique in detecting experimental brain infarcts and report our findings. Infarcts were produced in cats, gerbils, and rats by unilateral temporal and permanent cerebral vessel occlusion. After various time periods the animals were killed, and their brains were reacted with 2,3,5, triphenyl, 2H-tetrazolium chloride (TTC). The experimental and contralateral hemispheres were examined by light and electron microscopy. The TTC-stained tissue was correlated with histology. In some situations the histological condition of the tissue correlated well with the TTC staining results. Brain regions supplied by temporarily occluded vessels and judged infarcted by light and electron microscopy did not stain. In these regions less than 6% of the mitochondria were intact. In brain tissue from animals with permanent vessel occlusion (no reflow) mitochondria were intact despite the fact that other cellular organelles, such as nuclei, were destroyed. TTC stained such mitochondria and as a result could not distinguish infarcted brain in complete ischemia situations (no reflow). Another draw back to this staining procedure was 36 h after infarction macrophages with intact mitochondria would replace damage neurons and be stained. Under ideal conditions though this technique can detect irreversibly damaged brain as early as 2.5 h after artery occlusion.

Key words

Histochemical Delineation Infarcted Brain 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Arsenio-Nunes ML, Hossmann KA, Farkas-Bargeton E (1973) Ultrastructural and histochemical investigation of the cerebral cortex of cat during and after complete ischemia. Acta Neuropathol (Berl) 26:329–344Google Scholar
  2. 2.
    Aukland K, Bower BF, Berliner RW (1964) Measurement of local blood flow with hydrogen gas. Circ Res 14:164–187Google Scholar
  3. 3.
    Brierley JB, Meldrum BS, Brown AW (1973) The threshold and neuropathology of cerebral ‘anoxic-ischemic’ cell change. Arch Neurol 29:367–374Google Scholar
  4. 4.
    Brown AW, Brierley JB (1972) Anoxic-ischemic cell change in rat brain, light-microscopic and fine-structural observations. J Neurol Sci 16:59–84Google Scholar
  5. 5.
    Brown AW, Brierley JB (1973) The earliest alterations in rat neurons and astrocytes after anoxia-ischemia. Acta Neuropathol (Berl) 23:9–22Google Scholar
  6. 6.
    Derias NW, Adams CW (1978) Nitroblue tetrazolium test: early gross detection of human myocardial infarcts. Br J Exp Pathol 59:254–258Google Scholar
  7. 7.
    Dodson RF, Kawamura Y, Aoyagi M, Hartmann A, Cheung L (1973) A comparative evaluation of the ultrastructural changes following induced cerebral infarction in the squirrel monkey and baboon. Cytobios 8:175–182Google Scholar
  8. 8.
    Dodson RF, Aoyagi M, Hartmann A, Tagashira Y (1974) Acute cerebral infarction and hypotinsion: an ultrastructural study. J Neuropathol Exp Neurol 33:400–407Google Scholar
  9. 9.
    Dodson RF, Chu LW, Welch K, Achar VS (1977) Acute tissue response to cerebral ischemia in the gerbil, an ultrastructural study. J Neurol Sci 33:161–170Google Scholar
  10. 10.
    Fallon JT (1981) Postmortem histochemical techniques. In: Wagner GS (ed) Myocardial infarction measurement and intervention. Martinus Nijhoff, Boston, pp 373–384Google Scholar
  11. 11.
    Feldman S, Glagov S, Wissler RW, Hughes R (1976) Postmortem delineation of infarcted myocardium. Coronary perfusion with nitro blue tetrazolium. Arch Pathol Lab Med 100:55–58Google Scholar
  12. 12.
    Fernando DA, Lau KKH (1978) An electron microscopic study of the effects of acute ischemia in the brain. Acta Anat 100:241–249Google Scholar
  13. 13.
    Fishbein MC, Meerbaum S, Rit J, Lando U, Kanmatsuse K, Mercier JC, Corday E, Ganz W (1981) Early phase of acute myocardial infarct size quantitation: validation of the triphenyl tetrazolium chloride technique. Am Heart J 101:593–600Google Scholar
  14. 14.
    Garcia JH, Cox JV, Hudgins WR (1971) Ultrastructure of the microvasculature in experimental cerebral infarction. Acta Neuropathol (Berl) 18:273–285Google Scholar
  15. 15.
    Garcia JH, Kamijyo Y, Kalimo H, Tanaka J, Viloria JE, Trump BF (1975) Cerebral ischemia: the early structural changes and correlation of these with known metabolic and dynamic abnormalities. In: Wishnant JP, Sandok BA (eds) Cerebral vascular diseases. Grune and Stratton, New York, pp 313–323Google Scholar
  16. 16.
    Ginsberg MD, Graham DI, Welsh FA, Budd W (1978) Diffuse cerebral ischemia in the cat. III. Neuropathological sequelae of severe ischemia. Ann Neurol 5:350–358Google Scholar
  17. 17.
    Hager H, Hirschberger W, Scholtz W (1960) Electron-microscopic changes in brain tissue of Syrian hamster following acute hypoxia. Acrospace Med 34:379–389Google Scholar
  18. 18.
    Hamaya K, Sonobe H (1978) Macroscopic identification of cerebral infarction. VIIIth International Congress of Neuropathology. Washington, DC, [Abstr 124]Google Scholar
  19. 19.
    Hossmann KA, Sato K (1971) Effect of ischemia on the function of the sensorimotor cortex in cat. Electroencephalogy Clin Neurophysiol 30:535–545Google Scholar
  20. 20.
    Hossmann KA, Kleihues P (1973) Reversibility of ischemic brain damage. Arch Neurol 29:375–384Google Scholar
  21. 21.
    Hossmann KA, Zimmermann V (1974) Resuscitation of the monkey brain after 1 h of complete ischemia. I. Physiological and morphological observations. Brain Res 81:59–74Google Scholar
  22. 22.
    Jenkins LW, Povlishock JT, Lewett W, Miller JD, Becker DP (1981) The role of postischemic recirculation in the development of ischemic neuronal injury following complete cerebral ischemia. Acta Neuropathol (Berl) 55:205–220Google Scholar
  23. 23.
    Jennings RB (1976) Cell volume regulation in acute myocardial ischemie injury. Acta Med Scand 587:83–93Google Scholar
  24. 24.
    Jestadt R, Sandritter W (1959) Erfahrungen mit der TTC (Triphenyltetrazoliumchlorid)-Reaktion für die pathologisch-anatomische Diagnose des frischen Herzinfarktes. Kreislaufforsch 48:802–809Google Scholar
  25. 25.
    Kalimo H, Garcia JH, Kamijyo Y, Tanaka J, Trump BF (1977) The ultrastructure of ‘brain death’. II. Electron microscopy of feline cortex after complete ischemia. Virchows Arch [Cell Pathol] 25:207–220Google Scholar
  26. 26.
    Kalimo H, Olsson Y, Palajarvi L, Soderfeldt B (1982) Structural changes in brain tissue under hypoxic-ischemic conditions. J Cereb Blood Flow Metabol [Suppl 1] 2:19–22Google Scholar
  27. 27.
    Kleihues P, Hossmann KA (1971) Protein synthesis in the cat brain after prolonged cerebral ischemia. Brain Res 35:409–418Google Scholar
  28. 28.
    Levy DE, Brierley JB, Silverman DG, Plum F (1975) Brief hypoxia-ischemia initially damages neurons. Arch Neurol 32:450–456Google Scholar
  29. 29.
    McGee-Russel SM, Brown AW, Brierley JB (1970) A combined light and electron microscope study of early anoxic-ischemic cell changes in rat brain. Brain Res 20:193–200Google Scholar
  30. 30.
    Mittelman R, Szabo S, Heinsimar J, Reynolds ES, von Lichtenberg F (1975) Macroscopic histochemical detection of myocardial necrosis in the heart at autopsy by the triphenyl tetrazolium chloride (TTC) technique. Clin Res 23:198 [Abstr]Google Scholar
  31. 31.
    Nachlas MM, Tson KC, Souza ED, Chang CS, Seligman AM (1963) Cytochemical demonstration of succinic dehydrogenase by the use of a new p-nitrophenyl substituted ditetrazole. J Histochem Cytochem 5:420–436Google Scholar
  32. 32.
    Nachlas MM, Shnitka TK (1963) Macroscopic identification of early myocardial infarcts by alterations in dehydrogenase activity. Am J Pathol 42:379–405Google Scholar
  33. 33.
    Palajarvi L, Alihanka J, Kalimo H (1984) Significance of fluid for morphology of acute hypoxic-ischemic brain cell injury. Neuropathol Appl Neurobiol 10:43–52Google Scholar
  34. 34.
    Ramkissoon RA (1966) Macroscopic identification of early myocardial infarction by dehydrogenase alterations. J Clin Pathol 19:479–481Google Scholar
  35. 35.
    Sandritter W, Jestadt R (1958) Triphenyltetrazoliumchlorid (TTC) als Reduktionsindikator zur makroskopischen Diagnose des frischen Herzinfarktes. Zentralbl Allg Pathol 97:188–189Google Scholar
  36. 36.
    Schaper J, Mulch J, Winkler B, Schaper W (1979) Ultrastructural, functional, and biochemical criteria for estimation of reversibility of ischemic injury: a study on the effects of global ischemia on the isolated dog heart. J Molec Cell Cardiol 11:521–541Google Scholar
  37. 37.
    Schutz M, Silverstin PR, Vapalahti M (1973) Brain mitochondrial function after ischemia and hypoxia. Arch Neurol 29:408–416Google Scholar
  38. 38.
    Shnitka TK, Nachlas MM (1963) Histochemical alterations in ischemic heart muscle and early myocardial infarction. Am J Pathol 42:507–527Google Scholar
  39. 39.
    Trump BF, Croker BP, Mergner WJ (1971) The role of energy metabolism, ion, and watershifts in the pathogenesis of cell injury. In: Richter GW, Scarpelli DG (eds) Cell membranes: biological and pathological aspects. Williams and Wilkins, Baltimore, p 84Google Scholar
  40. 40.
    Trump BF, McDowell EM, Arstila AV (1980) Cellular reaction to injury. In: Hill RB, jr, La Via MF (eds) Principles of pathobiology. Oxford University Press, New Yrok, pp 30–47Google Scholar
  41. 41.
    Webster HF, Ames A (1965) Reversible and irreversible changes in the fine structure of nervous tissue during oxygen and glucose deprivation. J Cell Biol 26:885–909Google Scholar
  42. 42.
    Welsh FA, Ginsberg MD, Rieder W, Budd WW (1978) Diffuse cerebral ischemia in the cat. II. Regional metabolites during severe ischemia and recirculation. Ann Neurol 3:493–501Google Scholar
  43. 43.
    Westergaard E, Go G, Klatzo I, Spatz M (1976) Increased permeability of cerebral vessels to horseradish peroxidase induced by ischemia in Mongolian gerbils. Acta Neuropathol (Berl) 34:1–6Google Scholar
  44. 44.
    Zervas NT, Geyer RP, Han DH, Adams JF, Ropper AH, Kennedy SK, Heros RC, Varsos V, Borges L, Hedley-Whyte ET (1983) Can perfluochemicals reduce cerebral ischemia? In: Reivich M, Hutig HI (eds) Cerebrovascular diseases. Raven Press, New York, pp 409–419Google Scholar

Copyright information

© Springer-Verlag 1984

Authors and Affiliations

  • T. M. Liszczak
    • 1
    • 2
  • E. T. Hedley-Whyte
    • 1
    • 2
  • J. F. Adams
    • 1
    • 2
  • D. H. Han
    • 1
    • 2
  • V. S. Kolluri
    • 1
    • 2
  • F. X. Vacanti
    • 1
    • 2
  • R. C. Heros
    • 1
    • 2
  • N. T. Zervas
    • 1
    • 2
  1. 1.Neurosurgical Service and NeuropathologyMassachusetts General HospitalBostonUSA
  2. 2.Harvard Medical SchoolBostonUSA

Personalised recommendations