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Permanent increase of immunocytochemical reactivity for γ-aminobutyric acid (GABA), glutamic acid decarboxylase, mitochondrial enzymes, and glial fibrillary acidic protein in rat cerebral cortex damaged by early postnatal hypoxia-ischemia

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Abstract

A former study indicated that hypoxicischemic encephalopathy in rat sustained during early postnatal life may result in permanent epileptic activity in the baseline electroencephalogram. We, therefore, investigated whether the presumed higher firing frequency and metabolic activity of neurons in such hypoxia-damaged cortical areas would be reflected by an enhanced light microscopic immunoreactivity of γ-aminobutyric acid (GABA), the two isoforms of glutamic acid decarboxylase (GAD67 and GAD65), the mitochondrial enzymes cytochrome c oxidase and ATP synthase, and/or glial fibrillary acidic, protein (GFAP). To that end rat pups, 12–13 days of age, were unilaterally exposed to hypoxic-ischemic conditions and, after a survival period of 2 and 61/2 months, respectively, killed by perfusion fixation. After dissection of the brain, coronal vibratome sections of animals showing cortical damage were immunostained for the presence of the abovementioned antigens. Subsequent qualitative analysis revealed that the surroundings of cortical infarctions were unambiguously characterized by a disordered neural network containing numerous nerve cells, fibers and/or endings showing an enhanced immunoreactivity for GABA, both isoforms of glutamic acid decarboxylase, and cytochrome c oxidase and ATP synthase, while the astrocytes showed an enhanced immunoreactivity for GFAP. The diverse patterns of enhanced immunoreactivity suggested, furthermore, a wider low-to-high range of metabolic activities in both excitatory and inhibitory neurons.

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References

  1. André M, Vert P, Debrouille C (1978) Diagnostic et évolution de la souffrance cérébrale chez les nouveau-nés ayant presenté des signes d'hypoxie foetale. Arch Fr Pediatr 35: 23–36

    Google Scholar 

  2. Bakker HD, Scholte HR, Van den Bogert C, Ruitenbeek W, Jeneson JAL, Wanders RJA, Abeling NGGM, Dorland B, Sengers RCA, Van Gennip AH (1993) Deficiency of the adenine nucleotide translocator in muscle of a patient with myopathy and lactic acidosis: a new mitochondrial defect. Pediatr Res 33: 412–417

    Google Scholar 

  3. Beal MF, Hyman BT, Koroshetz W (1993) Do defects in mitochondrial energy metabolism underlie the pathology of neurodegenerative diseases? Trends Neurosci 16: 125–131

    Google Scholar 

  4. Bergamasco B, Benna P, Ferrero P, Gavinelli R (1984) Neonatal hypoxia and epileptic risk: a clinical prospective study. Epilepsia 25: 131–136

    Google Scholar 

  5. Brown JK (1976) Infants damaged during birth: perinatal asphyxia. In: Hull D (ed) Recent advances in paediatrics. Churchill Livingstone, Edinburgh, pp 57–88

    Google Scholar 

  6. Castiglioni AJ, Peterson SL, Sanabria EL, Tiffany-Castiglioni E (1990) Structural changes in astrocytes induced by seizures in a model of temporal lobe epilepsy. J Neurosci Res 26: 334–341

    Google Scholar 

  7. Chang Y-C, Gottlieb DI (1988) Characterization of the protein purified with monoclonal antibodies to glutamic acid decarboxylase. J Neurosci 8: 2123–2130

    Google Scholar 

  8. DeLeo J, Toth L, Schubert P, Rudolphi K, Kreutzberg GW (1987) Ischemia-induced neuronal cell death, calcium accumulation, and glial response in the hippocampus of the Mongolian gerbil and protection by propentofylline (HWA 285). J Cereb Blood Flow Metab 7: 745–751

    Google Scholar 

  9. Delpech B, Delpech A, Vidard MN, Girard N, Tayot J, Clément JC, Creissard P (1978) Glial fibrillary acidic protein in tumours of the central nervous system. Br J Cancer 37: 33–40

    Google Scholar 

  10. Eng LF (1985) Glial fibrillary acidic protein: the major protein of glial intermediate filaments in differentiated astrocytes. J Neuroimmunol 8: 203–214

    Google Scholar 

  11. Eng LF, Yu ACH, Lee YL (1992) Astrocytic response to injury. Prog Brain Res 94: 353–365

    Google Scholar 

  12. Erlander MG, Tillakaratne NJK, Feldblum S, Patel N, Tobin AJ (1991) Two genes encode distinct glutamate decarboxylases. Neuron 7: 91–100

    Google Scholar 

  13. Faddis BT, Vijayan VK (1988) Application of glial fibrillary acidic protein immunohistochemistry in the quantification of astrocytes in the rat brain. Am J Anat 1983: 316–322

    Google Scholar 

  14. Feldblum S, Ackermann RF, Tobin AJ (1990) Long-term increase of glutamate decarboxylase mRNA in a rat model of temporal lobe epilepsy. Neuron 5: 361–371

    Google Scholar 

  15. Feldblum S, Erlander MG, Tobin AJ (1993) Different distributions of GAD65 and GAD67 mRNAs suggest that the two glutamate decarboxylases play distinctive functional roles. J Neurosci Res 34: 689–706

    Google Scholar 

  16. Gabbott PLA, Stewart MG, Rose SPR (1986) The quantitative effects of dark-rearing and light exposure on the laminar composition and depth distribution of neurons and glia in the visual cortex (Area 17) of the rat. Exp Brain Res 64: 225–232

    Google Scholar 

  17. Gale K (1992) GABA and epilepsy: basic concepts from preclinical research. Epilepsia 33 [Suppl] 5: S3-S12

    Google Scholar 

  18. Gloor P, Fariello RG (1988) Generalized epilepsy: some of its cellular mechanisms differ from those of focal epilepsy. Trends Neurosci 11: 63–68

    Google Scholar 

  19. Gonzales C, Kaufman DL, Tobin AJ, Chesselet M-F (1991) Distribution of glutamic acid decarboxylase (M 1 67 000) in the basal ganglia of the rat: an immunohistochemical study with a selective cDNA-generated polyclonal antibody. J Neurocytol 20: 953–961

    Google Scholar 

  20. Grimaldi R, Zoli M, Agnati LF, Ferraguti F, Fuxe K, Toffano G, Zini I (1990) Effects of transient forebrain ischemia on peptidergic neurons and astroglial cells: evidence for recovery of peptide immunoreactivities in neocortex and striatum but not hippocampal formation. Exp Brain Res 82: 123–136

    Google Scholar 

  21. Hawrylak N, Chang F-LF, Greenough WT (1993) Astrocytic and synaptic response to kindling in hippocampal subfield CA1. I. Synaptogenesis and astrocytic process increases to in vivo kindling. Brain Res 603: 309–316

    Google Scholar 

  22. Hendry SHC (1991) Delayed reduction in GABA and GAD immunoreactivity of neurons in the adult monkey dorsal lateral geniculate nucleus following monocular deprivation or enucleation. Exp Brain Res 86: 47–59

    Google Scholar 

  23. Hendry SHC, Jones EG (1986) Reduction in number of immunostained GABAergic neurones in deprived-eye dominance columns of monkey area 17. Nature 320: 750–753

    Google Scholar 

  24. Hevner RF, Wong-Riley MRR (1991) Neuronal expression of nuclear and mitochondrial genes for cytochrome oxidase (CO) subunits analysed by in situ hybridization: comparison with CO activity and protein. J Neurosci 11: 1942–1958

    Google Scholar 

  25. Hevner RF, Wong-Riley MTT (1993) Mitochondrial and nuclear gene expression for cytochrome oxidase subunits are disproportionately regulated by functional activity in neurons. J Neurosci 13: 1805–1819

    Google Scholar 

  26. Hevner RF, Duff RS, Wong-Riley (1992) Coordination of ATP production and consumption in brain: parallel regulation of cytochrome oxidase and Na+K+-ATPase. Neurosci Lett 138: 188–192

    Google Scholar 

  27. Houser CR, Vaugh JE, Hendry SHC, Jones EG, Peters A (1984) GABA neurons in the cerebral cortex. In: Jones EG, Peters A (eds) Cerebral cortex, vol 2. Plenum Press, New York, pp 63–117

    Google Scholar 

  28. Kaufman DL, Houser CR, Tobin AJ (1991) Two forms of the γ-aminobutyric acid synthetic enzyme glutamate decarboxylase have distinct intraneuronal distributions and cofactor interactions. J Neurochem 56: 720–723

    Google Scholar 

  29. Kosaka T, Hama K (1986) Three-dimensional structure of astrocytes in the rat dentate gyrus. J Comp Neurol 249: 242–260

    Google Scholar 

  30. Levine S (1960) Anoxic-ischemic encephalopathy in rats. Am J Pathol 36: 1–14

    Google Scholar 

  31. Litwak J, Mercugliano M, Chesselet M-F, Oltmans GA (1990) Increased glutamic acid decarboxylase (GAD) mRNA and GAD activity in cerebellar Purkinje cells following lesion-induced increases in cell firing. Neurosci Lett 116: 179–183

    Google Scholar 

  32. Lopes da Silva FH, Kamphuis W, Van Neerven JMAM, Pijn JPM (1990) Cellular and network mechanisms in the kindling model of epilepsy: the role of GABAergic inhibition and the emergence of strange attractors. In: Roy John E, Valdes M (eds) Recent advances in basic and clinical neuroscience, machinery of the mind. Birkhäuser, Boston, pp 115–139

    Google Scholar 

  33. Martin DL, Rinvall K (1993) Regulation of g-aminobutyric acid synthesis in the brain. J Neurochem 60: 395–407

    Google Scholar 

  34. Martin SM, Landel HB, Lansing AJ, Vijayan VK (1991) Immunocytochemical double labeling of glial fibrillary acidic protein and transferrin permits the identification of astrocytes and oligodendrocytes in the rat brain. J Neuropathol Exp Neurol 50: 161–170

    Google Scholar 

  35. Meinecke DL, Peters A (1987) GABA immunoreactive neurons in rat visual cortex. J Comp Neurol 261: 388–404

    Google Scholar 

  36. Najlerahim A, Williams SF, Pearson RCA, Jefferys JGR (1992) Increased expression of GAD mRNA during the chronic epileptic syndrome due to intrahippocampal tetanus toxin. Exp Brain Res 90: 332–342

    Google Scholar 

  37. Norenberg MD, Hertz L, Schousboe A (eds) (1988) The biochemical pathology of astrocytes. Alan R. Liss, New York

    Google Scholar 

  38. Oertel WH, Schmechel DE, Tappaz ML, Kopin I (1981) Production of a specific antiserum to rat brain glutamic acid decarboxylase by injection of an antigen-antibody complex. Neuroscience 6: 2689–2700

    Google Scholar 

  39. Petito CK, Halaby IA (1993) Relationship between ischemia and ischemic neuronal necrosis to astrocyte expression of glial fibrillary acidic protein. Int J Dev Neurosci 11: 239–247

    Google Scholar 

  40. Petito CK, Morgello S, Felix JC, Holden LM (1988) Astrocytes in cerebral ischemia. In: Norenberg MD, Hertz L, Schousboe A (eds) The biochemical pathology of astrocytes. Alan R. Liss, New York, pp 79–90

    Google Scholar 

  41. Petito CK, Morgello S, Felix JC, Lesser ML (1990) The two patterns of reactive astrocytosis in postischemic rat brain. J Cereb Blood Flow Metab 10: 850–859

    Google Scholar 

  42. Petito CK, Chung M, Halaby IA, Cooper AJL (1992) Influence of the neuronal environment on the pattern of reactive astrocytosis following cerebral ischemia. Prog Brain Res 94: 381–387

    Google Scholar 

  43. Prince DA, Deisz RA, Thompson SM, Chagnac-Amitai Y (1992) Functional alterations in GABAergic inhibition during activity. In: Avanzani G, Engel J Jr, Fariello R, Heinemann U (eds.) Neurotransmitters in Epilepsy (Epilepsy Res [Suppl 8]. Elsevier, Amsterdam, pp 31–38

    Google Scholar 

  44. Rice JE, Vanucci RC, Brierley JB (1981) The influence of immaturity of hypoxic-ischemic brain damage in the rat. Ann Neurol 9: 131–141

    Google Scholar 

  45. Rigaud AS, Pinard E, Borredon J, Seylaz J (1990) Effect of chronic cervical sympathectomy on local cerebral blood flow during limbic seizures in rat. Brain Res 532: 347–350

    Google Scholar 

  46. Romijn HJ, Hofman MA, Gramsbergen A (1991) At what age is the developing cerebral cortex of the rat comparable to that of the full-term newborn human baby? Early Hum Dev 26: 61–67

    Google Scholar 

  47. Romijn HJ, Janszen AWJW, Van Voorst MJD, Buijs RM, Balázs R, Swaab DF (1992) Perinatal hypoxic ischemic encephalopathy affects the proportion of GABA-immunoreactive neurons in the cerebral cortex of the rat. Brain Res 592: 17–28

    Google Scholar 

  48. Romijn HJ, Van Marle J, Janszen AWJW (1993) Permanent increase of the GAD67/synaptophysin ratio in rat cerebral cortex nerve endings as a result of hypoxic-ischemic encephalopathy sustained in early postnatal life. A confocal laser scanning microscopic study. Brain Res 630: 315–329

    Google Scholar 

  49. Romijn HJ, Voskuyl RA, Coenen AML (1994) Hypoxic ischemic encephalopathy sustained in early postnatal life may result in premanent epileptic activity and an altered cortical convulsive threshold in rat. Epilepsy Res 17: 31–42

    Google Scholar 

  50. [Reference deleted]

  51. Schmidt-Kastner R, Wietasch K, Weigel H, Eysel UT (1993) Immunohistochemical staining for glial fibrillary acidic protein (GFAP) after deafferentation or ischemic infarction in rat visual system: features of reactive and damaged astrocytes. Int J Dev Neurosci 11: 157–174

    Google Scholar 

  52. Segovia J, Tillakaratne NJK, Whelan K, Tobin AJ, Gale K (1990) Parallel increases in striatal glutamic acid decarboxylase activity and mRNA levels in rats with lesions of the nigrostriatal pathway. Brain Res 529: 345–348

    Google Scholar 

  53. Silverstein F, Buchanan K, Johnston MV (1984) Pathogenesis of hypoxic-ischemic brain injury in a perinatal rodent model. Neurosci Lett 49: 271–277

    Google Scholar 

  54. Sirevaag AM, Greenough WT (1991) Plasticity of GFAP-immunoreactive astrocyte size and number in visual cortex of rats reared in complex environments. Brain Res 540: 273–278

    Google Scholar 

  55. Van den Bogert C, Pennings A, Dekker HL, Luciaková K, Boezeman JBM, Sinjorgo KMC (1991) Quantification of mitochondrial proteins in cultured cells by immuno-flow cytometry. Biochim Biophys Acta 1097: 87–94

    Google Scholar 

  56. Wasterlain C (1989) Epileptic seizures. In: Siegel GJ, et al (eds) Basic neurochemistry: molecular, cellular, and medical aspects, 4th edn. Raven Press, New York, pp 797–810

    Google Scholar 

  57. Watanabe K, Hara K, Miyazaki S, Hakamada S (1980) The role of perinatal brain injury in the genesis of childhood epilepsy. Folia Psychiatry Neurol Jpn 34: 227–232

    Google Scholar 

  58. Welker E, Soriano E, Dörfl J, Van der Loos H (1989) Plasticity in the barrel cortex of the adult mouse: transient increase of GAD-immunoreactivity following sensory stimulation. Exp Brain Res 78: 659–664

    Google Scholar 

  59. Wenzel J, Lammert G, Meyer U, Krug M (1991) The influence of long-term potentiation on the spatial relationship between astrocyte processes and potentiated synapses in the dentate gyrus neuropil of rat brain. Brain Res 560: 122–131

    Google Scholar 

  60. Wong-Riley MTT (1989) Cytochrome oxidase: an endogenous metabolic marker for neuronal activity. Trends Neurosci 12: 94–101

    Google Scholar 

  61. Yamamoto K, Yoshimine T, Homburger HA, Yanagihara T (1986) Immunohistochemical investigation of regional cerebral ischemia in the gerbil: occlusion of the posterior communicating artery. Brain Res 371: 244–252

    Google Scholar 

  62. Yu ACH, Hertz L, Norenberg MD, Syková E, Waxman SG (1992) Neuronal-astrocytic interactions. Implications for normal and pathological CNS function. Elsevier, Amsterdam

    Google Scholar 

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Authors and Affiliations

  1. Graduate School Neurosciences Amsterdam, Netherlands Institute for Brain Research, Meibergdreef 33, NL-1105 AZ, Amsterdam, The Netherlands

    H. J. Romijn & A. W. J. W. Janszen

  2. Graduate School Neurosciences Amsterdam, Department of Neurology, University of Amsterdam, Meibergdreef 9, NL-1105 AZ, Amsterdam, The Netherlands

    C. Van den Bogert

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Romijn, H.J., Janszen, A.W.J.W. & Van den Bogert, C. Permanent increase of immunocytochemical reactivity for γ-aminobutyric acid (GABA), glutamic acid decarboxylase, mitochondrial enzymes, and glial fibrillary acidic protein in rat cerebral cortex damaged by early postnatal hypoxia-ischemia. Acta Neuropathol 87, 612–627 (1994). https://doi.org/10.1007/BF00293323

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