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.
Similar content being viewed by others
References
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
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
Beal MF, Hyman BT, Koroshetz W (1993) Do defects in mitochondrial energy metabolism underlie the pathology of neurodegenerative diseases? Trends Neurosci 16: 125–131
Bergamasco B, Benna P, Ferrero P, Gavinelli R (1984) Neonatal hypoxia and epileptic risk: a clinical prospective study. Epilepsia 25: 131–136
Brown JK (1976) Infants damaged during birth: perinatal asphyxia. In: Hull D (ed) Recent advances in paediatrics. Churchill Livingstone, Edinburgh, pp 57–88
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
Chang Y-C, Gottlieb DI (1988) Characterization of the protein purified with monoclonal antibodies to glutamic acid decarboxylase. J Neurosci 8: 2123–2130
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
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
Eng LF (1985) Glial fibrillary acidic protein: the major protein of glial intermediate filaments in differentiated astrocytes. J Neuroimmunol 8: 203–214
Eng LF, Yu ACH, Lee YL (1992) Astrocytic response to injury. Prog Brain Res 94: 353–365
Erlander MG, Tillakaratne NJK, Feldblum S, Patel N, Tobin AJ (1991) Two genes encode distinct glutamate decarboxylases. Neuron 7: 91–100
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
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
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
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
Gale K (1992) GABA and epilepsy: basic concepts from preclinical research. Epilepsia 33 [Suppl] 5: S3-S12
Gloor P, Fariello RG (1988) Generalized epilepsy: some of its cellular mechanisms differ from those of focal epilepsy. Trends Neurosci 11: 63–68
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
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
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
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
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
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
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
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
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
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
Kosaka T, Hama K (1986) Three-dimensional structure of astrocytes in the rat dentate gyrus. J Comp Neurol 249: 242–260
Levine S (1960) Anoxic-ischemic encephalopathy in rats. Am J Pathol 36: 1–14
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
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
Martin DL, Rinvall K (1993) Regulation of g-aminobutyric acid synthesis in the brain. J Neurochem 60: 395–407
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
Meinecke DL, Peters A (1987) GABA immunoreactive neurons in rat visual cortex. J Comp Neurol 261: 388–404
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
Norenberg MD, Hertz L, Schousboe A (eds) (1988) The biochemical pathology of astrocytes. Alan R. Liss, New York
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
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
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
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
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
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
Rice JE, Vanucci RC, Brierley JB (1981) The influence of immaturity of hypoxic-ischemic brain damage in the rat. Ann Neurol 9: 131–141
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
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
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
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
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
[Reference deleted]
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
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
Silverstein F, Buchanan K, Johnston MV (1984) Pathogenesis of hypoxic-ischemic brain injury in a perinatal rodent model. Neurosci Lett 49: 271–277
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
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
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
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
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
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
Wong-Riley MTT (1989) Cytochrome oxidase: an endogenous metabolic marker for neuronal activity. Trends Neurosci 12: 94–101
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
Yu ACH, Hertz L, Norenberg MD, Syková E, Waxman SG (1992) Neuronal-astrocytic interactions. Implications for normal and pathological CNS function. Elsevier, Amsterdam
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
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
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00293323