Elsevier

Neuroscience

Volume 62, Issue 4, October 1994, Pages 1133-1146
Neuroscience

Quantitative ultrastructural localization of glutamate dehydrogenase in the rat cerebellar cortex

https://doi.org/10.1016/0306-4522(94)90349-2Get rights and content

Abstract

Glutamate dehydrogenase is one of the main enzymes involved in the formation and metabolism of the neurotransmitter glutamate. In the present study we investigated the enzyme ultrastructurally in the cerebellar cortex, a region rich in well defined glutamatergic neurons, by pre-embedding immunocytochemical staining (peroxidase-antiperoxidase), as well as by post-embedding immunogold labelling employing a new system for quantitation and for specificity testing under the conditions of the immunocytochemical procedure. A new antiserum against immunologically purified bovine liver glutamate dehydrogenase or antibodies isolated from this by affinity chromatography were used in rats fixed by perfusion with aldehydes.

The pre-embedding method displayed peroxidase reaction preferentially in mitochondria of astroglial cells (including the Bergmann glia). Mitochondria of neuronal tissue elements were usually free of peroxidase-reaction product. Extra-mitochondrial staining was not observed.

The post-embedding immunogold method was employed to overcome penetration problems and allow semiquantitative analysis of localization and specificity. The highest densities of gold particles were found over the mitochondria in astroglial cell elements (including the Bergmann glia). Mitochondria in cell bodies of Bergmann glia had a lower particle density than those in astrocytic processes. In the latter, analysis of frequency distribution revealed no evidence of a population of mitochondria lacking glutamate dehydrogenase, but suggested the presence of populations with different levels of immunoreactivity. Comparison with the labelling of embedded bovine liver glutamate dehydrogenase indicated that the enzyme constitutes a high proportion (10%) of the total matrix protein of these mitochondria. A weaker but significant labelling was found in oligodendrocytes of the white matter. The labelling of mitochondria in neuronal elements including glutamatergic mossy fibre terminals was of the order of 15% of that in astroglial mitochondria. No difference was detected between glutamatergic neurons (mossy and parallel fibres, granular cells) and non-glutamatergic neurons (Purkinje cells). The particle density over non-mitochondrial areas was very close to background over empty resin.

The results, obtained with different methods of tissue and antibody preparation, agree to show that the present form of glutamate dehydrogenase is restricted to mitochondria and preferentially localized in astrocytes.

References (48)

  • RotheF. et al.

    Immunohistochemical demonstration of glutamate dehydrogenase in the postnatally developing rat hippocampal formation and cerebellar cortex: comparison to activity staining

    Neuroscience

    (1990)
  • SchünzelG. et al.

    Topographic and quantitative characteristics of glutamate dehydrogenase of the hippocampus formation during the postnatal development of the rat brain

    Acta Histochem.

    (1982)
  • ShankR.P. et al.

    Present status and significance of the glutamine cycle in neural tissue

    Life Sci.

    (1981)
  • SomogyiP. et al.

    Quantification of immunogold labelling reveals enrichment of glutamate in mossy and parallel fibre terminals in cat cerebellum

    Neuroscience

    (1986)
  • SrereP.A.

    The infrastructure of the mitochondrial matrix

    Trends biochem. Sci.

    (1980)
  • Storm-MathisenJ. et al.

    Ultrastructural immunocytochemical observation on the localization, metabolism and transport of glutamate in normal and ischemic brain tissue

    Prog. Brain Res.

    (1992)
  • WolfG. et al.

    Glutamate dehydrogenase in aminoacidergic structures of the postnatally developing rat cerebellum

    Neurosci. Lett.

    (1987)
  • AokiC. et al.

    Glial glutamate dehydrogenase: ultrastructural localization and regional distribution in relation to the mitochondrial enzyme, cytochrome oxidase

    J. Neurosci. Res.

    (1987)
  • AokiC. et al.

    Regional distribution of astrocytes with intense immunoreactivity for glutamate dehydrogenase in rat brain: implications for neuron-glia interactions in glutamate transmission

    J. Neurosci.

    (1987)
  • BannerC. et al.

    Isolation of a human brain cDNA for glutamate dehydrogenase

    J. Neurochem.

    (1987)
  • BroszM.

    Estimating the number of normal distributed components and determining the initial values for the parameters of a mixture distribution

  • BuschH. et al.

    The Nucleolus

  • ColonA.D. et al.

    Purification and characterization of a soluble and a paniculate glutamate dehydrogenase from rat brain

    J. Neurochem.

    (1986)
  • DennisS.C. et al.

    The pathway of glutamate metabolism in rat brain mitochondria

    Biochem. J.

    (1977)
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