Abstract
In the internal granular layer of the cerebellar cortex the polysynaptic complexes called glomeruli consist mainly of homogeneous populations of glutamatergic and GABAergic synapses, both located on granule cell dendrites. A subcellular fraction enriched in glomeruli was prepared from rat cerebellum, and the distribution of GABAA and of benzodiazepine binding sites between membranes derived from this fraction (fraction G) and from a total cerebellar homogenate (fraction T) was studied. The benzodiazepine and GABA binding sites were measured by the binding of agonists [3H]flunitrazepam and [3H]muscimol, respectively. The results indicate that both binding sites are present, but only slightly enriched, in the glomerular synapses. We found a muscimol/flunitrazepam binding site ratio of two, which is consistent with the enrichement of muscimol binding sites in the granular layer shown by both autoradiographic with radioactive glutamatergic ligands and in situ hybridization experiments respectively.
Similar content being viewed by others
References
Viennot, F, Artault, J. C., Tholey, G., and Gombos, G. 1991a. An improved method for the preparation of rat cerebellar glomeruli. J. Neurosci. Meth. 38:51–62.
Viennot, F., de Barry, J., and Gombos G. 1991b. Non-NMDA excitatory amino acid receptors in a subcellular fraction enriched in cerebellar glomeruli. Neurochem. Res. 16:435–442.
Triller, A., Cluzeaud, F. and Korn, H. 1987. Gamma-aminobutyric acid-containing terminals can be apposed to glycine receptors at central synapses. J. Cell Biol. 104:947–956.
Bacon, E., and Viennot, F. 1990. Le système complexe des récepteurs GABA/benzodiazépine. M/S Med. Sci. 6:770–777.
Casalotti, S. O., Stephenson, F. A., and Barnard, E. A. 1986. Separate subunits for agonist and benzodiazepine binding in the γ-aminobytyric acidA receptor oligomer. J. Biol. Chem. 261:15013–15016.
Olsen, R. W. 1981. GABA-benzodiazepine-barbiturate receptor interactions. J. Neurochem. 37:1–13.
Shivers, B. D., Killisch, I., Sprengel, R., Sontheimer, H., Köhler, M., Schofield, P. R., and Seeburg, P. H. 1989. Two novel GABAA receptor subunits exist in distinct neuronal subpopulations. Neuron 3:327–337.
Wang, Y. J., Salvaterra, P., and Roberts, E. 1979. Characterization of [3H]muscimol binding to mouse brain membranes. Biochem. Pharmacol. 28:1123–1128.
Frostholm, A., and Rotter, A. 1987. The ontogeny of [3H]muscimol binding sites in the C57BL/6J mouse cerebellum. Dev. Brain Res. 37:157–166.
Matsumoto, R. R. 1989. GABA receptors: are cellular differences reflected in function? Brain Res. Rev. 14:203–225.
Olsen, R. W., McCabe, R. T., and Wamsley, J. K. 1990. GABAA receptor subtypes: autoradiographic comparison of GABA, benzodiazepine, and convulsant binding sites in the rat central nervous system. J. Chem. Neuroanat. 3:59–76.
Wisden, W., McNaughton, L. A., Darlison, M. G., Hunt, S. P., and Barnard, E. A. 1989. Differential distribution of GABAA receptor mRNAs in bovine cerebellum-localization of α2 mRNA in Bergmann glia layer. Neurosci. Lett. 106:7–12.
Molinoff, P. B., and Kravitz, E. A. 1968. The metabolism of γ-aminobutyric acid (GABA) in the Lobster nervous system-glutamic decarboxylase. J. Neurochem. 15:391–409.
Lowry, O. H., Rosebrough, N. J., Farr, A. L., and Randall, R. J. 1951. Protein measurement with the Folin phenol reagent. J. Biol. Chem. 193:265–275.
Syapin, P. J., and Rickman, D. W. 1981. Benzodiazepine receptor increase following repeated pentylenetetrazole injections. Eur. J. Pharmacol. 72:117–120.
Möhler, H., and Okada, T. 1977. Benzodiazepine receptors: demonstration in the central nervous system. Science 198:849–851.
Beaumont, K., Chilton, W. S., Yamamura, H. I., and Enna, S. J. 1978. Muscimol binding in rat brain: association with synaptic GABA receptors. Brain Res. 148:153–162.
Somogyi, P., Hodgson, A. J., Chubb, I. W., Penke, B., and Erdei, A. 1985. Antisera to gamma-aminobutyric acid. II. Immunocytochemical application to the central nervous system. J. Histochem. Cytochem. 33:240–248.
Sieghart, W., Eichinger, A., Richards, J. G., and Möhler, H. 1987. Photoaffinity labeling of benodiazepine receptor proteins with the partial inverse agonist [3H]RO 15-4513: a biochemical and autoradiographic study. J. Neurochem. 48:46–52.
Lüddens, H., Pritchett, D. B., Köhler, M., Killisch, I., Keinänen, K., Monyer, H., Sprengel, R., and Seeburg, P. H. 1990. Cerebellar GABAA receptor selective for a behavioural alcohol antagonist. Nature 346:648–651.
Sequier, J. M., Richards, J. G., Malherbe, P., Price, G. W., Mathews, S., and Möhler, H. 1988. Mapping of brain areas containing RNA homologous to cDNAs encoding the α and β subunits of the rat GABAA gamma-aminobutyrate receptor. Proc. Nat. Acad. Sci. USA 85:7815–7819.
Rotter, A., Gorenstein, C., and Frostholm, A. 1988. The localization of GABAA receptors in mice with mutations affecting the structure and connectivity of the cerebellum. Brain Res. 439:236–248.
Rotter, A., and Frostholm, A. 1988. Cerebellar benzodiazepine receptors: cellular localization and consequences of neurological mutations in mice. Brain Res. 444:133–146.
Zhang, J. H., Sato, M., and Tohyama, M. J. 1991. Region-specific expression of the mRNAs encoding β subunits (β1, β2 and β3) of GABAA receptor in the rat brain. J. Comp. Neurol. 303:637–657.
Geary II, W. A., Toga, A. W., and Wooten, G. F. 1985. Quantitative film autoradiography for tritium: methodological considerations. Brain Res. 337:99–108.
Somogyi, P., Takagi, H., and Richards, J. G. 1989. Subcellular localization of benzodiazepine/GABAA receptors in the cerebellum of rat, cat, and monkey using monoclonal antibodies. J. Neurosci. 9:2197–2209.
Palacios, J. M., Wamsley, J. K., and Kuhar, M. J. 1981. High affinity GABA receptors-autoradiographic localization. Brain Res. 222:285–307.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Viennot, F., Foucaud, B. & Gombos, G. Musciomol and flunitrazepam binding sites in a subcellular fraction enriched in rat cerebellar glomeruli. Neurochem Res 17, 683–686 (1992). https://doi.org/10.1007/BF00968005
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00968005