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Expression of glycine receptor subunits and gephyrin in single bipolar cells of the rat retina

Published online by Cambridge University Press:  02 June 2009

Ralf Enz
Affiliation:
Max-Planck-Institut für Hirnforschung, D-60528 Frankfurt, Germany
Joachim Bormann
Affiliation:
Max-Planck-Institut für Hirnforschung, D-60528 Frankfurt, Germany

Abstract

We studied the expression of glycine receptor (GlyR) subunits and gephyrin in the adult rat retina. Reverse transcribed RNA was amplified by polymerase chain reaction (RT-PCR) with primers designed to recognize GlyR α1, α2, α3, β subunits, and gephyrin. Using RNA isolated from the whole retina, signals for all four GlyR subunits and gephyrin could be observed. In rod bipolar cells, in contrast, we detected a subset of GlyR subunits, α1 and β, and no gephyrin. Patch-clamp recording employing two subtype-specific blockers of the GlyR, picrotoxinin and cyanotriphenylborate (CTB), indicated that the GlyR in rod bipolar cells is a heteromeric protein composed of the α1 and β subunit. Moreover, the absence of detectable amounts of gephyrin mRNA suggests that the anchor protein is not required for the function of GlyRs in rod bipolar cells.

Type
Research Articles
Copyright
Copyright © Cambridge University Press 1995

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References

Akaike, N. & Kaneda, M. (1989). Glycine-gated chloride currents in acutely isolated rat hypothalamic neurons. Journal of Neurophysiology 62, 14001409.CrossRefGoogle ScholarPubMed
Betz, H. (1992). Structure and function of inhibitory glycine receptors. Quarterly Reviews of Biophysics 25, 381394.Google Scholar
Bormann, J. (1992). U-tube drug application. In Practical Electrophysiological Methods, ed. Kettenmann, H. & Grantyn, R., pp. 136140. New York: Wiley-Liss.Google Scholar
Bormann, J., Hamill, O.P. & Sakmann, B. (1987). Mechanism of anion permeation through channels gated by glycine and γ-aminobutyric acid in mouse cultured spinal neurones. Journal of Physiology (London) 385, 243286.CrossRefGoogle ScholarPubMed
Bormann, J., Rundström, N., Betz, H. & Langosch, D. (1993). Residues within transmembrane segment M2 determine chloride conductance of glycine receptor homo- and hetero-oligomers. EMBO Journal 12, 37293737.CrossRefGoogle ScholarPubMed
Chomczynski, P. & Sacchi, N. (1987). Single-step method of RNA isolation by guanidinium thiocyanate-phenol-chloroform extraction. Analytical Biochemistry 162, 156159.Google Scholar
Greferath, U., Grünert, U. & Wässle, H. (1990). Rod bipolar cells in the mammalian retina show protein kinase C-like immunoreactivity. Journal of Comparative Neurology 301, 433442.CrossRefGoogle ScholarPubMed
Greferath, U., Brandstätter, J.H., Wässle, H., Kirsch, J., Kuhse, J. & Grünert, U. (1994). Differential expression of glycine receptor subunits in the retina of the rat: A study using immunohistochemistry and in situ hybridization. Visual Neuroscience 11, 721729.CrossRefGoogle Scholar
Grenningloh, G., Rienitz, A., Schmitt, B., Methfessel, C., Zensen, M., Beyrether, K., Gundelfinger, E.D. & Betz, H. (1987). The strychnine-binding subunit of the glycine receptor shows homology with nicotinic acetylcholine receptors. Nature 328, 215220.Google Scholar
Grenningloh, G., Pribilla, I., Prior, P., Multhaup, G., Beyrether, K., Taleb, O. & Betz, H. (1990). Cloning and expression of the 58 kd β subunit of the inhibitory glycine receptor. Neuron 4, 963970.CrossRefGoogle ScholarPubMed
Grünert, U. & Wässle, H. (1993). Immunocytochemical localization of glycine receptors in the mammalian retina. Journal of Comparative Neurology 335, 523537.CrossRefGoogle ScholarPubMed
Hamill, O.P., Marty, A., Neher, E., Sakmann, B. & Sigworth, F.J. (1981). Improved patch-clamp techniques for high resolution current recording from cells and cell-free membrane patches. Pflügers Archiv 391, 85100.CrossRefGoogle ScholarPubMed
Huba, R. & Hofmann, H.D. (1988). Tetanus toxin binding to isolated and cultured rat retinal glial cells. Glia 1, 156164.Google Scholar
Karschin, A. & Wässle, H. (1990). Voltage- and transmitter-gated currents in isolated rod bipolar cells of rat retina. Journal of Neurophysiology 63, 860876.Google Scholar
Kuhse, J., Schmieden, V. & Betz, H. (1990). Identification and functional expression of a novel binding subunit of the inhibitory glycine receptor. Journal of Biological Chemistry 265, 2231722320.CrossRefGoogle ScholarPubMed
Kuhse, J., Kuryatov, A., Maulet, Y., Malosio, M.L., Schmieden, V. & Betz, H. (1991). Alternative splicing generates two isoforms of the α2 subunit of the inhibitory glycine receptor. FEBS Letters 283, 7377.CrossRefGoogle Scholar
Langosch, D., Thomas, L. & Betz, H. (1988). Conserved quaternary structure of ligand gated ion channels: The postsynaptic glycine receptor is a pentamer. Proceedings of the National Academy of Sciences of the U.S.A. 85, 73947398.Google Scholar
Malosio, M., Pouey, M., Kuhse, J. & Betz, H. (1991). Widespread expression of glycine receptor subunit mRNAs in the adult and developing rat brain. EMBO Journal 10, 24012409.Google Scholar
Pfeiffer, F., Graham, D. & Betz, H. (1982). Purification by affinity chromatography of the glycine receptor of rat spinal cord. Journal of Biological Chemistry 257, 93899393.Google Scholar
Pribilla, I., Takagi, T., Langosch, D., Bormann, J. & Betz, H. (1992). The atypical M2 segment of the β subunit confers picrotoxinin resistance to inhibitory glycine receptor channels. EMBO Journal 11, 43054311.Google Scholar
Prior, P., Schmitt, B., Grenningloh, G., Pribilla, I., Multhaup, G., Beyrether, K., Maulet, Y., Werner, P., Langosch, D., Kirsch, A. & Betz, H. (1992). Primary structure and alternative splice variants of gephyrin, a putative glycine receptor-tubulin linker protein. Neuron 8, 11611170.Google Scholar
Rundström, N., Schmieden, V., Betz, H., Bormann, J. & Langosch, D. (1994). Cyanotriphenylborate: Subtype-specific blocker of glycine receptor chloride channels. Proceedings of the National Academy of Sciences of the U.S.A. 91, 89508954.Google Scholar
Sassoè-Pognetto, M., Wässle, H. & Grünert, U. (1994). Glycinergic synapses in the rod pathway of the rat retina: Cone bipolar cells express the α1 subunit of the glycine receptor. Journal of Neuroscience 14, 51315146.CrossRefGoogle Scholar
Schmieden, V., Grenningloh, G., Schofield, P.R. & Betz, H. (1989). Functional expression in Xenopus oocytes of the strychnine binding 48 kd subunit of the glycine receptor. EMBO Journal 8, 695700.Google Scholar
Shirasaki, T., Klee, M.R., Nakaye, T. & Akaike, N. (1991). Differential blockade of bicuculline and strychnine on GABA- and glycineinduced responses in dissociated rat hippocampal pyramidal cells. Brain Research 561, 7783.CrossRefGoogle ScholarPubMed
Suzuki, S., Tachibana, M. & Kaneko, A. (1990). Effects of glycine and GABA on isolated bipolar cells of the mouse retina. Journal of Physiology (London) 421, 645662.Google Scholar
Takahashi, T., Momiyama, A., Hirai, K., Hishinuma, F. & Akagi, A. (1992). Functional correlation of fetal and adult forms of glycine receptors with developmental changes in inhibitory synaptic receptor channels. Neuron 9, 11551161.Google Scholar
Udaykumar Epstein, J.S. & Hewlett, I.K. (1993). A novel method employing UNG to avoid carryover contamination in RNA-PCR. Nucleic Acids Research 21, 39173918.Google Scholar
Wässle, H., Yamashita, M., Greferath, U., Grünert, U. & Müller, F. (1991). The rod bipolar cell of the mammalian retina. Visual Neuroscience 7, 99112.Google Scholar