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Immunocytochemical localization of glycine and glycine receptors in the retina of the frog Rana ridibunda

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Abstract

Glycine and glycine receptors (GlyRs) were analyzed immunocytochemically in the retina of the frog Rana ridibunda. Glycine was localized to somata of glycinergic amacrine and interplexiform cells. Approximately 50% of the cells in the amacrine cell layer were found to be glycinergic. GlyRs of the inner plexiform layer (IPL) were localized to brightly fluorescent puncta, probably representing postsynaptic clusters of GlyRs. GlyR clusters were not evenly distributed across the IPL but showed patterns of stratification specific for the various GlyR subunits. Clusters containing the α1 subunit formed four narrow strata within the IPL. Clusters containing the α3 subunit were more abundant and covered the whole IPL, with a band of higher density in stratum 3. Clusters of GlyRs were also observed in the outer plexiform layer. Thus, several isoforms of synaptic GlyRs involved with different synapses and inhibitory circuits are present in the frog retina.

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References

  1. Ammermüller J, Kolb H (1996) Functional architecture of the turtle retina. Prog Retin Eye Res 15:393–433

  2. Brandstätter J, Koulen P, Wässle H (1998) Diversity of glutamate receptors in the mammalian retina. Vision Res 38:1385–1397

  3. Büttner C, Sadtler S, Leyendecker A, Laube B, Griffon N, Betz H, Schmalzing G (2001) Ubiquitination precedes internalization and proteolytic cleavage of plasma membrane-bound glycine receptors. J Biol Chem 276:42978–42985

  4. Cajal SR (1893) La rétine des vertébrés. Cellule 9:119–257

  5. Du J, Yang XL (2002) Bullfrog retinal bipolar cells may express heterogeneous glycine receptors at dendrites and axon terminals. Neurosci Lett 322:505–518

  6. Essrich C, Lorez M, Benson JA, Fritschy JM, Luscher B (1998) Postsynaptic clustering of major GABAA receptor subtypes requires the γ2 subunit and gephyrin. Nat Neurosci 1:563–571

  7. Feng G, Tintrup H, Kirsch J, Nichol MC, Kuhse J, Betz H, Sanes JR (1998) Dual requirement for gephyrin in glycine receptor clustering and molybdoenzyme activity. Science 282:1321–1324

  8. Fischer F, Kneussel M, Tintrup H, Haverkamp S, Rauen T, Betz H, Wässle H (2000) Reduced synaptic clustering of GABA and glycine receptors in the retina of the gephyrin null mutant mouse. J Comp Neurol 427:634–648

  9. Fritschy JM, Weinmann O, Wenzel A, Benke D (1998) Synapse-specific localization of NMDA and GABAA receptor subunits revealed by antigen-retrieval immunohistochemistry. J Comp Neurol 390:194–210

  10. Graham D, Pfeiffer F, Simler R, Betz H (1985) Purification and characterization of the glycine receptor of pig spinal cord. Biochemistry 24:990–994

  11. Greferath U, Brandstätter JH, Wässle H, Kirsch H, 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. Vis Neurosci 11:721–729

  12. Grenningloh G, Rienitz A, Schmitt B, Methfessel C, Zensen M, Beyreuther K, Gundelfinger ED, Betz H (1987) The strychnine-binding subunit of the glycine receptor shows homology with nicotinic acetylcholine receptors. Nature 328:215–220

  13. Grenningloh G, Schmieden V, Schofield PR, Seeburg PH, Siddique T, Mohandas TK, Becker C-M, Betz H (1990) Alpha subunit variants of the human glycine receptor: primary structures, functional expression and chromosomal localisation of the corresponding genes. EMBO J 9:771–776

  14. Han Y, Slaughter M (1998) Protein kinases modulate two glycine currents in salamander retinal ganglion cells. J Physiol (Lond) 508:681–690

  15. Han Y, Wu W (1999) Modulation of glycine receptors in retinal ganglion cells by zinc. Proc Natl Acad Sci USA 96:3234–3238

  16. Han Y, Zhang J, Slaughter M (1997) Partition of transient and sustained inhibitory glycinergic input to retinal ganglion cells. J Neurosci 17:3392–3400

  17. Harvey RJ, Betz H (2000) Structure, diversity, pharmacology, and pathology of glycine receptor chloride channels. In: Endo M, Kurachi Y, Mishina M (eds) Pharmacology of ionic channel function: activators and inhibitors. Springer, Berlin Heidelberg New York, pp 479–497

  18. Harvey RJ, Schmieden V, Holst A von, Laube B, Rohrer H, Betz H (2000) Glycine receptors containing the α4 subunit in the embryonic sympathetic nervous system, spinal cord and male genital ridge. Eur J Neurosci 12:994–1001

  19. Haverkamp S, Wässle H (2000) Immunocytochemical analysis of the mouse retina. J Comp Neurol 424:1–23

  20. Haverkamp S, Müller U, Harvey K, Harvey RJ, Betz H, Wässle H (2003) Diversity of glycine receptors in the mouse retina: localization of the α3 subunit. J Comp Neurol 465:524–539

  21. Kalloniatis M, Tomisich G (1999) Amino acid neurochemistry of the vertebrate retina. Prog Retin Eye Res 18:811–866

  22. Kneussel M, Brandstätter JH, Laube B, Stahl S, Müller U, Betz H (1999) Loss of postsynaptic GABAA receptor clustering in gephyrin-deficient mice. J Neurosci 19:9289–9297

  23. Kuhse J, Schmieden V, Betz H (1990) Identification and functional expression of a novel ligand binding subunit of the inhibitory glycine receptor. J Biol Chem 265:22317–22320

  24. Kupenova P, Vitanova L, Popova E, Mitova L (1997) Influence of picrotoxin and strychnine on the spectral sensitivity of the turtle ERG b- and d-wave. I. Dark adaptation. Acta Physiol Scand 159:217–225

  25. Laube B, Maksay G, Schemm R, Betz H (2002) Modulation of glycine receptor function: a novel approach for therapeutic intervention at inhibitory synapses? Trends Pharmacol Sci 23:519–527

  26. Legendre P (2001) The glycinergic inhibitory synapse. Cell Mol Life Sci 58:760–793

  27. MacNeil MA, Masland RH (1998) Extreme diversity among amacrine cells: implications for function. Neuron 20:971–982

  28. Maple BR, Wu SM (1998) Glycinergic synaptic inputs to bipolar cells in the salamander retina. J Physiol (Lond) 506:731–744

  29. Marc RE (1985) The role of glycine in retinal circuitry. In: Morgan WW (ed) Retinal transmitters and modulators: models for the brain. CRC, Boca Raton, pp 119–158

  30. Marc RE (1989) The role of glycine in the mammalian retina. Prog Retin Eye Res 8:67–107

  31. Marc RE, Lam DMK (1981) Glycinergic pathways in the goldfish retina. J Neurosci 1:152–165

  32. Matzenbach B, Maulet Y, Sefton L, Courtier B, Avner P, Guénet J-L, Betz H (1994) Structural analysis of mouse glycine receptor α subunits genes: identification and chromosomal localization of a novel variant, α4. J Biol Chem 269:2607–2612

  33. Menger N, Pow DV, Wässle H (1998) Glycinergic amacrine cells of the rat retina. J Comp Neurol 401:34–46

  34. Pfeiffer F, Simler R, Grenningloh G, Betz H (1984) Monoclonal antibodies and peptides mapping reveal structural similarities between the subunits of the glycine receptor of rat spinal cord. Proc Natl Acad Sci USA 81:7224–7227

  35. Popova E, Motova L, Vitanova L, Kupenova P (1997) Effect of glycinergic blockade on light responses of frog retinal ganglion cells. Comp Biochem Physiol 116:255–263

  36. Popova E, Mitova L, Vitanova L, Kupenova P (2000) Effect of 2-amino-4-phosphonobutyrate on the OFF-responses of frog retinal ganglion cells and local ERG after glycinergic blockade. Comp Biochem Physiol 126:139–151

  37. Pourcho RG (1996) Neurotransmitters in the retina. Curr Eye Res 15:797–803

  38. Pourcho RG, Goebel DJ (1985) A combined Golgi and autoradiographic study of (3H)-glycine-accumulating amacrine cells in the cat retina. J Comp Neurol 233:473–480

  39. Pourcho RG, Owczarzak MT (1991a) Connectivity of glycine immunoreactive amacrine cells in the cat retina. J Comp Neurol 307:549–561

  40. Pourcho RG, Owczarzak MT (1991b) Glycine receptor immunoreactivity is localized at amacrine synapses in cat retina. Vis Neurosci 7:611–618

  41. Pow DV (2001) Amino acids and their transporters in the retina. Neurochem Int 38:463–484

  42. Pow DV, Wright LL, Vaney DI (1995) Immunocytochemical detection of amino acid neurotransmitters in paraformaldehyde-fixed tissues. J Neurosci Methods 56:115–123

  43. Sassoè-Pognetto M, Wässle H (1997) Synaptogenesis in the rat retina: subcellular localization of glycine receptors, GABAA receptors, and the anchoring protein gephyrin. J Comp Neurol 381:158–174

  44. 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. J Neurosci 14:5131–5146

  45. Sassoè-Pognetto M, Kirsch J, Grünert U, Greferath U, Fritschy J-M, Möhler H, Betz H, Wässle H (1995) Colocalization of gephyrin and GABAA-receptor subunits in the rat retina. J Comp Neurol 357:1–14

  46. Schmitt B, Knaus P, Becker C-M, Betz H (1987) The M 93000 polypeptide of the postsynaptic glycine receptor complex is a peripheral membrane protein. Biochemistry 26:805–811

  47. Schröder S, Hoch W, Becker C-M, Grenningloh G, Betz H (1991) Mapping of antigenic epitopes on the α1 subunit of the inhibitory glycine receptor. Biochemistry 30:42–47

  48. Smiley JF, Basinger SF (1989) Glycine high-affinity uptake labels a subpopulation of somatostatin-like immunoreactive cells in the Rana pipiens retina. Brain Res 495:31–44

  49. Smiley JF, Yazulla S (1990) Glycinergic contacts in the outer plexiform layer of the Xenopus laevis retina characterized by antibodies to glycine, GABA and glycine receptors. J Comp Neurol 299:375–388

  50. Vaney DI (2002) Retinal neurons: cell types and coupled networks. Perspect Anal Philos 136:239–254

  51. Vannier C, Triller A (1997) Biology of the postsynaptic glycine receptor. Int Rev Cytol 176:201–244

  52. Vigh J, Banvölgyi T, Wilhelm M (2000) Amacrine cells of the anuran retina: morphology, chemical neuroanatomy, and physiology. Microsc Res Tech 50:373–383

  53. Vitanova L, Kupenova P, Popova E, Mitova L (1997) Influence of picrotoxin and strychnine on the spectral sensitivity of the turtle ERG b- and d-wave. II. Light adaptation. Acta Physiol Scand 159:227–235

  54. Wagner HJ, Wagner E (1988) Amacrine cells in the retina of a teleost fish, the roach (Rutilus rutilus): a Golgi study on differentiation and layering. Philos Trans R Soc Lond Biol 321:263–324

  55. Wässle H, Koulen P, Brandstätter JH, Fletcher EL, Becker C-M (1998) Glycine and GABA receptors in the mammalian retina. Vision Res 38:1411–1430

  56. Yang C-Y, Yazulla S (1988) Light microscopic localization of putative glycinergic neurons in the larval tiger salamander retina by immunocytochemical and autoradiographical methods. J Comp Neurol 272:343–357

  57. Yang C-Y, Lukasiewicz P, Maguire G, Werblin FS, Yazulla S (1991) Amacrine cells in the tiger salamander retina: morphology, physiology, and neurotransmitter identification. J Comp Neurol 312:19–32

  58. Yazulla S, Studholme KM (1990) Multiple subtypes of glycine-immunoreactive neurons in the goldfish retina: single- and double-label studies. Vis Neurosci 4:299–309

  59. Yazulla S, Studholme KM (1991a) Glycinergic interplexiform cells make synaptic contact with amacrine cell bodies in goldfish retina. J Comp Neurol 310:1–10

  60. Yazulla S, Studholme KM (1991b) Glycine-receptor immunoreactivity in retinal bipolar cells is postsynaptic to glycinergic and GABAergic amacrine cell synapses. J Comp Neurol 310:11–20

  61. Yazulla S, Studholme KM (2001) Neurochemical anatomy of the zebrafish retina as determined by immunocytochemistry. J Neurocytol 30:551–592

  62. Zucker CL, Ehinger B (1993) Synaptic connections involving immunoreactive glycine receptors in the turtle retina. Vis Neurosci 10:907–914

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Acknowledgments

We are grateful to F. Boij for technical assistance, to A. Staab for performing the Western blots, and to I. Odenthal for typing the manuscript.

Author information

Correspondence to Heinz Wässle.

Additional information

This work was supported by the Deutsche Forschungsgemeinschaft SFB269/B4

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Vitanova, L., Haverkamp, S. & Wässle, H. Immunocytochemical localization of glycine and glycine receptors in the retina of the frog Rana ridibunda . Cell Tissue Res 317, 227–235 (2004). https://doi.org/10.1007/s00441-004-0914-6

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Keywords

  • Retina
  • Glycine
  • Glycine receptors
  • Gephyrin
  • Frog Rana ridibunda (Anura)