Anatomical Science International

, Volume 86, Issue 1, pp 10–18 | Cite as

Development of an anatomical technique for visualizing the mode of climbing fiber innervation in Purkinje cells and its application to mutant mice lacking GluRδ2 and Cav2.1

Review Article

Abstract

In the adult cerebellum, a single climbing fiber (CF) innervates proximal dendrites of Purkinje cells (PCs). This monoinnervation is established by the developmental elimination of surplus CFs through homosynaptic competition among multiply innervating CFs and heterosynaptic competition between CFs and parallel fibers, i.e., granule cell axons innervating distal PC dendrites. Although the developmental process of CF monoinnervation and defects in it in mutant and experimental animal models have been extensively studied by electrophysiological techniques, for quite some time this subject was poorly understood from a morphological perspective due to a lack of neuroanatomical methods that could distinguish CFs with different neuronal origins. Soon after the identification of type 2 vesicular glutamate transporter (VGluT2) that selectively detects CF terminals in the molecular layer, we developed a novel method of combined anterograde tracer labeling and VGluT2 immunohistochemistry. This method enables us to identify the mode (mono vs. multiple) of CF innervation and the site of multiple innervation. Since then, we have applied this method to various kinds of gene-manipulated mice manifesting ataxia and other cerebellar phenotypes. In this review, we summarize experimental procedures for the combined tracer/VGluT2 labeling method, and then introduce what we have learned by applying this method in studies on the role of GluRδ2 and Cav2.1 in CF monoinnervation. This method has provided informative anatomical correlates to electrophysiological data and vice versa, and will extend our knowledge of the molecular and cellular mechanisms for the development, plasticity, degeneration, and repair of the CF–PC projection system.

Keywords

Purkinje cell Climbing fiber Cerebellum Anterograde tracer labeling Type 2 vesicular glutamate transporter 

References

  1. Aihara Y, Mashima H, Onda H, Hisano S, Kasuya H, Hori T, Yamada S, Tomura H, Yamada Y, Inoue I, Kojima I, Takeda J (2000) Molecular cloning of a novel brain-type Na(+)-dependent inorganic phosphate cotransporter. J Neurochem 74:2622–2625CrossRefPubMedGoogle Scholar
  2. Araki K, Meguro H, Kushiya E, Takayama C, Inoue Y, Mishina M (1993) Selective expression of the glutamate receptor channel delta 2 subunit in cerebellar Purkinje cells. Biochem Biophys Res Commun 197:1267–1276CrossRefPubMedGoogle Scholar
  3. Bravin M, Rossi F, Strata P (1995) Different climbing fibres innervate separate dendritic regions of the same Purkinje cell in hypogranular cerebellum. J Comp Neurol 357:395–407CrossRefPubMedGoogle Scholar
  4. Bravin M, Morando L, Vercelli A, Rossi F, Strata P (1999) Control of spine formation by electrical activity in the adult rat cerebellum. Proc Natl Acad Sci USA 96:1704–1709CrossRefPubMedGoogle Scholar
  5. Cesa R, Morando L, Strata P (2003) Glutamate receptor delta2 subunit in activity-dependent heterologous synaptic competition. J Neurosci 23:2363–2370PubMedGoogle Scholar
  6. Cesa R, Scelfo B, Strata P (2007) Activity-dependent presynaptic and postsynaptic structural plasticity in the mature cerebellum. J Neurosci 27:4603–4611CrossRefPubMedGoogle Scholar
  7. Crepel F, Delhaye-Bouchaud N, Dupont JL (1981) Fate of the multiple innervation of cerebellar Purkinje cells by climbing fibers in immature control, X-irradiated and hypothyroid rats. Brain Res 227:59–71Google Scholar
  8. Fremeau RT Jr, Troyer MD, Pahner I, Nygaard GO, Tran CH, Reimer RJ, Bellocchio EE, Fortin D, Storm-Mathisen J, Edwards RH (2001) The expression of vesicular glutamate transporters defines two classes of excitatory synapse. Neuron 31:247–260CrossRefPubMedGoogle Scholar
  9. Guastavino JM, Sotelo C, Damez-Kinselle I (1990) Hot-foot murine mutation: behavioral effects and neuroanatomical alterations. Brain Res 523:199–210CrossRefPubMedGoogle Scholar
  10. Hashimoto K, Kano M (2003) Functional differentiation of multiple climbing fiber inputs during synapse elimination in the developing cerebellum. Neuron 38:785–796CrossRefPubMedGoogle Scholar
  11. Hashimoto K, Ichikawa R, Takechi H, Inoue Y, Aiba A, Sakimura K, Mishina M, Hashikawa T, Konnerth A, Watanabe M, Kano M (2001) Roles of glutamate receptor delta 2 subunit (GluRdelta 2) and metabotropic glutamate receptor subtype 1 (mGluR1) in climbing fiber synapse elimination during postnatal cerebellar development. J Neurosci 21:9701–9712PubMedGoogle Scholar
  12. Hashimoto K, Ichikawa R, Kitamura K, Watanabe M, Kano M (2009) Translocation of a “winner” climbing fiber to the Purkinje cell dendrite and subsequent elimination of “losers” from the soma in developing cerebellum. Neuron 63:106–118CrossRefPubMedGoogle Scholar
  13. Herzog E, Bellenchi GC, Gras C, Bernard V, Ravassard P, Bedet C, Gasnier B, Giros B, El Mestikawy S (2001) The existence of a second vesicular glutamate transporter specifies subpopulations of glutamatergic neurons. J Neurosci 21:RC181PubMedGoogle Scholar
  14. Hirai H, Miyazaki T, Kakegawa W, Matsuda S, Mishina M, Watanabe M, Yuzaki M (2005a) Rescue of abnormal phenotypes of the delta2 glutamate receptor-null mice by mutant delta2 transgenes. EMBO Rep 6:90–95CrossRefPubMedGoogle Scholar
  15. Hirai H, Pang Z, Bao D, Miyazaki T, Li L, Miura E, Parris J, Rong Y, Watanabe M, Yuzaki M, Morgan JI (2005b) Cbln1 is essential for synaptic integrity and plasticity in the cerebellum. Nat Neurosci 8:1534–1541CrossRefPubMedGoogle Scholar
  16. Ichikawa R, Miyazaki T, Kano M, Hashikawa T, Tatsumi H, Sakimura K, Mishina M, Inoue Y, Watanabe M (2002) Distal extension of climbing fiber territory and multiple innervation caused by aberrant wiring to adjacent spiny branchlets in cerebellar Purkinje cells lacking glutamate receptor delta 2. J Neurosci 22:8487–8503PubMedGoogle Scholar
  17. Ikeda A, Miyazaki T, Kakizawa S, Okuno Y, Tsuchiya S, Myomoto A, Saito SY, Yamamoto T, Yamazaki T, Iino M, Tsujimoto G, Watanabe M, Takeshima H (2007) Abnormal features in mutant cerebellar Purkinje cells lacking junctophilins. Biochem Biophys Res Commun 363:835–839CrossRefPubMedGoogle Scholar
  18. Kakegawa W, Miyazaki T, Hirai H, Motohashi J, Mishina M, Watanabe M, Yuzaki M (2007a) Ca2+ permeability of the channel pore is not essential for the delta2 glutamate receptor to regulate synaptic plasticity and motor coordination. J Physiol 579:729–735CrossRefPubMedGoogle Scholar
  19. Kakegawa W, Kohda K, Yuzaki M (2007b) The delta2 “ionotropic” glutamate receptor functions as a non-ionotropic receptor to control cerebellar synaptic plasticity. J Physiol 584:89–96Google Scholar
  20. Kakizawa S, Miyazaki T, Yanagihara D, Iino M, Watanabe M, Kano M (2005) Maintenance of presynaptic function by AMPA receptor-mediated excitatory postsynaptic activity in adult brain. Proc Natl Acad Sci USA 102:19180–19185CrossRefPubMedGoogle Scholar
  21. Kano M, Rexhausen U, Dreessen J, Konnerth A (1992) Synaptic excitation produces a long-lasting rebound potentiation of inhibitory synaptic signals in cerebellar Purkinje cells. Nature 356:601–604CrossRefPubMedGoogle Scholar
  22. Kano M, Hashimoto K, Chen C, Abeliovich A, Aiba A, Kurihara H, Watanabe M, Inoue Y, Tonegawa S (1995) Impaired synapse elimination during cerebellar development in PKC gamma mutant mice. Cell 83:1223–1231CrossRefPubMedGoogle Scholar
  23. Kano M, Hashimoto K, Kurihara H, Watanabe M, Inoue Y, Aiba A, Tonegawa S (1997) Persistent multiple climbing fiber innervation of cerebellar Purkinje cells in mice lacking mGluR1. Neuron 18:71–79CrossRefPubMedGoogle Scholar
  24. Kano M, Hashimoto K, Watanabe M, Kurihara H, Offermanns S, Jiang H, Wu Y, Jun K, Shin HS, Inoue Y, Simon MI, Wu D (1998) Phospholipase cbeta4 is specifically involved in climbing fiber synapse elimination in the developing cerebellum. Proc Natl Acad Sci USA 95:15724–15729CrossRefPubMedGoogle Scholar
  25. Kashiwabuchi N, Ikeda K, Araki K, Hirano T, Shibuki K, Takayama C, Inoue Y, Kutsuwada T, Yagi T, Kang Y et al (1995) Impairment of motor coordination, Purkinje cell synapse formation, and cerebellar long-term depression in GluR delta 2 mutant mice. Cell 81:245–252CrossRefPubMedGoogle Scholar
  26. Konnerth A, Dreessen J, Augustine GJ (1992) Brief dendritic calcium signals initiate long-lasting synaptic depression in cerebellar Purkinje cells. Proc Natl Acad Sci USA 89:7051–7055CrossRefPubMedGoogle Scholar
  27. Kurihara H, Hashimoto K, Kano M, Takayama C, Sakimura K, Mishina M, Inoue Y, Watanabe M (1997) Impaired parallel fiber → Purkinje cell synapse stabilization during cerebellar development of mutant mice lacking the glutamate receptor delta2 subunit. J Neurosci 17:9613–9623PubMedGoogle Scholar
  28. Lalouette A, Lohof A, Sotelo C, Guenet J, Mariani J (2001) Neurobiological effects of a null mutation depend on genetic context: comparison between two hotfoot alleles of the delta-2 ionotropic glutamate receptor. Neuroscience 105:443–455CrossRefPubMedGoogle Scholar
  29. Landsend AS, Amiry-Moghaddam M, Matsubara A, Bergersen L, Usami S, Wenthold RJ, Ottersen OP (1997) Differential localization of delta glutamate receptors in the rat cerebellum: coexpression with AMPA receptors in parallel fiber-spine synapses and absence from climbing fiber-spine synapses. J Neurosci 17:834–842PubMedGoogle Scholar
  30. Lomeli H, Sprengel R, Laurie DJ, Kohr G, Herb A, Seeburg PH, Wisden W (1993) The rat delta-1 and delta-2 subunits extend the excitatory amino acid receptor family. FEBS Lett 315:318–322CrossRefPubMedGoogle Scholar
  31. Mandolesi G, Cesa R, Autuori E, Strata P (2009) An orphan ionotropic glutamate receptor: the delta2 subunit. Neuroscience 158:67–77CrossRefPubMedGoogle Scholar
  32. Mariani J (1982) Extent of multiple innervation of Purkinje cells by climbing fibers in the olivocerebellar system of weaver, reeler, and staggerer mutant mice. J Neurobiol 13:119–126CrossRefPubMedGoogle Scholar
  33. Matsuda K, Miura E, Miyazaki T, Kakegawa W, Emi K, Narumi S, Fukazawa Y, Ito-Ishida A, Kondo T, Shigemoto R, Watanabe M, Yuzaki M (2010) Cbln1 is a ligand for an orphan glutamate receptor delta2, a bidirectional synapse organizer. Science 328:363–368CrossRefPubMedGoogle Scholar
  34. Mintz IM, Venema VJ, Swiderek KM, Lee TD, Bean BP, Adams ME (1992) P-type calcium channels blocked by the spider toxin omega-Aga-IVA. Nature 355:827–829CrossRefPubMedGoogle Scholar
  35. Mintz IM, Sabatini BL, Regehr WG (1995) Calcium control of transmitter release at a cerebellar synapse. Neuron 15:675–688CrossRefPubMedGoogle Scholar
  36. Miyazaki T, Fukaya M, Shimizu H, Watanabe M (2003) Subtype switching of vesicular glutamate transporters at parallel fibre-Purkinje cell synapses in developing mouse cerebellum. Eur J Neurosci 17:2563–2572CrossRefPubMedGoogle Scholar
  37. Miyazaki T, Hashimoto K, Shin HS, Kano M, Watanabe M (2004) P/Q-type Ca2+ channel alpha1A regulates synaptic competition on developing cerebellar Purkinje cells. J Neurosci 24:1734–1743CrossRefPubMedGoogle Scholar
  38. Miyazaki T, Hashimoto K, Uda A, Sakagami H, Nakamura Y, Saito SY, Nishi M, Kume H, Tohgo A, Kaneko I, Kondo H, Fukunaga K, Kano M, Watanabe M, Takeshima H (2006) Disturbance of cerebellar synaptic maturation in mutant mice lacking BSRPs, a novel brain-specific receptor-like protein family. FEBS Lett 580:4057–4064CrossRefPubMedGoogle Scholar
  39. Miyazaki T, Yamasaki M, Takeuchi T, Sakimura K, Mishina M, Watanabe M (2010) Ablation of glutamate receptor GluRδ2 in adult Purkinje cells causes multiple innervation of climbing fibers by inducing aberrant invasion to parallel fiber innervation territory. J Neurosci 30:15196–15209CrossRefPubMedGoogle Scholar
  40. Morando L, Cesa R, Rasetti R, Harvey R, Strata P (2001) Role of glutamate delta-2 receptors in activity-dependent competition between heterologous afferent fibers. Proc Natl Acad Sci USA 98:9954–9959CrossRefPubMedGoogle Scholar
  41. Nishiyama H, Fukaya M, Watanabe M, Linden DJ (2007) Axonal motility and its modulation by activity are branch-type specific in the intact adult cerebellum. Neuron 56:472–487CrossRefPubMedGoogle Scholar
  42. Offermanns S, Hashimoto K, Watanabe M, Sun W, Kurihara H, Thompson RF, Inoue Y, Kano M, Simon MI (1997) Impaired motor coordination and persistent multiple climbing fiber innervation of cerebellar Purkinje cells in mice lacking Galphaq. Proc Natl Acad Sci USA 94:14089–14094CrossRefPubMedGoogle Scholar
  43. Palay S, Chan-Palay V (1974) Cerebellar cortex: cytology and organization. Springer, New York, pp 63–69 (see also pp 242–287)Google Scholar
  44. Regehr WG, Mintz IM (1994) Participation of multiple calcium channel types in transmission at single climbing fiber to Purkinje cell synapses. Neuron 12:605–613CrossRefPubMedGoogle Scholar
  45. Rossi F, van der Want JJ, Wiklund L, Strata P (1991) Reinnervation of cerebellar Purkinje cells by climbing fibres surviving a subtotal lesion of the inferior olive in the adult rat. II. Synaptic organization on reinnervated Purkinje cells. J Comp Neurol 308:536–554CrossRefPubMedGoogle Scholar
  46. Sotelo C, Hillman DE, Zamora AJ, Llinas R (1975) Climbing fiber deafferentation: its action on Purkinje cell dendritic spines. Brain Res 98:574–581CrossRefPubMedGoogle Scholar
  47. Stea A, Tomlinson WJ, Soong TW, Bourinet E, Dubel SJ, Vincent SR, Snutch TP (1994) Localization and functional properties of a rat brain alpha 1A calcium channel reflect similarities to neuronal Q- and P-type channels. Proc Natl Acad Sci USA 91:10576–10580CrossRefPubMedGoogle Scholar
  48. Sugihara I, Wu H, Shinoda Y (1999) Morphology of single olivocerebellar axons labeled with biotinylated dextran amine in the rat. J Comp Neurol 414:131–148CrossRefPubMedGoogle Scholar
  49. Sugihara I, Bailly Y, Mariani J (2000) Olivocerebellar climbing fibers in the granuloprival cerebellum: morphological study of individual axonal projections in the X-irradiated rat. J Neurosci 20:3745–3760PubMedGoogle Scholar
  50. Sugihara I, Wu HS, Shinoda Y (2001) The entire trajectories of single olivocerebellar axons in the cerebellar cortex and their contribution to cerebellar compartmentalization. J Neurosci 21:7715–7723Google Scholar
  51. Takayama C, Nakagawa S, Watanabe M, Mishina M, Inoue Y (1995) Light- and electron-microscopic localization of the glutamate receptor channel delta 2 subunit in the mouse Purkinje cell. Neurosci Lett 188:89–92CrossRefPubMedGoogle Scholar
  52. Takeuchi T, Miyazaki T, Watanabe M, Mori H, Sakimura K, Mishina M (2005) Control of synaptic connection by glutamate receptor delta2 in the adult cerebellum. J Neurosci 25:2146–2156CrossRefPubMedGoogle Scholar
  53. Tohgo A, Eiraku M, Miyazaki T, Miura E, Kawaguchi SY, Nishi M, Watanabe M, Hirano T, Kengaku M, Takeshima H (2006) Impaired cerebellar functions in mutant mice lacking DNER. Mol Cell Neurosci 31:326–333CrossRefPubMedGoogle Scholar
  54. Tomioka Y, Miyazaki T, Taharaguchi S, Yoshino S, Morimatsu M, Uede T, Ono E, Watanabe M (2008) Cerebellar pathology in transgenic mice expressing the pseudorabies virus immediate-early protein IE180. Eur J Neurosci 27:2115–2132CrossRefPubMedGoogle Scholar
  55. Uemura T, Kakizawa S, Yamasaki M, Sakimura K, Watanabe M, Iino M, Mishina M (2007) Regulation of long-term depression and climbing fiber territory by glutamate receptor delta2 at parallel fiber synapses through its C-terminal domain in cerebellar Purkinje cells. J Neurosci 27:12096–12108CrossRefPubMedGoogle Scholar
  56. Uemura T, Lee SJ, Yasumura M, Takeuchi T, Yoshida T, Ra M, Taguchi R, Sakimura K, Mishina M (2010) Trans-synaptic interaction of GluRdelta2 and Neurexin through Cbln1 mediates synapse formation in the cerebellum. Cell 141:1068–1079CrossRefPubMedGoogle Scholar
  57. Watanabe M (2008) Molecular mechanisms governing competitive synaptic wiring in cerebellar Purkinje cells. Tohoku J Exp Med 214:175–190CrossRefPubMedGoogle Scholar
  58. Watanabe M, Mishina M, Inoue Y (1994) Distinct spatiotemporal expressions of five NMDA receptor channel subunit mRNAs in the cerebellum. J Comp Neurol 343:513–519CrossRefPubMedGoogle Scholar
  59. Watanabe F, Miyazaki T, Takeuchi T, Fukaya M, Nomura T, Noguchi S, Mori H, Sakimura K, Watanabe M, Mishina M (2008) Effects of FAK ablation on cerebellar foliation, Bergmann glia positioning and climbing fiber territory on Purkinje cells. Eur J Neurosci 27:836–854CrossRefPubMedGoogle Scholar
  60. Woodward DJ, Hoffer BJ, Altman J (1974) Physiological and pharmacological properties of Purkinje cells in rat cerebellum degranulated by postnatal x-irradiation. J Neurobiol 5:283–304CrossRefPubMedGoogle Scholar
  61. Yamada K, Fukaya M, Shimizu H, Sakimura K, Watanabe M (2001) NMDA receptor subunits GluRepsilon1, GluRepsilon3 and GluRzeta1 are enriched at the mossy fibre-granule cell synapse in the adult mouse cerebellum. Eur J Neurosci 13:2025–2036CrossRefPubMedGoogle Scholar
  62. Yuzaki M (2009) New (but old) molecules regulating synapse integrity and plasticity: Cbln1 and the delta2 glutamate receptor. Neuroscience 162:633–643CrossRefPubMedGoogle Scholar

Copyright information

© Japanese Association of Anatomists 2010

Authors and Affiliations

  1. 1.Department of AnatomyHokkaido University Graduate School of MedicineSapporoJapan

Personalised recommendations