Brain Structure and Function

, Volume 219, Issue 6, pp 1901–1912 | Cite as

GABA is localized in dopaminergic synaptic vesicles in the rodent striatum

  • Mats Julius Stensrud
  • Maja Puchades
  • Vidar Gundersen
Original Article
First Online:
Received:
Accepted:

Abstract

Recently, electrophysiological evidence was given for inhibitory postsynaptic responses at dopaminergic striatal synapses. These responses were independent of the vesicular GABA transporter, VGAT, but dependent on the vesicular dopamine transporter VMAT2. The identity and the exact source of the released molecule, as well as the presence of the putative inhibitory transmitter in VMAT2 containing synaptic vesicles remain to be shown. To clarify this, in particular to determine whether GABA is responsible for the inhibitory response at dopaminergic synapses, we used the electron microscopic immunogold method to label in vivo perfusion fixed striatal tissue with antibodies recognising GABA, VGAT, VMAT2 and tyrosine hydroxylase. We show that about 13 % of tyrosine hydroxylase positive and 11 % of VMAT2 axonal terminals in the caudo-putamen contain significant labelling for GABA. Immunogold signals for tyrosine hydroxylase and VGAT was totally segregated into different pools of nerve terminals. Quantitative analyses of the distance between gold particles signalling GABA and synaptic vesicles showed that GABA was as closely associated with synaptic vesicles in tyrosine hydroxylase positive as in tyrosine hydroxylase negative nerve terminals. Likewise, in dopaminergic terminals GABA and VMAT2 immunogold particles showed a close spatial localization, strongly suggesting the presence of GABA in VMAT2 positive synaptic vesicles. Our results suggest that GABA is exocytosed together with dopamine from dopaminergic nerve terminals in the caudo-putamen through VGAT negative and VMAT2 positive synaptic vesicles.

Keywords

Nigrostriatal Electron microscopy Mouse Rat Caudoputamen Immunocytochemistry 
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Notes

Acknowledgments

This work was funded by grants from the Medical Faculty, University of Oslo, Norway. The authors thank Mrs. Grazyna Babinska for preparation of electron microscopic samples.

References

  1. Alilain WJ, Horn KP, Hu H, Dick TE, Silver J (2011) Functional regeneration of respiratory pathways after spinal cord injury. Nature 475(7355):196–200PubMedCentralPubMedCrossRefGoogle Scholar
  2. Arluison M, Dietl M, Thibault J (1984) Ultrastructural morphology of dopaminergic nerve terminals and synapses in the striatum of the rat using tyrosine hydroxylase immunocytochemistry: a topographical study. Brain Res Bull 13(2):269–285PubMedCrossRefGoogle Scholar
  3. Bennett BD, Bolam JP (1994) Synaptic input and output of parvalbumin-immunoreactive neurons in the neostriatum of the rat. Neuroscience 62(3):707–719PubMedCrossRefGoogle Scholar
  4. Bergersen L, Ruiz A, Bjaalie JG, Kullmann DM, Gundersen V (2003) GABA and GABAA receptors at hippocampal mossy fibre synapses. Eur J Neurosci 18(4):931–941PubMedCrossRefGoogle Scholar
  5. Bergersen LH, Storm-Mathisen J, Gundersen V (2008) Immunogold quantification of amino acids and proteins in complex subcellular compartments. Nat Protoc 3(1):144–152PubMedCrossRefGoogle Scholar
  6. Bergersen LH, Morland C, Ormel L, Rinholm JE, Larsson M, Wold JF, Roe AT, Stranna A, Santello M, Bouvier D, Ottersen OP, Volterra A, Gundersen V (2012) Immunogold detection of l-glutamate and d-serine in small synaptic-like microvesicles in adult hippocampal astrocytes. Cereb Cortex 22(7):1690–1697PubMedCrossRefGoogle Scholar
  7. Beucher A, Gjernes E, Collin C, Courtney M, Meunier A, Collombat P, Gradwohl G (2012) The homeodomain-containing transcription factors Arx and Pax4 control enteroendocrine subtype specification in mice. PLoS One 7(5):e36449PubMedCentralPubMedCrossRefGoogle Scholar
  8. Borisovska M, Bensen AL, Chong G, Westbrook GL (2013) Distinct modes of dopamine and GABA release in a dual transmitter neuron. J Neurosci Off J Soc Neurosci 33(5):1790–1796CrossRefGoogle Scholar
  9. Brunk I, Blex C, Rachakonda S, Holtje M, Winter S, Pahner I, Walther DJ, Ahnert-Hilger G (2006) The first luminal domain of vesicular monoamine transporters mediates G-protein-dependent regulation of transmitter uptake. J Biol Chem 281(44):33373–33385PubMedCrossRefGoogle Scholar
  10. Brunk I, Blex C, Sanchis-Segura C, Sternberg J, Perreau-Lenz S, Bilbao A, Hortnagl H, Baron J, Juranek J, Laube G, Birnbaumer L, Spanagel R, Ahnert-Hilger G (2008) Deletion of Go2alpha abolishes cocaine-induced behavioral sensitization by disturbing the striatal dopamine system. FASEB J Off Publ Fed Am Soc Exp Biol 22(10):3736–3746Google Scholar
  11. Burger PM, Hell J, Mehl E, Krasel C, Lottspeich F, Jahn R (1991) GABA and glycine in synaptic vesicles: storage and transport characteristics. Neuron 7(2):287–293PubMedCrossRefGoogle Scholar
  12. Chang YC, Gottlieb DI (1988) Characterization of the proteins purified with monoclonal antibodies to glutamic acid decarboxylase. J Neurosci Off J Soc Neurosci 8(6):2123–2130Google Scholar
  13. Chuhma N, Zhang H, Masson J, Zhuang X, Sulzer D, Hen R, Rayport S (2004) Dopamine neurons mediate a fast excitatory signal via their glutamatergic synapses. J Neurosci Off J Soc Neurosci 24(4):972–981CrossRefGoogle Scholar
  14. Conti F, Melone M, De Biasi S, Minelli A, Brecha NC, Ducati A (1998) Neuronal and glial localization of GAT-1, a high-affinity gamma-aminobutyric acid plasma membrane transporter, in human cerebral cortex: with a note on its distribution in monkey cortex. J Comp Neurol 396(1):51–63PubMedCrossRefGoogle Scholar
  15. Cowan RL, Wilson CJ, Emson PC, Heizmann CW (1990) Parvalbumin-containing GABAergic interneurons in the rat neostriatum. J Comp Neurol 302(2):197–205PubMedCrossRefGoogle Scholar
  16. Cserep C, Szonyi A, Veres JM, Nemeth B, Szabadits E, de Vente J, Hajos N, Freund TF, Nyiri G (2011) Nitric oxide signaling modulates synaptic transmission during early postnatal development. Cereb Cortex 21(9):2065–2074PubMedCentralPubMedCrossRefGoogle Scholar
  17. Fiorentino H, Kuczewski N, Diabira D, Ferrand N, Pangalos MN, Porcher C, Gaiarsa JL (2009) GABA(B) receptor activation triggers BDNF release and promotes the maturation of GABAergic synapses. J Neurosci Off J Soc Neurosci 29(37):11650–11661CrossRefGoogle Scholar
  18. Fortune T, Lurie DI (2009) Chronic low-level lead exposure affects the monoaminergic system in the mouse superior olivary complex. J Comp Neurol 513(5):542–558PubMedCentralPubMedCrossRefGoogle Scholar
  19. Fremeau RT Jr, Burman J, Qureshi T, Tran CH, Proctor J, Johnson J, Zhang H, Sulzer D, Copenhagen DR, Storm-Mathisen J, Reimer RJ, Chaudhry FA, Edwards RH (2002) The identification of vesicular glutamate transporter 3 suggests novel modes of signaling by glutamate. Proc Natl Acad Sci USA 99(22):14488–14493PubMedCentralPubMedCrossRefGoogle Scholar
  20. Fuxe K, Dahlstrom AB, Jonsson G, Marcellino D, Guescini M, Dam M, Manger P, Agnati L (2010) The discovery of central monoamine neurons gave volume transmission to the wired brain. Prog Neurobiol 90(2):82–100PubMedCrossRefGoogle Scholar
  21. Gaspar P, Berger B, Alvarez C, Vigny A, Henry JP (1985) Catecholaminergic innervation of the septal area in man: immunocytochemical study using TH and DBH antibodies. J Comp Neurol 241(1):12–33PubMedCrossRefGoogle Scholar
  22. Gerfen CR, Herkenham M, Thibault J (1987) The neostriatal mosaic: II. Patch- and matrix-directed mesostriatal dopaminergic and non-dopaminergic systems. J Neurosci Off J Soc Neurosci 7(12):3915–3934Google Scholar
  23. Gonchar Y, Pang L, Malitschek B, Bettler B, Burkhalter A (2001) Subcellular localization of GABA(B) receptor subunits in rat visual cortex. J Comp Neurol 431(2):182–197PubMedCrossRefGoogle Scholar
  24. Gonzalez-Hernandez T, Barroso-Chinea P, Acevedo A, Salido E, Rodriguez M (2001) Colocalization of tyrosine hydroxylase and GAD65 mRNA in mesostriatal neurons. Eur J Neurosci 13(1):57–67PubMedCrossRefGoogle Scholar
  25. Gras C, Herzog E, Bellenchi GC, Bernard V, Ravassard P, Pohl M, Gasnier B, Giros B, El Mestikawy S (2002) A third vesicular glutamate transporter expressed by cholinergic and serotoninergic neurons. J Neurosci Off J Soc Neurosci 22(13):5442–5451Google Scholar
  26. Gras C, Amilhon B, Lepicard EM, Poirel O, Vinatier J, Herbin M, Dumas S, Tzavara ET, Wade MR, Nomikos GG, Hanoun N, Saurini F, Kemel ML, Gasnier B, Giros B, El Mestikawy S (2008) The vesicular glutamate transporter VGLUT3 synergizes striatal acetylcholine tone. Nat Neurosci 11(3):292–300PubMedCrossRefGoogle Scholar
  27. Groves PM, Linder JC, Young SJ (1994) 5-hydroxydopamine-labeled dopaminergic axons: three-dimensional reconstructions of axons, synapses and postsynaptic targets in rat neostriatum. Neuroscience 58(3):593–604PubMedCrossRefGoogle Scholar
  28. Gundersen V (2008) Co-localization of excitatory and inhibitory transmitters in the brain. Acta Neurol Scand Suppl 188:29–33PubMedCrossRefGoogle Scholar
  29. Gundersen V, Chaudhry FA, Bjaalie JG, Fonnum F, Ottersen OP, Storm-Mathisen J (1998) Synaptic vesicular localization and exocytosis of L-aspartate in excitatory nerve terminals: a quantitative immunogold analysis in rat hippocampus. J Neurosci Off J Soc Neurosci 18(16):6059–6070Google Scholar
  30. Gundersen V, Fonnum F, Ottersen OP, Storm-Mathisen J (2001) Redistribution of neuroactive amino acids in hippocampus and striatum during hypoglycemia: a quantitative immunogold study. J Cereb Blood Flow Metab Off J Int Soc Cereb Blood Flow Metab 21(1):41–51CrossRefGoogle Scholar
  31. Gundersen V, Holten AT, Storm-Mathisen J (2004) GABAergic synapses in hippocampus exocytose aspartate on to NMDA receptors: quantitative immunogold evidence for co-transmission. Mol Cell Neurosci 26(1):156–165PubMedCrossRefGoogle Scholar
  32. Hattori T (1993) Conceptual history of the nigrostriatal dopamine system. Neurosci Res 16(4):239–262PubMedCrossRefGoogle Scholar
  33. Hirasawa H, Puopolo M, Raviola E (2009) Extrasynaptic release of GABA by retinal dopaminergic neurons. J Neurophysiol 102(1):146–158PubMedCentralPubMedCrossRefGoogle Scholar
  34. Hirasawa H, Betensky RA, Raviola E (2012) Corelease of dopamine and GABA by a retinal dopaminergic neuron. J Neurosci Off J Soci Neurosci 32(38):13281–13291CrossRefGoogle Scholar
  35. Kaufman DL, Houser CR, Tobin AJ (1991) Two forms of the gamma-aminobutyric acid synthetic enzyme glutamate decarboxylase have distinct intraneuronal distributions and cofactor interactions. J Neurochem 56(2):720–723PubMedCrossRefGoogle Scholar
  36. Kawaguchi Y (1992) Large aspiny cells in the matrix of the rat neostriatum in vitro: physiological identification, relation to the compartments and excitatory postsynaptic currents. J Neurophysiol 67(6):1669–1682PubMedGoogle Scholar
  37. Kawaguchi Y, Wilson CJ, Emson PC (1990) Projection subtypes of rat neostriatal matrix cells revealed by intracellular injection of biocytin. J Neurosci Off J Soci Neurosci 10(10):3421–3438Google Scholar
  38. Kiyokage E, Pan YZ, Shao Z, Kobayashi K, Szabo G, Yanagawa Y, Obata K, Okano H, Toida K, Puche AC, Shipley MT (2010) Molecular identity of periglomerular and short axon cells. J Neurosci Off J Soci Neurosci 30(3):1185–1196CrossRefGoogle Scholar
  39. Kosaka T, Kosaka K (2008) Tyrosine hydroxylase-positive GABAergic juxtaglomerular neurons are the main source of the interglomerular connections in the mouse main olfactory bulb. Neurosci Res 60(3):349–354PubMedCrossRefGoogle Scholar
  40. Kulik A, Vida I, Lujan R, Haas CA, Lopez-Bendito G, Shigemoto R, Frotscher M (2003) Subcellular localization of metabotropic GABA(B) receptor subunits GABA(B1a/b) and GABA(B2) in the rat hippocampus. J Neurosci Off J Soci Neurosci 23(35):11026–11035Google Scholar
  41. Liberia T, Blasco-Ibanez JM, Nacher J, Varea E, Lanciego JL, Crespo C (2013) Two types of periglomerular cells in the olfactory bulb of the macaque monkey (Macaca fascicularis). Brain Struct Funct 218(4):873–887PubMedCrossRefGoogle Scholar
  42. Liu S, Plachez C, Shao Z, Puche A, Shipley MT (2013) Olfactory bulb short axon cell release of GABA and dopamine produces a temporally biphasic inhibition-excitation response in external tufted cells. J Neurosci Off J Soc Neurosci 33(7):2916–2926CrossRefGoogle Scholar
  43. Mahendrasingam S, Wallam CA, Hackney CM (2003) Two approaches to double post-embedding immunogold labeling of freeze-substituted tissue embedded in low temperature Lowicryl HM20 resin. Brain Res Brain Res Protoc 11(2):134–141PubMedCrossRefGoogle Scholar
  44. Maher BJ, Westbrook GL (2008) Co-transmission of dopamine and GABA in periglomerular cells. J Neurophysiol 99(3):1559–1564PubMedCrossRefGoogle Scholar
  45. Mandemakers W, Snellinx A, O’Neill MJ, de Strooper B (2012) LRRK2 expression is enriched in the striosomal compartment of mouse striatum. Neurobiol Dis 48(3):582–593PubMedCrossRefGoogle Scholar
  46. McDowell KA, Hutchinson AN, Wong-Goodrich SJ, Presby MM, Su D, Rodriguiz RM, Law KC, Williams CL, Wetsel WC, West AE (2010) Reduced cortical BDNF expression and aberrant memory in Carf knock-out mice. J Neurosci Off J Soc Neurosci 30(22):7453–7465CrossRefGoogle Scholar
  47. Morland C, Nordengen K, Gundersen V (2012) Valproate causes reduction of the excitatory amino acid aspartate in nerve terminals. Neurosci Lett 527(2):100–104PubMedCrossRefGoogle Scholar
  48. Nadler JV, Vaca KW, White WF, Lynch GS, Cotman CW (1976) Aspartate and glutamate as possible transmitters of excitatory hippocampal afferents. Nature 260(5551):538–540PubMedCrossRefGoogle Scholar
  49. Olah S, Fule M, Komlosi G, Varga C, Baldi R, Barzo P, Tamas G (2009) Regulation of cortical microcircuits by unitary GABA-mediated volume transmission. Nature 461(7268):1278–1281PubMedCentralPubMedCrossRefGoogle Scholar
  50. Oprins A, Geuze HJ, Slot JW (1994) Cryosubstitution dehydration of aldehyde-fixed tissue: a favorable approach to quantitative immunocytochemistry. J Histochem Cytochem Off J Histochem Soc 42(4):497–503CrossRefGoogle Scholar
  51. Ormel L, Stensrud MJ, Chaudhry FA, Gundersen V (2012) A distinct set of synaptic-like microvesicles in atroglial cells contain VGLUT3. Glia 60(9):1289–1300PubMedCrossRefGoogle Scholar
  52. Rice ME, Cragg SJ (2008) Dopamine spillover after quantal release: rethinking dopamine transmission in the nigrostriatal pathway. Brain Res Rev 58(2):303–313PubMedCentralPubMedCrossRefGoogle Scholar
  53. Sesack SR, Aoki C, Pickel VM (1994) Ultrastructural localization of D2 receptor-like immunoreactivity in midbrain dopamine neurons and their striatal targets. J Neurosci Off J Soc Neurosci 14(1):88–106Google Scholar
  54. Sogn CJ, Puchades M, Gundersen V (2013) Rare contacts between synapses and microglial processes containing high levels of Iba1 and actin - a postembedding immunogold study in the healthy rat brain. Eur J Neurosci 38(1):2030–2040PubMedCrossRefGoogle Scholar
  55. Stensrud M, Chaudhry F, Leergaard T, Bjaalie JG, Gundersen V (2013) VGLUT3 in the rodent brain: vesicular co-localization with VGAT. J Comp Neurol. doi: 10.1002/cne.23331 PubMedGoogle Scholar
  56. Storm-Mathisen J, Leknes AK, Bore AT, Vaaland JL, Edminson P, Haug FM, Ottersen OP (1983) First visualization of glutamate and GABA in neurones by immunocytochemistry. Nature 301(5900):517–520PubMedCrossRefGoogle Scholar
  57. Stuber GD, Hnasko TS, Britt JP, Edwards RH, Bonci A (2010) Dopaminergic terminals in the nucleus accumbens but not the dorsal striatum corelease glutamate. J Neurosci Off J Soc Neurosci 30(24):8229–8233CrossRefGoogle Scholar
  58. Tecuapetla F, Patel JC, Xenias H, English D, Tadros I, Shah F, Berlin J, Deisseroth K, Rice ME, Tepper JM, Koos T (2010) Glutamatergic signaling by mesolimbic dopamine neurons in the nucleus accumbens. J Neurosci Off J Soc Neurosci 30(20):7105–7110CrossRefGoogle Scholar
  59. Tritsch NX, Ding JB, Sabatini BL (2012) Dopaminergic neurons inhibit striatal output through non-canonical release of GABA. Nature 490(7419):262–266PubMedCentralPubMedCrossRefGoogle Scholar
  60. Versteeg DH, Van Der Gugten J, De Jong W, Palkovits M (1976) Regional concentrations of noradrenaline and dopamine in rat brain. Brain Res 113(3):563–574PubMedCrossRefGoogle Scholar
  61. Vigot R, Barbieri S, Brauner-Osborne H, Turecek R, Shigemoto R, Zhang YP, Lujan R, Jacobson LH, Biermann B, Fritschy JM, Vacher CM, Muller M, Sansig G, Guetg N, Cryan JF, Kaupmann K, Gassmann M, Oertner TG, Bettler B (2006) Differential compartmentalization and distinct functions of GABAB receptor variants. Neuron 50(4):589–601PubMedCentralPubMedCrossRefGoogle Scholar
  62. Walker MC, Ruiz A, Kullmann DM (2001) Monosynaptic GABAergic signaling from dentate to CA3 with a pharmacological and physiological profile typical of mossy fiber synapses. Neuron 29(3):703–715PubMedCrossRefGoogle Scholar
  63. Yamaguchi T, Wang HL, Li X, Ng TH, Morales M (2011) Mesocorticolimbic glutamatergic pathway. J Neurosci Off J Soc Neurosci 31(23):8476–8490CrossRefGoogle Scholar
  64. Yung KK, Bolam JP, Smith AD, Hersch SM, Ciliax BJ, Levey AI (1995) Immunocytochemical localization of D1 and D2 dopamine receptors in the basal ganglia of the rat: light and electron microscopy. Neuroscience 65(3):709–730PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  • Mats Julius Stensrud
    • 1
  • Maja Puchades
    • 1
  • Vidar Gundersen
    • 1
    • 2
  1. 1.Glio and Neurotransmitter Group, Department of Anatomy, Institute for Basic Medical SciencesUniversity of OsloOsloNorway
  2. 2.Department of NeurologyOslo University HospitalOsloNorway

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