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Genetic absence of the vesicular inhibitory amino acid transporter differentially regulates respiratory and locomotor motor neuron development

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Abstract

During mid to late embryonic development (E13 to birth in mice), the neuromotor system is refined by reducing motor neuron (MN) numbers and establishing nascent synaptic connections onto and by MNs. Concurrently, the response to GABAergic and glycinergic synaptic activity switches from postsynaptic excitation to inhibition. Our previous studies on mutant mice lacking glycinergic transmission or deficient in GABA suggests that altered MN activity levels during this developmental period differentially regulates MN survival and muscle innervation for respiratory and non-respiratory motor pools. To determine if combined loss of GABAergic and glycinergic transmission plays a similar or exaggerated role, we quantified MN number and muscle innervation in two respiratory (hypoglossal and phrenic) and two locomotor (brachial and lumbar) motor pools, in mice lacking vesicular inhibitory amino acid transporter, which display absent or severely impaired GABAergic and glycinergic neurotransmission. For respiratory MNs, we observed significant decreases in MN number (−20 % hypoglossal and −36 % phrenic) and diaphragm axonal branching (−60 %). By contrast, for non-respiratory brachial and lumbar MNs, we observed increases in MN number (+62 % brachial and +84 % lumbar) and axonal branching for innervated muscles (+123 % latissimus dorsi for brachial and +61 % gluteal for lumbar). These results show that combined absence of GABAergic and glycinergic neurotransmission causes distinct regional changes in MN number and muscle innervation, which are dependent upon the motor function of the specific motor pool.

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References

  • Akerman CJ, Cline HT (2007) Refining the roles of GABAergic signaling during neural circuit formation. Trends Neurosci 30(8):382–389

    Article  CAS  PubMed  Google Scholar 

  • Allain AE, Bairi A, Meyrand P, Branchereau P (2004) Ontogenic changes of the GABAergic system in the embryonic mouse spinal cord. Brain Res 1000(1-2):134–147

    Article  CAS  PubMed  Google Scholar 

  • Allain AE, Bairi A, Meyrand P, Branchereau P (2006) Expression of the glycinergic system during the course of embryonic development in the mouse spinal cord and its co-localization with GABA immunoreactivity. J Comp Neurol 496(6):832–846

    Article  CAS  PubMed  Google Scholar 

  • Asada H, Kawamura Y, Maruyama K, Kume H, Ding RG, Kanbara N, Kuzume H, Sanbo M, Yagi T, Obata K (1997) Cleft palate and decreased brain gamma-aminobutyric acid in mice lacking the 67-kDa isoform of glutamic acid decarboxylase. Proc Natl Acad Sci USA 94(12):6496–6499

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Bacskai T, Fu Y, Sengul G, Rusznak Z, Paxinos G, Watson C (2013) Musculotopic organization of the motor neurons supplying forelimb and shoulder girdle muscles in the mouse. Brain Struct Funct 218(1):221–238

    Article  PubMed  Google Scholar 

  • Ballion B, Branchereau P, Chapron J, Viala D (2002) Ontogeny of descending serotonergic innervation and evidence for intraspinal 5-HT neurons in the mouse spinal cord. Brain Res Dev Brain Res 137(1):81–88

    Article  CAS  PubMed  Google Scholar 

  • Banks GB, Chau TN, Bartlett SE, Noakes PG (2001) Promotion of motoneuron survival and branching in rapsyn-deficient mice. J Comp Neurol 429(1):156–165

    Article  CAS  PubMed  Google Scholar 

  • Banks GB, Choy PT, Lavidis NA, Noakes PG (2003) Neuromuscular synapses mediate motor axon branching and motoneuron survival during the embryonic period of programmed cell death. Dev Biol 257(1):71–84

    Article  CAS  PubMed  Google Scholar 

  • Banks GB, Kanjhan R, Wiese S, Kneussel M, Wong LM, O’Sullivan G, Sendtner M, Bellingham MC, Betz H, Noakes PG (2005) Glycinergic and GABAergic synaptic activity differentially regulate motoneuron survival and skeletal muscle innervation. J Neurosci 25(5):1249–1259

    Article  CAS  PubMed  Google Scholar 

  • Ben-Ari Y (2002) Excitatory actions of gaba during development: the nature of the nurture. Nat Rev Neurosci 3(9):728–739

    Article  CAS  PubMed  Google Scholar 

  • Bhumbra GS, Moore NJ, Moroni M, Beato M (2012) Co-release of GABA does not occur at glycinergic synapses onto lumbar motoneurons in juvenile mice. Front Cell Neurosci 6:8

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Boyce RW, Dorph-Petersen KA, Lyck L, Gundersen HJ (2010) Design-based stereology: introduction to basic concepts and practical approaches for estimation of cell number. Toxicol Pathol 38(7):1011–1025

    Article  PubMed  Google Scholar 

  • Clarke PG, Oppenheim RW (1995) Neuron death in vertebrate development: in vitro methods. Methods Cell Biol 46:277–321

    Article  CAS  PubMed  Google Scholar 

  • Dahm LM, Landmesser LT (1988) The regulation of intramuscular nerve branching during normal development and following activity blockade. Dev Biol 130(2):621–644

    Article  CAS  PubMed  Google Scholar 

  • Dahm LM, Landmesser LT (1991) The regulation of synaptogenesis during normal development and following activity blockade. J Neurosci 11(1):238–255

    CAS  PubMed  Google Scholar 

  • Drummond GB (2009) Reporting ethical matters in the Journal of Physiology: standards and advice. J Physiol 587(Pt 4):713–719

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • 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(5392):1321–1324

    Article  CAS  PubMed  Google Scholar 

  • Ferraro E, Molinar F, Berghella L (2012) Molecular control of neuromuscular junction development. J Cachexia Sarcopenia Muscle 3(1):13–23

    Article  PubMed Central  PubMed  Google Scholar 

  • Fogarty MJ, Smallcombe KL, Yanagawa Y, Obata K, Bellingham MC, Noakes PG (2013) Genetic deficiency of GABA differentially regulates respiratory and non-respiratory motor neuron development. PLoS One 8(2):e56257. doi:10.1371/journal.pone.0056257

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Fujii M, Arata A, Kanbara-Kume N, Saito K, Yanagawa Y, Obata K (2007) Respiratory activity in brainstem of fetal mice lacking glutamate decarboxylase 65/67 and vesicular GABA transporter. Neuroscience 146(3):1044–1052

    Article  CAS  PubMed  Google Scholar 

  • Gao BX, Stricker C, Ziskind-Conhaim L (2001) Transition from GABAergic to glycinergic synaptic transmission in newly formed spinal networks. J Neurophysiol 86(1):492–502

    CAS  PubMed  Google Scholar 

  • Gundersen HJ, Bagger P, Bendtsen TF, Evans SM, Korbo L, Marcussen N, Moller A, Nielsen K, Nyengaard JR, Pakkenberg B et al (1988a) The new stereological tools: disector, fractionator, nucleator and point sampled intercepts and their use in pathological research and diagnosis. APMIS 96(10):857–881

    Article  CAS  PubMed  Google Scholar 

  • Gundersen HJ, Bendtsen TF, Korbo L, Marcussen N, Moller A, Nielsen K, Nyengaard JR, Pakkenberg B, Sorensen FB, Vesterby A et al (1988b) Some new, simple and efficient stereological methods and their use in pathological research and diagnosis. APMIS 96(5):379–394

    Article  CAS  PubMed  Google Scholar 

  • Hanson MG, Landmesser LT (2003) Characterization of the circuits that generate spontaneous episodes of activity in the early embryonic mouse spinal cord. J Neurosci 23(2):587–600

    CAS  PubMed  Google Scholar 

  • Kirsch J, Betz H (1993) Widespread expression of gephyrin, a putative glycine receptor-tubulin linker protein, in rat brain. Brain Res 621(2):301–310

    Article  CAS  PubMed  Google Scholar 

  • Kneussel M, Brandstatter JH, Laube B, Stahl S, Muller U, Betz H (1999) Loss of postsynaptic GABA(A) receptor clustering in gephyrin-deficient mice. J Neurosci 19(21):9289–9297

    CAS  PubMed  Google Scholar 

  • Kneussel M, Haverkamp S, Fuhrmann JC, Wang H, Wassle H, Olsen RW, Betz H (2000) The gamma-aminobutyric acid type A receptor (GABAAR)-associated protein GABARAP interacts with gephyrin but is not involved in receptor anchoring at the synapse. Proc Natl Acad Sci USA 97(15):8594–8599

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Lance-Jones C (1982) Motoneuron cell death in the developing lumbar spinal cord of the mouse. Brain Res 256(4):473–479

    Article  CAS  PubMed  Google Scholar 

  • Landmesser L (1992) The relationship of intramuscular nerve branching and synaptogenesis to motoneuron survival. J Neurobiol 23(9):1131–1139

    Article  CAS  PubMed  Google Scholar 

  • Levi S, Logan SM, Tovar KR, Craig AM (2004) Gephyrin is critical for glycine receptor clustering but not for the formation of functional GABAergic synapses in hippocampal neurons. J Neurosci 24(1):207–217

    Article  CAS  PubMed  Google Scholar 

  • Mantilla CB, Sieck GC (2008) Key aspects of phrenic motoneuron and diaphragm muscle development during the perinatal period. J Appl Physiol 104(6):1818–1827

    Article  PubMed Central  PubMed  Google Scholar 

  • Moody WJ, Bosma MM (2005) Ion channel development, spontaneous activity, and activity-dependent development in nerve and muscle cells. Physiol Rev 85(3):883–941

    Article  CAS  PubMed  Google Scholar 

  • Narayanan CH, Fox MW, Hamburger V (1971) Prenatal development of spontaneous and evoked activity in the rat (Rattus norvegicus albinus). Behaviour 40(1):100–134

    Article  CAS  PubMed  Google Scholar 

  • Nishimaru H, Kudo N (2000) Formation of the central pattern generator for locomotion in the rat and mouse. Brain Res Bull 53(5):661–669

    Article  CAS  PubMed  Google Scholar 

  • Nishimaru H, Iizuka M, Ozaki S, Kudo N (1996) Spontaneous motoneuronal activity mediated by glycine and GABA in the spinal cord of rat fetuses in vitro. J Physiol 497(Pt 1):131–143

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Noakes PG, Chin D, Kim SS, Liang S, Phillips WD (1999) Expression and localisation of dynamin and syntaxin during neural development and neuromuscular synapse formation. J Comp Neurol 410(4):531–540

    Article  CAS  PubMed  Google Scholar 

  • Oppenheim RW (1989) The neurotrophic theory and naturally occurring motoneuron death. Trends Neurosci 12(7):252–255

    Article  CAS  PubMed  Google Scholar 

  • Oppenheim RW (1991) Cell death during development of the nervous system. Annu Rev Neurosci 14:453–501

    Article  CAS  PubMed  Google Scholar 

  • Oppenheim RW, Nunez R (1982) Electrical stimulation of hindlimb increases neuronal cell death in chick embryo. Nature 295(5844):57–59

    Article  CAS  PubMed  Google Scholar 

  • Oppenheim RW, Prevette D, Houenou LJ, Pincon-Raymond M, Dimitriadou V, Donevan A, O’Donovan M, Wenner P, McKemy DD, Allen PD (1997) Neuromuscular development in the avian paralytic mutant crooked neck dwarf (cn/cn): further evidence for the role of neuromuscular activity in motoneuron survival. J Comp Neurol 381(3):353–372

    Article  CAS  PubMed  Google Scholar 

  • Oppenheim RW, Prevette D, D’Costa A, Wang S, Houenou LJ, McIntosh JM (2000) Reduction of neuromuscular activity is required for the rescue of motoneurons from naturally occurring cell death by nicotinic-blocking agents. J Neurosci 20(16):6117–6124

    CAS  PubMed  Google Scholar 

  • Oppenheim RW, Caldero J, Cuitat D, Esquerda J, Ayala V, Prevette D, Wang S (2003) Rescue of developing spinal motoneurons from programmed cell death by the GABA(A) agonist muscimol acts by blockade of neuromuscular activity and increased intramuscular nerve branching. Mol Cell Neurosci 22(3):331–343

    Article  CAS  PubMed  Google Scholar 

  • Oppenheim RW, Caldero J, Cuitat D, Esquerda J, McArdle JJ, Olivera BM, Prevette D, Teichert RW (2008) The rescue of developing avian motoneurons from programmed cell death by a selective inhibitor of the fetal muscle-specific nicotinic acetylcholine receptor. Dev Neurobiol 68(7):972–980

    Article  CAS  PubMed  Google Scholar 

  • Owens DF, Kriegstein AR (2002) Is there more to GABA than synaptic inhibition? Nat Rev Neurosci 3(9):715–727

    Article  CAS  PubMed  Google Scholar 

  • Paxinos G, Halliday G, Watson C, Koutcherov Y, Wang H (2006) Atlas of the developing mouse brain at E17.5, P0 and P6. Academic Press, New York

    Google Scholar 

  • Pittman R, Oppenheim RW (1979) Cell death of motoneurons in the chick embryo spinal cord. IV. Evidence that a functional neuromuscular interaction is involved in the regulation of naturally occurring cell death and the stabilization of synapses. J Comp Neurol 187(2):425–446

    Article  CAS  PubMed  Google Scholar 

  • Porcher C, Hatchett C, Longbottom RE, McAinch K, Sihra TS, Moss SJ, Thomson AM, Jovanovic JN (2011) Positive feedback regulation between gamma-aminobutyric acid type A (GABA(A)) receptor signaling and brain-derived neurotrophic factor (BDNF) release in developing neurons. J Biol Chem 286(24):21667–21677

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Ren J, Greer JJ (2003) Ontogeny of rhythmic motor patterns generated in the embryonic rat spinal cord. J Neurophysiol 89(3):1187–1195

    Article  PubMed  Google Scholar 

  • Saito K, Kakizaki T, Hayashi R, Nishimaru H, Furukawa T, Nakazato Y, Takamori S, Ebihara S, Uematsu M, Mishina M, Miyazaki J, Yokoyama M, Konishi S, Inoue K, Fukuda A, Fukumoto M, Nakamura K, Obata K, Yanagawa Y (2010) The physiological roles of vesicular GABA transporter during embryonic development: a study using knockout mice. Mol Brain 3:40

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Sibilla S, Ballerini L (2009) GABAergic and glycinergic interneuron expression during spinal cord development: dynamic interplay between inhibition and excitation in the control of ventral network outputs. Prog Neurobiol 89(1):46–60

    Article  CAS  PubMed  Google Scholar 

  • Singer JH, Berger AJ (2000) Development of inhibitory synaptic transmission to motoneurons. Brain Res Bull 53(5):553–560

    Article  CAS  PubMed  Google Scholar 

  • Tamamaki N, Yanagawa Y, Tomioka R, Miyazaki J, Obata K, Kaneko T (2003) Green fluorescent protein expression and colocalization with calretinin, parvalbumin, and somatostatin in the GAD67-GFP knock-in mouse. J Comp Neurol 467(1):60–79

    Article  CAS  PubMed  Google Scholar 

  • Tang J, Landmesser L (1993) Reduction of intramuscular nerve branching and synaptogenesis is correlated with decreased motoneuron survival. J Neurosci 13(7):3095–3103

    CAS  PubMed  Google Scholar 

  • Tsunekawa N, Arata A, Obata K (2005) Development of spontaneous mouth/tongue movement and related neural activity, and their repression in fetal mice lacking glutamate decarboxylase 67. Eur J Neurosci 21(1):173–178

    Article  PubMed  Google Scholar 

  • Waagepetersen HS, Sonnewald U, Schousboe A (1999) The GABA paradox: multiple roles as metabolite, neurotransmitter, and neurodifferentiative agent. J Neurochem 73(4):1335–1342

    Article  CAS  PubMed  Google Scholar 

  • Watson C, Paxinos G, Kayalioglu G (2008) The spinal cord. Academic Press, New York

    Google Scholar 

  • West MJ (1993) New stereological methods for counting neurons. Neurobiol Aging 14(4):275–285

    Article  CAS  PubMed  Google Scholar 

  • West MJ (1999) Stereological methods for estimating the total number of neurons and synapses: issues of precision and bias. Trends Neurosci 22(2):51–61

    Article  CAS  PubMed  Google Scholar 

  • Wojcik SM, Katsurabayashi S, Guillemin I, Friauf E, Rosenmund C, Brose N, Rhee JS (2006) A shared vesicular carrier allows synaptic corelease of GABA and glycine. Neuron 50(4):575–587

    Article  CAS  PubMed  Google Scholar 

  • Yvert B, Branchereau P, Meyrand P (2004) Multiple spontaneous rhythmic activity patterns generated by the embryonic mouse spinal cord occur within a specific developmental time window. J Neurophysiol 91(5):2101–2109

    Article  PubMed  Google Scholar 

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Acknowledgments

The work was funded by project Grant 568680 from the National Health and Medical Research Council, Australia, to PGN and MCB. MJF was supported by an Australian Postgraduate Award Scholarship from the Federal Government of Australia. YY was supported by a grant-in-aid for scientific research on innovative areas ‘Mesocropic Neurocircuitry’ and scientific research (B) from the Ministry of Education, Culture, Sports, Science and Technology, Japan and Takeda Science Foundation. The stereology system was purchased with the aid of an Australian Research Council Linkage Infrastructure, Equipment and Facilities Grant LE100100074. We thank Lynn Tolley for processing tissues for light microscopy. We also thank Maryam Shayegh, Luke Hammond and Mary White for their technical support.

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Authors and Affiliations

  1. School of Biomedical Sciences, University of Queensland, St. Lucia, Brisbane, QLD, 4072, Australia

    Matthew J. Fogarty, Mark C. Bellingham & Peter G. Noakes

  2. Queensland Brain Institute, University of Queensland, Brisbane, QLD, Australia

    Peter G. Noakes

  3. Gunma University Graduate School of Medicine, Maebashi, Japan

    Yuchio Yanagawa

  4. Japan Science and Technology Agency, CREST, Tokyo, Japan

    Yuchio Yanagawa

  5. Obata Research Unit, RIKEN Brain Science Institute, Saitama, Japan

    Kunihiko Obata

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  1. Matthew J. Fogarty
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  5. Peter G. Noakes
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Correspondence to Peter G. Noakes.

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Fogarty, M.J., Yanagawa, Y., Obata, K. et al. Genetic absence of the vesicular inhibitory amino acid transporter differentially regulates respiratory and locomotor motor neuron development. Brain Struct Funct 220, 525–540 (2015). https://doi.org/10.1007/s00429-013-0673-9

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