Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Tlx3 and Tlx1 are post-mitotic selector genes determining glutamatergic over GABAergic cell fates

Nature Neuroscience volume 7, pages 510–517 (2004)Cite this article

Abstract

Glutamatergic and GABAergic neurons mediate much of the excitatory and inhibitory neurotransmission, respectively, in the vertebrate nervous system. The process by which developing neurons select between these two cell fates is poorly understood. Here we show that the homeobox genes Tlx3 and Tlx1 determine excitatory over inhibitory cell fates in the mouse dorsal spinal cord. First, we found that Tlx3 was required for specification of, and expressed in, glutamatergic neurons. Both generic and region-specific glutamatergic markers, including VGLUT2 and the AMPA receptor Gria2, were absent in Tlx mutant dorsal horn. Second, spinal GABAergic markers were derepressed in Tlx mutants, including Pax2 that is necessary for GABAergic differentiation, Gad1/2 and Viaat that regulate GABA synthesis and transport, and the kainate receptors Grik2/3. Third, ectopic expression of Tlx3 was sufficient to suppress GABAergic differentiation and induce formation of glutamatergic neurons. Finally, excess GABA-mediated inhibition caused dysfunction of central respiratory circuits in Tlx3 mutant mice.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Relationship between Tlx3 or Pax2 expression and the glutamatergic or GABAergic neurotransmitter phenotype.
Figure 3: Reduction or loss of the glutamatergic marker in Tlx single- or double-null dorsal spinal cord.
Figure 2: Compromised development of GABAergic neurons in the Pax2 mutant dorsal horn.
Figure 4: Derepression of GABAergic markers in Tlx-null dorsal spinal cord.
Figure 5: Cell fate switch by ectopic expression of mouse Tlx3.
Figure 6: Regulation of AMPA and kainate receptors by Tlx genes and a summary of Tlx gene functions.
Figure 7: Inspiratory and pre-inspiratory neurons in wild-type (Tlx3+/+) and Tlx3 mutant (Tlx3−/−) mice, and transformation of the burst pattern of Tlx3−/− respiratory neuron by bicuculline.

Similar content being viewed by others

Accession codes

Accessions

GenBank/EMBL/DDBJ

References

  1. Bennett, M.R. & Balcar, V.J. Forty years of amino acid transmission in the brain. Neurochem. Int. 35, 269–280 (1999).

    Article  CAS  PubMed  Google Scholar 

  2. Bellocchio, E.E., Reimer, R.J., Fremeau, R.T.J. & Edwards, R.H. Uptake of glutamate into synaptic vesicles by an inorganic phosphate transporter. Science 289, 957–960 (2000).

    Article  CAS  PubMed  Google Scholar 

  3. Fremeau, R.T.J. et al. The expression of vesicular glutamate transporters defines two classes of excitatory synapse. Neuron 31, 247–260 (2001).

    Article  CAS  PubMed  Google Scholar 

  4. Polgar, E., Fowler, J.H., McGill, M.M. & Todd, A.J. The types of neuron which contain protein kinase C gamma in rat spinal cord. Brain Res. 833, 71–80 (1999).

    Article  CAS  PubMed  Google Scholar 

  5. Azkue, J.J. et al. Glutamate-like immunoreactivity in ascending spinofugal afferents to the rat periaqueductal grey. Brain Res. 790, 74–81 (1998).

    Article  CAS  PubMed  Google Scholar 

  6. Lu, Y. & Perl, E.R. A specific inhibitory pathway between substantia gelatinosa neurons receiving direct C-fiber input. J. Neurosci. 23, 8752–8758 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Melzack, R. & Wall, P.D. Pain mechanisms: a new theory. Science 150, 971–979 (1965).

    Article  CAS  PubMed  Google Scholar 

  8. Malcangio, M. & Bowery, N.G. GABA and its receptors in the spinal cord. Trends Pharmacol. Sci. 17, 457–462 (1996).

    Article  CAS  PubMed  Google Scholar 

  9. Kerchner, G.A., Wang, G.D., Qiu, C.S., Huettner, J.E. & Zhuo, M. Direct presynaptic regulation of GABA/glycine release by kainate receptors in the dorsal horn: an ionotropic mechanism. Neuron 32, 477–488 (2001).

    Article  CAS  PubMed  Google Scholar 

  10. Dickenson, A.H. Gate control theory of pain stands the test of time. Br. J. Anaesth. 88, 755–757 (2002).

    Article  CAS  PubMed  Google Scholar 

  11. Fode, C. et al. A role for neural determination genes in specifying the dorsoventral identity of telencephalic neurons. Genes Dev. 14, 67–80 (2000).

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Anderson, S.A., Kaznowski, C.E., Horn, C., Rubenstein, J.L. & McConnell, S.K. Distinct origins of neocortical projection neurons and interneurons in vivo. Cereb. Cortex 12, 702–709 (2002).

    Article  PubMed  Google Scholar 

  13. Muzio, L. et al. Conversion of cerebral cortex into basal ganglia in Emx2−/− Pax6(Sey/Sey) double-mutant mice. Nat. Neurosci. 5, 737–745 (2002).

    Article  CAS  PubMed  Google Scholar 

  14. Garcia-Bellido, A. Genetic control of wing disc development in Drosophila. Ciba. Found. Symp. 0, 161–182 (1975).

    CAS  PubMed  Google Scholar 

  15. Caspary, T. & Anderson, K.V. Patterning cell types in the dorsal spinal cord: what the mouse mutants say. Nat. Rev. Neurosci. 4, 289–297 (2003).

    Article  PubMed  Google Scholar 

  16. Goulding, M., Lanuza, G., Sapir, T. & Narayan, S. The formation of sensorimotor circuits. Curr. Opin. Neurobiol. 12, 508–515 (2002).

    Article  CAS  PubMed  Google Scholar 

  17. Helms, A.W. & Johnson, J.E. Specification of dorsal spinal cord interneurons. Curr. Opin. Neurobiol. 13, 42–49 (2003).

    Article  CAS  PubMed  Google Scholar 

  18. Qian, Y., Shirasawa, S., Chen, C.L., Cheng, L. & Ma, Q. Proper development of relay somatic sensory neurons and D2/D4 interneurons requires homeobox genes Rnx/Tlx-3 and Tlx-1. Genes Dev. 16, 1220–1233 (2002).

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  19. Qian, Y. et al. Formation of brainstem (nor)adrenergic centers and first-order relay visceral sensory neurons is dependent on homeodomain protein Rnx/Tlx3. Genes Dev. 15, 2533–2545 (2001).

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  20. Shirasawa, S. et al. Rnx deficiency results in congenital central hypoventilation. Nat. Genet. 24, 287–290 (2000).

    Article  CAS  PubMed  Google Scholar 

  21. Gross, M.K., Dottori, M. & Goulding, M. Lbx1 specifies somatosensory association interneurons in the dorsal spinal cord. Neuron 34, 535–549 (2002).

    Article  CAS  PubMed  Google Scholar 

  22. Muller, T. et al. The homeodomain factor Lbx1 distinguishes two major programs of neuronal differentiation in the dorsal spinal cord. Neuron 34, 551–562 (2002).

    Article  CAS  PubMed  Google Scholar 

  23. Stein, R., Orit, S. & Anderson, D.J. The induction of a neural-specific gene, SCG10, by nerve growth factor in PC12 cells is transcriptional, protein synthesis dependent, and glucocorticoid inhibitable. Dev. Biol. 127, 316–325 (1988).

    Article  CAS  PubMed  Google Scholar 

  24. Maricich, S.M. & Herrup, K. Pax-2 expression defines a subset of GABAergic interneurons and their precursors in the developing murine cerebellum. J. Neurobiol. 41, 281–294 (1999).

    Article  CAS  PubMed  Google Scholar 

  25. Erlander, M.G., Tillakaratne, N.J., Feldblum, S., Patel, N. & Tobin, A.J. Two genes encode distinct glutamate decarboxylases. Neuron 7, 91–100 (1991).

    Article  CAS  PubMed  Google Scholar 

  26. McIntire, S.L., Reimer, R.J., Schuske, K., Edwards, R.H. & Jorgensen, E.M. Identification and characterization of the vesicular GABA transporter. Nature 389, 870–876 (1997).

    Article  CAS  PubMed  Google Scholar 

  27. Kaneko, T. & Fujiyama, F. Complementary distribution of vesicular glutamate transporters in the central nervous system. Neurosci. Res. 42, 243–250 (2002).

    Article  CAS  PubMed  Google Scholar 

  28. Kullander, K. et al. Role of EphA4 and EphrinB3 in local neuronal circuits that control walking. Science 299, 1889–1892 (2003).

    Article  CAS  PubMed  Google Scholar 

  29. Bouchard, M., Pfeffer, P. & Busslinger, M. Functional equivalence of the transcription factors Pax2 and Pax5 in mouse development. Development 127, 3703–3713 (2000).

    CAS  PubMed  Google Scholar 

  30. Logan, C., Wingate, R.J.T., McKay, I.J. & Lumsden, A. Tlx-1 and Tlx-3 homeobox gene expression in cranial sensory ganglia and hindbrain of the chick embryo: markers of patterned connectivity. J. Neurosci. 18, 5389–5402 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Kerr, R.C., Maxwell, D.J. & Todd, A.J. GluR1 and GluR2/3 subunits of the AMPA-type glutamate receptor are associated with particular types of neuron in laminae I–III of the spinal dorsal horn of the rat. Eur. J. Neurosci. 10, 324–333 (1998).

    Article  CAS  PubMed  Google Scholar 

  32. Kerchner, G.A., Wilding, T.J., Huettner, J.E. & Zhuo, M. Kainate receptor subunits underlying presynaptic regulation of transmitter release in the dorsal horn. J. Neurosci. 22, 8010–8017 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Onimaru, H., Arata, A. & Homma, I. Neuronal mechanisms of respiratory rhythm generation: an approach using in vitro preparation. Jpn. J. Physiol. 47, 385–403 (1997).

    Article  CAS  PubMed  Google Scholar 

  34. Feldman, J.L., Mitchell, G.S. & Nattie, E.E. Breathing: rhythmicity, plasticity, chemosensitivity. Annu. Rev. Neurosci. 26, 239–266 (2003).

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  35. Suzue, T. Respiratory rhythm generation in the in vitro brain stem-spinal cord preparation of the neonatal rat. J. Physiol. 354, 173–183 (1984).

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  36. Ritter, B. & Zhang, W. Early postnatal maturation of GABAA-mediated inhibition in the brainstem respiratory rhythm-generating network of the mouse. Eur. J. Neurosci. 12, 2975–2984 (2000).

    Article  CAS  PubMed  Google Scholar 

  37. Mann, R.S. & Carroll, S.B. Molecular mechanisms of selector gene function and evolution. Curr. Opin. Genet. Dev. 12, 592–600 (2002).

    Article  CAS  PubMed  Google Scholar 

  38. Ma, W., Behar, T. & Barker, J.L. Transient expression of GABA immunoreactivity in the developing rat spinal cord. J. Comp. Neurol. 325, 271–290 (1992).

    Article  CAS  PubMed  Google Scholar 

  39. Schaffner, A.E., Behar, T., Nadi, S., Smallwood, V. & Barker, J.L. Quantitative analysis of transient GABA expression in embryonic and early postnatal rat spinal cord neurons. Brain Res. Dev. Brain Res. 72, 265–276 (1993).

    Article  CAS  PubMed  Google Scholar 

  40. Landis, S.C. Target regulation of neurotransmitter phenotype. Trends Neurosci. 13, 344–350 (1990).

    Article  CAS  PubMed  Google Scholar 

  41. Nabekura, J. et al. Developmental switch from GABA to glycine release in single central synaptic terminals. Nat. Neurosci. 7, 17–23 (2004).

    Article  CAS  PubMed  Google Scholar 

  42. Panganiban, G. & Rubenstein, J.L. Developmental functions of the Distal-less/Dlx homeobox genes. Development 129, 4371–4386 (2002).

    CAS  PubMed  Google Scholar 

  43. Blessing, W.W. The Lower Brainstem and Body Homeostasis (Oxford Univ. Press, New York, 1997).

    Google Scholar 

  44. Roberts, C.W., Shutter, J.R. & Korsmeyer, S.J. Hox11 controls the genesis of the spleen. Nature 368, 747–749 (1994).

    Article  CAS  PubMed  Google Scholar 

  45. Birren, S.J., Lo, L.C. & Anderson, D.J. Sympathetic neurons undergo a developmental switch in trophic dependence. Development 119, 597–610 (1993).

    CAS  PubMed  Google Scholar 

  46. Chen, Z.F. et al. The paired homeodomain protein DRG11 is required for the projection of cutaneous sensory afferent fibers to the dorsal spinal cord. Neuron 31, 59–73 (2001).

    Article  CAS  PubMed  Google Scholar 

  47. Morgan, B.A. & Fekete, D.M. Manipulating gene expression with replication-competent retroviruses. in Methods in Avian Embryology Vol. 51 (ed. Bronner-Fraser, M.E.) 185–218 (Academic, San Diego, 1996).

    Chapter  Google Scholar 

  48. Onimaru, H., Arata, A. & Homma, I. Primary respiratory rhythm generator in the medulla of brainstem-spinal cord preparation from newborn rat. Brain Res. 455, 314–324 (1988).

    Article  Google Scholar 

  49. Onimaru, H. & Homma, I. Whole cell recordings from respiratory neurons in the medulla of brainstem-spinal cord preparations isolated from newborn rats. Pflugers Arch. 420, 399–406 (1992).

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank S. Korsmeyer for providing Tlx3 and Tlx1 knockout mice. We are grateful to F. Guillemot, C. Schuuman, C. Stiles, M. Greenberg, G. Lemke and Z. He for critical comments or discussion. Q.M. is a Claudia Adams Barr Scholar and a Pew Scholar in Biomedical Sciences. P.A.G. is a Parker B. Francis Fellow in Pulmonary Medicine and C.C. is a Medical Foundation Fellow. This work was supported by grants from the National Institutes of Health to Q.M. and M.G., and in part by a Showa University Grant-in-Aid for Innovative Collaborative Research Projects from the Japanese Ministry of Education, Culture, Sports, Science and Technology to H.O.

Author information

Author notes

  1. Leping Cheng, Akiko Arata and Rumiko Mizuguchi: These authors contributed equally to this work.

Authors and Affiliations

  1. The Dana-Farber Cancer Institute, Harvard Medical School, 1 Jimmy Fund Way, Boston, 02115, Massachusetts, USA

    Leping Cheng, Ying Qian, Paul A Gray, Ping Luo, Chih-Li Chen & Qiufu Ma

  2. Department of Neurobiology, Harvard Medical School, 1 Jimmy Fund Way, Boston, 02115, Massachusetts, USA

    Leping Cheng, Ying Qian, Paul A Gray, Ping Luo, Chih-Li Chen & Qiufu Ma

  3. Laboratory for Memory and Learning, Brain Science Institute, RIKEN, Wako, 351-0198, Saitama, Japan

    Akiko Arata

  4. Molecular Neurobiology Laboratory, The Salk Institute for Biological Studies, 10010 North Torrey Pines Road, La Jolla, 92037, California, USA

    Rumiko Mizuguchi, Asanka Karunaratne & Martyn Goulding

  5. Center for Biotechnology, Showa University, Tokyo, 142-8555, Japan

    Satoru Arata

  6. Department of Pathology, International Medical Center of Japan, 1-21-1 Toyama, Shinjuku-ku, Tokyo, 162-8655, Japan

    Senji Shirasawa

  7. Research Institute of Molecular Pathology, Dr Bohr-Gasse 7, Vienna, A-1030, Austria

    Maxime Bouchard & Meinrad Busslinger

  8. Department of Physiology, Showa University School of Medicine, Tokyo, 142-8555, Japan

    Hiroshi Onimaru

Authors
  1. Leping Cheng
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Akiko Arata
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Rumiko Mizuguchi
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ying Qian
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Asanka Karunaratne
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Paul A Gray
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Satoru Arata
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Senji Shirasawa
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Maxime Bouchard
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Ping Luo
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Chih-Li Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Meinrad Busslinger
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Martyn Goulding
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. Hiroshi Onimaru
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. Qiufu Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Qiufu Ma.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Cheng, L., Arata, A., Mizuguchi, R. et al. Tlx3 and Tlx1 are post-mitotic selector genes determining glutamatergic over GABAergic cell fates. Nat Neurosci 7, 510–517 (2004). https://doi.org/10.1038/nn1221

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nn1221

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing