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.

  • Original Article
  • Published:

Lack of brain serotonin affects postnatal development and serotonergic neuronal circuitry formation

Molecular Psychiatry volume 18, pages 1106–1118 (2013)Cite this article

Subjects

Abstract

Despite increasing evidence suggests that serotonin (5-HT) can influence neurogenesis, neuronal migration and circuitry formation, the precise role of 5-HT on central nervous system (CNS) development is only beginning to be elucidated. Moreover, how changes in serotonin homeostasis during critical developmental periods may have etiological relevance to human mental disorders, remains an unsolved question. In this study we address the consequences of 5-HT synthesis abrogation on CNS development using a knock-in mouse line in which the tryptophan hydroxylase 2 (Tph2) gene is replaced by the eGFP reporter. We report that lack of brain 5-HT results in a dramatic reduction of body growth rate and in 60% lethality within the first 3 weeks after birth, with no gross anatomical changes in the brain. Thanks to the specific expression of the eGFP, we could highlight the serotonergic system independently of 5-HT immunoreactivity. We found that lack of central serotonin produces severe abnormalities in the serotonergic circuitry formation with a brain region- and time- specific effect. Indeed, we observed a striking reduction of serotonergic innervation to the suprachiasmatic and thalamic paraventricular nuclei, while a marked serotonergic hyperinnervation was found in the nucleus accumbens and hippocampus of Tph2∷eGFP mutants. Finally, we demonstrated that BDNF expression is significantly up-regulated in the hippocampus of mice lacking brain 5-HT, mirroring the timing of the appearance of hyperinnervation and thus unmasking a possible regulatory feedback mechanism tuning the serotonergic neuronal circuitry formation. On the whole, these findings reveal that alterations of serotonin levels during CNS development affect the proper wiring of the brain that may produce long-lasting changes leading to neurodevelopmental disorders.

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
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6

Similar content being viewed by others

References

  1. Gross C, Hen R . The developmental origins of anxiety. Nat Rev Neurosci 2004; 5: 545–552.

    Article  CAS  PubMed  Google Scholar 

  2. Lucki I . The spectrum of behaviors influenced by serotonin. Biol Psychiatry 1998; 44: 151–162.

    Article  CAS  PubMed  Google Scholar 

  3. Miczek KA, de Almeida RM, Kravitz EA, Rissman EF, de Boer SF, Raine A . Neurobiology of escalated aggression and violence. J Neurosci 2007; 27: 11803–11806.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Veenstra-VanderWeele J, Anderson GM, Cook EH . Pharmacogenetics and the serotonin system: initial studies and future directions. Eur J Pharmacol 2000; 410: 165–181.

    Article  CAS  PubMed  Google Scholar 

  5. Lidov HG, Molliver ME . Immunohistochemical study of the development of serotonergic neurons in the rat CNS. Brain Res Bull 1982; 9: 559–604.

    Article  CAS  PubMed  Google Scholar 

  6. Wallace JA, Lauder JM . Development of the serotonergic system in the rat embryo: an immunocytochemical study. Brain Res Bull 1983; 10: 459–479.

    Article  CAS  PubMed  Google Scholar 

  7. Bonnin A, Goeden N, Chen K, Wilson ML, King J, Shih JC et al. A transient placental source of serotonin for the fetal forebrain. Nature 2011; 472: 347–350.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Cote F, Fligny C, Bayard E, Launay JM, Gershon MD, Mallet J et al. Maternal serotonin is crucial for murine embryonic development. Proc Natl Acad Sci USA 2007; 104: 329–334.

    Article  CAS  PubMed  Google Scholar 

  9. Gaspar P, Cases O, Maroteaux L . The developmental role of serotonin: news from mouse molecular genetics. Nat Rev Neurosci 2003; 4: 1002–1012.

    Article  CAS  PubMed  Google Scholar 

  10. Lauder JM . Neurotransmitters as growth regulatory signals: role of receptors and second messengers. Trends Neurosci 1993; 16: 233–240.

    Article  CAS  PubMed  Google Scholar 

  11. Chugani DC, Muzik O, Behen M, Rothermel R, Janisse JJ, Lee J et al. Developmental changes in brain serotonin synthesis capacity in autistic and nonautistic children. Ann Neurol 1999; 45: 287–295.

    Article  CAS  PubMed  Google Scholar 

  12. Sodhi MS, Sanders-Bush E . Serotonin and brain development. Int Rev Neurobiol 2004; 59: 111–174.

    Article  CAS  PubMed  Google Scholar 

  13. Whitaker-Azmitia PM . Serotonin and brain development: role in human developmental diseases. Brain Res Bull 2001; 56: 479–485.

    Article  CAS  PubMed  Google Scholar 

  14. Cases O, Vitalis T, Seif I, De Maeyer E, Sotelo C, Gaspar P . Lack of barrels in the somatosensory cortex of monoamine oxidase A-deficient mice: role of a serotonin excess during the critical period. Neuron 1996; 16: 297–307.

    Article  CAS  PubMed  Google Scholar 

  15. Persico AM, Mengual E, Moessner R, Hall FS, Revay RS, Sora I et al. Barrel pattern formation requires serotonin uptake by thalamocortical afferents, and not vesicular monoamine release. J Neurosci 2001; 21: 6862–6873.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Riccio O, Potter G, Walzer C, Vallet P, Szabo G, Vutskits L et al. Excess of serotonin affects embryonic interneuron migration through activation of the serotonin receptor 6. Mol Psychiatry 2009; 14: 280–290.

    Article  CAS  PubMed  Google Scholar 

  17. Hendricks TJ, Fyodorov DV, Wegman LJ, Lelutiu NB, Pehek EA, Yamamoto B et al. Pet-1 ETS gene plays a critical role in 5-HT neuron development and is required for normal anxiety-like and aggressive behavior. Neuron 2003; 37: 233–247.

    Article  CAS  PubMed  Google Scholar 

  18. Hodges MR, Tattersall GJ, Harris MB, McEvoy SD, Richerson DN, Deneris ES et al. Defects in breathing and thermoregulation in mice with near-complete absence of central serotonin neurons. J Neurosci 2008; 28: 2495–2505.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Alenina N, Kikic D, Todiras M, Mosienko V, Qadri F, Plehm R et al. Growth retardation and altered autonomic control in mice lacking brain serotonin. Proc Natl Acad Sci USA 2009; 106: 10332–10337.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Gutknecht L, Waider J, Kraft S, Kriegebaum C, Holtmann B, Reif A et al. Deficiency of brain 5-HT synthesis but serotonergic neuron formation in Tph2 knockout mice. J Neural Transm 2008; 115: 1127–1132.

    Article  CAS  PubMed  Google Scholar 

  21. Migliarini S, Pacini G, Pasqualetti M . Generation of a Tph2/EGFP knockin mouse line for the study of the role of serotonin during the central nervous system development. Fundam Clin Pharmacol 2008; 22 (Suppl. 2): 129, (abstract SCP055).

    Google Scholar 

  22. Savelieva KV, Zhao S, Pogorelov VM, Rajan I, Yang Q, Cullinan E et al. Genetic disruption of both tryptophan hydroxylase genes dramatically reduces serotonin and affects behavior in models sensitive to antidepressants. PLoS One 2008; 3: e3301.

    Article  PubMed  PubMed Central  Google Scholar 

  23. Yadav VK, Oury F, Suda N, Liu ZW, Gao XB, Confavreux C et al. A serotonin- dependent mechanism explains the leptin regulation of bone mass, appetite, and energy expenditure. Cell 2009; 138: 976–989.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Migliarini S, Pacini G, Pelosi B, Errico F, Lunardi G, Usiello A et al. Tryptophan hydroxylase 2 (Tph2) knockout mice reveal a critical role for brain serotonin (5-HT) in postnatal development and in adult behaviour. Society for Neuroscience, Chicago 2009 (abstract 61.3/N36).

  25. Nichols J, Evans EP, Smith AG . Establishment of germ-line-competent embryonic stem (ES) cells using differentiation inhibiting activity. Development 1990; 110: 1341–1348.

    CAS  PubMed  Google Scholar 

  26. Lee EC, Yu D, Martinez de Velasco J, Tessarollo L, Swing DA, Court DL et al. A highly efficient Escherichia coli-based chromosome engineering system adapted for recombinogenic targeting and subcloning of BAC DNA. Genomics 2001; 73: 56–65.

    Article  CAS  PubMed  Google Scholar 

  27. Ren SY, Pasqualetti M, Dierich A, Le Meur M, Rijli FM . A Hoxa2 mutant conditional allele generated by Flp- and Cre-mediated recombination. Genesis 2002; 32: 105–108.

    Article  CAS  PubMed  Google Scholar 

  28. Rodriguez CI, Buchholz F, Galloway J, Sequerra R, Kasper J, Ayala R et al. High- efficiency deleter mice show that FLPe is an alternative to Cre-loxP. Nat Genet 2000; 25: 139–140.

    Article  CAS  PubMed  Google Scholar 

  29. Uyama O, Okamura N, Yanase M, Narita M, Kawabata K, Sugita M . Quantitative evaluation of vascular permeability in the gerbil brain after transient ischemia using Evans blue fluorescence. J Cereb Blood Flow Metab 1988; 8: 282–284.

    Article  CAS  PubMed  Google Scholar 

  30. Pasqualetti M, Nardi I, Ladinsky H, Marazziti D, Cassano GB . Comparative anatomical distribution of serotonin 1A, 1D alpha and 2A receptor mRNAs in human brain postmortem. Brain Res Mol Brain Res 1996; 39: 223–233.

    Article  CAS  PubMed  Google Scholar 

  31. Pasqualetti M, Neun R, Davenne M, Rijli FM . Retinoic acid rescues inner ear defects in Hoxa1 deficient mice. Nat Genet 2001; 29: 34–39.

    Article  CAS  PubMed  Google Scholar 

  32. Franklin KBJ, Paxinos G . The Mouse Brain in Stereotaxic Coordinates. Academic Press: Amsterdam, NL, 2008.

    Google Scholar 

  33. Pasqualetti M, Ren SY, Poulet M, LeMeur M, Dierich A, Rijli FM . A Hoxa2 knockin allele that expresses EGFP upon conditional Cre-mediated recombination. Genesis 2002; 32: 109–111.

    Article  CAS  PubMed  Google Scholar 

  34. Keith NM, Rowntree LG, Geraghty JT . A METHOD FOR THE DETERMINATION OF PLASMA AND BLOOD VOLUME. Arch Intern Med 1915; 16: 547–557.

    Article  CAS  Google Scholar 

  35. De Vitry F, Hamon M, Catelon J, Dubois M, Thibault J . Serotonin initiates and autoamplifies its own synthesis during mouse central nervous system development. Proc Natl Acad Sci USA 1986; 83: 8629–8633.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Galter D, Unsicker K . Sequential activation of the 5-HT1(A) serotonin receptor and TrkB induces the serotonergic neuronal phenotype. Mol Cell Neurosci 2000; 15: 446–455.

    Article  CAS  PubMed  Google Scholar 

  37. Whitaker-Azmitia PM, Azmitia EC . Stimulation of astroglial serotonin receptors produces culture media which regulates growth of serotonergic neurons. Brain Res 1989; 497: 80–85.

    Article  CAS  PubMed  Google Scholar 

  38. Lyons WE, Mamounas LA, Ricaurte GA, Coppola V, Reid SW, Bora SH et al. Brain- derived neurotrophic factor-deficient mice develop aggressiveness and hyperphagia in conjunction with brain serotonergic abnormalities. Proc Natl Acad Sci USA 1999; 96: 15239–15244.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Mamounas LA, Altar CA, Blue ME, Kaplan DR, Tessarollo L, Lyons WE . BDNF promotes the regenerative sprouting, but not survival, of injured serotonergic axons in the adult rat brain. J Neurosci 2000; 20: 771–782.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Mamounas LA, Blue ME, Siuciak JA, Altar CA . Brain-derived neurotrophic factor promotes the survival and sprouting of serotonergic axons in rat brain. J Neurosci 1995; 15: 7929–7939.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Audero E, Coppi E, Mlinar B, Rossetti T, Caprioli A, Banchaabouchi MA et al. Sporadic autonomic dysregulation and death associated with excessive serotonin autoinhibition. Science 2008; 321: 130–133.

    Article  CAS  PubMed  Google Scholar 

  42. Budnik V, Wu CF, White K . Altered branching of serotonin-containing neurons in Drosophila mutants unable to synthesize serotonin and dopamine. J Neurosci 1989; 9: 2866–2877.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Daubert EA, Condron BG . Serotonin: a regulator of neuronal morphology and circuitry. Trends Neurosci 2010; 33: 424–434.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Diefenbach TJ, Sloley BD, Goldberg JI . Neurite branch development of an identified serotonergic neuron from embryonic Helisoma: evidence for autoregulation by serotonin. Dev Biol 1995; 167: 282–293.

    Article  CAS  PubMed  Google Scholar 

  45. Haydon PG, McCobb DP, Kater SB . Serotonin selectively inhibits growth cone motility and synaptogenesis of specific identified neurons. Science 1984; 226: 561–564.

    Article  CAS  PubMed  Google Scholar 

  46. Sikich L, Hickok JM, Todd RD . 5-HT1A receptors control neurite branching during development. Brain Res Dev Brain Res 1990; 56: 269–274.

    Article  CAS  PubMed  Google Scholar 

  47. Ase AR, Reader TA, Hen R, Riad M, Descarries L . Regional changes in density of serotonin transporter in the brain of 5-HT1A and 5-HT1B knockout mice, and of serotonin innervation in the 5-HT1B knockout. J Neurochem 2001; 78: 619–630.

    Article  CAS  PubMed  Google Scholar 

  48. Dudok JJ, Groffen AJ, Witter MP, Voorn P, Verhage M . Chronic activation of the 5- HT(2) receptor reduces 5-HT neurite density as studied in organotypic slice cultures. Brain Res 2009; 1302: 1–9.

    Article  CAS  PubMed  Google Scholar 

  49. Hannon J, Hoyer D . Molecular biology of 5-HT receptors. Behav Brain Res 2008; 195: 198–213.

    Article  CAS  PubMed  Google Scholar 

  50. Bonnin A, Peng W, Hewlett W, Levitt P . Expression mapping of 5-HT1 serotonin receptor subtypes during fetal and early postnatal mouse forebrain development. Neuroscience 2006; 141: 781–794.

    Article  CAS  PubMed  Google Scholar 

  51. Lauder JM, Wilkie MB, Wu C, Singh S . Expression of 5-HT(2A), 5-HT(2B) and 5- HT(2C) receptors in the mouse embryo. Int J Dev Neurosci 2000; 18: 653–662.

    Article  CAS  PubMed  Google Scholar 

  52. Lein ES, Hawrylycz MJ, Ao N, Ayres M, Bensinger A, Bernard A et al. Genome-wide atlas of gene expression in the adult mouse brain. Nature 2007; 445: 168–176.

    CAS  PubMed  Google Scholar 

  53. Sze JY, Victor M, Loer C, Shi Y, Ruvkun G . Food and metabolic signalling defects in a Caenorhabditis elegans serotonin-synthesis mutant. Nature 2000; 403: 560–564.

    Article  CAS  PubMed  Google Scholar 

  54. Valles AM, White K . Development of serotonin-containing neurons in Drosophila mutants unable to synthesize serotonin. J Neurosci 1986; 6: 1482–1491.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Hill RA, Murray SS, Halley PG, Binder MD, Martin SJ, van den Buuse M . Brain-derived neurotrophic factor expression is increased in the hippocampus of 5-HT(2C) receptor knockout mice. Hippocampus 2011; 21: 434–445.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank Dr M Carta, Dr R Nisticò, Dr P Pelosi, Dr A Usiello and members of our laboratory for valuable discussions and comments on the manuscript. We thank the referees for their comments and helpful suggestions. We thank Dr A Simeone and Dr D Acampora for sharing protocols and advice. We acknowledge the Center for Nanotechnology Innovation for the use of the confocal microscope. We also thank the following for kind gifts of reagents: BayGenomics and MMRRC (E14Tg2a.4 ES cells), Dr D Court (DY380 cells), Dr S Dymecki (ACTB∷FLPe mouse line), Dr Y Bozzi (BDNF probe). This work was supported by Italian Ministry of Education, University and Research (MIUR) (Prin 2008, 200894SYW2) and Toscana Life Sciences Foundation (Orphan_0108 program) to MP. Grants supporting this work were Prin 2008 (200894SYW2) from Italian Ministry of Education, University and Research (MIUR), and Orphan_0108 program from Toscana Life Sciences Foundation to MP BayGenomics and MMRRC provided E14Tg2a.4 ES cells; Dr D Court provided DY380 cells; Dr S Dymecki provided ACTB∷FLPe mouse line; Dr Y Bozzi provided BDNF probe.

Author information

Author notes

  1. G Pacini and B Pelosi: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Biology, Unit of Cellular and Developmental Biology, University of Pisa, Pisa, Italy

    S Migliarini, G Pacini, B Pelosi & M Pasqualetti

  2. Department of Oncology, Sacro Cuore Hospital of Negrar, Verona, Italy

    G Lunardi

Authors
  1. S Migliarini
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. G Pacini
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. B Pelosi
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. G Lunardi
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. M Pasqualetti
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to M Pasqualetti.

Ethics declarations

Competing interests

The authors declare no conflict of interest.

Additional information

Supplementary information accompanies the paper on the Molecular Psychiatry website

Supplementary information

PowerPoint slides

Rights and permissions

Reprints and permissions

About this article

Cite this article

Migliarini, S., Pacini, G., Pelosi, B. et al. Lack of brain serotonin affects postnatal development and serotonergic neuronal circuitry formation. Mol Psychiatry 18, 1106–1118 (2013). https://doi.org/10.1038/mp.2012.128

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/mp.2012.128

Keywords

This article is cited by

Search

Advanced search

Quick links