Article | Published:

Cerebral organoids model human brain development and microcephaly

Nature volume 501, pages 373379 (19 September 2013) | Download Citation


The complexity of the human brain has made it difficult to study many brain disorders in model organisms, highlighting the need for an in vitro model of human brain development. Here we have developed a human pluripotent stem cell-derived three-dimensional organoid culture system, termed cerebral organoids, that develop various discrete, although interdependent, brain regions. These include a cerebral cortex containing progenitor populations that organize and produce mature cortical neuron subtypes. Furthermore, cerebral organoids are shown to recapitulate features of human cortical development, namely characteristic progenitor zone organization with abundant outer radial glial stem cells. Finally, we use RNA interference and patient-specific induced pluripotent stem cells to model microcephaly, a disorder that has been difficult to recapitulate in mice. We demonstrate premature neuronal differentiation in patient organoids, a defect that could help to explain the disease phenotype. Together, these data show that three-dimensional organoids can recapitulate development and disease even in this most complex human tissue.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.


  1. 1.

    & The cell biology of neurogenesis. Nature Rev. Mol. Cell Biol. 6, 777–788 (2005)

  2. 2.

    , & Contributions of cortical subventricular zone to the development of the human cerebral cortex. J. Comp. Neurol. 491, 109–122 (2005)

  3. 3.

    et al. OSVZ progenitors of human and ferret neocortex are epithelial-like and expand by integrin signaling. Nature Neurosci. 13, 690–699 (2010)

  4. 4.

    , , & Neurogenic radial glia in the outer subventricular zone of human neocortex. Nature 464, 554–561 (2010)

  5. 5.

    , , , & Unique morphological features of the proliferative zones and postmitotic compartments of the neural epithelium giving rise to striate and extrastriate cortex in the monkey. Cereb. Cortex 12, 37–53 (2002)

  6. 6.

    , & Oblique radial glial divisions in the developing mouse neocortex induce self-renewing progenitors outside the germinal zone that resemble primate outer subventricular zone progenitors. J. Neurosci. 31, 3683–3695 (2011)

  7. 7.

    , & Development and evolution of the human neocortex. Cell 146, 18–36 (2011)

  8. 8.

    & Cortical progenitor expansion, self-renewal and neurogenesis — a polarized perspective. Curr. Opin. Neurobiol. 21, 23–35 (2011)

  9. 9.

    , , & What primary microcephaly can tell us about brain growth. Trends Mol. Med. 12, 358–366 (2006)

  10. 10.

    , & Cdk5rap2 exposes the centrosomal root of microcephaly syndromes. Trends Cell Biol. 21, 470–480 (2011)

  11. 11.

    et al. CDK5RAP2 regulates centriole engagement and cohesion in mice. Dev. Cell 18, 913–926 (2010)

  12. 12.

    et al. Cdk5rap2 regulates centrosome function and chromosome segregation in neuronal progenitors. Development 137, 1907–1917 (2010)

  13. 13.

    et al. Mutations in mouse Aspm (abnormal spindle-like microcephaly associated) cause not only microcephaly but also major defects in the germline. Proc. Natl Acad. Sci. USA 107, 16595–16600 (2010)

  14. 14.

    et al. MCPH1 regulates the neuroprogenitor division mode by coupling the centrosomal cycle with mitotic entry through the Chk1-Cdc25 pathway. Nature Cell Biol. 13, 1325–1334 (2011)

  15. 15.

    et al. Single Lgr5 stem cells build crypt–villus structures in vitro without a mesenchymal niche. Nature 459, 262–265 (2009)

  16. 16.

    et al. Self-formation of functional adenohypophysis in three-dimensional culture. Nature 480, 57–62 (2011)

  17. 17.

    et al. Self-formation of optic cups and storable stratified neural retina from human ESCs. Cell Stem Cell 10, 771–785 (2012)

  18. 18.

    et al. Self-organizing optic-cup morphogenesis in three-dimensional culture. Nature 472, 51–56 (2011)

  19. 19.

    & Self-formation of layered neural structures in three-dimensional culture of ES cells. Curr. Opin. Neurobiol. 22, 768–777 (2012)

  20. 20.

    et al. Self-organized formation of polarized cortical tissues from ESCs and its active manipulation by extrinsic signals. Cell Stem Cell 3, 519–532 (2008)

  21. 21.

    et al. Subregional specification of embryonic stem cell-derived ventral telencephalic tissues by timed and combinatory treatment with extrinsic signals. J. Neurosci. 31, 1919–1933 (2011)

  22. 22.

    et al. Ontogeny-recapitulating generation and tissue integration of ES cell-derived Purkinje cells. Nature Neurosci. 13, 1171–1180 (2010)

  23. 23.

    et al. Modeling human cortical development in vitro using induced pluripotent stem cells. Proc. Natl Acad. Sci. USA 109, 12770–12775 (2012)

  24. 24.

    & Differentiation of neuroepithelia from human embryonic stem cells. Methods Mol. Biol. 549, 51–58 (2009)

  25. 25.

    Mapping the human brain: past, present, and future. Trends Neurosci. 18, 471–474 (1995)

  26. 26.

    et al. Tbr1 regulates regional and laminar identity of postmitotic neurons in developing neocortex. Proc. Natl Acad. Sci. USA 107, 13129–13134 (2010)

  27. 27.

    et al. MicroRNA-mediated conversion of human fibroblasts to neurons. Nature 476, 228–231 (2011)

  28. 28.

    et al. An essential switch in subunit composition of a chromatin remodeling complex during neural development. Neuron 55, 201–215 (2007)

  29. 29.

    & Cell biological regulation of division fate in vertebrate neuroepithelial cells. Dev. Dyn. 240, 1865–1879 (2011)

  30. 30.

    & Cleavage orientation and the asymmetric inheritance of Notch1 immunoreactivity in mammalian neurogenesis. Cell 82, 631–641 (1995)

  31. 31.

    et al. Neuroepithelial progenitors undergo LGN-dependent planar divisions to maintain self-renewability during mammalian neurogenesis. Nature Cell Biol. 10, 93–101 (2008)

  32. 32.

    et al. Neuroepithelial stem cell proliferation requires LIS1 for precise spindle orientation and symmetric division. Cell 132, 474–486 (2008)

  33. 33.

    et al. Mouse inscuteable induces apical-basal spindle orientation to facilitate intermediate progenitor generation in the developing neocortex. Neuron 72, 269–284 (2011)

  34. 34.

    Proliferative characteristics of the ependymal layer during the early development of the mouse neocortex: a pilot study based on recording the number, location and plane of cleavage of mitotic figures. J. Anat. 116, 67–91 (1973)

  35. 35.

    Quantitative studies of mitoses in fetal rat brain: orientations of the spindles. Brain Res. 428, 143–146 (1987)

  36. 36.

    , , & Mitotic spindle orientation predicts outer radial glial cell generation in human neocortex. Nature Commun 4, 1665 (2013)

  37. 37.

    et al. Tbr1 regulates differentiation of the preplate and layer 6. Neuron 29, 353–366 (2001)

  38. 38.

    & Making sense of the multiple MAP-2 transcripts and their role in the neuron. Mol. Neurobiol. 16, 149–162 (1998)

  39. 39.

    Cajal-Retzius cells, Reelin, and the formation of layers. Curr. Opin. Neurobiol. 8, 570–575 (1998)

  40. 40.

    & Nucleokinesis in neuronal migration. Neuron 46, 383–388 (2005)

  41. 41.

    et al. An intrinsic mechanism of corticogenesis from embryonic stem cells. Nature 455, 351–357 (2008)

  42. 42.

    & Growth and targeting of subplate axons and establishment of major cortical pathways. J. Neurosci. 12, 1194–1211 (1992)

  43. 43.

    Further tales of the midline. Curr. Opin. Neurobiol. 21, 68–75 (2011)

  44. 44.

    , , & The functional microarchitecture of the mouse barrel cortex. PLoS Biol. 5, e189 (2007)

  45. 45.

    et al. A centrosomal mechanism involving CDK5RAP2 and CENPJ controls brain size. Nature Genet. 37, 353–355 (2005)

  46. 46.

    et al. A novel nonsense CDK5RAP2 mutation in a Somali child with primary microcephaly and sensorineural hearing loss. Am. J. Med. Genet. 158A, 2577–2582 (2012)

  47. 47.

    & Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126, 663–676 (2006)

  48. 48.

    , & Generation of germline-competent induced pluripotent stem cells. Nature 448, 313–317 (2007)

  49. 49.

    et al. A ROCK inhibitor permits survival of dissociated human embryonic stem cells. Nature Biotechnol. 25, 681–686 (2007)

  50. 50.

    & Directed differentiation of neural-stem cells and subtype-specific neurons from hESCs. Methods Mol. Biol. 636, 123–137 (2010)

  51. 51.

    & Electroporation and RNA interference in the rodent retina in vivo and in vitro. Proc. Natl Acad. Sci. USA 101, 16–22 (2004)

  52. 52.

    et al. Retinoic acid from the meninges regulates cortical neuron generation. Cell 139, 597–609 (2009)

  53. 53.

    , & Culture of mouse embryonic stem cells. Curr. Protoc. Stem Cell Biol. 1, Unit–1C.4 (2008)

Download references


We are grateful to members of the Knoblich laboratory for technical expertise and feedback, A. Peer, P. Moeseneder and N. Corsini for experimental support and M. Repic for help with establishing organoid electroporations. We also thank the Stem Cell and BioOptics core facilities of IMBA/IMP for technical support. We would especially like to thank the patients and their families for participating in this study. We would also like to thank S. McGurk for providing control MRI images. M.A.L. received funding from an EMBO post-doctoral fellowship and a Helen Hay Whitney post-doctoral fellowship. Work in A.P.J.’s laboratory is supported by the Medical Research Council, a starter grant from the European Research Council (ERC) and the Lister Institute for Preventative Medicine. This research was also supported in part by Wellcome Trust grant WT098051. Work in J.A.K.’s laboratory is supported by the Austrian Academy of Sciences, the Austrian Science Fund (FWF) (projects Z153-B09 and I552-B19) and an advanced grant from ERC.

Author information


  1. Institute of Molecular Biotechnology of the Austrian Academy of Science (IMBA), Vienna 1030, Austria

    • Madeline A. Lancaster
    • , Magdalena Renner
    • , Daniel Wenzel
    • , Josef M. Penninger
    •  & Juergen A. Knoblich
  2. MRC Human Genetics Unit, Institute of Genetics and Molecular Medicine, University of Edinburgh, Edinburgh EH4 2XU, UK

    • Carol-Anne Martin
    • , Louise S. Bicknell
    •  & Andrew P. Jackson
  3. Wellcome Trust Sanger Institute, Cambridge CB10 1SA, UK

    • Matthew E. Hurles
  4. Department of Clinical Genetics, St. George’s University, London SW17 0RE, UK

    • Tessa Homfray


  1. Search for Madeline A. Lancaster in:

  2. Search for Magdalena Renner in:

  3. Search for Carol-Anne Martin in:

  4. Search for Daniel Wenzel in:

  5. Search for Louise S. Bicknell in:

  6. Search for Matthew E. Hurles in:

  7. Search for Tessa Homfray in:

  8. Search for Josef M. Penninger in:

  9. Search for Andrew P. Jackson in:

  10. Search for Juergen A. Knoblich in:


M.A.L. and J.A.K. conceived the project and experimental design and wrote the manuscript. M.A.L. performed experiments and analysed data. M.R., C.-A.M. and D.W. performed experiments and analysed data under the supervision of J.A.K., J.M.P. and A.P.J. L.S.B., M.E.H. and T.H. performed patient diagnosis and provided MRIs coordinated by A.P.J. J.A.K. directed and supervised the project.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Juergen A. Knoblich.

Extended data

Supplementary information

PDF files

  1. 1.

    Supplementary Information

    This file contains Supplementary Text.


  1. 1.

    Interkinetic nuclear migration in cerebral organoids

    Live imaging of GFP electroporated organoid revealing movement of nuclei along apical and basal processes of RG. Arrow marks one RG in particular with clear IKNM. Time shown in hrs:min.

  2. 2.

    Calcium surges in neurons of cerebral organoids

    Live imaging of Fluo-4 signal in a human cerebral organoid revealing spontaneous calcium surges in individual neurons (arrows). Time shown in min:sec.

  3. 3.

    False colour heat map of spontaneous neural activity

    False colour heat map of a zoomed in region of Supplemental Video 2 showing spontaneous calcium surges. Time shown in min:sec.

About this article

Publication history





Further reading


By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.