Neurochemical Research

, Volume 36, Issue 4, pp 583–593 | Cite as

Automated Tracing of Horizontal Neuron Processes During Retinal Development

  • Ryan A. KerekesEmail author
  • Rodrigo A. P. Martins
  • Denise Davis
  • Mahmut Karakaya
  • Shaun Gleason
  • Michael A. Dyer
Original Paper


In the developing mammalian retina, horizontal neurons undergo a dramatic reorganization of their processes shortly after they migrate to their appropriate laminar position. This is an important process because it is now understood that the apical processes are important for establishing the regular mosaic of horizontal cells in the retina and proper reorganization during lamination is required for synaptogenesis with photoreceptors and bipolar neurons. However, this process is difficult to study because the analysis of horizontal neuron anatomy is labor intensive and time-consuming. In this paper, we present a computational method for automatically tracing the three-dimensional (3-D) dendritic structure of horizontal retinal neurons in two-photon laser scanning microscope (TPLSM) imagery. Our method is based on 3-D skeletonization and is thus able to preserve the complex structure of the dendritic arbor of these cells. We demonstrate the effectiveness of our approach by comparing our tracing results against two sets of semi-automated traces over a set of 10 horizontal neurons ranging in age from P1 to P5. We observe an average agreement level of 81% between our automated trace and the manual traces. This automated method will serve as an important starting point for further refinement and optimization.


Horizontal retinal neuron Retinal development Automated tracing Segmentation algorithm 



Supported by grants from the National Institutes of Health (R01EY018599 and R01EY014867); Cancer Center Support CA 21765 from the National Cancer Institute; and grants from the American Cancer Society, the Pew Charitable Trust, Macular Vision Research Foundation and the American Lebanese Syrian Associated Charities. Dr. Dyer is a Howard Hughes Medical Institute Early Career Investigator.


  1. 1.
    Livesey FJ, Cepko CL (2001) Vertebrate neural cell-fate determination: lessons from the retina. Nat Rev Neurosci 2:109–118PubMedCrossRefGoogle Scholar
  2. 2.
    Donovan SL, Dyer MA (2005) Regulation of proliferation during central nervous system development. Semin Cell Dev Biol 16:407–421PubMedCrossRefGoogle Scholar
  3. 3.
    Cepko CL, Austin CP, Yang X et al (1996) Cell fate determination in the vertebrate retina. Proc Natl Acad Sci USA 93:589–595PubMedCrossRefGoogle Scholar
  4. 4.
    Dyer MA (2003) Regulation of proliferation, cell fate specification and differentiation by the homeodomain proteins Prox1, Six3, and Chx10 in the developing retina. Cell Cycle 2:350–357PubMedCrossRefGoogle Scholar
  5. 5.
    Dyer MA, Cepko CL (2001) Regulating proliferation during retinal development. Nat Rev Neurosci 2:333–342PubMedCrossRefGoogle Scholar
  6. 6.
    Belliveau MJ, Cepko CL (1999) Extrinsic and intrinsic factors control the genesis of amacrine and cone cells in the rat retina. Development 126:555–566PubMedGoogle Scholar
  7. 7.
    Donovan SL, Dyer MA (2004) Developmental defects in Rb-deficient retinae. Vision Res 44:3323–3333PubMedCrossRefGoogle Scholar
  8. 8.
    Johnson DA, Donovan SL, Dyer MA (2006) Mosaic deletion of Rb arrests rod differentiation and stimulates ectopic synaptogenesis in the mouse retina. J Comp Neurol 498:112–128PubMedCrossRefGoogle Scholar
  9. 9.
    Al-Kofahi O, Radke RJ, Roysam B et al (2006) Automated semantic analysis of changes in image sequences of neurons in culture. IEEE Trans Biomed Eng 53:1109–1123PubMedCrossRefGoogle Scholar
  10. 10.
    Xiong G, Zhou X, Degterev A et al (2006) Automated neurite labeling and analysis in fluorescence microscopy images. Cytometry A 69:494–505PubMedGoogle Scholar
  11. 11.
    Lee PC, Ching YT, Chang HM, Chiang AS (2008) A semi-automatic method for neuron centerline extraction in confocal microscopic image stack. IEEE International Symposium on Biomedical Imaging 5, 3Google Scholar
  12. 12.
    Losavio BE et al (2008) Live neuron morphology automatically reconstructed from multiphoton and confocal imaging data. J Neurophysiol 100:2422–2429PubMedCrossRefGoogle Scholar
  13. 13.
    Chan TF, Vese LA (2001) Active contours without edges. IEEE Trans Image Process 10:266–277PubMedCrossRefGoogle Scholar
  14. 14.
    Chen Z, Molloi S (2003) Automatic 3D vascular tree construction in CT angiography. Comput Med Imaging Graph 27:469–479PubMedCrossRefGoogle Scholar
  15. 15.
    Borgefors G, Nyström I, Di Baja GS (1999) Computing skeletons in three dimensions. Pattern Recogn 32:1225–1236CrossRefGoogle Scholar
  16. 16.
    Ma CM, Sonka M (1996) A fully parallel 3D thinning algorithm and its applications. Comput Vision Image Underst 64:420–433CrossRefGoogle Scholar
  17. 17.
    He X, Kischell M, Rioult M et al (1998) Three-dimensional thinning algorithm that peels the outmost layer with application to neuron tracing. J Comput Assisted Microsc 10:123–135CrossRefGoogle Scholar
  18. 18.
    Huckfeldt RM, Schubert T, Morgan JL et al (2009) Transient neurites of retinal horizontal cells exhibit columnar tiling via homotypic interactions. Nat Neurosci 12:35–43PubMedCrossRefGoogle Scholar
  19. 19.
    Dyer MA, Cepko CL (2000) Control of Muller glial cell proliferation and activation following retinal injury. Nat Neurosci 3:873–880PubMedCrossRefGoogle Scholar
  20. 20.
    Jones BW et al (2003) Retinal remodeling triggered by photoreceptor degenerations. J Comp Neurol 464:1–16PubMedCrossRefGoogle Scholar
  21. 21.
    Marc RE, Jones BW (2003) Retinal remodeling in inherited photoreceptor degenerations. Mol Neurobiol 28:139–147PubMedCrossRefGoogle Scholar
  22. 22.
    Marc RE, Jones BW, Watt CB et al (2003) Neural remodeling in retinal degeneration. Prog Retin Eye Res 22:607–655PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC (outside the USA)  2011

Authors and Affiliations

  • Ryan A. Kerekes
    • 1
    Email author
  • Rodrigo A. P. Martins
    • 2
  • Denise Davis
    • 3
  • Mahmut Karakaya
    • 4
  • Shaun Gleason
    • 1
  • Michael A. Dyer
    • 3
    • 5
    • 6
  1. 1.Measurement Science and Systems Engineering DivisionOak Ridge National LaboratoryOak RidgeUSA
  2. 2.Instituto de Biofísica Carlos Chagas FilhoUniversidade Federal do Rio de JaneiroRio de JaneiroBrazil
  3. 3.Department of Developmental NeurobiologySt. Jude Children’s Research HospitalMemphisUSA
  4. 4.Department of Computer Science and Electrical EngineeringUniversity of TennesseeKnoxvilleUSA
  5. 5.Department of OphthalmologyUniversity of Tennessee Health Sciences CenterMemphisUSA
  6. 6.Howard Hughes Medical InstituteBaltimoreUSA

Personalised recommendations