Skip to main content
SpringerLink
Log in

Visualizing and Manipulating Olfactory Cilia Through Viral Delivery Coupled with En Face Imaging of Intact OE

  • Protocol
  • First Online:
The Olfactory System

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2710))

Abstract

Olfactory cilia are the obligate transducers of the odorant signal, and thus their study has been a focus of investigation in the olfactory field. Various methodologies have been established to visualize the cilia of olfactory sensory neurons; however, these approaches are limited to static imaging and often lack the ability to resolve individual cilia projecting from solitary neurons in the postnatal mouse. Here we detail a procedure of the visualization of olfactory cilia by ectopic expression of fluorescently tagged proteins. The procedure can be used for the observation and manipulation of the olfactory cilia and ciliary proteins in both static and dynamic conditions.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 139.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Hardcover Book
USD 179.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Similar content being viewed by others

References

  1. Bloom G (1954) Studies on the olfactory epithelium of the frog and the toad with the aid of light and electron microscopy. Z Zellforsch Mikrosk Anat 41(1):89–100. https://doi.org/10.1007/BF00340285

    Article  CAS  PubMed  Google Scholar 

  2. Menco BP, Farbman AI (1985) Genesis of cilia and microvilli of rat nasal epithelia during pre-natal development. I. Olfactory epithelium, qualitative studies. J Cell Sci 78:283–310

    Article  CAS  PubMed  Google Scholar 

  3. Menco BP, Farbman AI (1985) Genesis of cilia and microvilli of rat nasal epithelia during pre-natal development. II. Olfactory epithelium, a morphometric analysis. J Cell Sci 78:311–336

    Article  CAS  PubMed  Google Scholar 

  4. Okano M, Weber AF, Frommes SP (1967) Electron microscopic studies on the distal border of the canine olfactory epithelium. J Ultrastruct Res 17(5):487–502. https://doi.org/10.1016/s0022-5320(67)80137-0

    Article  CAS  PubMed  Google Scholar 

  5. Kavoi B, Makanya A, Hassanali J, Carlsson HE, Kiama S (2010) Comparative functional structure of the olfactory mucosa in the domestic dog and sheep. Ann Anat 192(5):329–337. https://doi.org/10.1016/j.aanat.2010.07.004

    Article  PubMed  Google Scholar 

  6. Kratzing JE (1975) The fine structure of the olfactory and vomeronasal organs of a lizard (Tiliqua scincoides scincoides). Cell Tissue Res 156(2):239–252. https://doi.org/10.1007/BF00221807

    Article  CAS  PubMed  Google Scholar 

  7. Hansen A (2007) Olfactory and solitary chemosensory cells: two different chemosensory systems in the nasal cavity of the American alligator, Alligator mississippiensis. BMC Neurosci 8:64. https://doi.org/10.1186/1471-2202-8-64

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Schulte E, Holl A (1971) Ultrastructure of the olfactory epithelium of Calamoichthys calabaricus J. A. Smoth (Pisces, Brachiopterygii). Z Zellforsch Mikrosk Anat 120(2):261–279. https://doi.org/10.1007/BF00335539

    Article  CAS  PubMed  Google Scholar 

  9. Hansen A, Zielinski BS (2005) Diversity in the olfactory epithelium of bony fishes: development, lamellar arrangement, sensory neuron cell types and transduction components. J Neurocytol 34(3–5):183–208. https://doi.org/10.1007/s11068-005-8353-1

    Article  CAS  PubMed  Google Scholar 

  10. Green WW, Uytingco CR, Ukhanov K, Kolb Z, Moretta J, McIntyre JC et al (2018) Peripheral gene therapeutic rescue of an olfactory ciliopathy restores sensory input, axonal pathfinding, and odor-guided behavior. J Neurosci 38(34):7462–7475. https://doi.org/10.1523/JNEUROSCI.0084-18.2018

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Uytingco CR, Williams CL, Xie C, Shively DT, Green WW, Ukhanov K et al (2019) BBS4 is required for intraflagellar transport coordination and basal body number in mammalian olfactory cilia. J Cell Sci 132(5). https://doi.org/10.1242/jcs.222331

  12. Williams CL, Uytingco CR, Green WW, McIntyre JC, Ukhanov K, Zimmerman AD et al (2017) Gene therapeutic reversal of peripheral olfactory impairment in Bardet-Biedl syndrome. Mol Ther 25(4):904–916. https://doi.org/10.1016/j.ymthe.2017.02.006

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. McIntyre JC, Davis EE, Joiner A, Williams CL, Tsai IC, Jenkins PM et al (2012) Gene therapy rescues cilia defects and restores olfactory function in a mammalian ciliopathy model. Nat Med 18(9):1423–1428. https://doi.org/10.1038/nm.2860

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Ukhanov K, Uytingco C, Green W, Zhang L, Schurmans S, Martens JR (2022) INPP5E controls ciliary localization of phospholipids and the odor response in olfactory sensory neurons. J Cell Sci 135(5). https://doi.org/10.1242/jcs.258364

  15. Xie C, Habif JC, Uytingco CR, Ukhanov K, Zhang L, de Celis C et al (2021) Gene therapy rescues olfactory perception in a clinically relevant ciliopathy model of Bardet-Biedl syndrome. FASEB J 35(9):e21766. https://doi.org/10.1096/fj.202100627R

    Article  CAS  PubMed  Google Scholar 

  16. Oberland S, Neuhaus EM (2014) Whole mount labeling of cilia in the main olfactory system of mice. J Vis Exp 94. https://doi.org/10.3791/52299

  17. Alevra M, Schwartz P, Schild D (2012) Direct measurement of diffusion in olfactory cilia using a modified FRAP approach. PLoS One 7(7):e39628. https://doi.org/10.1371/journal.pone.0039628

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Jenkins PM, Hurd TW, Zhang L, McEwen DP, Brown RL, Margolis B et al (2006) Ciliary targeting of olfactory CNG channels requires the CNGB1b subunit and the kinesin-2 motor protein, KIF17. Curr Biol 16(12):1211–1216. https://doi.org/10.1016/j.cub.2006.04.034

    Article  CAS  PubMed  Google Scholar 

  19. Delling M, DeCaen PG, Doerner JF, Febvay S, Clapham DE (2013) Primary cilia are specialized calcium signalling organelles. Nature 504(7479):311–314. https://doi.org/10.1038/nature12833

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Fan S, Fogg V, Wang Q, Chen XW, Liu CJ, Margolis B (2007) A novel Crumbs3 isoform regulates cell division and ciliogenesis via importin beta interactions. J Cell Biol 178(3):387–398. https://doi.org/10.1083/jcb.200609096

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Hansen JN, Kaiser F, Klausen C, Stüven B, Chong R, Bönigk W et al (2020) Nanobody-directed targeting of optogenetic tools to study signaling in the primary cilium. elife 9. https://doi.org/10.7554/eLife.57907

  22. Iwadate Y (2003) Photolysis of caged calcium in cilia induces ciliary reversal in Paramecium caudatum. J Exp Biol 206(Pt 7):1163–1170. https://doi.org/10.1242/jeb.00219

    Article  CAS  PubMed  Google Scholar 

  23. Williams CL, McIntyre JC, Norris SR, Jenkins PM, Zhang L, Pei Q et al (2014) Direct evidence for BBSome-associated intraflagellar transport reveals distinct properties of native mammalian cilia. Nat Commun 5:5813. https://doi.org/10.1038/ncomms6813

    Article  CAS  PubMed  Google Scholar 

  24. Xie C, Habif JC, Ukhanov K, Uytingco CR, Zhang L, Campbell RJ et al (2022) Reversal of ciliary mechanisms of disassembly rescues olfactory dysfunction in ciliopathies. JCI Insight. https://doi.org/10.1172/jci.insight.158736

  25. Uytingco CR, Martens JR (2019) Intranasal delivery of adenoviral and AAV vectors for transduction of the mammalian peripheral olfactory system. Methods Mol Biol 1950:283–297. https://doi.org/10.1007/978-1-4939-9139-6_17

    Article  CAS  PubMed  Google Scholar 

  26. Corey EA, Ukhanov K, Bobkov YV, McIntyre JC, Martens JR, Ache BW (2021) Inhibitory signaling in mammalian olfactory transduction potentially mediated by Gα. Mol Cell Neurosci 110:103585. https://doi.org/10.1016/j.mcn.2020.103585

    Article  CAS  PubMed  Google Scholar 

  27. McIntyre JC, Joiner AM, Zhang L, Iñiguez-Lluhí J, Martens JR (2015) SUMOylation regulates ciliary localization of olfactory signaling proteins. J Cell Sci 128(10):1934–1945. https://doi.org/10.1242/jcs.164673

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Zhao H, Ivic L, Otaki JM, Hashimoto M, Mikoshiba K, Firestein S (1998) Functional expression of a mammalian odorant receptor. Science 279(5348):237–242. https://doi.org/10.1126/science.279.5348.237

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

We acknowledge Dr. Kirill Ukhanov for providing a representative whole-mount staining image and technical assistance. We thank Dr. Cedric Uytingco for providing a representative TIRF image. This work was supported by the National Institute of Deafness and Other Communication Disorders R01DC019345 (JRM), T32DC015994 (JCH), and F31DC020638 (JCH).

Author information

Author notes

  1. Authors “Julien C. Habif” and “Chao Xie” have equally contributed to this chapter.

Authors and Affiliations

  1. Department of Pharmacology and Therapeutics, University of Florida College of Medicine, Gainesville, FL, USA

    Julien C. Habif, Chao Xie & Jeffrey R. Martens

  2. Center for Smell and Taste, University of Florida College of Medicine, Gainesville, FL, USA

    Julien C. Habif, Chao Xie & Jeffrey R. Martens

Authors
  1. Julien C. Habif
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Chao Xie
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jeffrey R. Martens
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jeffrey R. Martens .

Editor information

Editors and Affiliations

  1. Department of Head and Neck Surgery & Communication Sciences, Duke University School of Medicine, Durham, NC, USA

    Bradley J. Goldstein

  2. Department of Molecular Genetics and Microbiology, Duke University School of Medicine, Durham, NC, USA

    Hiroaki Matsunami

Rights and permissions

Reprints and permissions

Copyright information

© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Habif, J.C., Xie, C., Martens, J.R. (2023). Visualizing and Manipulating Olfactory Cilia Through Viral Delivery Coupled with En Face Imaging of Intact OE. In: Goldstein, B.J., Matsunami, H. (eds) The Olfactory System. Methods in Molecular Biology, vol 2710. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3425-7_1

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-3425-7_1

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3424-0

  • Online ISBN: 978-1-0716-3425-7

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation