Skip to main content
SpringerLink
Log in

HiRet/NeuRet Vectors: Lentiviral System for Highly Efficient Gene Transfer Through Retrograde Axonal Transport

  • Protocol
  • First Online:
Vectorology for Optogenetics and Chemogenetics

Part of the book series: Neuromethods ((NM,volume 195))

  • 379 Accesses

Abstract

Lentiviral vectors are used for a wide range of research applications in the field of neuroscience. Notably, lentiviral tropism can be manipulated by genetically engineering envelope glycoproteins that are essential for vector transduction. The structural and functional analyses of specific neural pathways are crucial to understanding the neural mechanisms underlying brain functions controlled through complex neural circuits. Viral vectors that produce gene transfer via retrograde axonal transport offer a powerful tool for the analysis of neural pathways. We have succeeded in developing novel types of lentiviral vectors for retrograde gene transfer, named “highly efficient retrograde gene transfer” (HiRet) and “neuron-specific retrograde gene transfer” (NeuRet) vectors, by pseudotyping human immunodeficiency virus type 1 with fusion envelope glycoproteins composed of rabies virus glycoprotein segments and vesicular stomatitis virus glycoprotein segments. HiRet/NeuRet vectors show highly efficient retrograde gene transfer in diverse neural pathways in animal models. These vectors have been harnessed for the analysis of specific neural pathways in combination with various genetic approaches for neuromodulation, including optogenetics and chemogenetics. In this chapter, we describe experimental procedures for producing HiRet/NeuRet vectors and injecting them into brain regions, as well as summarize points important to conduct the experiments smoothly and effectively.

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 109.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 139.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 219.99
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

References

  1. Naldini L, Blömer U, Gage FH et al (1996) Efficient transfer, integration, and sustained long-term expression of the transgene in adult rat brains injected with a lentiviral vector. Proc Natl Acad Sci U S A 93:11382–11388

    Article  CAS  Google Scholar 

  2. Mitrophanous K, Yoon S, Rohll J et al (1999) Stable gene transfer to the nervous system using a non-primate lentiviral vector. Gene Ther 6:1808–1818

    Article  CAS  Google Scholar 

  3. Butler SL, Johnson EP, Bushman FD (2002) Human immunodeficiency virus cDNA metabolism: notable stability of two-long terminal repeat circles. J Virol 76:3739–3747

    Article  CAS  Google Scholar 

  4. Thomas CE, Ehrhardt A, Kay MA (2003) Progress and problems with the use of viral vectors for gene therapy. Nat Rev Genet 4:346–358

    Article  CAS  Google Scholar 

  5. Kafri T (2004) Gene delivery by lentivirus vectors an overview. Methods Mol Biol 246:367–390

    CAS  Google Scholar 

  6. Wong LF, Goodhead L, Prat C et al (2006) Lentivirus-mediated gene transfer to the central nervous system: therapeutic and research applications. Hum Gene Ther 17:1–9

    Article  CAS  Google Scholar 

  7. Cockrell AS, Kafri T (2007) Gene delivery by lentivirus vectors. Mol Biotechnol 36:184–204

    Article  CAS  Google Scholar 

  8. Nectow AR, Nestler EJ (2020) Viral tools for neuroscience. Nat Rev Neurosci 21:669–681

    Article  CAS  Google Scholar 

  9. Cronin J, Zhang XY, Reiser J (2005) Altering the tropism of lentiviral vectors through pseudotyping. Curr Gene Ther 5:387–398

    Article  CAS  Google Scholar 

  10. Kobayashi K, Inoue KI, Tanabe S et al (2017) Pseudotyped lentiviral vectors for retrograde gene delivery into target brain regions. Front Neuroanat 11:65

    Article  Google Scholar 

  11. Kobayashi K, Kato S, Kobayashi K (2018) Genetic manipulation of specific neural circuits by use of a viral vector system. J Neural Transm (Vienna) 125:67–75

    Article  Google Scholar 

  12. Kato S, Kobayashi K (2020) Pseudotyped lentiviral vectors for tract-targeting and application for the functional control of selective neural circuits. J Neurosci Methods 344:108854

    Article  CAS  Google Scholar 

  13. Joshi S, Joshi RL (1996) Molecular biology of human immunodeficiency virus type-1. Transfus Sci 17:351–378

    Article  CAS  Google Scholar 

  14. Nielsen MH, Pedersen FS, Kjems J (2005) Molecular strategies to inhibit HIV-1 replication. Retrovirology 2:10

    Article  Google Scholar 

  15. Pluta K, Kacprzak MM (2009) Use of HIV as a gene transfer vector. Acta Biochim Pol 56:531–595

    Article  CAS  Google Scholar 

  16. Haery L, Deverman BE, Matho KS et al (2019) Adeno-associated virus technologies and methods for targeted neuronal manipulation. Front Neuroanat 13:93

    Article  CAS  Google Scholar 

  17. Callaway EM, Luo L (2015) Monosynaptic circuit tracing with glycoprotein-deleted rabies viruses. J Neurosci 35:8979–8985

    Article  CAS  Google Scholar 

  18. Junyent F, Kremer EJ (2015) CAV-2--why a canine virus is a neurobiologist's best friend. Curr Opin Pharmacol 24:86–93

    Article  CAS  Google Scholar 

  19. Pomeranz LE, Reynolds AE, Hengartner CJ (2005) Molecular biology of pseudorabies virus: impact on neurovirology and veterinary medicine. Microbiol Mol Biol Rev 69:462–500

    Article  CAS  Google Scholar 

  20. Fenno LE, Mattis J, Ramakrishnan C et al (2014) Targeting cells with single vectors using multiple-feature Boolean logic. Nat Methods 11:763–772

    Article  CAS  Google Scholar 

  21. Kato S, Kobayashi K, Inoue K et al (2011) A lentiviral strategy for highly efficient retrograde gene transfer by pseudotyping with fusion envelope glycoprotein. Hum Gene Ther 22:197–206

    Article  CAS  Google Scholar 

  22. Kato S, Kuramochi M, Takasumi K et al (2011) Neuron-specific gene transfer through retrograde transport of lentiviral vector pseudotyped with a novel type of fusion envelope glycoprotein. Hum Gene Ther 22:1511–1523

    Article  CAS  Google Scholar 

  23. Kato S, Kobayashi K, Kobayashi K (2014) Improved transduction efficiency of a lentiviral vector for neuron-specific retrograde gene transfer by optimizing the junction of fusion envelope glycoprotein. J Neurosci Methods 227:151–158

    Article  CAS  Google Scholar 

  24. Kato S, Sugawara M, Kobayashi K et al (2019) Enhancement of the transduction efficiency of a lentiviral vector for neuron-specific retrograde gene delivery through the point mutation of fusion glycoprotein type E. J Neurosci Methods 311:147–155

    Article  CAS  Google Scholar 

  25. Tanabe S, Inoue KI, Tsuge H et al (2017) The use of an optimized chimeric envelope glycoprotein enhances the efficiency of retrograde gene transfer of a pseudotyped lentiviral vector in the primate brain. Neurosci Res 120:45–52

    Article  CAS  Google Scholar 

  26. Tanabe S, Uezono S, Tsuge H et al (2019) A note on retrograde gene transfer efficiency and inflammatory response of lentiviral vectors pseudotyped with FuG-E vs. FuG-B2 glycoproteins. Sci Rep 9:3567

    Article  Google Scholar 

  27. Matsuda T, Hiyama TY, Niimura F et al (2017) Distinct neural mechanisms for the control of thirst and salt appetite in the subfornical organ. Nat Neurosci 20:230–241

    Article  CAS  Google Scholar 

  28. Morishima M, Kobayashi K, Kato S et al (2017) Segregated excitatory-inhibitory recurrent subnetworks in layer 5 of the rat frontal cortex. Cereb Cortex 27:5846–5857

    Article  Google Scholar 

  29. Kato S, Fukabori R, Nishizawa K et al (2018) Action selection and flexible switching controlled by the intralaminar thalamic neurons. Cell Rep 22:2370–2382

    Article  CAS  Google Scholar 

  30. Nomura K, Hiyama TY, Sakuta H et al (2019) [Na+] increases in body fluids sensed by central Nax induce sympathetically mediated blood pressure elevations via H+-dependent activation of ASIC1a. Neuron 101:60–75

    Article  CAS  Google Scholar 

  31. Isa K, Sooksawate T, Kobayashi K et al (2020) Dissecting the tectal output channels for orienting and defense responses. eNeuro 7:ENEURO.0271-20.2020

    Article  Google Scholar 

  32. Ishida A, Kobayashi K, Ueda Y et al (2019) Dynamic interaction between cortico-brainstem pathways during training-induced recovery in stroke model rats. J Neurosci 39:7306–7320

    Article  CAS  Google Scholar 

  33. Hayashi T, Akikawa R, Kawasaki K et al (2020) Macaques exhibit implicit gaze bias anticipating others' false-belief-driven actions via medial prefrontal cortex. Cell Rep 30:4433–4444

    Article  CAS  Google Scholar 

  34. Kato S, Kuramochi M, Kobayashi K et al (2011) Selective neural pathway targeting reveals key roles of thalamostriatal projection in the control of visual discrimination. J Neurosci 31:17169–17179

    Article  CAS  Google Scholar 

  35. Kobayashi K, Sano H, Kato S et al (2016) Survival of corticostriatal neurons by Rho/Rho-kinase signaling pathway. Neurosci Lett 630:45–52

    Article  CAS  Google Scholar 

  36. Paxinos G, Franklin KBJ (2008) The mouse brain in stereotaxic coordinates, 3rd edn. Academic Press, San Diego

    Google Scholar 

  37. Kato S, Inoue K, Kobayashi K et al (2007) Efficient gene transfer via retrograde transport in rodent and primate brains using a human immunodeficiency virus type 1-based vector pseudotyped with rabies virus glycoprotein. Hum Gene Ther 18:1141–1151

    Article  CAS  Google Scholar 

  38. Inoue K, Koketsu D, Kato S et al (2012) Immunotoxin-mediated tract targeting in the primate brain: selective elimination of the cortico-subthalamic “hyperdirect” pathway. PLoS One 7:e39149

    Article  CAS  Google Scholar 

  39. Kinoshita M, Matsui R, Kato S et al (2012) Genetic dissection of the circuit for hand dexterity in primates. Nature 487:235–238

    Article  CAS  Google Scholar 

  40. Sooksawate T, Isa K, Matsui R et al (2013) Viral vector-mediated selective and reversible blockade of the pathway for visual orienting in mice. Front Neural Circuits 7:162

    Article  CAS  Google Scholar 

  41. Ishida A, Isa K, Umeda T et al (2016) Causal link between the cortico-rubral pathway and functional recovery through forced impaired limb use in rats with stroke. J Neurosci 36:455–467

    Article  Google Scholar 

  42. Tohyama T, Kinoshita M, Kobayashi K et al (2017) Contribution of propriospinal neurons to recovery of hand dexterity after corticospinal tract lesions in monkeys. Proc Natl Acad Sci U S A 114:604–609

    Article  CAS  Google Scholar 

  43. Kinoshita M, Kato R, Isa K et al (2019) Dissecting the circuit for blindsight to reveal the critical role of pulvinar and superior colliculus. Nat Commun 10:135

    Article  Google Scholar 

  44. Vancraeyenest P, Arsenault JT, Li X et al (2020) Selective mesoaccumbal pathway inactivation affects motivation but not reinforcement-based learning in macaques. Neuron 108:568–581

    Article  CAS  Google Scholar 

Download references

Acknowledgments

This work was supported by a grant-in-aid from Japan Agency for Medical Research and Development (JP17dm0207052 to Ka.K.). We thank Dr. R. Fukabori for providing photographs and Dr. Y. Iguchi for his critical reading of this manuscript.

Author information

Authors and Affiliations

  1. Section of Viral Vector Development, National Institute for Physiological Sciences, Okazaki, Japan

    Kenta Kobayashi

  2. SOKENDAI (The Graduate University for Advanced Studies), Hayama, Japan

    Kenta Kobayashi

  3. Department of Molecular Genetics, Institute of Biomedical Sciences, Fukushima Medical University School of Medicine, Fukushima, Japan

    Shigeki Kato & Kazuto Kobayashi

Authors
  1. Kenta Kobayashi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Shigeki Kato
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Kazuto Kobayashi
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Kazuto Kobayashi .

Editor information

Editors and Affiliations

  1. NIMH, National Institutes of Health, Behesda, MD, USA

    Mark A.G. Eldridge

  2. Department of Neurology, School of Medicine, and Emory National Primate Research Center, Emory University, Atlanta, GA, USA

    Adriana Galvan

Rights and permissions

Reprints and permissions

Copyright information

© 2023 This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Kobayashi, K., Kato, S., Kobayashi, K. (2023). HiRet/NeuRet Vectors: Lentiviral System for Highly Efficient Gene Transfer Through Retrograde Axonal Transport. In: Eldridge, M.A., Galvan, A. (eds) Vectorology for Optogenetics and Chemogenetics. Neuromethods, vol 195. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2918-5_2

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-2918-5_2

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2917-8

  • Online ISBN: 978-1-0716-2918-5

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation