Abstract
Viral vector-mediated gene transfer, especially using adeno-associated viral (AAV) vectors, is a powerful strategy to manipulate gene expression in vivo in injured neurons. In this chapter we provide two methods to efficiently transduce dorsal root ganglion (DRG) neurons in vivo using AAV. We describe detailed procedures to perform direct injections into specific DRG and delivery via the intrathecal space to transduce the lumbar DRG. Finally, we discuss the specific advantages and disadvantages of these two methods of delivery. The main advantages of direct injection are that high transduction rates can be achieved in specific ganglia (L4/L5) with low amounts (μl) of a viral vector stock; however, the procedure is complex, invasive, and time-consuming. Intrathecal injection has the advantage of being a fast and simple method to transduce multiple DRG bilaterally, and involves no surgical manipulation of the DRG. However, intrathecal delivery does require much larger amounts of viral stock (10–20 μl) and has the disadvantage that viral particles will leak from the cerebrospinal fluid to the spinal cord and/or peripheral tissues.
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References
Gaudet D, Methot J, Dery S et al (2013) Efficacy and long-term safety of alipogene tiparvovec (AAV1-LPLS447X) gene therapy for lipoprotein lipase deficiency: an open-label trial. Gene Ther 20:361–369
Kaplitt MG, Feigin A, Tang C et al (2007) Safety and tolerability of gene therapy with an adeno-associated virus (AAV) borne GAD gene for Parkinson’s disease: an open label, phase I trial. Lancet 369:2097–2105
Hermens WT, Ter BO, Dijkhuizen PA et al (1999) Purification of recombinant adeno-associated virus by iodixanol gradient ultracentrifugation allows rapid and reproducible preparation of vector stocks for gene transfer in the nervous system. Hum Gene Ther 10:1885–1891
Kaplitt MG, Leone P, Samulski RJ et al (1994) Long-term gene expression and phenotypic correction using adeno-associated virus vectors in the mammalian brain. Nat Genet 8:148–154
McCown TJ, Xiao X, Li J et al (1996) Differential and persistent expression patterns of CNS gene transfer by an adeno-associated virus (AAV) vector. Brain Res 713:99–107
Peel AL, Zolotukhin S, Schrimsher GW et al (1997) Efficient transduction of green fluorescent protein in spinal cord neurons using adeno-associated virus vectors containing cell type-specific promoters. Gene Ther 4:16–24
Hoke A (2006) Mechanisms of Disease: what factors limit the success of peripheral nerve regeneration in humans? Nat Clin Pract Neurol 2:448–454
Macgillavry HD, Stam FJ, Sassen MM et al (2009) NFIL3 and cAMP response element-binding protein form a transcriptional feedforward loop that controls neuronal regeneration-associated gene expression. J Neurosci 29:15542–15550
Stam FJ, Macgillavry HD, Armstrong NJ et al (2007) Identification of candidate transcriptional modulators involved in successful regeneration after nerve injury. Eur J Neurosci 25:3629–3637
Geeven G, Macgillavry HD, Eggers R et al (2011) LLM3D: a log-linear modeling-based method to predict functional gene regulatory interactions from genome-wide expression data. Nucleic Acids Res 39:5313–5327
Arthur-Farraj PJ, Latouche M, Wilton DK et al (2012) c-Jun reprograms Schwann cells of injured nerves to generate a repair cell essential for regeneration. Neuron 75:633–647
Li S, Liu Q, Wang Y et al (2013) Differential gene expression profiling and biological process analysis in proximal nerve segments after sciatic nerve transection. PLoS One 8:e57000
Verhaagen J, Van Kesteren RE, Bossers KA et al (2012) Molecular target discovery for neural repair in the functional genomics era. Handb Clin Neurol 109:595–616
Beutler AS, Reinhardt M (2009) AAV for pain: steps towards clinical translation. Gene Ther 16:461–469
Mingozzi F, High KA (2011) Therapeutic in vivo gene transfer for genetic disease using AAV: progress and challenges. Nat Rev Genet 12:341–355
Ginn SL, Alexander IE, Edelstein ML et al (2013) Gene therapy clinical trials worldwide to 2012 - an update. J Gene Med 15:65–77
Mason MR, Ehlert EM, Eggers R et al (2010) Comparison of AAV serotypes for gene delivery to dorsal root ganglion neurons. Mol Ther 18:715–724
Naldini L, Blomer U, Gallay P et al (1996) In vivo gene delivery and stable transduction of nondividing cells by a lentiviral vector. Science 272:263–267
Yu H, Fischer G, Jia G et al (2011) Lentiviral gene transfer into the dorsal root ganglion of adult rats. Mol Pain 7:63
Storek B, Harder NM, Banck MS et al (2006) Intrathecal long-term gene expression by self-complementary adeno-associated virus type 1 suitable for chronic pain studies in rats. Mol Pain 2:4
Storek B, Reinhardt M, Wang C et al (2008) Sensory neuron targeting by self-complementary AAV8 via lumbar puncture for chronic pain. Proc Natl Acad Sci U S A 105:1055–1060
Towne C, Pertin M, Beggah AT et al (2009) Recombinant adeno-associated virus serotype 6 (rAAV2/6)-mediated gene transfer to nociceptive neurons through different routes of delivery. Mol Pain 5:52
Xu Q, Chou B, Fitzsimmons B et al (2012) In vivo gene knockdown in rat dorsal root ganglia mediated by self-complementary adeno-associated virus serotype 5 following intrathecal delivery. PLoS One 7:e32581
Malkmus SA, Yaksh TL (2004) Intrathecal catheterization and drug delivery in the rat. Methods Mol Med 99:109–121
Yaksh TL, Rudy TA (1976) Chronic catheterization of the spinal subarachnoid space. Physiol Behav 17:1031–1036
Vulchanova L, Schuster DJ, Belur LR et al (2010) Differential adeno-associated virus mediated gene transfer to sensory neurons following intrathecal delivery by direct lumbar puncture. Mol Pain 6:31
Parikh P, Hao Y, Hosseinkhani M et al (2011) Regeneration of axons in injured spinal cord by activation of bone morphogenetic protein/Smad1 signaling pathway in adult neurons. Proc Natl Acad Sci U S A 108:E99–E107
Asato F, Butler M, Blomberg H et al (2001) Distribution of intrathecal catheter positions in rat. Acta Anaesthesiol Scand 45:608–611
Acknowledgements
The authors are grateful for research funding provided by the International Spinal Research Trust and the Dutch Organization for Scientific Research (NWO).
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Fagoe, N.D., Eggers, R., Verhaagen, J., Mason, M.R.J. (2015). Gene Delivery to Neurons of the Dorsal Root Ganglia Using Adeno-Associated Viral Vectors. In: Bo, X., Verhaagen, J. (eds) Gene Delivery and Therapy for Neurological Disorders. Neuromethods, vol 98. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2306-9_7
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DOI: https://doi.org/10.1007/978-1-4939-2306-9_7
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