Abstract
Adeno-associated virus serotype 9 (AAV9)-mediated gene transfer has been reported in central nervous system (CNS) and peripheral tissues. The current study compared the pattern of expression of Green Fluorescent Protein (GFP) across the mouse CNS and selected peripheral tissues after intrathecal (i.t.) or intravenous (i.v.) delivery of equivalent doses of single-stranded AAV9 vector. After i.t. delivery, GFP immunoreactivity (-ir) was observed in spinal neurons, primary afferent fibers and corresponding primary sensory neurons at all spinal levels. Robust transduction was seen in small and large dorsal root ganglion (DRG) neurons as well as trigeminal and vagal primary afferent neurons. Transduction efficiency in sensory ganglia was substantially lower in i.v. treated mice. In brain, i.v. delivery yielded GFP-immunoreactivity (-ir) primarily in spinal trigeminal tract, pituitary, and scattered isolated neurons and astrocytes. In contrast, after i.t. delivery, GFP-ir was widespread throughout CNS, with greater intensity and more abundant neuropil-like staining at 6 weeks compared to 3 weeks. Brain regions with prominent GFP-ir included cranial nerve nuclei, ventral pons, cerebellar cortex, hippocampus, pituitary, choroid plexus, and selected nuclei of midbrain, thalamus and hypothalamus. In cortex, GFP-ir was associated with blood vessels, and was seen in both neurons and astrocytes. In the periphery, GFP-ir in colon and ileum was present in the enteric nervous system in both i.v. and i.t. treated mice. Liver and adrenal cortex, but not adrenal medulla, also showed abundant GFP-ir after both routes of delivery. In summary, i.t. delivery yielded higher transduction efficiency in sensory neurons and the CNS. The observation of comparable gene transfer to peripheral tissues using the two routes indicates that a component of i.t. delivered vector is redistributed from the subarachnoid space to the systemic circulation.
Highlights
Adeno-associated virus serotype 9 (AAV9) vector has engendered considerable interest in its therapeutic development for treating neurological disorders (Dayton et al, 2012), in part because it has been heralded as capable of traversing the blood brain barrier to target the central nervous system (CNS), where it has been shown to transduce astrocytes and neurons (Foust et al, 2009; Gray et al, 2011; Samaranch et al, 2013)
I.t. delivery of full-strength AAV9-green fluorescent protein (GFP) resulted in substantial transduction of small-diameter dorsal root ganglion (DRG) neurons (64 ± 17% of all small-diameter neurons, range 27–94%; Figure 4B), which was illustrated by the frequent colocalization of GFP-ir with the markers of peptidergic and non-peptidergic small-diameter neurons, CGRP and P2X3 (Figures 2A–C)
The present analysis of the biodistribution of single-stranded AAV9-GFP following intrathecal or intravenous delivery yielded several important observations related to AAV9-mediated gene transfer in the central and peripheral nervous system
Summary
Adeno-associated virus serotype 9 (AAV9) vector has engendered considerable interest in its therapeutic development for treating neurological disorders (Dayton et al, 2012), in part because it has been heralded as capable of traversing the blood brain barrier to target the central nervous system (CNS), where it has been shown to transduce astrocytes and neurons (Foust et al, 2009; Gray et al, 2011; Samaranch et al, 2013). Whereas the intravenous route of delivery of AAV9 has been examined extensively and successfully utilized in models of neurological disorders (Foust et al., 2009; Fu et al, 2011; Ruzo et al, 2012; Garg et al, 2013; Shen et al, 2013; Yamashita et al, 2013; Elmallah et al, 2014), AAV9 administration into the CSF has been employed less frequently in rodents (Snyder et al, 2011; Haurigot et al, 2013; Hirai et al, 2014), and the biodistribution of the vector following intrathecal delivery has not been fully characterized This direct CNS route of administration offers advantages such as reduction in the vector dose required for CNS transduction, and potentially restricted distribution, minimizing off-target effects (Gray et al, 2011, 2013; Snyder et al, 2011; Samaranch et al, 2013). AAV9-mediated transduction of primary sensory neurons has been previously reported in rodents and Frontiers in Neuroanatomy www.frontiersin.org
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