Widespread neuron-specific transgene expression in brain and spinal cord following synapsin promoter-driven AAV9 neonatal intracerebroventricular injection.

Neurosci Lett. 2014 Jul 25;576:73-8.

McLean JR1, Smith GA1, Rocha EM1, Hayes MA1, Beagan JA1, Hallett PJ1, Isacson O2.

1Neuroregeneration Institute, McLean Hospital/Harvard Medical School, Belmont, MA, USA.

2Neuroregeneration Institute, McLean Hospital/Harvard Medical School, Belmont, MA, USA. Electronic address: isacson@hms.harvard.edu.


Adeno-associated viral (AAV) gene transfer holds great promise for treating a wide-range of neurodegenerative disorders. The AAV9 serotype crosses the blood-brain barrier and shows enhanced transduction efficiency compared to other serotypes, thus offering advantageous targeting when global transgene expression is required. Neonatal intravenous or intracerebroventricular (i.c.v.) delivery of recombinant AAV9 (rAAV9) have recently proven effective for modeling and treating several rodent models of neurodegenerative disease, however, the technique is associated with variable cellular tropism, making tailored gene transfer a challenge. In the current study, we employ the human synapsin 1 (hSYN1) gene promoter to drive neuron-specific expression of green fluorescent protein (GFP) after neonatal i.c.v. injection of rAAV9 in mice. We observed widespread GFP expression in neurons throughout the brain, spinal cord, and peripheral nerves and ganglia at 6 weeks-of-age. Region-specific quantification of GFP expression showed high neuronal transduction rates in substantia nigra pars reticulata (43.9±5.4%), motor cortex (43.5±3.3%), hippocampus (43.1±2.7%), cerebellum (29.6±2.3%), cervical spinal cord (24.9±3.9%), and ventromedial striatum (16.9±4.3%), among others. We found that 14.6±2.2% of neuromuscular junctions innervating the gastrocnemius muscle displayed GFP immunoreactivity. GFP expression was identified in several neuronal sub-types, including nigral tyrosine hydroxylase (TH)-positive dopaminergic cells, striatal dopamine- and cAMP-regulated neuronal phosphoprotein (DARPP-32)-positive neurons, and choline acetyltransferase (ChAT)-positive motor neurons. These results build on contemporary gene transfer techniques, demonstrating that the hSYN1 promoter can be used with rAAV9 to drive robust neuron-specific transgene expression throughout the nervous system. Copyright © 2014 The Authors. Published by Elsevier Ireland Ltd..

KEYWORDS: AAV9; ALS; Alzheimer’s disease; Gene delivery; Neurodegeneration; Parkinson’s disease

PMID: 24882721



AAV-mediated gene transfer is increasingly recognized as a promising therapeutic tool for a wide-range of neurodegenerative disorders. AAV vectors can be pseudotyped with natural or engineered capsids to facilitate different transduction efficiencies and cellular tropism in the nervous system, and, depending on the promoter and/or transgene-of-interest, may be directed towards long-term, cell-type-specific expression or knockdown.

The trans blood-brain barrier capacity and high transduction rate of AAV9 make this an attractive serotype for performing widespread gene delivery in the nervous system. This is particularly evident after intravenous or intracerebroventricular (i.c.v.) injection of AAV9 in neonatal rodents [1-4]. Recently, these delivery methods were successfully used to target neurodegenerative processes in mouse models of spinal muscular atrophy [5-10] and amyotrophic lateral sclerosis [11]. However, since most studies to date have used strong, ubiquitous promoters, such as CMV and CMV/chicken-β-actin, transgene expression will occur in a non-cell-specific manner.

Our lab is interested in developing novel neurodegenerative models and therapeutics using AAV-mediated gene transfer in rodents. Often, we require neuron-specific transgene expression to help us gain a clearer understanding of specific cellular processes when under target engagement. We have successfully employed the human synapsin (hSYN) promoter to drive neuron-specific transgene expression following intraparenchymal injection of rats with AAV2 [12], an ideal serotype for confining transgene expression to a circumscribed population of cells.

In the current study, we show that the hSYN promoter can be used with AAV9 to achieve robust, neuron-specific GFP expression throughout the CNS and PNS of adult mice, including the sciatic nerve, DRG, and NMJs. Interestingly, while we performed i.c.v. injections at post-natal day 2 – an age associated with preferential astrocytic tropism for AAV9 – we observed some of the highest neuron-specific transduction rates, to date, without GFP expression in astrocytes or microglia, thus showing that the hSYN promoter confers neuronal specificity.