Local and retrograde gene transfer into primate neuronal pathways via adeno-associated virus serotype 8 and 9

Abstract

Viral vector-mediated gene transfer has become increasingly valuable for primate brain research, in particular for application of genetic methods (e.g. optogenetics) to study neuronal circuit functions. Neuronal cell tropisms and infection patterns are viable options for obtaining viral vector-mediated transgene delivery that is selective for particular neuronal pathways. For example, several types of viral vectors can infect axon terminals (retrograde infections), which enables targeted transgene delivery to neurons that directly project to a particular viral injection region. Although recent studies in rodents have demonstrated that adeno-associated virus serotype 8 (AAV8) and 9 (AAV9) efficiently transduce neurons, the tropisms and infection patterns remain poorly understood in primate brains. Here, we constructed recombinant AAV8 or AAV9, which expressed an enhanced green fluorescent protein (EGFP) gene driven by a ubiquitous promoter (AAV8-EGFP and AAV9-EGFP, respectively), and stereotaxically injected it into several brain regions in marmosets and macaque monkeys. Immunohistochemical analyses revealed almost exclusive colocalization of EGFP fluorescence via AAV9-mediated gene transfer with a neuron-specific marker, indicating endogenous neuronal tropism of AAV9, which was consistent with our previous results utilizing AAV8. Injections of either AAV8-EGFP or AAV9-EGFP into the marmoset striatum resulted in EGFP expression in local striatal neurons as a result of local infection, as well as expression in dopaminergic neurons of the substantia nigra via retrograde transport along nigrostriatal axonal projections. Retrograde infections were also observed in the frontal cortex and thalamus, which are known to have direct projections to the striatum. These local and retrograde gene transfers were further demonstrated in the geniculocortical pathway of the marmoset visual system. These findings indicate promising capabilities of AAV8 and AAV9 to deliver molecular tools into a range of primate neural systems in pathway-specific manners through their neuronal tropisms and infection patterns.