Efficient Labeling of Neurons and Identification of Postsynaptic Sites Using Adeno-Associated Virus Vector

Abstract

Adeno-associated virus (AAV) is a small, non-enveloped, and single-stranded DNA virus with minimal potential hazard to animals including human beings. Owing to the efficient and long-term gene transduction of neuronal and glial cells without pathology, AAV vectors are one of the most prevalent tools for in vivo gene transfer in neuroscience research and have made great contribution to the progress in understanding of neuronal circuits and functions. AAV-mediated gene transduction in combination with Cre-expressing animals is now widely used to label specific groups of neurons. With this approach, we have anterogradely labeled axons from several groups of neurons with different neurotransmitters, such as serotonergic, cholinergic, and neocortical GABAergic neurons, and analyzed their axonal projections to elucidate neuronal connections in the central nervous system. In this chapter, we first describe the detailed protocol of production, purification, and titration of AAV vectors. We then demonstrate visualization of cortical neurons by stereotaxic injection of a prepared AAV vector into the mouse neocortex for qualitative and quantitative analyses of excitatory postsynaptic sites on individual pyramidal cells by confocal laser-scanning microscopy.