Viral transduction of the neonatal brain delivers controllable genetic mosaicism for visualising and manipulating neuronal circuits <i>in vivo</i>

Abstract

The neonatal intraventricular injection of adeno-associated virus has been shown to transduce neurons widely throughout the brain, but its full potential for experimental neuroscience has not been adequately explored. We report a detailed analysis of the method's versatility with an emphasis on experimental applications where tools for genetic manipulation are currently lacking. Viral injection into the neonatal mouse brain is fast, easy, and accesses regions of the brain including the cerebellum and brainstem that have been difficult to target with other techniques such as electroporation. We show that viral transduction produces an inherently mosaic expression pattern that can be exploited by varying the titer to transduce isolated neurons or densely-packed populations. We demonstrate that the expression of virally-encoded proteins is active much sooner than previously believed, allowing genetic perturbation during critical periods of neuronal plasticity, but is also long-lasting and stable, allowing chronic studies of aging. We harness these features to visualise and manipulate neurons in the hindbrain that have been recalcitrant to approaches commonly applied in the cortex. We show that viral labeling aids the analysis of postnatal dendritic maturation in cerebellar Purkinje neurons by allowing individual cells to be readily distinguished, and then demonstrate that the same sparse labeling allows live in vivo imaging of mature Purkinje neurons at a resolution sufficient for complete analytical reconstruction. Given the rising availability of viral constructs, packaging services, and genetically modified animals, these techniques should facilitate a wide range of experiments into brain development, function, and degeneration.