Abstract
A brain region typically receives inputs from multiple upstream areas. However, currently, no method is available to selectively dissect neurons that receive monosynaptic inputs from two upstream regions. Here, we developed a method to genetically label such neurons with a single gene of interest in mice by combining the anterograde transsynaptic spread of adeno-associated virus serotype 1 (AAV1) with intersectional gene expression. Injections of AAV1 expressing either Cre or Flpo recombinases and the Cre/Flpo double-dependent AAV into two upstream regions and the downstream region, respectively, were used to label postsynaptic neurons receiving inputs from the two upstream regions. We demonstrated this labelling in two distinct circuits: the retina/primary visual cortex to the superior colliculus and the bilateral motor cortex to the dorsal striatum. Systemic delivery of the intersectional AAV allowed the unbiased detection of the labelled neurons throughout the brain. This strategy may help analyse the interregional integration of information in the brain.