Extensive mapping of axonal connections, cell types, and gene expression has provided rich data about the brain’s complex organization. When combined with whole-brain functional connectivity imaging, these maps have shed light on the association between brain structure and function. However, it remains unclear how these structural elements support information flow and processing in neural circuits, and how they influence behavior. Combining these multimodal imaging with cell-specific optogenetic stimulation allows in vivo interrogation of a specific circuit to understand the functional and causal roles of these cellular and connectomic substrates on brain network dynamics. This article describes methods for conducting such experiments and comparing functional, connectomic, cell-type, and genomic data in mice. These combined tools could provide new insights into the brain’s structure–function relationship, circuit mechanisms of brain disorders, and the development of potential interventions.