Protocol for correlating in vivo calcium imaging with spatial transcriptomics in the mouse brain

A central challenge in neuroscience research is to elucidate how distinct neuronal types coordinate to produce complex behaviors, which requires linking the molecular identity of neuronal types with their connectivity and activity in behaving animals. Here, we present a protocol for integrating spatial gene-expression profiling with in vivo calcium imaging in the mouse brain. We describe detailed procedures for tissue recovery after in vivo imaging, multiplexed gene-expression detection, and the computational steps to register these two datasets. For complete details on the use and execution of this protocol, please refer to Yamazaki et al.¹.