TY - JOUR T1 - Light microscopy based approach for mapping connectivity with molecular specificity JF - bioRxiv DO - 10.1101/2020.02.24.963538 SP - 2020.02.24.963538 AU - Fred Y. Shen AU - Margaret M. Harrington AU - Logan A. Walker AU - Hon Pong Jimmy Cheng AU - Edward S. Boyden AU - Dawen Cai Y1 - 2020/01/01 UR - http://biorxiv.org/content/early/2020/02/25/2020.02.24.963538.abstract N2 - Mapping neuroanatomy is a foundational goal towards understanding brain function. Electron microscopy (EM) has been the gold standard for connectivity analysis because nanoscale resolution is necessary to unambiguously resolve chemical and electrical synapses. However, molecular information that specifies cell types is often lost in EM reconstructions. To address this, we devised a light microscopy approach for connectivity analysis of defined cell types called spectral connectomics. We combined multicolor genetic labeling (Brainbow) of neurons with a multi-round immunostaining Expansion Microscopy (miriEx) strategy to simultaneously interrogate morphology, molecular markers, and connectivity in the same brain section. We applied our multimodal profiling strategy to directly link inhibitory neuron cell types with their network morphologies. Furthermore, we showed that correlative Brainbow and endogenous synaptic machinery immunostaining can be used to define putative synaptic connections between spectrally unique neurons, as well as map putative inhibitory and excitatory inputs. We envision that spectral connectomics can be applied routinely in neurobiology labs to gain insights into normal and pathophysiological neuroanatomy across multiple animals and time points. ER -