Cheng, H. P. J. | Search bioRxiv

bioRxiv2020

Bitbow: a digital format of Brainbow enables highly efficient neuronal lineage tracing and morphology reconstruction in single brains

Identifying the cellular origins and mapping the dendritic and axonal arbors of neurons have been century old quests to understand the heterogeneity among these brain cells. Classical chemical and genetic methods take advantage of light microscopy and sparse labeling to unambiguously, albeit inefficiently, trace a few neuronal lineages or reconstruct their morphologies in each sampled brain. To improve the analysis throughput, we designed Bitbow, a digital format of Brainbow which exponentially expands the color palette to provide tens of thousands of spectrally resolved unique labels. We generated transgenic Bitbow Drosophila lines, established statistical tools, and streamlined sample preparation, image processing and data analysis pipelines to allow conveniently mapping neural lineages, studying neuronal morphology and revealing neural network patterns with an unprecedented speed, scale and resolution.

neuroscience↗

bioRxiv2020

Light microscopy based approach for mapping connectivity with molecular specificity

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.

neuroscience↗