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One of the major challenges in large scale optical imaging of neuronal activity is to simultaneously achieve sufficient temporal and spatial resolution across a large volume. Here, we introduce sparse decomposition light-field microscopy (SDLFM), a computational imaging technique based on light-field microscopy (LFM) that takes algorithmic advantage of the high temporal resolution of LFM and the inherent temporal sparsity of spikes to improve effective spatial resolution and signal-to-noise ratios (SNRs). With increased effective spatial resolution and SNRs, neuronal activity at the single-cell level can be recovered over a large volume. We demonstrate the single-cell imaging capability of SDLFM within vivoimaging of neuronal activity of whole brains of larval zebrafish with estimated lateral and axial resolutions of$\sim <\#comment/>3.5\text{µ<\#comment/>}\mathrm{m}$and$\sim <\#comment/>7.4\text{µ<\#comment/>}\mathrm{m}$, respectively, acquired at volumetric imaging rates up to 50 Hz. We also show that SDLFM increases the quality of neural imaging in adult fruit flies.

Methods for highly multiplexed RNA imaging are limited in spatial resolution and thus in their ability to localize transcripts to nanoscale and subcellular compartments. We adapt expansion microscopy, which physically expands biological specimens, for long-read untargeted and targeted in situ RNA sequencing. We applied untargeted expansion sequencing (ExSeq) to the mouse brain, which yielded the readout of thousands of genes, including splice variants. Targeted ExSeq yielded nanoscale-resolution maps of RNAs throughout dendrites and spines in the neurons of the mouse hippocampus, revealing patterns across multiple cell types, layer-specific cell types across the mouse visual cortex, and the organization and position-dependent states of tumor and immune cells in a human metastatic breast cancer biopsy. Thus, ExSeq enables highly multiplexed mapping of RNAs from nanoscale to system scale.