@article {70285, title = {Whole-brain co-mapping of gene expression and neuronal activity at cellular resolution in behaving zebrafish}, journal = {bioRxiv}, year = {2026}, month = {2026 Feb 10}, abstract = {

The brain{\textquoteright}s capabilities rely on both the molecular properties of individual cells and their interactions across brain-wide networks. However, relating gene expression to activity in individual neurons across the entire brain remains elusive. Here we developed an experimental-computational platform, WARP, for whole-brain imaging of neuronal activity during behavior, expansion-assisted spatial transcriptomics, and cellular-level registration of these two modalities. Through joint analysis of whole-brain neuronal activity during multiple behaviors, cellular gene expression, and anatomy, we identified functions of molecularly defined populations {\textemdash} including luminance coding in a cckb-pou4f2 midbrain population and task-structured activity in pvalb7-eomesa hippocampal-like neurons {\textemdash} and defined over 2,000 other function-gene-anatomy subpopulations. Analysis of this unprecedented multimodal dataset also revealed that most gene-matched neurons showed stronger activity correlations, highlighting a brain-wide role for gene expression in functional organization. WARP establishes a foundational platform and open-access dataset for cross-experiment discovery, high-throughput function-to-gene mapping, unification of cell biology and systems neuroscience, and scalable circuit modeling at the whole-brain scale.

}, doi = {10.64898/2026.02.07.704095}, url = {https://www.biorxiv.org/content/early/2026/02/10/2026.02.07.704095}, author = {Marquez Legorreta, Emmanuel and Fleishman, Greg M. and Hesselink, Luuk W. and Eddison, Mark and Smeets, Kasper and Stringer, Carsen and Keller, Philipp J. and Narayan, Sujatha and Chen, Alex B. and Mensh, Brett D. and Sternson, Scott M. and Englitz, Bernhard and Tillberg, Paul W. and Ahrens, Misha B.} } @article {70084, title = {Imaging cellular activity simultaneously across all organs of a vertebrate reveals body-wide circuits}, journal = {bioRxiv}, year = {2025}, month = {2025 August 22}, abstract = {

All cells in an animal collectively ensure, moment-to-moment, the survival of the whole organism in the face of environmental stressors1,2. Physiology seeks to elucidate the intricate network of interactions that sustain life, which often span multiple organs, cell types, and timescales, but a major challenge lies in the inability to simultaneously record time-varying cellular activity throughout the entire body.We developed WHOLISTIC, a method to image second-timescale, time-varying intracellular dynamics across cell-types of the vertebrate body. By advancing and integrating volumetric fluorescence microscopy, machine learning, and pancellular transgenic expression of calcium sensors in transparent young Danio rerio (zebrafish) and adult Danionella, the method enables real-time recording of cellular dynamics across the organism. Calcium is a universal intracellular messenger, with a large array of cellular processes depending on changes in calcium concentration across varying time-scales, making it an ideal proxy of cellular activity3.Using this platform to screen the dynamics of all cells in the body, we discovered unexpected responses of specific cell types to stimuli, such as chondrocyte reactions to cold, meningeal responses to ketamine, and state-dependent activity, such as oscillatory ependymal-cell activity during periods of extended motor quiescence. At the organ scale, the method uncovered pulsating traveling waves along the kidney nephron. At the multi-organ scale, we uncovered muscle synergies and independencies, as well as muscle-organ interactions. Integration with optogenetics allowed us to all-optically determine the causal direction of brain-body interactions. At the whole-organism scale, the method captured the rapid brainstem-controlled redistribution of blood flow across the body.Finally, we advanced Whole-Body Expansion Microscopy4 to provide ground-truth molecular and ultrastructural anatomical context, explaining the spatiotemporal structure of activity captured by WHOLISTIC. Together, these innovations establish a new paradigm for systems biology, bridging cellular and organismal physiology, with broad implications for both fundamental research and drug discovery.

}, doi = {10.1101/2025.08.20.670374}, url = {https://www.biorxiv.org/content/early/2025/08/22/2025.08.20.670374.1}, author = {Ruetten, Virginia M. S. and Zheng, Wei and Siwanowicz, Igor and Mensh, Brett D. and Eddison, Mark and Hu, Amy and Chi, Yunfeng and Lemire, Andrew L. and Guo, Caiying and Kadobianskyi, Mykola and Renz, Marc and Lelek-Greskovic, Sara and He, Yisheng and Close, Kari and Ihrke, Gudrun and Dev, Aparna and Petruncio, Alyson and Wan, Yinan and Engert, Florian and Fishman, Mark C. and Judkewitz, Benjamin and Rubinov, Mikail and Keller, Philipp J. and Satou, Chie and Yu, Guoqiang and Tillberg, Paul W. and Sahani, Maneesh and Ahrens, Misha B.} } @article {69803, title = {TEMI: Tissue Expansion Mass Spectrometry Imaging}, journal = {Nat Methods}, year = {2025}, month = {2025 Apr 22}, abstract = {

The spatial distribution of diverse biomolecules in multicellular organisms is essential for their physiological functions. High-throughput in situ mapping of biomolecules is crucial for both basic and medical research, and requires high scanning speed, spatial resolution, and chemical sensitivity. Here, we developed a Tissue Expansion method compatible with matrix-assisted laser desorption/ionization Mass spectrometry Imaging (TEMI). TEMI reaches single-cell spatial resolution without sacrificing voxel throughput and enables the profiling of hundreds of biomolecules, including lipids, metabolites, peptides (proteins), and N-glycans. Using TEMI, we mapped the spatial distribution of biomolecules across various mammalian tissues and uncovered metabolic heterogeneity in tumors. TEMI can be easily adapted and broadly applied in biological and medical research, to advance spatial multi-omics profiling.

Preprint:\ 10.1101/2025.02.22.639343

}, doi = {10.1101/2025.02.22.639343}, url = {https://doi.org/ 10.1038/s41592-025-02664-9.}, author = {Zhang, Hua and Ding, Lang and Hu, Amy and Shi, Xudong and Huang, Penghsuan and Lu, Haiyan and Tillberg, Paul W. and Wang, Meng C. and Li, Lingjun} } @article {69710, title = {Discovery of neuronal cell types by pairing whole cell reconstructions with RNA expression profiles}, journal = {bioRxiv}, year = {2024}, month = {12/2024}, abstract = {

Effective classification of neuronal cell types requires both molecular and morphological descriptors to be collected in situ at single cell resolution. However, current spatial transcriptomics techniques are not compatible with imaging workflows that successfully reconstruct the morphology of complete axonal projections. Here, we introduce a new methodology that combines tissue clearing, submicron whole-brain two photon imaging, and Expansion-Assisted Iterative Fluorescence In Situ Hybridization (EASI-FISH) to assign molecular identities to fully reconstructed neurons in the mouse brain, which we call morphoFISH. We used morphoFISH to molecularly identify a previously unknown population of cingulate neurons projecting ipsilaterally to the dorsal striatum and contralaterally to higher-order thalamus. By pairing whole-brain morphometry, improved techniques for nucleic acid preservation and spatial gene expression, morphoFISH offers a quantitative solution for discovery of multimodal cell types and complements existing techniques for characterization of increasingly fine-grained cellular heterogeneity in brain circuits.Competing Interest StatementThe authors have declared no competing interest.

}, doi = {10.1101/2024.12.30.630829}, url = {https://www.biorxiv.org/content/early/2024/12/31/2024.12.30.630829}, author = {The MouseLight Project Team and Ferreira, Tiago A. and Eddison, Mark and Copeland, Monique and Lay, Mary and Tenshaw, Emily and Weldon, Monet and Schauder, David and Olbris, Donald J. and Rokicki, Konrad and Spruston, Nelson and Tillberg, Paul W. and Korff, Wyatt and Dudman, Joshua T.} }