Multispectral tracing in densely labeled mouse brain with nTracer

Roossien, Douglas H.; Sadis, Benjamin V; Yan, Yan; Webb, John; Min, Lia; Dizaji, Aslan S.; Bogart, Luke J; Mazuski, Cristina; Huth, Robert S.; Stecher, Johanna S.; Akula, Sriakhila; Shen, Fred Y.; Li, Ye; Xiao, Tingxin; Vandenbrink, Madeleine; Lichtman, Jeff W.; Hensch, Takao K.; Herzog, Erik D.; Cai, Dawen

doi:10.1093/bioinformatics/btz084

Cited by 23 publications

(36 citation statements)

References 15 publications

(15 reference statements)

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“…S6 ). In agreement with past findings, PV expressing neurons appear to have more complex branching patterns compared with SOM expressing neurons, despite having similar dendritic lengths 25,26 . This type of dense reconstruction enables study of how different neuronal cell types interact with each other anatomically, making it a powerful tool for network morphology analysis.…”

Section: Resultssupporting

confidence: 92%

“…Moreover, our approach towards spectral connectomics allowed us to eliminate false positives that would not be possible with diffraction-limited LM. Neuronal contacts have previously been used as correlates for synaptic sites 26,30 , but we show an example of a close apposition lacking synaptic machinery that would be falsely classified as a connection ( Fig. 3g, k ).…”

Section: Resultsmentioning

confidence: 92%

“…Brainbow AAVs labeled 2 out of 4 (50%) PV neurons, 20 out of 31 (70%) PV/Calb neurons, 7 out of 7 (100%) SOM neurons, and 24 out of 28 (86%) SOM/Calb neurons within a 590 x 404 x 160 μm 3 volume. Following identification of each Brainbow neuron by its molecular subtype, we reconstructed its dendritic morphology using nTracer 26 , an ImageJ/Fiji plugin for tracing multispectral datasets ( Fig. 2g-h, Movie S1 ).…”

Section: Resultsmentioning

confidence: 99%

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Light microscopy based approach for mapping connectivity with molecular specificity

Shen

Harrington

Walker

et al. 2020

Preprint

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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 m ult i-r ound i mmunostaining Ex pansion 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.

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Section: Resultssupporting

confidence: 92%

Section: Resultsmentioning

confidence: 92%

Section: Resultsmentioning

confidence: 99%

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Light microscopy based approach for mapping connectivity with molecular specificity

Shen

Harrington

Walker

et al. 2020

Preprint

Self Cite

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“…TraceMontage was tested under various scenarios to demonstrate its utility. Test microscope images were collected from Brainbow-labeled mouse Hippocampus or primary visual cortex (V1) samples, as indicated in (Roossien et al, 2019;Roossien and Cai, 2017), and split into overlapping tiles to simulate the result of a two-exposure imaging experiment. Each tile was independently traced with nTracer by separate, trained biologists (see Acknowledgements), followed by merging with TraceMontage.…”

Section: Testing and Demonstrationmentioning

confidence: 99%

“…More recently, the transgenic Brainbow techniques (Cai et al, 2013;Lichtman et al, 2008;Livet et al, 2007) have facilitated high throughput neuronal imaging at the microscale resolution, using the combinatorial expression of several fluorescent proteins in neurons to color-tag individual cells and fluorescence microscopy to visualize them. This allows dense and unique labeling of neuronal processes, which permits short-to-long-range circuit tracing of multiple neurons of the same sample by the software nTracer (Roossien et al, 2019).…”

Section: Introductionmentioning

confidence: 99%

TraceMontage: a Method for Merging Multiple Independent Neuronal Traces

Dizaji

Walker

Cai

2019

Preprint

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Background: The ability to reconstruct neuronal networks, local microcircuits, or the entire connectome is a central goal of modern neuroscience. Recently, advancements in sample preparation (e.g., sample expansion and Brainbow labeling) and optical (e.g., confocal and light sheet) techniques have enabled the imaging of increasingly large neural systems with high contrast. Tracing neuronal structures from these images proves challenging, however, necessitating tools that integrate multiple neuronal traces, potentially derived by various methods, into one combined (montaged) result.New Method: Here, we present TraceMontage, an ImageJ/Fiji plugin for the combination of multiple neuron traces of a single image, either redundantly or non-redundantly. Internally, it uses graph theory to connect topological patterns in the 3-D spatial coordinates of neuronal trees. The software generates a single output tracing file containing the montage traces of the input tracing files and provides several measures of consistency analysis among multiple tracers. Results and Comparison to existing method(s):To our knowledge, our software is the first dedicated method for the combination of tracing results. Combining multiple tracers increases the accuracy and speed of tracing of densely-labeled samples by harnessing collaborative effort. This utility is demonstrated using fluorescence microscope images from the hippocampus and primary visual cortex (V1) in Brainbow-labeled mice.Conclusions: TraceMontage provides researchers the ability to combine neuronal tracing data generated by either the same or different method(s). As datasets become larger, the ability to trace images in this parallel manner will help connectomics scale to increasingly larger neural systems.

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Multicolor strategies for investigating clonal expansion and tissue plasticity

Dumas

Clavreul

Michon

et al. 2022

Cell. Mol. Life Sci.

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Understanding the generation of complexity in living organisms requires the use of lineage tracing tools at a multicellular scale. In this review, we describe the different multicolor strategies focusing on mouse models expressing several fluorescent reporter proteins, generated by classical (MADM, Brainbow and its multiple derivatives) or acute (StarTrack, CLoNe, MAGIC Markers, iOn, viral vectors) transgenesis. After detailing the multi-reporter genetic strategies that serve as a basis for the establishment of these multicolor mouse models, we briefly mention other animal and cellular models (zebrafish, chicken, drosophila, iPSC) that also rely on these constructs. Then, we highlight practical applications of multicolor mouse models to better understand organogenesis at single progenitor scale (clonal analyses) in the brain and briefly in several other tissues (intestine, skin, vascular, hematopoietic and immune systems). In addition, we detail the critical contribution of multicolor fate mapping strategies in apprehending the fine cellular choreography underlying tissue morphogenesis in several models with a particular focus on brain cytoarchitecture in health and diseases. Finally, we present the latest technological advances in multichannel and in-depth imaging, and automated analyses that enable to better exploit the large amount of data generated from multicolored tissues.

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Multispectral tracing in densely labeled mouse brain with nTracer

Cited by 23 publications

References 15 publications

Light microscopy based approach for mapping connectivity with molecular specificity

Light microscopy based approach for mapping connectivity with molecular specificity

TraceMontage: a Method for Merging Multiple Independent Neuronal Traces

Multicolor strategies for investigating clonal expansion and tissue plasticity

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