Oligodendrocyte precursor cells engulf synapses during circuit remodeling in mice

Oligodendrocyte precursor cells (OPCs) give rise to myelinating oligodendrocytes throughout life, but the functions of OPCs are not limited to oligodendrogenesis. Here we show that OPCs contribute to thalamocortical presynapse elimination in the developing and adult mouse visual cortex. OPC-mediated synapse engulfment increases in response to sensory experience during neural circuit refinement. Our data suggest that OPCs may regulate synaptic connectivity in the brain independently of oligodendrogenesis.


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Data analysis
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Life sciences study design
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Sample size
Sample sizes were based off of Schafer et al. ( 2012) and prior experience with these approaches.
Data exclusions One data point in the P20 microglial engulfment of VGluT2 (Fig. 1D) was a strong outlier (greater than 3 times the interquartile range) and therefore excluded.We also excluded strong outliers from the data shown in Figure 1F,L, and Extended Data Figure 3B by using the Graphpad Prism function "Identify outliers" based upon the ROUT method (robust regression and outlier removal) with a Q (maximum desired false discovery rate) = 1 %.Outlier detection and removal was decided a priori as a means of providing more accurate data given the potential for noise in quantifying fluorescence imaging data.

Replication
For all imaging datasets, we have analyzed a minimum of three images per mouse over a set of three mice.For data where individual cells are plotted, multiple cells were taken from a minimum of three images per mouse (and a minimum of three mice per condition where applicable) and used as representative data points.Furthermore, littermate controls were used whenever possible.For in vivo imaging, a minimum of three mice were used for analysis to create representative oligodendrocyte precursor cell (OPC) volumes.In vivo imaging mice were subject to exclusion based off of window integrity as well as signal to noise (SNR) of fluorescent protein emissions.Both single volumes and time-lapse volumes were subject to exclusion based off of motion artifact as well as overt changes to SNR over the imaging session.For flow cytometry datasets three independent replicates with one mouse per condition within a given replicate were used.Each independent replicate was analyzed separately and then pooled for data presentation.
Randomization Mice were either selected for the study because of correct genotype or simply ordered from Jackson Laboratory.Mice were randomly assigned to groups while ensuring an even distribution of sexes.

Blinding
Experimenters were blinded to conditions for quantitative imaging experiments.One experimenter harvested the tissue and assigned it a randomized label before providing the blinded tissue to another experimenter for analysis.After data acquisition and processing, the data were plotted in Graphpad and then samples were unblinded.

Wild animals
The study did not involve wild animals.
Field-collected samples The study did not involve animals collected from the field.

Ethics oversight
Institutional Animal Care and Use Committee at Cold Spring Harbor Laboratory, protocol # 20-3 Note that full information on the approval of the study protocol must also be provided in the manuscript.

Flow Cytometry Plots
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All plots are contour plots with outliers or pseudocolor plots.
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nature portfolio | reporting summary
March 2021

Software
The BD FACSDiva was used to collect the data and CytoexplorerR was used to analyze and generate the the plots and UMAP.

Cell population abundance
Nothing was sorted and/or collected

Gating strategy
The cells were initially distinguished from debris by the expression of CD45 and PDGFRA proteins.Next, the CD45lo and PDGFRAhi population were separately gated as follows: single cells1 → single cells2 → viable cells → OPCs (PDGFRAhi A2B5hi) and microglia (Cd11bhi CD45lo).An independent fluorescence minus one (FMO) control was used as a baseline to set the gate for the negative and positive events related to SYNAPSIN and SNAP25.The highest mean fluorescence intensity in the microglia population was used to set the gate for the high positive events in OPCs.
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