Data from: Microglial motility is modulated by neuronal activity and correlates with dendritic spine plasticity in the hippocampus of awake mice
Data files
Mar 27, 2026 version files 10.70 GB
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Figure1_Awake_TTX_stack.tif
31.10 MB
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Figure1_Awake_TTX.tif
1.64 MB
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Figure1_figure_supplement1_Calcium_CA1_anesthesia.tif
446.91 MB
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Figure1_figures_supplement1_Calcium_CA1_awake.tif
446.91 MB
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Figure2_DREADD_Microglia_Mot_hM3DGq.tif
47.52 MB
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Figure2_DREADD_Microglia_Mot_hM4DGi.tif
51.90 MB
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Figure2_DREADD_Microglia_Mot_Vehicle.tif
51.90 MB
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Figure2_Figure_supplement2_Calcium_Shaffer_Collaterals_hM3Gq_CNO.tif
3.67 GB
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Figure2_Figure_supplement2_Calcium_Shaffer_Collaterals_hM3Gq_DMSO.tif
3.67 GB
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Figure3_Stack_anesthesia_d48h.tif
352.90 MB
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Figure3_Stack_anesthesia_tp0.tif
352.92 MB
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Figure3_stack_awake_48h.tif
340.34 MB
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Figure3_stack_awake_tp0.tif
321.64 MB
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Figure3_stack_d48h.tif
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Figure3_Stack1_Dendrite_cut_out.tif
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Figure3_Stack1_OV.tif
352.90 MB
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Figure4_gain_d48h.tif
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Figure4_gain_tp0.tif
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Figure4_gain_tp1.tif
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Figure4_gain_tp2.tif
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Figure4_gain_tp3.tif
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Figure4_gain_tp4.tif
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Figure4_gain_tp5.tif
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Figure4_gain_tp6.tif
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Figure4_gain_tp7.tif
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Figure4_gain_tp8.tif
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Figure4_loss_d48h.tif
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Figure4_loss_tp0.tif
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Figure4_loss_tp1.tif
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Figure4_loss_tp2.tif
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Figure4_loss_tp3.tif
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Figure4_loss_tp4.tif
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Figure4_loss_tp5.tif
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Figure4_loss_tp6.tif
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Figure4_loss_tp7.tif
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Figure4_loss_tp8.tif
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README.md
3.99 KB
Abstract
Microglia, the resident immune cells of the brain, play a complex role in health and disease. They actively survey the brain parenchyma by physically interacting with other cells and structurally shaping the brain. Yet the mechanisms underlying microglial motility and significance for synapse stability, especially in the hippocampus during adulthood, remain widely unresolved. Here we investigated the effect of neuronal activity on microglial motility and the implications for the formation and survival of dendritic spines on hippocampal CA1 neurons in vivo. We used repetitive two-photon in vivo imaging in the hippocampus of awake and anesthetized mice to simultaneously study the motility of microglia and their interaction with dendritic spines. We found that CA3 to CA1 input is sufficient to modulate microglial process motility. Simultaneously, more dendritic spines emerged in mice after awake compared to anesthetized imaging. Interestingly, the rate of microglial contacts with individual dendritic spines and dendrites was associated with the stability, removal, and emergence of dendritic spines. These results suggest that microglia might sense neuronal activity via neurotransmitter release and actively participate in synaptic rewiring of the hippocampal neural network during adulthood. Further, this study has profound relevance for hippocampal learning and memory processes.
Dataset DOI: 10.5061/dryad.63xsj3v68
Description of the data and file structure
Longitudinal in vivo two-photon imaging datasets of microglial dynamics and dendritic spine remodeling in mouse hippocampal CA1 under awake and anesthetized conditions. Includes time-lapse z-stack images (GFP/YFP), processed projections, and metadata with imaging parameters and experimental conditions.
Files and variables
File: Figure1_Awake_TTX.tif
Description: Timelapse of microglial motility in str. radiatum after topical TTX application.
File: Figure1_Awake_TTX_stack.tif
Description: 3D Stack of microglia in Str. Radiatum after topical TTX application.
File: Figure1_Awake_Microglia_Mot.tif
Description: 3D Stack of microglia in Str. Radiatum under awake conditions.
File: Figure2_DREADD_Microglia_Mot_hM3DGq.tif
Description: 3D Stack of Microglia in Str. Radiatum after CNO activation of hM3DGq.
File: Figure2_DREADD_Microglia_Mot_hM4DGi.tif
Description: 3D Stack of Microglia in Str. Radiatum after CNO activation of hM4DGi.
File: Figure2_DREADD_Microglia_Mot_Vehicle.tif
Description: 3D Stack of Microglia in Str. Radiatum after sham injetion/vehicle treatment.
File: Figure1_figure_supplement1_Calcium_CA1_anesthesia.tif
Description: GCamP6 labeled neurons in Str.Pyramidale CA1 under anesthesia.
File: Figure1_figures_supplement1_Calcium_CA1_awake.tif
Description: GCamP6 labeled neurons in Str.Pyramidale CA1 in an awake mouse.
File: Figure3_Stack_anesthesia_d48h.tif
Description: Same neurons and denrites as in Figure3_Stack_anesthesia_tp0 but after 48h.
File: Figure3_stack_d48h.tif
Description: Cut out dendrite 48 hours after recording
File: Figure3_Stack_anesthesia_tp0.tif
Description: 3D Stack of Neurons and dendrites in CA1 region of the hippocampus.
File: Figure3_Stack1_Dendrite_cut_out.tif
Description: Cut out dendrite from Figure3_Stack1_OV.
File: Figure4_gain_tp0-8.tif
Description: Cropped out 3D Stack of Microglia (green) and Dendrites/Neurons and individual dendritic spines (Magenta), tp0-8, 5 minutes between each z-stack
File: Figure4_gain_d48h.tif
Description: Same dendrites/neurons as in Figure4_gain_tp0-8 after 48h.
File: Figure3_stack_awake_tp0.tif
Description: Initial 3D Stack of pyramidal neurons in CA1 Str. Pyramidale and Dendrites in Str. Radiatum.
File: Figure3_stack_awake_48h.tif
Description: 3D Stack of CA1 pyramdial Neurons and dendrites in Str. radiatum 48 hours after initial recording (tp0).
File: Figure4_loss_tp0-8.tif
Description: Cropped out 3D Stack of Microglia (green) and Dendrites/Neurons and individual dendritic spines (Magenta), tp0-8, 5 minutes between each z-stack
File: Figure4_loss_d48h.tif
Description: Same dendrites/neurons as in Figure4_loss_tp0-8 after 48h.
File: Figure3_Stack1_OV.tif
Description: 3D Overview stack of neurons and denrites from str.pyramidale into str.radiatum in a Thy1-YFP mouse.
File: Figure2_Figure_supplement2_Calcium_Shaffer_Collaterals_hM3Gq_CNO.tif
Description: GCaMP6 labeled axons image in str. radiatum following DREADD injection into CA3 pyramidal neurons, CNO condition.
File: Figure2_Figure_supplement2_Calcium_Shaffer_Collaterals_hM3Gq_DMSO.tif
Description: GCaMP6 labeled axons image in str. radiatum following DREADD injection into CA3 pyramidal neurons, DMSO condition.
Code/software
ImageJ/Fiji is sufficient. Image Analysis and processing was performed using ImageJ and Zeiss ZEN.
Access information
Other publicly accessible locations of the data:
- None.
Data was derived from the following sources:
- None.
Mice were fixed to a custom-made head-holder and mounted under the microscope on a heating plate (37°C) to keep the body temperature at optimum. Images were acquired with a Nikon 16x water immersion objective (NA 0.8) with a working distance of 3 mm. For awake imaging, we used a spinning disk made of styrofoam (r = 10 cm, h = 7 cm), or a custom-made linear treadmill, which allowed for imaging of running, head-fixed mice that control the speed of the spinning disk or the treadmill independently. Time-lapse images were acquired at 100 μm x 100 μm in x, y-direction with a pixel size of 0.088 μm/pixel. The z-steps were 1 µm spaced for anesthetized recordings and spanned 100 µm.The acquired z-stacks started in the CA1 pyramidal layer spanning 100 µm deep into SR. To allow for the correction of motion-artifacts, oversampling in z was performed. Z-step sizes were set to 0.2 µm/step achieving an oversampling of 5x per micrometer. This allowed us to manually remove any distorted or out-of-focus frames by carefully scrolling through each z-stack individually. Due to this oversampling and subsequent image removal stable maximum intensity projections could be generated similar to anesthetized conditions. Time-lapse images were acquired every 5 minutes for a period of 45 minutes. Image acquisition was carried out on an upright TrimScope II (LaVision Biotech) equipped with a Ti:Sa laser (Cameleon Ultra II, Coherent) that was tuned to 920 nm to allow for simultaneous excitation of GFP and YFP with a maximum output power of 50 mW to prevent photo damage. GFP fluorescence was acquired with a 480/40 BP Filterset. YFP was separated from GFP by a 510 LP filter and detected using a 535/30 BP Filterset.
