Microglial motility is modulated by neuronal activity and correlates with dendritic spine plasticity in the hippocampus of awake mice

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 individua..., 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 ..., , , # Data from: Microglial motility is modulated by neuronal activity and correlates with dendritic spine plasticity in the hippocampus of awake mice

Dataset DOI: [10.5061/dryad.63xsj3v68](https://doi.org/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.

#### Fil...,