Microglial motility is modulated by neuronal activity and correlates with dendritic spine plasticity in the hippocampus of awake mice
- Neuroimmunology and Imaging Group, German Center for Neurodegenerative Diseases, Germany
- Light Microscopy Facility, German Center for Neurodegenerative Diseases, Germany
Figures
Figure 1 with 6 supplements
Increased microglial motility in the hippocampus of awake mice.
Experimental paradigm of repetitive microglia imaging in dorsal CA1 (stratum radiatum [SR]) in the same mice under varying conditions. (b) Schematic of the accessible hippocampal layers in CA1 …
Figure 1—figure supplement 1
Isoflurane anesthesia decreases neuronal activity in the hippocampus.
(a) Exemplary image of Ca2+ imaging (GCaMP6m) in CamKII+ cells of the CA1 region in the dorsal CA1 of the hippocampus in an awake mouse averaged over 60 s at 30 Hz sampling rate. (b) Ca2+ imaging …
Figure 1—figure supplement 2
Examples of microglia in CA1 under anesthesia, awake, and tetrodotoxin (TTX)-treated conditions.
(a–c) Exemplary in vivo images of microglia from stratum radiatum (SR) of dorsal CA1 under (a) isoflurane anesthesia, (b) awake, and (c) awake but after topical TTX treatment. We did not observe …
Figure 1—video 1
Microglial motility in an anesthetized mouse.
Microglia fine process motility recorded in the hippocampus of a mouse under isoflurane anesthesia (1% in O2) using two-photon microscopy over 45 min. Z-stacks were acquired 5 min apart each …
Figure 1—video 2
Microglial motility in an awake mouse.
Microglia fine process motility recorded in the hippocampus of an awake mouse head-fixed on a circular treadmill using two-photon microscopy. Z-stacks were taken 5 min apart each spanning 100 µm …
Figure 1—video 3
Calcium imaging of CA1 pyramidal neurons under isoflurane anesthesia.
Exemplary video of Ca2+ imaging (GCaMP6m) in CamKII+ cells in the CA1 region of the hippocampus in an anesthetized mouse (isoflurane, 1% in O2) over 60 s. Scale bar: 50 µm.
Figure 1—video 4
Calcium imaging of CA1 pyramidal cells in an awake mouse.
Exemplary video of Ca2+ imaging (GCaMP6m) in CamKII+ cells in the CA1 region of the hippocampus of an awake mouse over 60 s. Scale bar: 50 µm.
Figure 2 with 4 supplements
Increasing or decreasing CA3 input to CA1 reciprocally changes microglial motility.
(a, e) Adeno-associated virus (AAV) injections into the CA3 region of the hippocampus with CamKII-driven Cre-recombinase expression in neurons. This allows for the expression of loxP-flanked …
Figure 2—figure supplement 1
Microglial motility in awake mice depends on neuronal activity.
(a) Injection of two adeno-associated viruses (AVVs) into the CA1 and CA3 region of the hippocampus with CamKII-driven Cre-recombinase expression in neurons (first AAV). This allows for the …
Figure 2—figure supplement 2
Elevated Ca2+ event frequency in Schaffer collaterals following DREADD (designer receptor exclusively activated by designer drug) activation.
(a) Schematic illustrating stereotactic injections into the CA3 region of the hippocampus with a combined injection of AAV9-CamKII0.4-Cre-SC40 together with AAV1-Syn-Flex-GCaMP6m and a …
Figure 2—video 1
Ca2+ imaging of Schaffer collaterals in CA1 of vehicle-treated hM3D(Gq)-expressing CA3 neurons in awake mice.
Imaging of axonal fibers in stratum radiatum (SR) after a co-injection of AAV9-CamKII0.4-Cre-SC40 together with AAV1-Syn-Flex-GCaMP6m and a AAV2-hSyn-DIO-hM3D(Gq)-DREADD into the CA3 cell layer. …
Figure 2—video 2
Ca2+ imaging of Schaffer collaterals in CA1 of clozapine N-oxide (CNO)-treated hM3D(Gq)-expressing CA3 neurons in awake mice.
Imaging of axonal fibers in stratum radiatum (SR) after a co-injection of AAV9-CamKII0.4-Cre-SC40 together with AAV1-Syn-Flex-GCaMP6m and AAV2-hSyn-DIO-hM3D(Gq)-DREADD into the CA3 cell layer. Video …
Figure 3 with 2 supplements
Increased structural plasticity of dendritic spines in the hippocampus of awake mice.
(a) Mice were imaged twice with a 2-day interval, first in anesthesia and subsequently awake on a spinning disk. (b) We acquired z-stacks containing radial oblique dendrites in stratum radiatum of …
Figure 3—figure supplement 1
Clustered structural plasticity of spines associated with microglia contact frequency.
(a) Schematic illustrating clustered spine turnover quantification. Three different classes of clustered events (within a distance of 4 µm) were defined. Lost: If two neighboring spines were lost; …
Figure 3—figure supplement 2
Increased structural plasticity of dendritic spines in the hippocampus of awake mice, statistics over dendrites.
No changes of spine density after imaging in anesthesia (1.09 ± 0.04 μm–1 at d0 vs. 1.07 ± 0.04 μm–1 at d2; Wilcoxon matched-pairs signed rank test, p=0.2759). (b) Elevated spine density after awake …
Figure 4 with 2 supplements
Microglia contact frequency is associated with spine stability in the hippocampus.
(a) Schematic illustrating the analysis between microglia contacts and spine stability in stratum radiatum of dorsal CA1: direct spine contacts (circle) can lead to a stable (blue arrow) or lost …
Figure 4—figure supplement 1
Microglia contact frequency associated with spine stability in the hippocampus, statistics over dendrites.
(a, b) Direct microglia contacts (a) and contacts along the dendritic shaft (b) in anesthetized vs. awake mice unpaired t-test (a,b); p<0.0001 (a), p=0.0964 (b). (c, d) Contact rate in anesthesia (c)…
Figure 4—video 1
Microglia interacting with dendrites and spines in the hippocampus of an awake mouse.
Microglia (green) engage in contact with the dendritic shaft and individual dendritic spines (magenta). Regions of colocalization of microglia and neuronal structures appear white. Z-stacks were …
Tables
Table 1
Mouse number and gender used in the experiments.
| Mouse number Figure 4 | #Spines analyzed for contacts, anesthesia | #Spines analyzed for contacts, awake |
|---|---|---|
| 1. | 157 | 152 |
| 2. | 133 | 110 |
| 3. | 125 | 125 |
| 4. | 130 | 145 |
Table 2
Number of spines analyzed for contacts.
| Figure | Condition | Total number of mice | Number of male mice | Number of female mice |
|---|---|---|---|---|
| Figure 1d | Anesthesia | 6 | 4 | 2 |
| Figure 1d | Awake | 6 | 4 | 2 |
| Figure 1d | TTX | 4 | 3 | 1 |
| Figure 1, Figure 1—figure supplement 1e | Ca2+, CA1, An vs. Aw | 3 | 3 | 0 |
| Figure 2k | CA3, hM4D(Gi) | 4 | 2 | 2 |
| Figure 2l | CA3, hM3D(Gq) | 4 | 1 | 3 |
| Figure 2, Figure 2—figure supplement 1g | CA1 + CA3, hM4D(Gi) | 5 | 2 | 3 |
| Figure 2, Figure 2—figure supplement 2g | CA3, Ca2+, hM3D(Gq) | 3 | 0 | 3 |
| Figure 3c–f | Structural plasticity, An vs. Aw | 4 | 2 | 2 |
| Figure 3, Figure 3—figure supplement 1 | Cluster | 4 | 2 | 2 |
| Figure 4i–l | Microglial contact rates | 4 | 2 | 2 |
