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Time-lapse microscopy was carried out on the same microscope using a heated incubator, heated stage plate and CO2 controller (Pecon) to maintain the cells.
Z1 inverse microscope equipped with a 37 °C heated stage and CO2 chamber (40× objective, oil immersion, NA = 1.1).
Images were acquired using a Zeiss Axio Observer Z1 equipped with a 100× objective, heated stage and CO2 controller.
The heated stage consists of an Inheco CPAC Ultraflat HT microplate heater with a temperature range of 4°C to 110°C (Inheco).
Animals were then placed in an air-tight box in the imaging chamber (with heated stage, 37°C) at a distance allowing for whole body imaging in a single field of view.
After 60 min, images were acquired from 200 μm Z-stacks by intravital confocal microscopy (ICM) using an upright Nikon A1R laser scanning confocal microscope equipped with a resonance scanner, motorized and heated stage.
The SPQ-loaded cells were then mounted on a LSM510 Meta confocal microscope (Zeiss, Milan, Italy) with a 37 °C heated stage and perfused with iodide buffer for 5 to 8 min.
Cells were heated to 42.5°C using the heated stage and objective and visualized at 100× plan apochromat oil N.A1.3.
The bathing artificial CSF was heated to 32 36°C using a sleeve heater and heated stage (Warner Instruments).
Devices were placed within a 5% CO2-regulated chamber on a heated stage at 37 °C.
FRAP was performed using a confocal microscope (FV-3000; Olympus), equipped with a heated stage and cover filled with humidified 5% CO2 and 95% air.
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