Brain diseases such as autism and Alzheimer's disease (each inflicting >1% of the world population) involve a large network of genes displaying subtle changes in their expression. Abnormalities in intraneuronal transport have been linked to genetic risk factors found in patients, suggesting the relevance of measuring this key biological process. However, current techniques are not sensitive enough to detect minor abnormalities. Here we report a sensitive method to measure the changes in intraneuronal transport induced by brain-disease-related genetic risk factors using fluorescent nanodiamonds (FNDs). We show that the high brightness, photostability and absence of cytotoxicity allow FNDs to be tracked inside the branches of dissociated neurons with a spatial resolution of 12 nm and a temporal resolution of 50 ms. As proof of principle, we applied the FND tracking assay on two transgenic mouse lines that mimic the slight changes in protein concentration (∼30%) found in the brains of patients. In both cases, we show that the FND assay is sufficiently sensitive to detect these changes.
Genetically encoded voltage indicators (GEVIs) are emerging optical tools
for acquiring brain-wide cell-type-specific functional data at unparalleled
temporal resolution. To broaden the application of GEVIs in high-speed
multispectral imaging, we used a high-throughput strategy to develop
voltage-activated red neuronal activity monitor (VARNAM), a fusion of the fast
Acetabularia opsin and the bright red fluorophore mRuby3.
Imageable under the modest illumination intensities required by bright green
probes (<50 mW mm−2), VARNAM is readily usable in vivo.
VARNAM can be combined with blue-shifted optical tools to enable
cell-type-specific all-optical electrophysiology and dual-color spike imaging in
acute brain slices and live Drosophila. With enhanced
sensitivity to subthreshold voltages, VARNAM resolves postsynaptic potentials in
slices and cortical and hippocampal rhythms in freely behaving mice. Together,
VARNAM lends a new hue to the optical toolbox, opening the door to high-speed in
vivo multispectral functional imaging.
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