Abstract
A combination of genetically-encoded calcium indicators and micro-optics has enabled monitoring of large-scale dynamics of neuronal activity from behaving animals. In these studies, wide-field microscopy is often used to visualize neural activity. However, this method lacks optical sectioning capability, and therefore its axial resolution is generally poor. At present, it is unclear whether wide-field microscopy can visualize activity of densely packed small neurons at cellular resolution. To examine the applicability of wide-field microscopy for small-sized neurons, we recorded calcium activity of dentate granule cells having a small soma diameter of approximately 10 micrometers. Using a combination of high numerical aperture (0.8) objective lens and independent component analysis-based image segmentation technique, activity of putative single granule cell activity was separated from wide-field calcium imaging data. The result encourages wider application of wide-field microscopy in in vivo neurophysiology.
Highlights
To record neural activity in living animals, extracellular electrophysiological recording has been widely used [1,2,3]
In in vivo calcium imaging with wide field fluorescence microscopy, gradient refractive index (GRIN) lens is often used as objective lens
In each XZ plane contains one or more GCaMP6f-labeled neurons, and the shape of individual neurons were clearly visualized (S2 Fig). These results suggest that the spatial resolution of this imaging setup is sufficient for cellular-level calcium imaging of dentate granule cells (GCs)
Summary
To record neural activity in living animals, extracellular electrophysiological recording has been widely used [1,2,3] This technique offers high temporal resolution and low invasion of brain tissue, but recording the same neurons for a long period of time (days to weeks) and to determine their precise location and cell types are difficult. Two-photon microscope using wide-field excitation or multi-beam scanning technique offers wide field of view at high frame rate [6,7].
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