Significance: Light-field microscopy (LFM) enables high signal-to-noise ratio (SNR) and light efficient volume imaging at fast frame rates. Voltage imaging with genetically encoded voltage indicators (GEVIs) stands to particularly benefit from LFM’s volumetric imaging capability due to high required sampling rates and limited probe brightness and functional sensitivity.
Aim: We demonstrate subcellular resolution GEVI light-field imaging in acute mouse brain slices resolving dendritic voltage signals in three spatial dimensions.
Approach: We imaged action potential-induced fluorescence transients in mouse brain slices sparsely expressing the GEVI VSFP-Butterfly 1.2 in wide-field microscopy (WFM) and LFM modes. We compared functional signal SNR and localization between different LFM reconstruction approaches and between LFM and WFM.
Results: LFM enabled three-dimensional (3-D) localization of action potential-induced fluorescence transients in neuronal somata and dendrites. Nonregularized deconvolution decreased SNR with increased iteration number compared to synthetic refocusing but increased axial and lateral signal localization. SNR was unaffected for LFM compared to WFM.
Conclusions: LFM enables 3-D localization of fluorescence transients, therefore eliminating the need for structures to lie in a single focal plane. These results demonstrate LFM’s potential for studying dendritic integration and action potential propagation in three spatial dimensions.
Surface plasmon resonance (SPR) at planar metal/dielectric interfaces and localised SPR (LSPR) for metal nanoparticles have both been extensively studied, but it is less clear what happens to the optical properties of surface plasmon-polaritons (SPPs) at the micrometer scale. This paper characterises the angular responses of microscale gold patterns on a glass substrate using an SPR imaging configuration using a high numerical aperture objective lens. The potential use for biosensing is also discussed.
The propagation length of a SPP equals approximately 10 μm, so the geometries that are investigated are 10 × 16, 10 × 10, 6 × 16, and 6 × 6 μm sized rectangles with 2 μm spacings. The gold patterns are photolithographically produced using an image reversal process. Bright-field microscopy is used to investigate their morphology and stylus profilometry is used to check the gold thickness. The SPR responses were compared to planar gold film to see if they were characteristic and confirm the relation to SPR.
All the gold microstructures present plasmonic properties. SPR dips are observed for all samples, however, they are not as sensitive and are wider than that of planar gold film. This phenomenon becomes more pronounced as the length of the gold structure decreases, because of the spatial constriction of the propagating SPP. The process suggests that the SPR technique can be successfully implemented to detect individual action potentials. Further work is required in order to achieve a reliable process and investigate the capabilities and sensitivity of the proposed technique.