Calcium Imaging Using a Miniature Structured Illumination Microscope

Widefield miniature microscopes enable neuroscience researchers to record neuronal activity in freely moving animals. However, these devices are limited to poor signal-to-background ratio (SBR) due to the lack of optical sectioning capability. In previous work, our group developed a miniature structured illumination microscope (SIMScope3D) that uses periodic structured illumination (SI) to overcome this limitation [1]. The SIMScope3D can provide optically sectioned recordings with an axial resolution of 18 μm but was limited to recordings of slow temporal dynamics using traditional OS-SIM reconstruction algorithms [2], [3]. In recent work, our group demonstrated a technique that applies HiLo reconstruction, which is more robust for time-dependent signals, using three periodic SI patterns [3], [4]. Pseudo-HiLo (pHiLo) enables a more accurate reconstruction of time-dependent signals with reduced noise compared to OS-SIM [3]. In this work, we utilize the SIMScope3D to record calcium activity from cells expressing GCaMP6f in the CA1 hippocampal region. We perform pseudo-Widefield (pWF), OS-SIM and pHiLo reconstructions and compare the fidelity of extracted ΔF/F time courses. Results indicate that the optical sectioning capabilities of the SIMScope3D may be extended to recordings of functional calcium activity, expanding the toolkit for researchers who utilize miniature microscopes for experiments with freely moving animals. Further development of sensitive, high-speed cameras, combined with advances in image reconstruction algorithms, will provide the groundwork for future experiments that utilize genetically encoded voltage indicators (GEVIs) with freely moving animals.