Calcium image analysis in the moving gut.

Background: The neural control of gastrointestinal muscle relies on circuit activity whose underlying motifs remain limited by small-sample calcium imaging recordings confounded by motion artifact, paralytics, and muscle dissections. We present a sequence of resources to register images from moving preparations and identify out-of-focus events in widefield fluorescent microscopy.

Methods: Our algorithm uses piecewise rigid registration with pathfinding to correct movements associated with smooth muscle contractions.

Decontaminate Traces From Fluorescence Calcium Imaging Videos Using Targeted Non-negative Matrix Factorization.

Fluorescence microscopy and genetically encoded calcium indicators help understand brain function by recording large-scale videos in assorted animal models. Extracting the fluorescent transients that represent active periods of individual neurons is a key step when analyzing imaging videos. Non-specific calcium sources and background adjacent to segmented neurons contaminate the neurons' temporal traces with false transients.

Rational engineering of ratiometric calcium sensors with bright green and red fluorescent proteins.

Ratiometric genetically encoded calcium indicators (GECIs) record neural activity with high brightness while mitigating motion-induced artifacts. Recently developed ratiometric GECIs primarily employ cyan and yellow-fluorescent fluorescence resonance energy transfer pairs, and thus fall short in some applications that require deep tissue penetration and resistance to photobleaching. We engineered a set of green-red ratiometric calcium sensors that fused two fluorescent proteins and calcium sensing domain within an alternate configuration.

Hybrid light-sheet and light-field microscope for high resolution and large volume neuroimaging.

Large scale simultaneous recording of fast patterns of neural activity remains challenging. Volumetric imaging modalities such as scanning-beam light-sheet microscopy (LSM) and wide-field light-field microscopy (WFLFM) fall short of the goal due to their complex calibration procedure, low spatial resolution, or high-photobleaching. Here, we demonstrate a hybrid light-sheet light-field microscopy (LSLFM) modality that yields high spatial resolution with simplified alignment of the imaging plane and the excitation plane.