To Deconvolve, or Not to Deconvolve: Inferences of Neuronal Activities using Calcium Imaging Data

With the increasing popularity of calcium imaging data in neuroscience research, methods for analyzing calcium trace data are critical to address various questions. The observed calcium traces are either analyzed directly or deconvolved to spike trains to infer neuronal activities. When both approaches are applicable, it is unclear whether deconvolving calcium traces is a necessary step. In this article, we compare the performance of using calcium traces or their deconvolved spike trains for three common analyses: clustering, principal component analysis (PCA), and population decoding. Our simulations and applications to real data suggest that the estimated spike data outperform calcium trace data for both clustering and PCA. Although calcium trace data show higher predictability than spike data at each time point, spike history or cumulative spike counts is comparable to or better than calcium traces in population decoding

Practical considerations in an era of multicolor optogenetics

The ability to control synaptic communication is indispensable to modern neuroscience. Until recently, only single-pathway manipulations were possible due to limited availability of opsins activated by distinct wavelengths. However, extensive protein engineering and screening efforts have drastically expanded the optogenetic toolkit, ushering in an era of multicolor approaches for studying neural circuits. Nonetheless, opsins with truly discrete spectra are scarce. Experimenters must therefore take care to avoid unintended cross-activation of optogenetic tools (crosstalk). Here, we demonstrate the multidimensional nature of crosstalk in a single model synaptic pathway, testing stimulus wavelength, irradiance, duration, and opsin choice. We then propose a “lookup table” method for maximizing the dynamic range of opsin responses on an experiment-by-experiment basis

Time-varying ℓ0\ell_0 optimization for Spike Inference from Multi-Trial Calcium Recordings

Optical imaging of genetically encoded calcium indicators is a powerful tool to record the activity of a large number of neurons simultaneously over a long period of time from freely behaving animals. However, determining the exact time at which a neuron spikes and estimating the underlying firing rate from calcium fluorescence data remains challenging, especially for calcium imaging data obtained from a longitudinal study. We propose a multi-trial time-varying $\ell_0$ penalized method to jointly detect spikes and estimate firing rates by robustly integrating evolving neural dynamics across trials. Our simulation study shows that the proposed method performs well in both spike detection and firing rate estimation. We demonstrate the usefulness of our method on calcium fluorescence trace data from two studies, with the first study showing differential firing rate functions between two behaviors and the second study showing evolving firing rate function across trials due to learning

Simultaneous mesoscopic and two-photon imaging of neuronal activity in cortical circuits.

Spontaneous and sensory-evoked activity propagates across varying spatial scales in the mammalian cortex, but technical challenges have limited conceptual links between the function of local neuronal circuits and brain-wide network dynamics. We present a method for simultaneous cellular-resolution two-photon calcium imaging of a local microcircuit and mesoscopic widefield calcium imaging of the entire cortical mantle in awake mice. Our multi-scale approach involves a microscope with an orthogonal axis design where the mesoscopic objective is oriented above the brain and the two-photon objective is oriented horizontally, with imaging performed through a microprism. We also introduce a viral transduction method for robust and widespread gene delivery in the mouse brain. These approaches allow us to identify the behavioral state-dependent functional connectivity of pyramidal neurons and vasoactive intestinal peptide-expressing interneurons with long-range cortical networks. Our imaging system provides a powerful strategy for investigating cortical architecture across a wide range of spatial scales

ATP depletion induces translocation of STIM1 to puncta and formation of STIM1–ORAI1 clusters: translocation and re-translocation of STIM1 does not require ATP

Depletion of the endoplasmic reticulum (ER) calcium store triggers translocation of stromal interacting molecule one (STIM1) to the sub-plasmalemmal region and formation of puncta—structures in which STIM1 interacts and activates calcium channels. ATP depletion induced the formation of STIM1 puncta in PANC1, RAMA37, and HeLa cells. The sequence of events triggered by inhibition of ATP production included a rapid decline of ATP, depletion of phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) and a slow calcium leak from the ER followed by formation of STIM1 puncta. STIM1 puncta induced by ATP depletion were co-localized with clusters of ORAI1 channels. STIM1–ORAI1 clusters that developed as a result of ATP depletion were very poor mediators of Ca2+ influx. Re-translocation of STIM1 from puncta back to the ER was observed during total ATP depletion. We can therefore conclude that STIM1 translocation and re-translocation as well as formation of STIM1–ORAI1 clusters occur in an ATP-independent fashion and under conditions of PI(4,5)P2 depletion

InsP3 receptors and Orai channels in pancreatic acinar cells: co-localization and its consequences

Orai1 proteins have been recently identified as subunits of SOCE (store-operated Ca2+ entry) channels. In primary isolated PACs (pancreatic acinar cells), Orai1 showed remarkable co-localization and co-immunoprecipitation with all three subtypes of IP3Rs (InsP3 receptors). The co-localization between Orai1 and IP3Rs was restricted to the apical part of PACs. Neither co-localization nor co-immunoprecipitation was affected by Ca2+ store depletion. Importantly we also characterized Orai1 in basal and lateral membranes of PACs. The basal and lateral membranes of PACs have been shown previously to accumulate STIM1 (stromal interaction molecule 1) puncta as a result of Ca2+ store depletion. We therefore conclude that these polarized secretory cells contain two pools of Orai1: an apical pool that interacts with IP3Rs and a basolateral pool that interacts with STIM1 following the Ca2+ store depletion. Experiments on IP3R knockout animals demonstrated that the apical Orai1 localization does not require IP3Rs and that IP3Rs are not necessary for the activation of SOCE. However, the InsP3-releasing secretagogue ACh (acetylcholine) produced a negative modulatory effect on SOCE, suggesting that activated IP3Rs could have an inhibitory effect on this Ca2+ entry mechanism

STIM1, Orai1 and store operated calcium entry in pancreatic acinar cells

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