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Article
| Open Access
Geometric transformation adaptive optics (GTAO) for volumetric deep brain imaging through gradient-index lenses
The GRIN lenses widely used for deep brain functional imaging suffer from a small measurement field of view due to strong fourth-order astigmatism. Here the authors report Geometric Transformation Adaptive Optics (GTAO) that corrects field-dependent astigmatism and enables large-volume in vivo imaging of deep mouse brain through 0.5 mm GRIN lenses.
- Yuting Li
- , Zongyue Cheng
- & Meng Cui
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Article
| Open Access
Multiplex translaminar imaging in the spinal cord of behaving mice
Fluorescence imaging of the spinal cord poses challenges, including depth of imaging. Here the authors describe a custom microscope and chronically implanted microprism that enables multicolor translaminar imaging of sensory and motor evoked activity in behaving mice, and show that spinal astrocytes show sensorimotor program-dependent calcium excitation.
- Pavel Shekhtmeyster
- , Erin M. Carey
- & Axel Nimmerjahn
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Article
| Open Access
Tiltable objective microscope visualizes selectivity for head motion direction and dynamics in zebrafish vestibular system
Signals about head orientation and movement in the vestibular periphery are fundamental to the sense of balance and motion, but difficult to measure systematically during head motion. Here, the authors build a microscope that visualizes neural activity in hair cells and vestibular ganglion cells during 360° head tilt and vibration in zebrafish larvae, and reveal a topographic organization of direction- and static/dynamic stimulus-selective responses.
- Masashi Tanimoto
- , Ikuko Watakabe
- & Shin-ichi Higashijima
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Article
| Open Access
Capture at the ER-mitochondrial contacts licenses IP3 receptors to stimulate local Ca2+ transfer and oxidative metabolism
The formation and dissolution of ER-Mitochondria contacts is unclear. Here, authors show that the IP3 receptor traffics in and out of the contacts and, when trapped, improves calcium signaling to stimulate energy metabolism.
- Máté Katona
- , Ádám Bartók
- & György Hajnóczky
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Article
| Open Access
Optical gearbox enabled versatile multiscale high-throughput multiphoton functional imaging
The authors develop an optical gearbox to accelerate the laser scanning microscopes, achieving flexible adjustment of imaging frame rate from tens of Hz to 1 kHz. The technology is validated through in vivo functional imaging of mice brains.
- Jianian Lin
- , Zongyue Cheng
- & Meng Cui
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Article
| Open Access
Absolute measurement of cellular activities using photochromic single-fluorophore biosensors and intermittent quantification
Biosensors often report relative rather than absolute values. Here the authors report a method that utilises the photochromic properties of biosensors to provide an absolute measure of the analyte concentration or activity: photochromism-enabled absolute quantification (PEAQ) biosensing.
- Franziska Bierbuesse
- , Anaïs C. Bourges
- & Peter Dedecker
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Article
| Open Access
A deep-learning approach for online cell identification and trace extraction in functional two-photon calcium imaging
Processing of two-photon calcium imaging data is generally time-consuming, especially for large fields of view. Here, the authors present CITE-On, a tool based on a convolutional neural network, enabling online automatic cell identification, segmentation, identity tracking, and trace extraction.
- Luca Sità
- , Marco Brondi
- & Tommaso Fellin
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Article
| Open Access
Quantification of bone marrow interstitial pH and calcium concentration by intravital ratiometric imaging
The fate of hematopoietic stem cells can be controlled by factors such as calcium ion concentration. Here the authors report an intravital ratiometric analysis method to measure extracellular calcium ion concentrations and absolute pH in mouse bone marrow.
- S-C. A. Yeh
- , J. Hou
- & C. P. Lin
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Article
| Open Access
A turquoise fluorescence lifetime-based biosensor for quantitative imaging of intracellular calcium
Currently, genetically encoded calcium indicators are not suitable for direct quantification. Here the authors engineer a fluorescence lifetime imaging calcium biosensor, Turquoise Calcium Fluorescence LIfeTime Sensor (Tq-Ca-FLITS), and measure intracellular calcium concentrations in human-derived organoids.
- Franka H. van der Linden
- , Eike K. Mahlandt
- & Joachim Goedhart
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Article
| Open Access
Flexible simultaneous mesoscale two-photon imaging of neural activity at high speeds
Functional brain imaging with two-photon microscopy is limited by a tradeoff between imaging area and acquisition speed. Here, the authors present Quadroscope, a flexible microscope which allows for simultaneous video rate acquisition of four independently targetable brain regions across 5 mm.
- Mitchell Clough
- , Ichun Anderson Chen
- & Jerry L. Chen
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Article
| Open Access
Computational optical sectioning with an incoherent multiscale scattering model for light-field microscopy
Light-field microscopy provides volumetric imaging at high speeds, but suffers from degradation in scattering tissue. Here, the authors present an incoherent multiscale scattering model which allows for quantitative 3D reconstruction in complex environments, and demonstrate dynamic imaging in vivo.
- Yi Zhang
- , Zhi Lu
- & Qionghai Dai
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Article
| Open Access
Wide field light-sheet microscopy with lens-axicon controlled two-photon Bessel beam illumination
Here, the authors present a two-photon light-sheet microscopy with an extended Bessel beam for a tunable field of view and reduced photodamage. They demonstrate long-term imaging of cellular dynamics over the whole body of medaka fish.
- Sota Takanezawa
- , Takashi Saitou
- & Takeshi Imamura
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Article
| Open Access
Quantification of FRET-induced angular displacement by monitoring sensitized acceptor anisotropy using a dim fluorescent donor
The FRET efficiency usually predominantly depends on the proximity of donor and acceptor. Here the authors report an anisotropy-based mode of FRET detection, FRET-induced Angular Displacement Evaluation via Dim donor (FADED), to allow quantification of the relative angle between donor and acceptor.
- Danai Laskaratou
- , Guillermo Solís Fernández
- & Hideaki Mizuno
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Article
| Open Access
The cell-wide web coordinates cellular processes by directing site-specific Ca2+ flux across cytoplasmic nanocourses
Although calcium signals are known to be critical for many cellular processes, how signaling elicits specific functions remains unclear. In visually striking work, Duan et al. reveal that networks of cytoplasmic nanocourses orchestrate cell activity by directing site-specific calcium signals.
- Jingxian Duan
- , Jorge Navarro-Dorado
- & A. Mark Evans
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Article
| Open Access
Improved methods for marking active neuron populations
Methods to directly label active neurons are still lacking. Here the authors develop CaMPARI2, a photoconvertible fluorescent protein sensor for neuronal activity with improved brightness and calcium binding kinetics, as well as an antibody to amplify the activated sensor signal in fixed samples.
- Benjamien Moeyaert
- , Graham Holt
- & Eric R. Schreiter
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Article
| Open Access
Super-wide-field two-photon imaging with a micro-optical device moving in post-objective space
Recording the activity of neurons over large brain regions requires expanding the field of view of the optics without losing on spatial and temporal resolution. Here, the authors report a micro-opto-mechanical device that enables two-photon imaging across distant motor areas around 6 mm apart in the mouse.
- Shin-Ichiro Terada
- , Kenta Kobayashi
- & Masanori Matsuzaki
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Article
| Open Access
Probing the gating mechanism of the mechanosensitive channel Piezo1 with the small molecule Yoda1
Piezo ion channels transduce mechanical stimuli into biological signals but the underlying mechanism has remained elusive. Here, the authors use the selective agonist Yoda1 to identify molecular determinants of Piezo activation, providing mechanistic insights into Piezo-mediated mechanotransduction.
- Jerome J. Lacroix
- , Wesley M. Botello-Smith
- & Yun Luo
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Article
| Open Access
Two-photon imaging of neuronal activity in motor cortex of marmosets during upper-limb movement tasks
Marmosets are an important model organism in neuroscience but there has only been limited success in training them on behavioral tasks. Here the authors report their ability to train marmosets in various motor tasks and simultaneously image neural dynamics in motor cortex with 2-photon imaging.
- Teppei Ebina
- , Yoshito Masamizu
- & Masanori Matsuzaki
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Article
| Open Access
Quantitative optical nanophysiology of Ca2+ signaling at inner hair cell active zones
Quantitatively studying components of the presynapse requires high resolution optical methods. Here the authors use confocal microscopy as well as 2D- and 3D-STED nanoscopy to quantify the number and activity of active zone Ca2+ channels in inner hair cells.
- Jakob Neef
- , Nicolai T. Urban
- & Tobias Moser
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Article
| Open Access
Ratiometric Matryoshka biosensors from a nested cassette of green- and orange-emitting fluorescent proteins
Single fluorescent protein biosensors are susceptible to expression and instrumental artifacts. Here Ast et al. describe a dual fluorescent protein design whereby a reference fluorescent protein is nested within a reporter fluorescent protein to control for such artifacts while preserving sensitivity and dynamic range.
- Cindy Ast
- , Jessica Foret
- & Wolf B. Frommer
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A long Stokes shift red fluorescent Ca2+ indicator protein for two-photon and ratiometric imaging
Current calcium-sensitive probes based on red fluorescent proteins are unsuitable for two-photon excitation at the near-infrared wavelengths commonly used for green fluorescent probes. Wu et al. use a structure-guided approach to engineer a red fluorescent probe with optimal two-photon excitation at these wavelengths.
- Jiahui Wu
- , Ahmed S. Abdelfattah
- & Robert E. Campbell
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Article
| Open Access
Imaging intraorganellar Ca2+ at subcellular resolution using CEPIA
The use of intracellular calcium sensors provides important information about the dynamics of calcium signalling in cells. Here Suzuki et al. develop organelle-targeted sensors to simultaneously measure calcium concentrations in ER and mitochondria, and uncover novel insights into calcium flux in mitochondria.
- Junji Suzuki
- , Kazunori Kanemaru
- & Masamitsu Iino
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Biocompatibility of a genetically encoded calcium indicator in a transgenic mouse model
Calcium-sensing fluorescent proteins such as TN-XXL are valuable tools for studying cellular function but, when expressed in mice, may affect animal physiology and behaviour. The authors of this paper create transgenic mice expressing TN-XXL and show that long-term expression of TN-XXL is tolerated well.
- Stephan Direnberger
- , Marsilius Mues
- & Oliver Griesbeck
Mesoscopic calcium imaging in a head-unrestrained male non-human primate using a lensless microscope