A5047211354- Advanced Fluorescence Microscopy Techniques
- Neuroscience and Neuropharmacology Research
- Photoreceptor and optogenetics research
- Neural dynamics and brain function
- Retinal Development and Disorders
- Neuroscience and Neural Engineering
- Receptor Mechanisms and Signaling
- Analytical Chemistry and Sensors
- Advanced biosensing and bioanalysis techniques
- Photoacoustic and Ultrasonic Imaging
- Neurobiology and Insect Physiology Research
- bioluminescence and chemiluminescence research
- Cellular transport and secretion
- Cell Image Analysis Techniques
- Advanced MRI Techniques and Applications
- Computational Physics and Python Applications
- Autoimmune Neurological Disorders and Treatments
- Advanced Chemical Sensor Technologies
- Power Transformer Diagnostics and Insulation
- Optical Coherence Tomography Applications
- Photochromic and Fluorescence Chemistry
- Gene Regulatory Network Analysis
- Atomic and Subatomic Physics Research
- Zebrafish Biomedical Research Applications
- Radiation Detection and Scintillator Technologies
Janelia Research Campus
2016-2025
Howard Hughes Medical Institute
2016-2025
Allen Institute
2022-2025
Allen Institute for Neural Dynamics
2022-2025
Helix (United States)
2018-2024
Allen Institute for Brain Science
2022-2024
North Seattle College
2024
University of British Columbia
2012-2021
Huazhong University of Science and Technology
2021
George Washington University Virginia Campus
2021
Genetically encoded voltage indicators (GEVIs) enable monitoring of neuronal activity at high spatial and temporal resolution. However, the utility existing GEVIs has been limited by brightness photostability fluorescent proteins rhodopsins. We engineered a GEVI, called Voltron, that uses bright photostable synthetic dyes instead protein-based fluorophores, thereby extending number neurons imaged simultaneously in vivo factor 10 enabling imaging for significantly longer durations relative to...
Two-photon imaging and optogenetic stimulation rely on high illumination powers, particularly for state-of-the-art applications that target deeper structures, achieve faster measurements, or probe larger brain areas. However, little information is available heating resulting damage induced by high-power in the brain. In current study we used thermocouple probes quantum dot nanothermometers to measure temperature changes two-photon microscopy neocortex of awake anaesthetized mice. We...
Abstract The fluorescent glutamate indicator iGluSnFR enables imaging of neurotransmission with genetic and molecular specificity. However, existing variants exhibit low in vivo signal-to-noise ratios, saturating activation kinetics exclusion from postsynaptic densities. Using a multiassay screen bacteria, soluble protein cultured neurons, we generated improved ratios kinetics. We developed surface display constructs that improve iGluSnFR’s nanoscopic localization to postsynapses. resulting...
Abstract Here we design and optimize a genetically encoded fluorescent indicator, iAChSnFR, for the ubiquitous neurotransmitter acetylcholine, based on bacterial periplasmic binding protein. iAChSnFR shows large fluorescence changes, rapid rise decay kinetics, insensitivity to most cholinergic drugs. revealed transients in variety of slice vivo preparations mouse, fish, fly worm. will be useful study acetylcholine all animals.
Abstract l -Lactate, traditionally considered a metabolic waste product, is increasingly recognized as an important intercellular energy currency in mammals. To enable investigations of the emerging roles shuttling -lactate, we now report intensiometric green fluorescent genetically encoded biosensor for extracellular -lactate. This biosensor, designated eLACCO1.1, enables cellular resolution imaging -lactate cultured mammalian cells and brain tissue.
L-Lactate is increasingly appreciated as a key metabolite and signaling molecule in mammals. However, investigations of the inter- intra-cellular dynamics L-lactate are currently hampered by limited selection performance L-lactate-specific genetically encoded biosensors. Here we now report spectrally functionally orthogonal pair high-performance biosensors: green fluorescent extracellular biosensor, designated eLACCO2.1, red intracellular R-iLACCO1. eLACCO2.1 exhibits excellent membrane...
We examined how correlated firing controls axon remodeling, using in vivo time-lapse imaging and electrophysiological analysis of individual retinal ganglion cell (RGC) axons that were visually stimulated either synchronously or asynchronously relative to neighboring inputs the Xenopus laevis optic tectum. RGCs out synchrony rapidly lost ability drive tectal postsynaptic partners while their grew added many new branches. In contrast, activated produced fewer branches, but these more stable....
Abstract Imaging membrane voltage from genetically defined cells offers the unique ability to report spatial and temporal dynamics of electrical signaling at cellular circuit levels. Here, we present a general approach engineer electrochromic fluorescence resonance energy transfer (eFRET) encoded indicators (GEVIs) with positive-going response depolarization through rational manipulation native proton transport pathway in microbial rhodopsins. We transform state-of-the-art eFRET GEVI Voltron...
Genetically encodable calcium ion (Ca2+) indicators (GECIs) based on green fluorescent proteins (GFP) are powerful tools for imaging of cell signaling and neural activity in model organisms. Following almost 2 decades steady improvements the Aequorea victoria GFP-based GCaMP series GECIs, performance most recent generation (i.e., jGCaMP7) may have reached its practical limit due to inherent properties GFP. In an effort sustain progression toward ever-improved we undertook development a new...
Voltage imaging enables monitoring neural activity at sub-millisecond and sub-cellular scale, unlocking the study of subthreshold activity, synchrony, network dynamics with unprecedented spatio-temporal resolution. However, high data rates (>800MB/s) low signal-to-noise ratios create bottlenecks for analyzing such datasets. Here we present VolPy, an automated scalable pipeline to pre-process voltage VolPy features motion correction, memory mapping, segmentation, denoising spike extraction,...
Potassium ion (K + ) plays a critical role as an essential electrolyte in all biological systems. Genetically-encoded fluorescent K biosensors are promising tools to further improve our understanding of -dependent processes under normal and pathological conditions. Here, we report the crystal structure previously reported genetically-encoded biosensor, GINKO1, -bound state. Using structure-guided optimization directed evolution, have engineered improved designated GINKO2, with higher...
Fluorescent dyes that are bright, stable, small, and biocompatible needed for high-sensitivity two-photon imaging, but the combination of these traits has been elusive. We identified a class squaraine derivatives with large action cross-sections (up to 10,000 GM) at near-infrared wavelengths critical in vivo imaging. demonstrate biocompatibility stability red-emitting squaraine-rotaxane (SeTau-647) by imaging dye-filled neurons over 5 days, utility sensitive subcellular synthesizing specific...
Abstract Genetically encoded calcium ion (Ca 2+ ) indicators (GECIs) are widely-used molecular tools for functional imaging of Ca dynamics and neuronal activities on a single cell level. Here we report the design development two new far-red fluorescent GECIs, FR-GECO1a FR-GECO1c, based monomeric protein mKelly. We characterized these GECIs as purified proteins assessed their performance when expressed in cultured neurons. FR-GECOs have excitation emission maxima at ~ 596 nm 644 nm,...
Abstract The fluorescent glutamate indicator iGluSnFR enables imaging of neurotransmission with genetic and molecular specificity. However, existing variants exhibit saturating activation kinetics are excluded from post-synaptic densities, limiting their ability to distinguish synaptic extrasynaptic glutamate. Using a multi-assay screen in bacteria, soluble protein, cultured neurons, we generated novel improved signal-to-noise ratios. We also developed surface display constructs that improve...
We compared performance of recently developed silicon photomultipliers (SiPMs) to GaAsP photomultiplier tubes (PMTs) for two-photon imaging neural activity. Despite higher dark counts, SiPMs match or exceed the signal-to-noise ratio PMTs at photon rates encountered in typical calcium experiments due their low pulse height variability. At during high-speed voltage imaging, substantially outperform PMTs.
Abstract Proper brain function requires the assembly and of diverse populations neurons glia. Single cell gene expression studies have mostly focused on characterization neuronal diversity; however, recent revealed substantial diversity glial cells, particularly astrocytes. To better understand types their roles in neurobiology, we built a new suite adeno-associated viral (AAV)-based genetic tools to enable access astrocytes oligodendrocytes. These oligodendrocyte astrocyte enhancer-AAVs are...
Abstract Identifying the input-output operations of neurons requires measurements synaptic transmission simultaneously at many a neuron’s thousands inputs in intact brain. To facilitate this goal, we engineered and screened 3365 variants fluorescent protein glutamate indicator iGluSnFR3 neuron culture, selected mouse visual cortex. Two have high sensitivity, fast activation (< 2 ms) deactivation times tailored for recording large populations synapses (iGluSnFR4s, 153 or rapid dynamics...
<title>Abstract</title> Identifying the input-output operations of neurons requires measurements synaptic transmission simultaneously at many a neuron’s thousands inputs in intact brain. To facilitate this goal, we engineered and screened 3365 variants fluorescent protein glutamate indicator iGluSnFR3 neuron culture, selected mouse visual cortex. Two have high sensitivity, fast activation (< 2 ms) deactivation times tailored for recording large populations synapses (iGluSnFR4s, 153 or...
The fine control of synaptic function requires robust trans-synaptic molecular interactions. However, it remains poorly understood how bridges change to reflect the functional states synapse. Here, we develop optical tools visualize in firing synapses behavior two proteins, LGI1 and ADAM23, find that neuronal activity acutely rearranges their abundance at cleft. Surprisingly, is primarily not secreted, as described elsewhere, but exo- endocytosed through its interaction with ADAM23....