Sub-millisecond Temporal Resolution Research Articles

Direct simultaneous measurement of electrocaloric effect and hysteresis loss heating in ferroelectrics

Electrocaloric effect and loss-induced self-heating are simultaneously investigated in single-crystalline relaxor 0.9Pb(Mg1/3Nb2/3)O3-0.1PbTiO3 by a direct, high-resolution method. Transients of the total temperature change for few-cycle electric field pulses allow to distinguish and individually determine the contributions from electrocaloric effect and self-heating with millikelvin temperature and submillisecond temporal resolution. Simultaneous dielectric measurements make the comparison of observed self-heating to hysteresis losses possible, where very good agreement is found. The loss factor as a figure of merit for electrocaloric materials is directly obtained from the combined determination of caloric temperature change and losses.

Simulation study on high spatio-temporal resolution acousto-electrophysiological neuroimaging.

Objective.Acousto-electrophysiological neuroimaging (AENI) is a technique hypothesized to record electrophysiological activity of the brain with millimeter spatial and sub-millisecond temporal resolution. This improvement is obtained by tagging areas with focused ultrasound (fUS). Due to mechanical vibration with respect to the measuring electrodes, the electrical activity of the marked region will be modulated onto the ultrasonic frequency. The region's electrical activity can subsequently be retrieved via demodulation of the measured signal. In this study, the feasibility of this hypothesized technique is tested.Approach.This is done by calculating the forward electroencephalography response under quasi-static assumptions. The head is simplified as a set of concentric spheres. Two sizes are evaluated representing human and mouse brains. Moreover, feasibility is assessed for wet and dry transcranial, and for cortically placed electrodes. The activity sources are modeled by dipoles, with their current intensity profile drawn from a power-law power spectral density.Results.It is shown that mechanical vibration modulates the endogenous activity onto the ultrasonic frequency. The signal strength depends non-linearly on the alignment between dipole orientation, vibration direction and recording point. The strongest signal is measured when these three dependencies are perfectly aligned. The signal strengths are in the pV-range for a dipole moment of 5 nAm and ultrasonic pressures within Food and Drug Administration (FDA)-limits. The endogenous activity can then be accurately reconstructed via demodulation. Two interference types are investigated: vibrational and static. Depending on the vibrational interference, it is shown that millimeter resolution signal detection is possible also for deep brain regions. Subsequently, successful demodulation depends on the static interference, that at MHz-range has to be sub-picovolt.Significance.Our results show that mechanical vibration is a possible underlying mechanism of acousto-electrophyisological neuroimaging. This paper is a first step towards improved understanding of the conditions under which AENI is feasible.

Quantum diamond microscope for dynamic imaging of magnetic fields

Wide-field imaging of magnetic signals using ensembles of nitrogen-vacancy (NV) centers in diamond has garnered increasing interest due to its combination of micron-scale resolution, millimeter-scale field of view, and compatibility with diverse samples from across the physical and life sciences. Recently, wide-field NV magnetic imaging based on the Ramsey protocol has achieved uniform and enhanced sensitivity compared to conventional measurements. Here, we integrate the Ramsey-based protocol with spin-bath driving to extend the NV spin dephasing time and improve magnetic sensitivity. We also employ a high-speed camera to enable dynamic wide-field magnetic imaging. We benchmark the utility of this quantum diamond microscope (QDM) by imaging magnetic fields produced from a fabricated wire phantom. Over a 270 × 270 μm2 field of view, a median per-pixel magnetic sensitivity of 4.1(1) nT /Hz is realized with a spatial resolution ≲ 10 μm and sub-millisecond temporal resolution. Importantly, the spatial magnetic noise floor can be reduced to the picotesla scale by time-averaging and signal modulation, which enables imaging of a magnetic-field pattern with a peak-to-peak amplitude difference of about 300 pT. Finally, we discuss potential new applications of this dynamic QDM in studying biomineralization and electrically active cells.

Cinematographic study ofstochastic chemical events at atomic resolution.

The advent of single-molecule atomic-resolution time-resolved electron microscopy (SMART-EM) has created a new field of 'cinematic chemistry,' allowing for the cinematographic recording of dynamic behaviors of organic and inorganic molecules and their assembly. However, the limited electron dose per frame of video images presents a major challenge in SMART-EM. Recent advances in direct electron counting cameras and techniques to enhance image quality through the implementation of a denoising algorithm have enabled the tracking of stochastic molecular motions and chemical reactions with sub-millisecond temporal resolution and sub-angstrom localization precision. This review showcases the development of dynamic molecular imaging using the SMART-EM technique, highlighting insights into nanomechanical behavior during molecular shuttle motion, pathways of multistep chemical reactions, and elucidation of crystallization processes at the atomic level.

The ATPase cycle of the AAA-ATPase protein transporter Bcs1 is highly concerted.

Bcs1, a homo-heptameric transmembrane AAA-ATPase that belongs to the superfamily of AAA proteins, acts as translocation machinery for the folded Rieske iron-sulfur protein (ISP, a key subunit of the mitochondrial complex-III) across the inner mitochondrial membrane. Cryo-EM structures depicted a ring-shaped transporter with two cavities, a cylindrical one formed by the ATPase domains on the matrix surface and a conical one formed by the transmembrane helices. Structures in different nucleotide states (ATPγS, ADP, APO) provided intuitive snapshots of the functional cycle, but the kinetics and structural transitions of the ATPase cycle remain entirely unknown. Here, using high-speed atomic force microscopy (HS-AFM) and HS-AFM line scanning (HS-AFM-LS), we characterized the ATPase cycle of Bcs1 at the single molecule level with sub-millisecond temporal resolution. We determined, in conditions where the molecules are in a continuous ATPase cycle turnover, that the ATP-bound conformational has a lifetime of ∼3700 ms while the lifetime of ADP/APO conformation is much shorter, ∼400 ms. By monitoring the conformational state of the two Bcs1 protomers on the two opposite sides of the ring at microsecond temporal resolution, we found that ATP-binding and -hydrolysis coupled conformational changes are highly concerted. We propose that the opening of the cone cavity to the intermembrane surface through the straightening of the transmembrane helices in the membrane is energetically costly and membrane elasticity drives the resealing of the gate. Overall, our results establish a working mechanism for the ATPase cycle of Bcs1 at the single molecule level and propose a mechanism for how Bcs1 can translocate a folded protein.

Dynamic correlations in lipid bilayer membranes over finite time intervals.

Recent single-molecule measurements [Schoch et al., Proc. Natl. Acad. Sci. U. S. A. 118, e2113202118 (2021)] have observed dynamic lipid-lipid correlations in membranes with submicrometer spatial resolution and submillisecond temporal resolution. While short from an instrumentation standpoint, these length and time scales remain long compared to microscopic molecular motions. Theoretical expressions are derived to infer experimentally measurable correlations from the two-body diffusion matrix appropriate for membrane-bound bodies coupled by hydrodynamic interactions. The temporal (and associated spatial) averaging resulting from finite acquisition times has the effect of washing out correlations as compared to naive predictions (i.e., the bare elements of the diffusion matrix), which would be expected to hold for instantaneous measurements. The theoretical predictions are shown to be in excellent agreement with Brownian dynamics simulations of experimental measurements. Numerical results suggest that the experimental measurement of membrane protein diffusion, in complement to lipid diffusion measurements, might help to resolve the experimental ambiguities encountered for certain black lipid membranes.

Machine learning-aided real-time detection of keyhole pore generation in laser powder bed fusion.

Porosity defects are currently a major factor that hinders the widespread adoption of laser-based metal additive manufacturing technologies. One common porosity occurs when an unstable vapor depression zone (keyhole) forms because of excess laser energy input. With simultaneous high-speed synchrotron x-ray imaging and thermal imaging, coupled with multiphysics simulations, we discovered two types of keyhole oscillation in laser powder bed fusion of Ti-6Al-4V. Amplifying this understanding with machine learning, we developed an approach for detecting the stochastic keyhole porosity generation events with submillisecond temporal resolution and near-perfect prediction rate. The highly accurate data labeling enabled by operando x-ray imaging allowed us to demonstrate a facile and practical way to adopt our approach in commercial systems.

Single-molecule imaging for probing the electrochemical interface

Single-molecule fluorescence microscopy (SMFM) has gained increasing attention in studying redox processes at the electrochemical interface due to its unique ability to probe individual redox molecules with sub-millisecond temporal resolution and nanometer spatial resolution. Here, we wish to highlight some of the major microscopy techniques which have been successfully used in imaging single molecules at the electrode surface. In addition to the common practice of detecting a change in the fluorescence signal caused by a faradaic reaction, one can also label and image single nanobubbles generated on the electrode surface with single-molecule fluorescence.

Resolving the Heat of Trimethylaluminum and Water Atomic Layer Deposition Half-Reactions.

Atomic layer deposition (ALD) is a surface synthesis technique that is characterized by self-limiting reactions between gas-phase precursors and a solid substrate. Although ALD processes have been demonstrated that span the periodic table, a greater understanding of the surface chemistry that affords ALD is necessary to enable greater precision, including area- and site-selective growth. We offer new insight into the thermodynamics and kinetics of the trimethylaluminum (TMA) and H2O ALD half-reactions with calibrated and time-resolved in situ pyroelectric calorimetry. The half-reactions produce 3.46 and 2.76 eV/Al heat, respectively, which is greater than the heat predicted by computational models based on crystalline Al2O3 substrates and closely aligned with the heat predicted by standard heats of formation. The pyroelectric thin-film calorimeter offers submillisecond temporal resolution that uniquely and clearly resolves precursor delivery and reaction kinetics. Both half-reactions are observed to exhibit multiple kinetic rates, with average TMA half-reaction rates at least 2 orders of magnitude faster than the H2O half-reaction kinetics. Comparing the experimental heat with published computational literature and additional first-principles modeling highlights the need to refine our models and mechanistic understanding of even the most ubiquitous ALD reactions.

Fast in-situ synchrotron X-ray imaging of the interfacial reaction during self-propagating exothermic reactive bonding

Self-Propagating Exothermic Reactive (SPER) bonding with lead-free solders is potentially attractive for microelectronics assembly due to its highly localised heating and minimal thermal loading of the components and substrates. The transient dynamics of melting, wetting, solidification and defect formation during SPER bonding were observed using in-situ synchrotron X-ray imaging with sub-millisecond temporal resolution and the results were further analysed using electron microscopy and thermal modelling. In-situ imaging revealed the preferential melting of the solder and subsequent wetting of the substrate. Numerous air bubbles were observed to form at the bonding interface. The distribution of these bubbles was found to vary with the thermal conductivity and wettability of the substrates. These bubbles appear to reduce the effectiveness of bonding by promoting the formation of cracks and voids within the solder joint. Our results show that metallisation layers on the bonding substrate can influence the dynamics of melting for the solder materials and thereby directly influence the reliability of SPER interconnects.

Ultrafast Force-Clamp Spectroscopy of Microtubule-Binding Proteins.

Optical trapping has been instrumental for deciphering translocation mechanisms of the force-generating cytoskeletal proteins. However, studies of the dynamic interactions between microtubules (MTs) and MT-associated proteins (MAPs) with no motor activity are lagging. Investigating the motility of MAPs that can diffuse along MT walls is a particular challenge for optical-trapping assays because thermally driven motions rely on weak and highly transient interactions. Three-bead, ultrafast force-clamp (UFFC) spectroscopy has the potential to resolve static and diffusive translocations of different MAPs with sub-millisecond temporal resolution and sub-nanometer spatial precision. In this report, we present detailed procedures for implementing UFFC, including setup of the optical instrument and feedback control, immobilization and functionalization of pedestal beads, and preparation of MT dumbbells. Example results for strong static interactions were generated using the Kinesin-7 motor CENP-E in the presence of AMP-PNP. Time resolution for MAP-MT interactions in the UFFC assay is limited by the MT dumbbell relaxation time, which is significantly longer than reported for analogous experiments using actin filaments. UFFC, however, provides a unique opportunity for quantitative studies on MAPs that glide along MTs under a dragging force, as illustrated using the kinetochore-associated Ska complex.

Pre-steady-state Kinetic Analysis of Amino Acid Transporter SLC6A14 Reveals Rapid Turnover Rate and Substrate Translocation.

SLC6A14 (solute carrier family 6 member 14) is an amino acid transporter, driven by Na+ and Cl− co-transport, whose structure, function, and molecular and kinetic mechanism have not been well characterized. Its broad substrate selectivity, including neutral and cationic amino acids, differentiates it from other SLC6 family members, and its proposed involvement in nutrient transport in several cancers suggest that it could become an important drug target. In the present study, we investigated SLC6A14 function and its kinetic mechanism after expression in human embryonic kidney (HEK293) cells, including substrate specificity and voltage dependence under various ionic conditions. We applied rapid solution exchange, voltage jumps, and laser photolysis of caged alanine, allowing sub-millisecond temporal resolution, to study SLC6A14 steady state and pre-steady state kinetics. The results highlight the broad substrate specificity and suggest that extracellular chloride enhances substrate transport but is not required for transport. As in other SLC6 family members, Na+ binding to the substrate-free transporter (or conformational changes associated with it) is electrogenic and is likely rate limiting for transporter turnover. Transient current decaying with a time constant of <1ms is also observed after rapid amino acid application, both in forward transport and homoexchange modes, indicating a slightly electrogenic, but fast and not rate-limiting substrate translocation step. Our results, which are consistent with kinetic modeling, suggest rapid transporter turnover rate and substrate translocation with faster kinetics compared with other SLC6 family members. Together, these results provided novel information on the SLC6A14 transport cycle and mechanism, expanding our understanding of SLC6A14 function.

Ultrafast four-dimensional imaging of cardiac mechanical wave propagation with sparse optoacoustic sensing

Propagation of electromechanical waves in excitable heart muscles follows complex spatiotemporal patterns holding the key to understanding life-threatening arrhythmias and other cardiac conditions. Accurate volumetric mapping of cardiac wave propagation is currently hampered by fast heart motion, particularly in small model organisms. Here we demonstrate that ultrafast four-dimensional imaging of cardiac mechanical wave propagation in entire beating murine heart can be accomplished by sparse optoacoustic sensing with high contrast, ∼115-µm spatial and submillisecond temporal resolution. We extract accurate dispersion and phase velocity maps of the cardiac waves and reveal vortex-like patterns associated with mechanical phase singularities that occur during arrhythmic events induced via burst ventricular electric stimulation. The newly introduced cardiac mapping approach is a bold step toward deciphering the complex mechanisms underlying cardiac arrhythmias and enabling precise therapeutic interventions.

Recent insights into the relative timing of myosin's powerstroke and release of phosphate.

Myosin is a motor enzyme that converts the chemical energy in ATP into mechanical work to drive a myriad of intracellular processes, from muscle contraction to vesicular transport. Key steps in the transduction of energy are the force-generating powerstroke, and the release of phosphate (Pi ) from the nucleotide-binding site. Both events occur rapidly after binding to actin, making it difficult to determine which event occurs first. Early efforts suggested that these events occur simultaneously; however, recent findings indicate that they are separate and distinct events that occur at different rates. High-resolution crystal structures of myosin captured in intermediate states of the ATPase cycle suggest that when Pi is in the active site it prevents the powerstroke from occurring, leading to the hypothesis that Pi -release precedes the powerstroke. However, advances in functional assays, enabling sub-millisecond temporal and nanometer spatial resolution, are challenging this hypothesis. For example, Föster Resonance Energy Transfer (FRET) based assays, as well as single molecule laser trap assays, suggest the opposite; that the powerstroke occurs prior to the release of Pi from myosin's active site. This review provides some historical context and then highlights recent reports that reveal exciting new insight into this fundamental mechanism of energy transduction by this prototypical motor enzyme.

Voltage-Gated Potassium Channels Ensure Action Potential Shape Fidelity in Distal Axons.

The initiation and propagation of the action potential (AP) along an axon allows neurons to convey information rapidly and across distant sites. Although AP properties have typically been characterized at the soma and proximal axon, knowledge of the propagation of APs toward distal axonal domains of mammalian CNS neurons remains limited. We used genetically encoded voltage indicators (GEVIs) to image APs with submillisecond temporal resolution simultaneously at different locations along the long axons of dissociated hippocampal neurons from rat embryos of either sex. We found that APs became sharper and showed remarkable fidelity as they traveled toward distal axons, even during a high-frequency train. Blocking voltage-gated potassium channels (Kv) with 4-AP resulted in an increase in AP width in all compartments, which was stronger at distal locations and exacerbated during AP trains. We conclude that the higher levels of Kv channel activity in distal axons serve to sustain AP fidelity, conveying a reliable digital signal to presynaptic boutons.SIGNIFICANCE STATEMENT The AP represents the electrical signal carried along axons toward distant presynaptic boutons where it culminates in the release of neurotransmitters. The nonlinearities involved in this process are such that small changes in AP shape can result in large changes in neurotransmitter release. Since axons are remarkably long structures, any distortions that APs suffer along the way have the potential to translate into a significant modulation of synaptic transmission, particularly in distal domains. To avoid these issues, distal axons have ensured that signals are kept remarkably constant and insensitive to modulation during a train, despite the long distances traveled. Here, we uncover the mechanisms that allow distal axonal domains to provide a reliable and faithful digital signal to presynaptic terminals.

Visualization of Human Aortic Valve Dynamics Using Magnetic Resonance Imaging with Sub-Millisecond Temporal Resolution.

Visualization of aortic valve dynamics is important in diagnosing valvular diseases but is challenging to perform with magnetic resonance imaging (MRI) due to the limited temporal resolution. To develop an MRI technique with sub-millisecond temporal resolution and demonstrate its application in visualizing rapid aortic valve opening and closing in human subjects in comparison with echocardiography and conventional MRI techniques. Prospective. Twelve healthy subjects. 3 T; gradient-echo-train-based sub-millisecond periodic event encoded imaging (get-SPEEDI) and balanced steady-state free precession (bSSFP). Images were acquired using get-SPEEDI with a temporal resolution of 0.6 msec. get-SPEEDI was triggered by an electrocardiogram so that each echo in the gradient echo train corresponded to an image at a specific time point, providing a time-resolved characterization of aortic valve dynamics. For comparison, bSSFP was also employed with 12 msec and 24 msec temporal resolutions, respectively. The durations of the aortic valve rapid opening (Tro ), rapid closing (Trc ), and the maximal aortic valve area (AVA) normalized to height were measured with all three temporal resolutions. M-mode echocardiograms with a temporal resolution of 0.8 msec were obtained for further comparison. Parameters were compared between the three sequences, together with the echocardiography results, with a Mann-Whitney U test. Significantly shorter Tro (mean ± SD: 27.5 ± 6.7 msec) and Trc (43.8 ± 11.6 msec) and larger maximal AVA/height (2.01 ± 0.29 cm2 /m) were measured with get-SPEEDI compared to either bSSFP sequence (Tro of 56.3 ± 18.8 and 63.8 ± 20.2 msec; Trc of 68.2 ± 16.6 and 72.8 ± 18.2 msec; maximal AVA/height of 1.63 ± 0.28 and 1.65 ± 0.32 cm2 /m for 12 msec and 24 msec temporal resolutions, respectively, P < 0.05). In addition, the get-SPEEDI results were more consistent with those measured using echocardiography, especially for Tro (29.0 ± 4.1 msec, P=0.79) and Trc (41.6 ± 4.3 msec, P=0.16). DATA CONCLUSION: get-SPEEDI allows for visualization of human aortic valve dynamics and provided values closer to those measured using echocardiography than the bSSFP sequences. 1 TECHNICAL EFFICACY STAGE: 1.

Magnetic resonance imaging with submillisecond temporal resolution.

To demonstrate an MRI technique-Submillisecond Periodic Event Encoded Dynamic Imaging (SPEEDI)-for capturing cyclic dynamic events with submillisecond temporal resolution. The SPEEDI technique is based on an FID or an echo signal in which each time point in the signal is used to sample a distinct k-space raster, followed by repeated FIDs or echoes to produce the remaining k-space data in each k-space raster. All acquisitions are synchronized with a cyclic event, resulting in a set of time-resolved images of the cyclic event with a temporal resolution determined by the dwell time. In SPEEDI, spatial encoding is accomplished by phase encoding. The SPEEDI technique was demonstrated in two experiments at 3 T to (1) visualize fast-changing electric currents that mimicked the waveform of an action potential, and (2) characterize rapidly decaying eddy currents in an MRI system, with a temporal resolution of 0.2 ms and 0.4 ms, respectively. In both experiments, compressed sensing was incorporated to reduce the scan times. Phase difference maps related to the dynamics of electric currents or eddy currents were then obtained. In the first experiment, time-resolved phase maps resulting from the action potential-mimicking current waveform were successfully obtained and agreed well with theoretical calculations (normalized RMS error = 0.07). In the second experiment, spatially resolved eddy current phase maps revealed time constants (27.1 ± 0.2 ms, 41.1 ± 3.5 ms, and 34.8 ± 0.7 ms) that matched well with those obtained from an established method using point sources (26.4 ms, 41.2 ms and 34.8 ms). For both experiments, phase maps from fully sampled and compressed-sensing-accelerated k-space data exhibited a high structural similarity (> 0.8) despite a two-fold to three-fold acceleration. We have illustrated that SPEEDI can provide submillisecond temporal resolution. This capability will likely lead to future exploration of ultrafast, cyclic biomedical processes using MRI.

Limits of subjective and objective vection for ultra-high frame rate visual displays

Large-field optic flow generates the illusory percept of self-motion, termed ‘vection’. Smoother visual motion displays generate a more compelling subjective sense of vection and objective postural responses, as well as a greater sense of immersiveness for the user observing the visual display. Research suggests that the function linking frame rate and vection asymptotes at 60 frames per second (FPS), but previous studies have used only moderate frame rates that do not approach the limits of human perception. Here, we measure vection using subjective and objective (mean frequency and path length of postural centre-of-pressure (COP) excursions) responses following the presentation of high-contrast optic flow stimuli at slow and fast speeds and low and ultra-high frame rates. We achieve this using a novel rendering method implemented with a projector capable of sub-millisecond temporal resolution in order to simulate refresh rates ranging from very low (15 FPS) to ultra-high frame rates (480 FPS). The results suggest that subjective vection was experienced most strongly at 60 FPS. Below and above 60 FPS, subjective vection is generally weaker, shorter, and starts later, although this pattern varied slightly according to the speed of stimuli. For objective measures, while the frequency of postural sway was unaffected by frame rate, COP path length was greatest for 480 FPS stimuli. Together, our results support diminishing returns for vection above 60 FPS and provide insight into the use of high frame rate for enhancing the user experience in visual displays.

MEG for Greater Sensitivity and More Precise Localization in Epilepsy.

Magnetoencephalography is the noninvasive measurement of miniscule magnetic fields produced by brain electrical currents, and is used most fruitfully to evaluate epilepsy patients. While other modalities infer brain function indirectly by measuring changes in blood flow, metabolism, and oxygenation, magnetoencephalography measures neuronal and synaptic function directly with submillisecond temporal resolution. The brain's magnetic field is recorded by neuromagnetometers surrounding the head in a helmet-shaped sensor array. Because magnetic signals are not distorted by anatomy, magnetoencephalography allows for a more accurate measurement and localization of brain activities than electroencephalography. Magnetoencephalography has become an indispensable part of the armamentarium at epilepsy centers.

Three-dimensional fuzzy graphene ultra-microelectrodes for subcellular electrical recordings

Microelectrode arrays (MEAs) have enabled investigation of cellular networks at sub-millisecond temporal resolution. However, current MEAs are limited by the large electrode footprint since reducing the electrode's geometric area to sub-cellular dimensions leads to a significant increase in impedance thus affecting its recording capabilities. We report a breakthrough ultra-microelectrodes platform by leveraging the outstanding surface-to-volume ratio of nanowire-templated out-of-plane synthesized three-dimensional fuzzy graphene (NT-3DFG). The enormous surface area of NT-3DFG leads to 140-fold reduction in electrode impedance compared to bare Au microelectrodes, thus enabling scaling down the geometric size by 625-fold to ca. 2 μm × 2 μm. The out-of-plane morphology of NT-3DFG leads to a tight seal with the cell membrane thus enabling recording of electrical signals with high signal-to-noise ratio (SNR) of > 6. This work highlights the possibility to push the limits of the conventional MEA technology to enable electrophysiological investigation at sub-cellular level without the need of any surface coatings. This presented approach would greatly impact our basic understanding of signal transduction within a single cell as well as complex cellular assemblies.