Double-µPeriscope, a tool for multilayer optical recordings, optogenetic stimulations or both.

Intelligent behavior and cognitive functions in mammals depend on cortical microcircuits made up of a variety of excitatory and inhibitory cells that form a forest-like complex across six layers. Mechanistic understanding of cortical microcircuits requires both manipulation and monitoring of multiple layers and interactions between them. However, existing techniques are limited as to simultaneous monitoring and stimulation at different depths without damaging a large volume of cortical tissue. Here, we present a relatively simple and versatile method for delivering light to any two cortical layers simultaneously. The method uses a tiny optical probe consisting of two microprisms mounted on a single shaft. We demonstrate the versatility of the probe in three sets of experiments: first, two distinct cortical layers were optogenetically and independently manipulated; second, one layer was stimulated while the activity of another layer was monitored; third, the activity of thalamic axons distributed in two distinct cortical layers was simultaneously monitored in awake mice. Its simple-design, versatility, small-size, and low-cost allow the probe to be applied widely to address important biological questions.

Optogenetic therapy: high spatiotemporal resolution and pattern discrimination compatible with vision restoration in non-human primates.

Vision restoration is an ideal medical application for optogenetics, because the eye provides direct optical access to the retina for stimulation. Optogenetic therapy could be used for diseases involving photoreceptor degeneration, such as retinitis pigmentosa or age-related macular degeneration. We describe here the selection, in non-human primates, of a specific optogenetic construct currently tested in a clinical trial. We used the microbial opsin ChrimsonR, and showed that the AAV2.7m8 vector had a higher transfection efficiency than AAV2 in retinal ganglion cells (RGCs) and that ChrimsonR fused to tdTomato (ChR-tdT) was expressed more efficiently than ChrimsonR. Light at 600 nm activated RGCs transfected with AAV2.7m8 ChR-tdT, from an irradiance of 1015 photons.cm-2.s-1. Vector doses of 5 × 1010 and 5 × 1011 vg/eye transfected up to 7000 RGCs/mm2 in the perifovea, with no significant immune reaction. We recorded RGC responses from a stimulus duration of 1 ms upwards. When using the recorded activity to decode stimulus information, we obtained an estimated visual acuity of 20/249, above the level of legal blindness (20/400). These results lay the groundwork for the ongoing clinical trial with the AAV2.7m8 - ChR-tdT vector for vision restoration in patients with retinitis pigmentosa.

Phasic Activation of Dorsal Raphe Serotonergic Neurons Increases Pupil Size.

Transient variations in pupil size (PS) under constant luminance are coupled to rapid changes in arousal state,1-3 which have been interpreted as vigilance,4 salience,5 or a surprise signal.6-8 Neural control of such fluctuations presumably involves multiple brain regions5,9-11 and neuromodulatory systems,3,12,13 but it is often associated with phasic activity of the noradrenergic system.9,12,14,15 Serotonin (5-HT), a neuromodulator also implicated in aspects of arousal16 such as sleep-wake transitions,17 motivational state regulation,18 and signaling of unexpected events,19 seems to affect PS,20-24 but these effects have not been investigated in detail. Here we show that phasic 5-HT neuron stimulation causes transient PS changes. We used optogenetic activation of 5-HT neurons in the dorsal raphe nucleus (DRN) of head-fixed mice performing a foraging task. 5-HT-driven modulations of PS were maintained throughout the photostimulation period and sustained for a few seconds after the end of stimulation. We found no evidence that the increase in PS with activation of 5-HT neurons resulted from interactions of photostimulation with behavioral variables, such as locomotion or licking. Furthermore, we observed that the effect of 5-HT on PS depended on the level of environmental uncertainty, consistent with the idea that 5-HT could report a surprise signal.19 These results advance our understanding of the neuromodulatory control of PS, revealing a tight relationship between phasic activation of 5-HT neurons and changes in PS.

Stimulus-Driven Retinal Intrinsic Signal Optical Imaging in Mouse Demonstrates a Dominant Rod-Driven Component.

Purpose: The primary hypotheses tested are that (1) there exist stimulus-driven intrinsic optical signals in the mouse retina similar to those previously observed in other species, and (2) these optical signals require an intact rod photoreceptor phototransduction cascade. Methods: We used 38 wild-type C57BL6J mice and 18 genetic knockout Gnat1-/- mice to study the light-evoked retinal intrinsic response. A custom mouse fundus camera delivered visual stimuli and collected mouse retinal imaging data of changes in retinal reflectance for further analysis. The retina was stimulated in the high-mesopic range with a 505-nm light-emitting diode while also being illuminated with 780-nm near-infrared light. Results: Wild-type C57BL6J mice yielded retinal imaging signals that typically showed a stimulus-driven decrease in retinal reflectance of ∼0.1%, with a time course of several seconds. The signals exhibit spatial specificity in the retina. Overall, the mouse imaging signals are similar in sign and time course to those reported in other mammalian species but are of lower amplitude. In contrast, functional retinal imaging of Gnat1-/- mice that lack a functional rod transducin yielded no such stimulus-driven signals. Conclusions: Previous studies have not shown which pathway component is essential for the generation of these imaged signals. The absence of the intrinsic signal responses in Gnat1-/- knockout mice indicates that a functional rod transducin is likely to be necessary for generating the retinal intrinsic signals. These studies, to the best of our knowledge, demonstrate for the first time in vivo mouse retinal functional imaging signals similar to those previously shown in other mammalian species.

Perception of the difference between past and present stimulus: A rare orientation illusion may indicate incidental access to prediction error-like signals.

A popular model for sensory processing, known as predictive coding, proposes that incoming signals are iteratively compared with top-down predictions along a hierarchical processing scheme. At each step, error signals arising from differences between actual input and prediction are forwarded and recurrently minimized by updating internal models to finally be "explained away". However, the neuronal mechanisms underlying such computations and their limitations in processing speed are largely unknown. Further, it remains unclear at which step of cortical processing prediction errors are explained away, if at all. In the present study, human subjects briefly viewed the superposition of two orthogonally oriented gratings followed by abrupt removal of one orientation after either 33 or 200 milliseconds. Instead of strictly seeing the remaining orientation, observers report rarely but highly significantly an illusory percept of the arithmetic difference between previous and actual orientations. Previous findings in cats using the identical paradigm suggest that such difference signals are inherited from first steps of visual cortical processing. In light of early modeling accounts of predictive coding, in which visual neurons were interpreted as residual error detectors signaling the difference between actual input and its temporal prediction based on past input, our data may indicate continued access to residual errors. Such strategy permits time-critical perceptual decision making across a spectrum of competing internal signals up to the highest levels of processing. Thus, the occasional appearance of a prediction error-like illusory percept may uncover maintained flexibility at perceptual decision stages when subjects cope with highly dynamic and ambiguous visual stimuli.

Accuracy and precision of stimulus timing and reaction times with Unreal Engine and SteamVR.

The increasing interest in Virtual Reality (VR) as a tool for neuroscientific research contrasts with the current lack of established toolboxes and standards. In several recent studies, game engines like Unity or Unreal Engine were used. It remains to be tested whether these software packages provide sufficiently precise and accurate stimulus timing and time measurements that allow inferring ongoing mental and neural processes. We here investigated the precision and accuracy of the timing mechanisms of Unreal Engine 4 and SteamVR in combination with the HTC Vive VR system. In a first experiment, objective external measures revealed that stimulus durations were highly accurate. In contrast, in a second experiment, the assessment of the precision of built-in timing procedures revealed highly variable reaction time measurements and inaccurate determination of stimulus onsets. Hence, we developed a new software-based method that allows precise and accurate reaction time measurements with Unreal Engine and SteamVR. Instead of using the standard timing procedures implemented within Unreal Engine, time acquisition was outsourced to a background application. Timing benchmarks revealed that the newly developed method allows reaction time measurements with a precision and accuracy in the millisecond range. Overall, the present results indicate that the HTC Vive together with Unreal Engine and SteamVR can achieve high levels of precision and accuracy both concerning stimulus duration and critical time measurements. The latter can be achieved using a newly developed routine that allows not only accurate reaction time measures but also provides precise timing parameters that can be used in combination with time-sensitive functional measures such as electroencephalography (EEG) or transcranial magnetic stimulation (TMS).

Active inference as a unifying, generic and adaptive framework for a P300-based BCI.

OBJECTIVE: Going adaptive is a major challenge for the field of brain-computer interface (BCI). This entails a machine that optimally articulates inference about the user's intentions and its own actions. Adaptation can operate over several dimensions which calls for a generic and flexible framework. APPROACH: We appeal to one of the most comprehensive computational approach to brain (adaptive) functions: the active inference (AI) framework. It entails an explicit (probabilistic) model of the user that the machine interacts with, here involved in a P300-spelling task. This takes the form of a discrete input-output state-space model establishing the link between the machine's (i) observations-a P300 or error potential for instance, (ii) representations-of the user intentions to spell or pause, and (iii) actions-to flash, spell or switch-off the application. MAIN RESULTS: Using simulations with real EEG data from 18 subjects, results demonstrate the ability of AI to yield a significant increase in bit rate (17%) over state-of-the-art approaches, such as dynamic stopping. SIGNIFICANCE: Thanks to its flexibility, this one model enables to implement optimal (dynamic) stopping but also optimal flashing (i.e. active sampling), automated error correction, and switching off when the user does not look at the screen anymore. Importantly, this approach enables the machine to flexibly arbitrate between all these possible actions. We demonstrate AI as a unifying and generic framework to implement a flexible interaction behaviour in a given BCI context.

Enhancement and Suppression Flexibly Guide Attention.

Previous research suggests that observers can suppress salient-but-irrelevant stimuli in a top-down manner. However, one question left unresolved is whether such suppression is, in fact, solely due to distractor-feature suppression or whether it instead also reflects some degree of target-feature enhancement. The present study (N = 60) addressed this issue. On search trials (70% of trials), participants searched for a shape target when an irrelevant color singleton was either present or absent; performance was better when a color singleton was present. On interleaved probe trials (30% of trials), participants searched for a letter target. Responses were faster for the letter on a target-colored item than on a neutral-colored item, whereas responses were slower for the letter on a distractor-colored item than on a neutral-colored item. The results demonstrate that target-feature enhancement and distractor-feature suppression contribute to attentional guidance independently; enhancement and suppression flexibly guide attention as the occasion demands.

Transparent multispectral photodetectors mimicking the human visual system.

Compact and lightweight photodetection elements play a critical role in the newly emerging augmented reality, wearable and sensing technologies. In these technologies, devices are preferred to be transparent to form an optical interface between a viewer and the outside world. For this reason, it is of great value to create detection platforms that are imperceptible to the human eye directly onto transparent substrates. Semiconductor nanowires (NWs) make ideal photodetectors as their optical resonances enable parsing of the multi-dimensional information carried by light. Unfortunately, these optical resonances also give rise to strong, undesired light scattering. In this work, we illustrate how a new optical resonance arising from the radiative coupling between arrayed silicon NWs can be harnessed to remove reflections from dielectric interfaces while affording spectro-polarimetric detection. The demonstrated transparent photodetector concept opens up promising platforms for transparent substrates as the base for opto-electronic devices and in situ optical measurement systems.

An arbitrary-spectrum spatial visual stimulator for vision research.

Visual neuroscientists require accurate control of visual stimulation. However, few stimulator solutions simultaneously offer high spatio-temporal resolution and free control over the spectra of the light sources, because they rely on off-the-shelf technology developed for human trichromatic vision. Importantly, consumer displays fail to drive UV-shifted short wavelength-sensitive photoreceptors, which strongly contribute to visual behaviour in many animals, including mice, zebrafish and fruit flies. Moreover, many non-mammalian species feature more than three spectral photoreceptor types. Here, we present a flexible, spatial visual stimulator with up to six arbitrary spectrum chromatic channels. It combines a standard digital light processing engine with open source hard- and software that can be easily adapted to the experimentalist's needs. We demonstrate the capability of this general visual stimulator experimentally in the in vitro mouse retinal whole-mount and the in vivo zebrafish. With this work, we intend to start a community effort of sharing and developing a common stimulator design for vision research.

Interneuronal correlations at longer time scales predict decision signals for bistable structure-from-motion perception.

Perceptual decisions are thought to depend on the activation of task-relevant neurons, whose activity is often correlated in time. Here, we examined how the temporal structure of shared variability in neuronal firing relates to perceptual choices. We recorded stimulus-selective neurons from visual area V5/MT while two monkeys (Macaca mulatta) made perceptual decisions about the rotation direction of structure-from-motion cylinders. Interneuronal correlations for a perceptually ambiguous cylinder stimulus were significantly higher than those for unambiguous cylinders or for random 2D motion during passive viewing. Much of the difference arose from correlations at relatively long timescales (hundreds of milliseconds). Choice-related neural activity (quantified as choice probability; CP) for ambiguous cylinders was positively correlated with interneuronal correlations and was specifically associated with their long timescale component. Furthermore, the slope of the long timescale - but not the instantaneous - component of the correlation predicted higher CPs towards the end of the trial i.e. close to the decision. Our results suggest that the perceptual stability of structure-from-motion cylinders may be controlled by enhanced interneuronal correlations on longer timescales. We propose this as a potential signature of top-down influences onto V5/MT processing that shape and stabilize the appearance of 3D-motion percepts.

Ventral midbrain stimulation induces perceptual learning and cortical plasticity in primates.

Practice improves perception and enhances neural representations of trained visual stimuli, a phenomenon known as visual perceptual learning (VPL). While attention to task-relevant stimuli plays an important role in such learning, Pavlovian stimulus-reinforcer associations are sufficient to drive VPL, even subconsciously. It has been proposed that reinforcement facilitates perceptual learning through the activation of neuromodulatory centers, but this has not been directly confirmed in primates. Here, we paired task-irrelevant visual stimuli with microstimulation of a dopaminergic center, the ventral tegmental area (VTA), in macaques. Pairing VTA microstimulation with a task-irrelevant visual stimulus increased fMRI activity and improved classification of fMRI activity patterns selectively for the microstimulation-paired stimulus. Moreover, pairing VTA microstimulation with a task-irrelevant visual stimulus improved the subject's capacity to discriminate that stimulus. This is the first causal demonstration of the role of neuromodulatory centers in VPL in primates.

Multifunctional multi-shank neural probe for investigating and modulating long-range neural circuits in vivo.

Investigation and modulation of neural circuits in vivo at the cellular level are very important for studying functional connectivity in a brain. Recently, neural probes with stimulation capabilities have been introduced, and they provided an opportunity for studying neural activities at a specific region in the brain using various stimuli. However, previous methods have a limitation in dissecting long-range neural circuits due to inherent limitations on their designs. Moreover, the large size of the previously reported probes induces more significant tissue damage. Herein, we present a multifunctional multi-shank MEMS neural probe that is monolithically integrated with an optical waveguide for optical stimulation, microfluidic channels for drug delivery, and microelectrode arrays for recording neural signals from different regions at the cellular level. In this work, we successfully demonstrated the functionality of our probe by confirming and modulating the functional connectivity between the hippocampal CA3 and CA1 regions in vivo.

Moving base driving simulators' potential for carsickness research.

We investigated whether motion sickness analogous to carsickness can be studied in a moving base simulator, despite the limited motion envelope. Importantly, to avoid simulator sickness, vision outside the simulator cabin was restricted. Participants (N = 16) were exposed blindfolded to 15-min lateral sinusoidal motion at 0.2 Hz and 0.35 Hz on separate days. These conditions were selected to realize optimal provocativeness of the stimulus given the simulator's maximum displacement and knowledge on frequency-acceleration interactions for motion sickness. Average motion sickness on an 11-point scale was 2.21 ±â€¯1.97 for 0.2 Hz and 1.93 ±â€¯1.94 for 0.35 Hz. The motion sickness increase over time was comparable to that found in studies using actual vehicles. We argue that motion base simulators can be used to incite motion sickness analogous to carsickness, provided considerable restrictions on vision. Future research on carsickness, potentially more prevalent in autonomous vehicles, could benefit from employing simulators.

Instrumentation for electrodiagnostic studies.

The instrument and accessories are an important part of the electrodiagnostic (EDX) testing. Their functional understanding is useful to recognize and reduce various artifacts and noise/interference in the signal. In this review, we will describe the technical specifications of various components of the instrument, and their effect on signals and noise. This will be illustrated using example of electromyography and nerve conduction studies. However the same principles also apply to other modalities of testing. We will also provide general strategies to reduce noise and artifacts, followed by some modality specific examples.

Methods for Presenting Real-world Objects Under Controlled Laboratory Conditions.

Our knowledge of human object vision is based almost exclusively on studies in which the stimuli are presented in the form of computerized two-dimensional (2-D) images. In everyday life, however, humans interact predominantly with real-world solid objects, not images. Currently, we know very little about whether images of objects trigger similar behavioral or neural processes as do real-world exemplars. Here, we present methods for bringing the real-world into the laboratory. We detail methods for presenting rich, ecologically-valid real-world stimuli under tightly-controlled viewing conditions. We describe how to match closely the visual appearance of real objects and their images, as well as novel apparatus and protocols that can be used to present real objects and computerized images on successively interleaved trials. We use a decision-making paradigm as a case example in which we compare willingness-to-pay (WTP) for real snack foods versus 2-D images of the same items. We show that WTP increases by 6.6% for food items displayed as real objects versus high-resolution 2-D colored images of the same foods -suggesting that real foods are perceived as being more valuable than their images. Although presenting real object stimuli under controlled conditions presents several practical challenges for the experimenter, this approach will fundamentally expand our understanding of the cognitive and neural processes that underlie naturalistic vision.

Effects of stimulation frequency and stimulation waveform on steady-state visual evoked potentials using a computer monitor.

OBJECTIVE: A visual stimulator plays a vital part in brain-computer interfaces (BCIs) based on steady-state visual evoked potential (SSVEP). The properties of visual stimulation, such as frequency, color, and waveform, will influence SSVEP-based BCI performance to some extent. Recently, the computer monitor serves as a visual stimulator that is widespread in SSVEP-based BCIs because of its great flexibility in generating visual stimuli. However, stimulation properties based on a computer monitor have received very little attention. For a better comprehension of SSVEPs, this study explored the stimulation effects of waveforms and frequencies, when evoking SSVEPs through a computer monitor. APPROACH: This study utilized the approximation methods to realize sine- and square-wave temporal modulations at 18 stimulation frequencies ranging from 6 to 40 Hz on a conventional 120 Hz LCD screen. We collected electroencephalogram (EEG) datasets from 12 healthy subjects and compared the signal-to-noise ratios (SNRs), amplitudes, and topographic mapping of SSVEPs evoked by these two temporal modulation flickers (sine- and square-wave). In addition, a BCI experiment with two nine-target BCIs (i.e. low-frequency BCI and high-frequency BCI) was implemented to compare the two stimulation waveforms in terms of BCI performance. MAIN RESULTS: For both sine- and square-wave stimulation conditions, strong SSVEPs over the occipital area were observed for each stimulation frequency. SSVEP amplitudes at the stimulation frequency exhibited a global peak in the low-frequency band. The second harmonic SSVEP frequency-response functions showed the largest amplitude at 6 Hz and fell sharply for higher frequencies. In the BCI experiment, the classification performance of the square-wave stimuli was notably higher than that of the sine-wave stimuli when using shorter data lengths. SIGNIFICANCE: These results suggested that the square-wave flicker was more efficient at implementing high-speed BCIs based on SSVEP when using a computer monitor as a visual stimulator.

Interactions between egocentric and allocentric spatial coding of sounds revealed by a multisensory learning paradigm.

Although sound position is initially head-centred (egocentric coordinates), our brain can also represent sounds relative to one another (allocentric coordinates). Whether reference frames for spatial hearing are independent or interact remained largely unexplored. Here we developed a new allocentric spatial-hearing training and tested whether it can improve egocentric sound-localisation performance in normal-hearing adults listening with one ear plugged. Two groups of participants (N = 15 each) performed an egocentric sound-localisation task (point to a syllable), in monaural listening, before and after 4-days of multisensory training on triplets of white-noise bursts paired with occasional visual feedback. Critically, one group performed an allocentric task (auditory bisection task), whereas the other processed the same stimuli to perform an egocentric task (pointing to a designated sound of the triplet). Unlike most previous works, we tested also a no training group (N = 15). Egocentric sound-localisation abilities in the horizontal plane improved for all groups in the space ipsilateral to the ear-plug. This unexpected finding highlights the importance of including a no training group when studying sound localisation re-learning. Yet, performance changes were qualitatively different in trained compared to untrained participants, providing initial evidence that allocentric and multisensory procedures may prove useful when aiming to promote sound localisation re-learning.

A mm-Sized Free-Floating Wirelessly Powered Implantable Optical Stimulation Device.

This paper presents a mm-sized, free-floating, wirelessly powered, implantable optical stimulation (FF-WIOS) device for untethered optogenetic neuromodulation. A resonator-based three-coil inductive link creates a homogeneous magnetic field that continuously delivers sufficient power (>2.7 mW) at an optimal carrier frequency of 60 MHz to the FF-WIOS in the near field without surpassing the specific absorption rate limit, regardless of the position of the FF-WIOS in a large brain area. Forward data telemetry carries stimulation parameters by on-off-keying the power carrier at a data rate of 50 kb/s to selectively activate a 4 × 4 µLED array. Load-shift-keying back telemetry controls the wireless power transmission by reporting the FF-WIOS received power level in a closed-loop power control mechanism. LEDs typically require high instantaneous power to emit sufficient light for optical stimulation. Thus, a switched-capacitor-based stimulation architecture is used as an energy storage buffer with one off-chip capacitor to receive charge directly from the inductive link and deliver it to the selected µLED at the onset of stimulation. The FF-WIOS system-on-a-chip prototype, fabricated in a 0.35-µm standard CMOS process, charges a 10-µF capacitor up to 5 V with 37% efficiency and passes instantaneous current spikes up to 10 mA in the selected µLED, creating a bright exponentially decaying flash with minimal wasted power. An in vivo experiment was conducted to verify the efficacy of the FF-WIOS by observing light-evoked local field potentials and immunostained tissue response from the primary visual cortex (V1) of two anesthetized rats.

A flexible geometry for panoramic visual and optogenetic stimulation during behavior and physiology.

BACKGROUND: To study visual processing, it is necessary to precisely control visual stimuli while recording neural and behavioral responses. It can be important to present stimuli over a broad area of the visual field, which can be technically difficult. NEW METHOD: We present a simple geometry that can be used to display panoramic stimuli. A single digital light projector generates images that are reflected by mirrors onto flat screens that surround an animal. It can be used for behavioral and neurophysiological measurements, so virtually identical stimuli can be presented. Moreover, this geometry permits light from the projector to be used to activate optogenetic tools. RESULTS: Using this geometry, we presented panoramic visual stimulation to Drosophila in three paradigms. We presented drifting contrast gratings while recording walking and turning speed. We used the same projector to activate optogenetic channels during visual stimulation. Finally, we used two-photon microscopy to record responses in direction-selective cells to drifting gratings. COMPARISON WITH EXISTING METHOD(S): Existing methods have typically required custom hardware or curved screens, while this method requires only flat back projection screens and a digital light projector. The projector generates images in real time and does not require pre-generated images. Finally, while many setups are large, this geometry occupies a 30 × 20 cm footprint with a 25 cm height. CONCLUSIONS: This flexible geometry enables measurements of behavioral and neural responses to panoramic stimuli. This allows moderate throughput behavioral experiments with simultaneous optogenetic manipulation, with easy comparisons between behavior and neural activity using virtually identical stimuli.