Each of our eyes normally sees a slightly different image of the world around us. The brain can combine these two images into a single coherent representation. However, when the eyes are presented with images that are sufficiently different from each other, an interesting thing happens: Rather than fusing the two images into a combined conscious percept, what transpires is a pattern of perceptual alternations where one image dominates awareness while the other is suppressed; dominance alternates between the two images, typically every few seconds. This perceptual phenomenon is known as binocular rivalry. Binocular rivalry is considered useful for studying perceptual selection and awareness in both human and animal models, because unchanging visual input to each eye leads to alternations in visual awareness and perception. To create a binocular rivalry stimulus, all that is necessary is to present each eye with a different image at the same perceived location. There are several ways of doing this, but newcomers to the field are often unsure which method would best suit their specific needs. The purpose of this article is to describe a number of inexpensive and straightforward ways to create and use binocular rivalry. We detail methods that do not require expensive specialized equipment and describe each method's advantages and disadvantages. The methods described include the use of red-blue goggles, mirror stereoscopes and prism goggles.
20 Related JoVE Articles!
Simultaneous Multicolor Imaging of Biological Structures with Fluorescence Photoactivation Localization Microscopy
Institutions: University of Maine.
Localization-based super resolution microscopy can be applied to obtain a spatial map (image) of the distribution of individual fluorescently labeled single molecules within a sample with a spatial resolution of tens of nanometers. Using either photoactivatable (PAFP) or photoswitchable (PSFP) fluorescent proteins fused to proteins of interest, or organic dyes conjugated to antibodies or other molecules of interest, fluorescence photoactivation localization microscopy (FPALM) can simultaneously image multiple species of molecules within single cells. By using the following approach, populations of large numbers (thousands to hundreds of thousands) of individual molecules are imaged in single cells and localized with a precision of ~10-30 nm. Data obtained can be applied to understanding the nanoscale spatial distributions of multiple protein types within a cell. One primary advantage of this technique is the dramatic increase in spatial resolution: while diffraction limits resolution to ~200-250 nm in conventional light microscopy, FPALM can image length scales more than an order of magnitude smaller. As many biological hypotheses concern the spatial relationships among different biomolecules, the improved resolution of FPALM can provide insight into questions of cellular organization which have previously been inaccessible to conventional fluorescence microscopy. In addition to detailing the methods for sample preparation and data acquisition, we here describe the optical setup for FPALM. One additional consideration for researchers wishing to do super-resolution microscopy is cost: in-house setups are significantly cheaper than most commercially available imaging machines. Limitations of this technique include the need for optimizing the labeling of molecules of interest within cell samples, and the need for post-processing software to visualize results. We here describe the use of PAFP and PSFP expression to image two protein species in fixed cells. Extension of the technique to living cells is also described.
Basic Protocol, Issue 82, Microscopy, Super-resolution imaging, Multicolor, single molecule, FPALM, Localization microscopy, fluorescent proteins
Long-term Behavioral Tracking of Freely Swimming Weakly Electric Fish
Institutions: University of Ottawa, University of Ottawa, University of Ottawa.
Long-term behavioral tracking can capture and quantify natural animal behaviors, including those occurring infrequently. Behaviors such as exploration and social interactions can be best studied by observing unrestrained, freely behaving animals. Weakly electric fish (WEF) display readily observable exploratory and social behaviors by emitting electric organ discharge (EOD). Here, we describe three effective techniques to synchronously measure the EOD, body position, and posture of a free-swimming WEF for an extended period of time. First, we describe the construction of an experimental tank inside of an isolation chamber designed to block external sources of sensory stimuli such as light, sound, and vibration. The aquarium was partitioned to accommodate four test specimens, and automated gates remotely control the animals' access to the central arena. Second, we describe a precise and reliable real-time EOD timing measurement method from freely swimming WEF. Signal distortions caused by the animal's body movements are corrected by spatial averaging and temporal processing stages. Third, we describe an underwater near-infrared imaging setup to observe unperturbed nocturnal animal behaviors. Infrared light pulses were used to synchronize the timing between the video and the physiological signal over a long recording duration. Our automated tracking software measures the animal's body position and posture reliably in an aquatic scene. In combination, these techniques enable long term observation of spontaneous behavior of freely swimming weakly electric fish in a reliable and precise manner. We believe our method can be similarly applied to the study of other aquatic animals by relating their physiological signals with exploratory or social behaviors.
Neuroscience, Issue 85, animal tracking, weakly electric fish, electric organ discharge, underwater infrared imaging, automated image tracking, sensory isolation chamber, exploratory behavior
Automated, Quantitative Cognitive/Behavioral Screening of Mice: For Genetics, Pharmacology, Animal Cognition and Undergraduate Instruction
Institutions: Rutgers University, Koç University, New York University, Fairfield University.
We describe a high-throughput, high-volume, fully automated, live-in 24/7 behavioral testing system for assessing the effects of genetic and pharmacological manipulations on basic mechanisms of cognition and learning in mice. A standard polypropylene mouse housing tub is connected through an acrylic tube to a standard commercial mouse test box. The test box has 3 hoppers, 2 of which are connected to pellet feeders. All are internally illuminable with an LED and monitored for head entries by infrared (IR) beams. Mice live in the environment, which eliminates handling during screening. They obtain their food during two or more daily feeding periods by performing in operant (instrumental) and Pavlovian (classical) protocols, for which we have written protocol-control software and quasi-real-time data analysis and graphing software. The data analysis and graphing routines are written in a MATLAB-based language created to simplify greatly the analysis of large time-stamped behavioral and physiological event records and to preserve a full data trail from raw data through all intermediate analyses to the published graphs and statistics within a single data structure. The data-analysis code harvests the data several times a day and subjects it to statistical and graphical analyses, which are automatically stored in the "cloud" and on in-lab computers. Thus, the progress of individual mice is visualized and quantified daily. The data-analysis code talks to the protocol-control code, permitting the automated advance from protocol to protocol of individual subjects. The behavioral protocols implemented are matching, autoshaping, timed hopper-switching, risk assessment in timed hopper-switching, impulsivity measurement, and the circadian anticipation of food availability. Open-source protocol-control and data-analysis code makes the addition of new protocols simple. Eight test environments fit in a 48 in x 24 in x 78 in cabinet; two such cabinets (16 environments) may be controlled by one computer.
Behavior, Issue 84, genetics, cognitive mechanisms, behavioral screening, learning, memory, timing
Measurement of Coherence Decay in GaMnAs Using Femtosecond Four-wave Mixing
Institutions: Dalhousie University, University of Notre Dame.
The application of femtosecond four-wave mixing to the study of fundamental properties of diluted magnetic semiconductors ((s,p)-d hybridization, spin-flip scattering) is described, using experiments on GaMnAs as a prototype III-Mn-V system. Spectrally-resolved and time-resolved experimental configurations are described, including the use of zero-background autocorrelation techniques for pulse optimization. The etching process used to prepare GaMnAs samples for four-wave mixing experiments is also highlighted. The high temporal resolution of this technique, afforded by the use of short (20 fsec) optical pulses, permits the rapid spin-flip scattering process in this system to be studied directly in the time domain, providing new insight into the strong exchange coupling responsible for carrier-mediated ferromagnetism. We also show that spectral resolution of the four-wave mixing signal allows one to extract clear signatures of (s,p)-d hybridization in this system, unlike linear spectroscopy techniques. This increased sensitivity is due to the nonlinearity of the technique, which suppresses defect-related contributions to the optical response. This method may be used to measure the time scale for coherence decay (tied to the fastest scattering processes) in a wide variety of semiconductor systems of interest for next generation electronics and optoelectronics.
Physics, Issue 82, Four-wave mixing, spin-flip scattering, ultrafast, GaMnAs, diluted magnetic semiconductor, photon echo, dephasing, GaAs, low temperature grown semiconductor, exchange, ferromagnetic
Testing Visual Sensitivity to the Speed and Direction of Motion in Lizards
Institutions: Macquarie University.
Testing visual sensitivity in any species provides basic information regarding behaviour, evolution, and ecology. However, testing specific features of the visual system provide more empirical evidence for functional applications. Investigation into the sensory system provides information about the sensory capacity, learning and memory ability, and establishes known baseline behaviour in which to gauge deviations (Burghardt, 1977). However, unlike mammalian or avian systems, testing for learning and memory in a reptile species is difficult. Furthermore, using an operant paradigm as a psychophysical measure of sensory ability is likewise as difficult. Historically, reptilian species have responded poorly to conditioning trials because of issues related to motivation, physiology, metabolism, and basic biological characteristics. Here, I demonstrate an operant paradigm used a novel model lizard species, the Jacky dragon (Amphibolurus muricatus) and describe how to test peripheral sensitivity to salient speed and motion characteristics. This method uses an innovative approach to assessing learning and sensory capacity in lizards. I employ the use of random-dot kinematograms (RDKs) to measure sensitivity to speed, and manipulate the level of signal strength by changing the proportion of dots moving in a coherent direction. RDKs do not represent a biologically meaningful stimulus, engages the visual system, and is a classic psychophysical tool used to measure sensitivity in humans and other animals. Here, RDKs are displayed to lizards using three video playback systems. Lizards are to select the direction (left or right) in which they perceive dots to be moving. Selection of the appropriate direction is reinforced by biologically important prey stimuli, simulated by computer-animated invertebrates.
Neuroscience, Issue 2, Visual sensitivity, motion perception, operant conditioning, speed, coherence, Jacky dragon (Amphibolurus muricatus)
Dynamic Visual Tests to Identify and Quantify Visual Damage and Repair Following Demyelination in Optic Neuritis Patients
Institutions: Hadassah Hebrew-University Medical Center.
In order to follow optic neuritis patients and evaluate the effectiveness of their treatment, a handy, accurate and quantifiable tool is required to assess changes in myelination at the central nervous system (CNS). However, standard measurements, including routine visual tests and MRI scans, are not sensitive enough for this purpose. We present two visual tests addressing dynamic monocular and binocular functions which may closely associate with the extent of myelination along visual pathways. These include Object From Motion (OFM) extraction and Time-constrained stereo protocols. In the OFM test, an array of dots compose an object, by moving the dots within the image rightward while moving the dots outside the image leftward or vice versa. The dot pattern generates a camouflaged object that cannot be detected when the dots are stationary or moving as a whole. Importantly, object recognition is critically dependent on motion perception. In the Time-constrained Stereo protocol, spatially disparate images are presented for a limited length of time, challenging binocular 3-dimensional integration in time. Both tests are appropriate for clinical usage and provide a simple, yet powerful, way to identify and quantify processes of demyelination and remyelination along visual pathways. These protocols may be efficient to diagnose and follow optic neuritis and multiple sclerosis patients.
In the diagnostic process, these protocols may reveal visual deficits that cannot be identified via current standard visual measurements. Moreover, these protocols sensitively identify the basis of the currently unexplained continued visual complaints of patients following recovery of visual acuity. In the longitudinal follow up course, the protocols can be used as a sensitive marker of demyelinating and remyelinating processes along time. These protocols may therefore be used to evaluate the efficacy of current and evolving therapeutic strategies, targeting myelination of the CNS.
Medicine, Issue 86, Optic neuritis, visual impairment, dynamic visual functions, motion perception, stereopsis, demyelination, remyelination
Methods to Explore the Influence of Top-down Visual Processes on Motor Behavior
Institutions: Rutgers University, Rutgers University, Rutgers University, Rutgers University, Rutgers University.
Kinesthetic awareness is important to successfully navigate the environment. When we interact with our daily surroundings, some aspects of movement are deliberately planned, while others spontaneously occur below conscious awareness. The deliberate component of this dichotomy has been studied extensively in several contexts, while the spontaneous component remains largely under-explored. Moreover, how perceptual processes modulate these movement classes is still unclear. In particular, a currently debated issue is whether the visuomotor system is governed by the spatial percept produced by a visual illusion or whether it is not affected by the illusion and is governed instead by the veridical percept. Bistable percepts such as 3D depth inversion illusions (DIIs) provide an excellent context to study such interactions and balance, particularly when used in combination with reach-to-grasp movements. In this study, a methodology is developed that uses a DII to clarify the role of top-down processes on motor action, particularly exploring how reaches toward a target on a DII are affected in both deliberate and spontaneous movement domains.
Behavior, Issue 86, vision for action, vision for perception, motor control, reach, grasp, visuomotor, ventral stream, dorsal stream, illusion, space perception, depth inversion
Cortical Source Analysis of High-Density EEG Recordings in Children
Institutions: UCL Institute of Child Health, University College London.
EEG is traditionally described as a neuroimaging technique with high temporal and low spatial resolution. Recent advances in biophysical modelling and signal processing make it possible to exploit information from other imaging modalities like structural MRI that provide high spatial resolution to overcome this constraint1
. This is especially useful for investigations that require high resolution in the temporal as well as spatial domain. In addition, due to the easy application and low cost of EEG recordings, EEG is often the method of choice when working with populations, such as young children, that do not tolerate functional MRI scans well. However, in order to investigate which neural substrates are involved, anatomical information from structural MRI is still needed. Most EEG analysis packages work with standard head models that are based on adult anatomy. The accuracy of these models when used for children is limited2
, because the composition and spatial configuration of head tissues changes dramatically over development3
In the present paper, we provide an overview of our recent work in utilizing head models based on individual structural MRI scans or age specific head models to reconstruct the cortical generators of high density EEG. This article describes how EEG recordings are acquired, processed, and analyzed with pediatric populations at the London Baby Lab, including laboratory setup, task design, EEG preprocessing, MRI processing, and EEG channel level and source analysis.
Behavior, Issue 88, EEG, electroencephalogram, development, source analysis, pediatric, minimum-norm estimation, cognitive neuroscience, event-related potentials
Transcranial Magnetic Stimulation for Investigating Causal Brain-behavioral Relationships and their Time Course
Institutions: University College London.
Transcranial magnetic stimulation (TMS) is a safe, non-invasive brain stimulation technique that uses a strong electromagnet in order to temporarily disrupt information processing in a brain region, generating a short-lived “virtual lesion.” Stimulation that interferes with task performance indicates that the affected brain region is necessary to perform the task normally. In other words, unlike neuroimaging methods such as functional magnetic resonance imaging (fMRI) that indicate correlations between brain and behavior, TMS can be used to demonstrate causal brain-behavior relations. Furthermore, by varying the duration and onset of the virtual lesion, TMS can also reveal the time course of normal processing. As a result, TMS has become an important tool in cognitive neuroscience. Advantages of the technique over lesion-deficit studies include better spatial-temporal precision of the disruption effect, the ability to use participants as their own control subjects, and the accessibility of participants. Limitations include concurrent auditory and somatosensory stimulation that may influence task performance, limited access to structures more than a few centimeters from the surface of the scalp, and the relatively large space of free parameters that need to be optimized in order for the experiment to work. Experimental designs that give careful consideration to appropriate control conditions help to address these concerns. This article illustrates these issues with TMS results that investigate the spatial and temporal contributions of the left supramarginal gyrus (SMG) to reading.
Behavior, Issue 89,
Transcranial magnetic stimulation, virtual lesion, chronometric, cognition, brain, behavior
Trajectory Data Analyses for Pedestrian Space-time Activity Study
Institutions: Kean University, University of Wisconsin-Madison.
It is well recognized that human movement in the spatial and temporal dimensions has direct influence on disease transmission1-3
. An infectious disease typically spreads via contact between infected and susceptible individuals in their overlapped activity spaces. Therefore, daily mobility-activity information can be used as an indicator to measure exposures to risk factors of infection. However, a major difficulty and thus the reason for paucity of studies of infectious disease transmission at the micro scale arise from the lack of detailed individual mobility data. Previously in transportation and tourism research detailed space-time activity data often relied on the time-space diary technique, which requires subjects to actively record their activities in time and space. This is highly demanding for the participants and collaboration from the participants greatly affects the quality of data4
Modern technologies such as GPS and mobile communications have made possible the automatic collection of trajectory data. The data collected, however, is not ideal for modeling human space-time activities, limited by the accuracies of existing devices. There is also no readily available tool for efficient processing of the data for human behavior study. We present here a suite of methods and an integrated ArcGIS desktop-based visual interface for the pre-processing and spatiotemporal analyses of trajectory data. We provide examples of how such processing may be used to model human space-time activities, especially with error-rich pedestrian trajectory data, that could be useful in public health studies such as infectious disease transmission modeling.
The procedure presented includes pre-processing, trajectory segmentation, activity space characterization, density estimation and visualization, and a few other exploratory analysis methods. Pre-processing is the cleaning of noisy raw trajectory data. We introduce an interactive visual pre-processing interface as well as an automatic module. Trajectory segmentation5
involves the identification of indoor and outdoor parts from pre-processed space-time tracks. Again, both interactive visual segmentation and automatic segmentation are supported. Segmented space-time tracks are then analyzed to derive characteristics of one's activity space such as activity radius etc.
Density estimation and visualization are used to examine large amount of trajectory data to model hot spots and interactions. We demonstrate both density surface mapping6
and density volume rendering7
. We also include a couple of other exploratory data analyses (EDA) and visualizations tools, such as Google Earth animation support and connection analysis. The suite of analytical as well as visual methods presented in this paper may be applied to any trajectory data for space-time activity studies.
Environmental Sciences, Issue 72, Computer Science, Behavior, Infectious Diseases, Geography, Cartography, Data Display, Disease Outbreaks, cartography, human behavior, Trajectory data, space-time activity, GPS, GIS, ArcGIS, spatiotemporal analysis, visualization, segmentation, density surface, density volume, exploratory data analysis, modelling
Perceptual and Category Processing of the Uncanny Valley Hypothesis' Dimension of Human Likeness: Some Methodological Issues
Institutions: University of Zurich.
Mori's Uncanny Valley Hypothesis1,2
proposes that the perception of humanlike characters such as robots and, by extension, avatars (computer-generated characters) can evoke negative or positive affect (valence) depending on the object's degree of visual and behavioral realism along a dimension of human likeness
) (Figure 1
). But studies of affective valence of subjective responses to variously realistic non-human characters have produced inconsistent findings 3, 4, 5, 6
. One of a number of reasons for this is that human likeness is not perceived as the hypothesis assumes. While the DHL can be defined following Mori's description as a smooth linear change in the degree of physical humanlike similarity, subjective perception of objects along the DHL can be understood in terms of the psychological effects of categorical perception (CP) 7
. Further behavioral and neuroimaging investigations of category processing and CP along the DHL and of the potential influence of the dimension's underlying category structure on affective experience are needed. This protocol therefore focuses on the DHL and allows examination of CP. Based on the protocol presented in the video as an example, issues surrounding the methodology in the protocol and the use in "uncanny" research of stimuli drawn from morph continua to represent the DHL are discussed in the article that accompanies the video. The use of neuroimaging and morph stimuli to represent the DHL in order to disentangle brain regions neurally responsive to physical human-like similarity from those responsive to category change and category processing is briefly illustrated.
Behavior, Issue 76, Neuroscience, Neurobiology, Molecular Biology, Psychology, Neuropsychology, uncanny valley, functional magnetic resonance imaging, fMRI, categorical perception, virtual reality, avatar, human likeness, Mori, uncanny valley hypothesis, perception, magnetic resonance imaging, MRI, imaging, clinical techniques
The Gateway to the Brain: Dissecting the Primate Eye
Institutions: University of Montreal, University of Montreal, Universite du Quebec a Trois-Rivieres.
The visual system in humans is considered the gateway to the world and plays a principal role in the plethora of sensory, perceptual and cognitive processes. It is therefore not surprising that quality of vision is tied to quality of life . Despite widespread clinical and basic research surrounding the causes of visual disorders, many forms of visual impairments, such as retinitis pigmentosa and macular degeneration, lack effective treatments. Non-human primates have the closest general features of eye development to that of humans. Not only do they have a similar vascular anatomy, but amongst other mammals, primates have the unique characteristic of having a region in the temporal retina specialized for high visual acuity, the fovea1
. Here we describe a general technique for dissecting the primate retina to provide tissue for retinal histology, immunohistochemistry, laser capture microdissection, as well as light and electron microscopy. With the extended use of the non-human primate as a translational model, our hope is that improved understanding of the retina will provide insights into effective approaches towards attenuating or reversing the negative impact of visual disorders on the quality of life of affected individuals.
Neuroscience, Issue 27, Non-human primate, eye, retina, dissection, retina ganglion cells, cornea
A Novel Approach for Documenting Phosphenes Induced by Transcranial Magnetic Stimulation
Institutions: Boston University School of Medicine, Beth Israel Deaconess Med Center, Centre National de la Recherche Scientifique (CNRS).
Stimulation of the human visual cortex produces a transient perception of light, known as a phosphene. Phosphenes are induced by invasive electrical stimulation of the occipital cortex, but also by non-invasive Transcranial Magnetic Stimulation (TMS)1
of the same cortical regions. The intensity at which a phosphene is induced (phosphene threshold) is a well established measure of visual cortical excitability and is used to study cortico-cortical interactions, functional organization 2
, susceptibility to pathology 3,4
and visual processing 5-7
. Phosphenes are typically defined by three characteristics: they are observed in the visual hemifield contralateral to stimulation; they are induced when the subject s eyes are open or closed, and their spatial location changes with the direction of gaze 2
. Various methods have been used to document phosphenes, but a standardized methodology is lacking. We demonstrate a reliable procedure to obtain phosphene threshold values and introduce a novel system for the documentation and analysis of phosphenes. We developed the Laser Tracking and Painting system (LTaP), a low cost, easily built and operated system that records the location and size of perceived phosphenes in real-time. The LTaP system provides a stable and customizable environment for quantification and analysis of phosphenes.
Neuroscience, Issue 38, Transcranial Magnetic Stimulation (TMS), Phosphenes, Occipital, Human visual cortex, Threshold
Membrane Potentials, Synaptic Responses, Neuronal Circuitry, Neuromodulation and Muscle Histology Using the Crayfish: Student Laboratory Exercises
Institutions: University of Kentucky, University of Toronto.
The purpose of this report is to help develop an understanding of the effects caused by ion gradients across a biological membrane. Two aspects that influence a cell's membrane potential and which we address in these experiments are: (1) Ion concentration of K+
on the outside of the membrane, and (2) the permeability of the membrane to specific ions. The crayfish abdominal extensor muscles are in groupings with some being tonic (slow) and others phasic (fast) in their biochemical and physiological phenotypes, as well as in their structure; the motor neurons that innervate these muscles are correspondingly different in functional characteristics. We use these muscles as well as the superficial, tonic abdominal flexor muscle to demonstrate properties in synaptic transmission. In addition, we introduce a sensory-CNS-motor neuron-muscle circuit to demonstrate the effect of cuticular sensory stimulation as well as the influence of neuromodulators on certain aspects of the circuit. With the techniques obtained in this exercise, one can begin to answer many questions remaining in other experimental preparations as well as in physiological applications related to medicine and health. We have demonstrated the usefulness of model invertebrate preparations to address fundamental questions pertinent to all animals.
Neuroscience, Issue 47, Invertebrate, Crayfish, neurophysiology, muscle, anatomy, electrophysiology
Correlating Behavioral Responses to fMRI Signals from Human Prefrontal Cortex: Examining Cognitive Processes Using Task Analysis
Institutions: Centre for Vision Research, York University, Centre for Vision Research, York University.
The aim of this methods paper is to describe how to implement a neuroimaging technique to examine complementary brain processes engaged by two similar tasks. Participants' behavior during task performance in an fMRI scanner can then be correlated to the brain activity using the blood-oxygen-level-dependent signal. We measure behavior to be able to sort correct trials, where the subject performed the task correctly and then be able to examine the brain signals related to correct performance. Conversely, if subjects do not perform the task correctly, and these trials are included in the same analysis with the correct trials we would introduce trials that were not only for correct performance. Thus, in many cases these errors can be used themselves to then correlate brain activity to them. We describe two complementary tasks that are used in our lab to examine the brain during suppression of an automatic responses: the stroop1
and anti-saccade tasks. The emotional stroop paradigm instructs participants to either report the superimposed emotional 'word' across the affective faces or the facial 'expressions' of the face stimuli1,2
. When the word and the facial expression refer to different emotions, a conflict between what must be said and what is automatically read occurs. The participant has to resolve the conflict between two simultaneously competing processes of word reading and facial expression. Our urge to read out a word leads to strong 'stimulus-response (SR)' associations; hence inhibiting these strong SR's is difficult and participants are prone to making errors. Overcoming this conflict and directing attention away from the face or the word requires the subject to inhibit bottom up processes which typically directs attention to the more salient stimulus. Similarly, in the anti-saccade task3,4,5,6
, where an instruction cue is used to direct only attention to a peripheral stimulus location but then the eye movement is made to the mirror opposite position. Yet again we measure behavior by recording the eye movements of participants which allows for the sorting of the behavioral responses into correct and error trials7
which then can be correlated to brain activity. Neuroimaging now allows researchers to measure different behaviors of correct and error trials that are indicative of different cognitive processes and pinpoint the different neural networks involved.
Neuroscience, Issue 64, fMRI, eyetracking, BOLD, attention, inhibition, Magnetic Resonance Imaging, MRI
Radio Frequency Identification and Motion-sensitive Video Efficiently Automate Recording of Unrewarded Choice Behavior by Bumblebees
Institutions: University of Ottawa.
We present two methods for observing bumblebee choice behavior in an enclosed testing space. The first method consists of Radio Frequency Identification (RFID) readers built into artificial flowers that display various visual cues, and RFID tags (i.e.
, passive transponders) glued to the thorax of bumblebee workers. The novelty in our implementation is that RFID readers are built directly into artificial flowers that are capable of displaying several distinct visual properties such as color, pattern type, spatial frequency (i.e.
, “busyness” of the pattern), and symmetry (spatial frequency and symmetry were not manipulated in this experiment). Additionally, these visual displays in conjunction with the automated systems are capable of recording unrewarded
choice behavior. The second method consists of recording choice behavior at artificial flowers using motion-sensitive high-definition camcorders. Bumblebees have number tags glued to their thoraces for unique identification. The advantage in this implementation over RFID is that in addition to observing landing behavior, alternate measures of preference such as hovering and antennation may also be observed. Both automation methods increase experimental control, and internal validity by allowing larger scale studies that take into account individual differences. External validity is also improved because bees can freely enter and exit the testing environment without constraints such as the availability of a research assistant on-site. Compared to human observation in real time, the automated methods are more cost-effective and possibly less error-prone.
Neuroscience, Issue 93, bumblebee, unlearned behaviors, floral choice, visual perception, Bombus spp, information processing, radio-frequency identification, motion-sensitive video
Institutions: University of Utah.
A limitation of traditional full-field electroretinograms (ERG) for the diagnosis of retinopathy is lack of sensitivity. Generally, ERG results are normal unless more than approximately 20% of the retina is affected. In practical terms, a patient might be legally blind as a result of macular degeneration or other scotomas and still appear normal, according to traditional full field ERG. An important development in ERGs is the multifocal ERG (mfERG). Erich Sutter adapted the mathematical sequences called binary m-sequences enabling the isolation from a single electrical signal an electroretinogram representing less than each square millimeter of retina in response to a visual stimulus1
Results that are generated by mfERG appear similar to those generated by flash ERG. In contrast to flash ERG, which best generates data appropriate for whole-eye disorders. The basic mfERG result is based on the calculated mathematical average of an approximation of the positive deflection component of traditional ERG response, known as the b-wave1
. Multifocal ERG programs measure electrical activity from more than a hundred retinal areas per eye, in a few minutes. The enhanced spatial resolution enables scotomas and retinal dysfunction to be mapped and quantified.
In the protocol below, we describe the recording of mfERGs using a bipolar speculum contact lens.
Components of mfERG systems vary between manufacturers. For the presentation of visible stimulus, some suitable CRT monitors are available but most systems have adopted the use of flat-panel liquid crystal displays (LCD). The visual stimuli depicted here, were produced by a LCD microdisplay subtending 35 - 40 degrees horizontally and 30 - 35 degrees vertically of visual field, and calibrated to produce multifocal flash intensities of 2.7 cd s m-2
. Amplification was 50K. Lower and upper bandpass limits were 10 and 300 Hz. The software packages used were VERIS versions 5 and 6.
Medicine, Issue 58, Multifocal electroretinogram, mfERG, electroretinogram, ERG
Functional Mapping with Simultaneous MEG and EEG
Institutions: MGH - Massachusetts General Hospital.
We use magnetoencephalography (MEG) and electroencephalography (EEG) to locate and determine the temporal evolution in brain areas involved in the processing of simple sensory stimuli. We will use somatosensory stimuli to locate the hand somatosensory areas, auditory stimuli to locate the auditory cortices, visual stimuli in four quadrants of the visual field to locate the early visual areas. These type of experiments are used for functional mapping in epileptic and brain tumor patients to locate eloquent cortices. In basic neuroscience similar experimental protocols are used to study the orchestration of cortical activity. The acquisition protocol includes quality assurance procedures, subject preparation for the combined MEG/EEG study, and acquisition of evoked-response data with somatosensory, auditory, and visual stimuli. We also demonstrate analysis of the data using the equivalent current dipole model and cortically-constrained minimum-norm estimates. Anatomical MRI data are employed in the analysis for visualization and for deriving boundaries of tissue boundaries for forward modeling and cortical location and orientation constraints for the minimum-norm estimates.
JoVE neuroscience, Issue 40, neuroscience, brain, MEG, EEG, functional imaging
Enabling High Grayscale Resolution Displays and Accurate Response Time Measurements on Conventional Computers
Institutions: The Ohio State University, University of Southern California, University of Southern California, University of Southern California, The Ohio State University.
Display systems based on conventional computer graphics cards are capable of generating images with 8-bit gray level resolution. However, most experiments in vision research require displays with more than 12 bits of luminance resolution. Several solutions are available. Bit++ 1
and DataPixx 2
use the Digital Visual Interface (DVI) output from graphics cards and high resolution (14 or 16-bit) digital-to-analog converters to drive analog display devices. The VideoSwitcher 3
described here combines analog video signals from the red and blue channels of graphics cards with different weights using a passive resister network 4
and an active circuit to deliver identical video signals to the three channels of color monitors. The method provides an inexpensive way to enable high-resolution monochromatic displays using conventional graphics cards and analog monitors. It can also provide trigger signals that can be used to mark stimulus onsets, making it easy to synchronize visual displays with physiological recordings or response time measurements.
Although computer keyboards and mice are frequently used in measuring response times (RT), the accuracy of these measurements is quite low. The RTbox is a specialized hardware and software solution for accurate RT measurements. Connected to the host computer through a USB connection, the driver of the RTbox is compatible with all conventional operating systems. It uses a microprocessor and high-resolution clock to record the identities and timing of button events, which are buffered until the host computer retrieves them. The recorded button events are not affected by potential timing uncertainties or biases associated with data transmission and processing in the host computer. The asynchronous storage greatly simplifies the design of user programs. Several methods are available to synchronize the clocks of the RTbox and the host computer. The RTbox can also receive external triggers and be used to measure RT with respect to external events.
Both VideoSwitcher and RTbox are available for users to purchase. The relevant information and many demonstration programs can be found at https://lobes.usc.edu/.
Neuroscience, Issue 60, VideoSwitcher, Visual stimulus, Luminance resolution, Contrast, Trigger, RTbox, Response time
Laser Capture Microdissection of Mammalian Tissue
Institutions: University of California, Irvine (UCI).
Laser capture microscopy, also known as laser microdissection (LMD), enables the user to isolate small numbers of cells or tissues from frozen or formalin-fixed, paraffin-embedded tissue sections. LMD techniques rely on a thermo labile membrane placed either on top of, or underneath, the tissue section. In one method, focused laser energy is used to melt the membrane onto the underlying cells, which can then be lifted out of the tissue section. In the other, the laser energy vaporizes the foil along a path "drawn" on the tissue, allowing the selected cells to fall into a collection device. Each technique allows the selection of cells with a minimum resolution of several microns. DNA, RNA, protein, and lipid samples may be isolated and analyzed from micro-dissected samples. In this video, we demonstrate the use of the Leica AS-LMD laser microdissection instrument in seven segments, including an introduction to the principles of LMD, initializing the instrument for use, general considerations for sample preparation, mounting the specimen and setting up capture tubes, aligning the microscope, adjusting the capture controls, and capturing tissue specimens. Laser-capture micro-dissection enables the investigator to isolate samples of pure cell populations as small as a few cell-equivalents. This allows the analysis of cells of interest that are free of neighboring contaminants, which may confound experimental results.
Issue 8, Basic Protocols, Laser Capture Microdissection, Microdissection Techniques, Leica