In this video article we present a detailed demonstration of a highly efficient method for generating terahertz waves. Our technique is based on photoconduction, which has been one of the most commonly used techniques for terahertz generation 1-8. Terahertz generation in a photoconductive emitter is achieved by pumping an ultrafast photoconductor with a pulsed or heterodyned laser illumination. The induced photocurrent, which follows the envelope of the pump laser, is routed to a terahertz radiating antenna connected to the photoconductor contact electrodes to generate terahertz radiation. Although the quantum efficiency of a photoconductive emitter can theoretically reach 100%, the relatively long transport path lengths of photo-generated carriers to the contact electrodes of conventional photoconductors have severely limited their quantum efficiency. Additionally, the carrier screening effect and thermal breakdown strictly limit the maximum output power of conventional photoconductive terahertz sources. To address the quantum efficiency limitations of conventional photoconductive terahertz emitters, we have developed a new photoconductive emitter concept which incorporates a plasmonic contact electrode configuration to offer high quantum-efficiency and ultrafast operation simultaneously. By using nano-scale plasmonic contact electrodes, we significantly reduce the average photo-generated carrier transport path to photoconductor contact electrodes compared to conventional photoconductors 9. Our method also allows increasing photoconductor active area without a considerable increase in the capacitive loading to the antenna, boosting the maximum terahertz radiation power by preventing the carrier screening effect and thermal breakdown at high optical pump powers. By incorporating plasmonic contact electrodes, we demonstrate enhancing the optical-to-terahertz power conversion efficiency of a conventional photoconductive terahertz emitter by a factor of 50 10.
21 Related JoVE Articles!
Intact Histological Characterization of Brain-implanted Microdevices and Surrounding Tissue
Institutions: Purdue University, Purdue University.
Research into the design and utilization of brain-implanted microdevices, such as microelectrode arrays, aims to produce clinically relevant devices that interface chronically with surrounding brain tissue. Tissue surrounding these implants is thought to react to the presence of the devices over time, which includes the formation of an insulating "glial scar" around the devices. However, histological analysis of these tissue changes is typically performed after explanting the device, in a process that can disrupt the morphology of the tissue of interest.
Here we demonstrate a protocol in which cortical-implanted devices are collected intact in surrounding rodent brain tissue. We describe how, once perfused with fixative, brains are removed and sliced in such a way as to avoid explanting devices. We outline fluorescent antibody labeling and optical clearing methods useful for producing an informative, yet thick tissue section. Finally, we demonstrate the mounting and imaging of these tissue sections in order to investigate the biological interface around brain-implanted devices.
Neurobiology, Issue 72, Neuroscience, Biomedical Engineering, Medicine, Central Nervous System, Brain, Neuroglia, Neurons, Immunohistochemistry (IHC), Histocytological Preparation Techniques, Microscopy, Confocal, nondestructive testing, bioengineering (man-machine systems), bionics, histology, brain implants, microelectrode arrays, immunohistochemistry, neuroprosthetics, brain machine interface, microscopy, thick tissue, optical clearing, animal model
Whole Mount Immunolabeling of Olfactory Receptor Neurons in the Drosophila Antenna
Institutions: Doshisha University, RIKEN Brain Science Institute, RIKEN Brain Science Institute.
Odorant molecules bind to their target receptors in a precise and coordinated manner. Each receptor recognizes a specific signal and relays this information to the brain. As such, determining how olfactory information is transferred to the brain, modifying both perception and behavior, merits investigation. Interestingly, there is emerging evidence that cellular transduction and transcriptional factors are involved in the diversification of olfactory receptor neuron. Here we provide a robust whole mount immunological labeling method to assay in vivo olfactory receptor neuron organization. Using this method, we identified all olfactory receptor neurons with anti-ELAV antibody, a known pan-neural marker and Or49a-mCD8::GFP, an olfactory receptor neuron specifically expressed in Nba neuron using anti-GFP antibody.
Neuroscience, Issue 87,
Developmental biology, Drosophila, Whole mount immunolabeling, olfactory receptor neurons, antennae, sensory organ
Time Multiplexing Super Resolving Technique for Imaging from a Moving Platform
Institutions: Bar-Ilan University, Kfar Saba, Israel.
We propose a method for increasing the resolution of an object and overcoming the diffraction limit of an optical system installed on top of a moving imaging system, such as an airborne platform or satellite. The resolution improvement is obtained in a two-step process. First, three low resolution differently defocused images are being captured and the optical phase is retrieved using an improved iterative Gerchberg-Saxton based algorithm. The phase retrieval allows to numerically back propagate the field to the aperture plane. Second, the imaging system is shifted and the first step is repeated. The obtained optical fields at the aperture plane are combined and a synthetically increased lens aperture is generated along the direction of movement, yielding higher imaging resolution. The method resembles a well-known approach from the microwave regime called the Synthetic Aperture Radar (SAR) in which the antenna size is synthetically increased along the platform propagation direction. The proposed method is demonstrated through laboratory experiment.
Physics, Issue 84, Superresolution, Fourier optics, Remote Sensing and Sensors, Digital Image Processing, optics, resolution
Electrophysiological Measurements from a Moth Olfactory System
Institutions: University of California, Davis.
Insect olfactory systems provide unique opportunities for recording odorant-induced responses in the forms of electroantennograms (EAG) and single sensillum recordings (SSR), which are summed responses from all odorant receptor neurons (ORNs) located on the antenna and from those housed in individual sensilla, respectively. These approaches have been exploited for getting a better understanding of insect chemical communication. The identified stimuli can then be used as either attractants or repellents in management strategies for insect pests.
Neuroscience, Issue 49, Insect Olfaction, Electroantennogram (EAG), Single Sensillum Recordings (SSR), navel orangeworm
Single Sensillum Recordings in the Insects Drosophila melanogaster and Anopheles gambiae
Institutions: Rockefeller University.
The sense of smell is essential for insects to find foods, mates, predators, and oviposition sites3
. Insect olfactory sensory neurons (OSNs) are enclosed in sensory hairs called sensilla, which cover the surface of olfactory organs. The surface of each sensillum is covered with tiny pores, through which odorants pass and dissolve in a fluid called sensillum lymph, which bathes the sensory dendrites of the OSNs housed in a given sensillum. The OSN dendrites express odorant receptor (OR) proteins, which in insects function as odor-gated ion channels4, 5
. The interaction of odorants with ORs either increases or decreases the basal firing rate of the OSN. This neuronal activity in the form of action potentials embodies the first representation of the quality, intensity, and temporal characteristics of the odorant6, 7
Given the easy access to these sensory hairs, it is possible to perform extracellular recordings from single OSNs by introducing a recording electrode into the sensillum lymph, while the reference electrode is placed in the lymph of the eye or body of the insect. In Drosophila
, sensilla house between one and four OSNs, but each OSN typically displays a characteristic spike amplitude. Spike sorting techniques make it possible to assign spiking responses to individual OSNs. This single sensillum recording (SSR) technique monitors the difference in potential between the sensillum lymph and the reference electrode as electrical spikes that are generated by the receptor activity on OSNs1, 2, 8
. Changes in the number of spikes in response to the odorant represent the cellular basis of odor coding in insects. Here, we describe the preparation method currently used in our lab to perform SSR on Drosophila melanogaster
and Anopheles gambiae
, and show representative traces induced by the odorants in a sensillum-specific manner.
JoVE Neuroscience, Issue 36, electrophysiology, sensory neuron, insect, olfaction, extracellular recording
Construction of Microdrive Arrays for Chronic Neural Recordings in Awake Behaving Mice
Institutions: North Shore LIJ Health System, Hofstra North Shore LIJ School of Medicine.
State-of-the-art electrophysiological recordings from the brains of freely behaving animals allow researchers to simultaneously examine local field potentials (LFPs) from populations of neurons and action potentials from individual cells, as the animal engages in experimentally relevant tasks. Chronically implanted microdrives allow for brain recordings to last over periods of several weeks. Miniaturized drives and lightweight components allow for these long-term recordings to occur in small mammals, such as mice. By using tetrodes, which consist of tightly braided bundles of four electrodes in which each wire has a diameter of 12.5 μm, it is possible to isolate physiologically active neurons in superficial brain regions such as the cerebral cortex, dorsal hippocampus, and subiculum, as well as deeper regions such as the striatum and the amygdala. Moreover, this technique insures stable, high-fidelity neural recordings as the animal is challenged with a variety of behavioral tasks. This manuscript describes several techniques that have been optimized to record from the mouse brain. First, we show how to fabricate tetrodes, load them into driveable tubes, and gold-plate their tips in order to reduce their impedance from MΩ to KΩ range. Second, we show how to construct a custom microdrive assembly for carrying and moving the tetrodes vertically, with the use of inexpensive materials. Third, we show the steps for assembling a commercially available microdrive (Neuralynx VersaDrive) that is designed to carry independently movable tetrodes. Finally, we present representative results of local field potentials and single-unit signals obtained in the dorsal subiculum of mice. These techniques can be easily modified to accommodate different types of electrode arrays and recording schemes in the mouse brain.
Behavior, Issue 77, Neuroscience, Neurobiology, Anatomy, Physiology, Biomedical Engineering, Brain, Amygdala, Hippocampus, Electrodes, Implanted, Microelectrodes, Action Potentials, Neurosciences, Neurophysiology, Neuroscience, brain, mouse, in vivo electrophysiology, tetrodes, microdrive, chronic recordings, local field potential, dorsal subiculum, animal model
Using Insect Electroantennogram Sensors on Autonomous Robots for Olfactory Searches
Institutions: Centre National de la Recherche Scientifique (CNRS), Institut d'Ecologie et des Sciences de l'Environnement de Paris, Institut Pasteur.
Robots designed to track chemical leaks in hazardous industrial facilities1
or explosive traces in landmine fields2
face the same problem as insects foraging for food or searching for mates3
: the olfactory search is constrained by the physics of turbulent transport4
. The concentration landscape of wind borne odors is discontinuous and consists of sporadically located patches. A pre-requisite to olfactory search is that intermittent odor patches are detected. Because of its high speed and sensitivity5-6
, the olfactory organ of insects provides a unique opportunity for detection. Insect antennae have been used in the past to detect not only sex pheromones7
but also chemicals that are relevant to humans, e.g.
, volatile compounds emanating from cancer cells8
or toxic and illicit substances9-11
. We describe here a protocol for using insect antennae on autonomous robots and present a proof of concept for tracking odor plumes to their source. The global response of olfactory neurons is recorded in situ
in the form of electroantennograms (EAGs). Our experimental design, based on a whole insect preparation, allows stable recordings within a working day. In comparison, EAGs on excised antennae have a lifetime of 2 hr. A custom hardware/software interface was developed between the EAG electrodes and a robot. The measurement system resolves individual odor patches up to 10 Hz, which exceeds the time scale of artificial chemical sensors12
. The efficiency of EAG sensors for olfactory searches is further demonstrated in driving the robot toward a source of pheromone. By using identical olfactory stimuli and sensors as in real animals, our robotic platform provides a direct means for testing biological hypotheses about olfactory coding and search strategies13
. It may also prove beneficial for detecting other odorants of interests by combining EAGs from different insect species in a bioelectronic nose configuration14
or using nanostructured gas sensors that mimic insect antennae15
Neuroscience, Issue 90, robotics, electroantennogram, EAG, gas sensor, electronic nose, olfactory search, surge and casting, moth, insect, olfaction, neuron
Designing a Bio-responsive Robot from DNA Origami
Institutions: Bar-Ilan University.
Nucleic acids are astonishingly versatile. In addition to their natural role as storage medium for biological information1
, they can be utilized in parallel computing2,3
, recognize and bind molecular or cellular targets4,5
, catalyze chemical reactions6,7
, and generate calculated responses in a biological system8,9
. Importantly, nucleic acids can be programmed to self-assemble into 2D and 3D structures10-12
, enabling the integration of all these remarkable features in a single robot linking the sensing of biological cues to a preset response in order to exert a desired effect.
Creating shapes from nucleic acids was first proposed by Seeman13
, and several variations on this theme have since been realized using various techniques11,12,14,15
. However, the most significant is perhaps the one proposed by Rothemund, termed scaffolded DNA origami16
. In this technique, the folding of a long (>7,000 bases) single-stranded DNA 'scaffold'
is directed to a desired shape by hundreds of short complementary strands termed 'staples'
. Folding is carried out by temperature annealing ramp. This technique was successfully demonstrated in the creation of a diverse array of 2D shapes with remarkable precision and robustness. DNA origami was later extended to 3D as well17,18
The current paper will focus on the caDNAno 2.0 software19
developed by Douglas and colleagues. caDNAno is a robust, user-friendly CAD tool enabling the design of 2D and 3D DNA origami shapes with versatile features. The design process relies on a systematic and accurate abstraction scheme for DNA structures, making it relatively straightforward and efficient.
In this paper we demonstrate the design of a DNA origami nanorobot that has been recently described20
. This robot is 'robotic' in the sense that it links sensing to actuation, in order to perform a task. We explain how various sensing schemes can be integrated into the structure, and how this can be relayed to a desired effect. Finally we use Cando21
to simulate the mechanical properties of the designed shape. The concept we discuss can be adapted to multiple tasks and settings.
Bioengineering, Issue 77, Genetics, Biomedical Engineering, Molecular Biology, Medicine, Genomics, Nanotechnology, Nanomedicine, DNA origami, nanorobot, caDNAno, DNA, DNA Origami, nucleic acids, DNA structures, CAD, sequencing
Voltage Biasing, Cyclic Voltammetry, & Electrical Impedance Spectroscopy for Neural Interfaces
Institutions: Purdue University, University of Wisconsin-Madison, University of Michigan , Purdue University.
Electrical impedance spectroscopy (EIS) and cyclic voltammetry (CV) measure properties of the electrode-tissue interface without additional invasive procedures, and can be used to monitor electrode performance over the long term. EIS measures electrical impedance at multiple frequencies, and increases in impedance indicate increased glial scar formation around the device, while cyclic voltammetry measures the charge carrying capacity of the electrode, and indicates how charge is transferred at different voltage levels. As implanted electrodes age, EIS and CV data change, and electrode sites that previously recorded spiking neurons often exhibit significantly lower efficacy for neural recording. The application of a brief voltage pulse to implanted electrode arrays, known as rejuvenation, can bring back spiking activity on otherwise silent electrode sites for a period of time. Rejuvenation alters EIS and CV, and can be monitored by these complementary methods. Typically, EIS is measured daily as an indication of the tissue response at the electrode site. If spikes are absent in a channel that previously had spikes, then CV is used to determine the charge carrying capacity of the electrode site, and rejuvenation can be applied to improve the interface efficacy. CV and EIS are then repeated to check the changes at the electrode-tissue interface, and neural recordings are collected. The overall goal of rejuvenation is to extend the functional lifetime of implanted arrays.
Neuroscience, Issue 60, neuroprosthesis, electrode-tissue interface, rejuvenation, neural engineering, neuroscience, neural implant, electrode, brain-computer interface, electrochemistry
The Use of Magnetic Resonance Spectroscopy as a Tool for the Measurement of Bi-hemispheric Transcranial Electric Stimulation Effects on Primary Motor Cortex Metabolism
Institutions: University of Montréal, McGill University, University of Minnesota.
Transcranial direct current stimulation (tDCS) is a neuromodulation technique that has been increasingly used over the past decade in the treatment of neurological and psychiatric disorders such as stroke and depression. Yet, the mechanisms underlying its ability to modulate brain excitability to improve clinical symptoms remains poorly understood 33
. To help improve this understanding, proton magnetic resonance spectroscopy (1
H-MRS) can be used as it allows the in vivo
quantification of brain metabolites such as γ-aminobutyric acid (GABA) and glutamate in a region-specific manner 41
. In fact, a recent study demonstrated that 1
H-MRS is indeed a powerful means to better understand the effects of tDCS on neurotransmitter concentration 34
. This article aims to describe the complete protocol for combining tDCS (NeuroConn MR compatible stimulator) with 1
H-MRS at 3 T using a MEGA-PRESS sequence. We will describe the impact of a protocol that has shown great promise for the treatment of motor dysfunctions after stroke, which consists of bilateral stimulation of primary motor cortices 27,30,31
. Methodological factors to consider and possible modifications to the protocol are also discussed.
Neuroscience, Issue 93, proton magnetic resonance spectroscopy, transcranial direct current stimulation, primary motor cortex, GABA, glutamate, stroke
Simulation, Fabrication and Characterization of THz Metamaterial Absorbers
Institutions: University of Glasgow.
Metamaterials (MM), artificial materials engineered to have properties that may not be found in nature, have been widely explored since the first theoretical1
and experimental demonstration2
of their unique properties. MMs can provide a highly controllable electromagnetic response, and to date have been demonstrated in every technologically relevant spectral range including the optical3
, near IR4
, mid IR5
bands. Applications include perfect lenses10
, invisibility cloaks13
. We have recently developed single band16
, dual band17
THz metamaterial absorber devices capable of greater than 80% absorption at the resonance peak. The concept of a MM absorber is especially important at THz frequencies where it is difficult to find strong frequency selective THz absorbers19
. In our MM absorber the THz radiation is absorbed in a thickness of ~ λ/20, overcoming the thickness limitation of traditional quarter wavelength absorbers. MM absorbers naturally lend themselves to THz detection applications, such as thermal sensors, and if integrated with suitable THz sources (e.g.
QCLs), could lead to compact, highly sensitive, low cost, real time THz imaging systems.
Materials Science, Issue 70, Physics, Engineering, Metamaterial, terahertz, sensing, fabrication, clean room, simulation, FTIR, spectroscopy
Flat-floored Air-lifted Platform: A New Method for Combining Behavior with Microscopy or Electrophysiology on Awake Freely Moving Rodents
Institutions: University of Helsinki, Neurotar LTD, University of Eastern Finland, University of Helsinki.
It is widely acknowledged that the use of general anesthetics can undermine the relevance of electrophysiological or microscopical data obtained from a living animal’s brain. Moreover, the lengthy recovery from anesthesia limits the frequency of repeated recording/imaging episodes in longitudinal studies. Hence, new methods that would allow stable recordings from non-anesthetized behaving mice are expected to advance the fields of cellular and cognitive neurosciences. Existing solutions range from mere physical restraint to more sophisticated approaches, such as linear and spherical treadmills used in combination with computer-generated virtual reality. Here, a novel method is described where a head-fixed mouse can move around an air-lifted mobile homecage and explore its environment under stress-free conditions. This method allows researchers to perform behavioral tests (e.g.
, learning, habituation or novel object recognition) simultaneously with two-photon microscopic imaging and/or patch-clamp recordings, all combined in a single experiment. This video-article describes the use of the awake animal head fixation device (mobile homecage), demonstrates the procedures of animal habituation, and exemplifies a number of possible applications of the method.
Empty Value, Issue 88, awake, in vivo two-photon microscopy, blood vessels, dendrites, dendritic spines, Ca2+ imaging, intrinsic optical imaging, patch-clamp
Insertion of Flexible Neural Probes Using Rigid Stiffeners Attached with Biodissolvable Adhesive
Institutions: Lawrence Livermore National Laboratory, University of California, San Francisco.
Microelectrode arrays for neural interface devices that are made of biocompatible thin-film polymer are expected to have extended functional lifetime because the flexible material may minimize adverse tissue response caused by micromotion. However, their flexibility prevents them from being accurately inserted into neural tissue. This article demonstrates a method to temporarily attach a flexible microelectrode probe to a rigid stiffener using biodissolvable polyethylene glycol (PEG) to facilitate precise, surgical insertion of the probe. A unique stiffener design allows for uniform distribution of the PEG adhesive along the length of the probe. Flip-chip bonding, a common tool used in microelectronics packaging, enables accurate and repeatable alignment and attachment of the probe to the stiffener. The probe and stiffener are surgically implanted together, then the PEG is allowed to dissolve so that the stiffener can be extracted leaving the probe in place. Finally, an in vitro
test method is used to evaluate stiffener extraction in an agarose gel model of brain tissue. This approach to implantation has proven particularly advantageous for longer flexible probes (>3 mm). It also provides a feasible method to implant dual-sided flexible probes. To date, the technique has been used to obtain various in vivo
recording data from the rat cortex.
Bioengineering, Issue 79, Nervous System Diseases, Surgical Procedures, Operative, Investigative Techniques, Nonmetallic Materials, Engineering (General), neural interfaces, polymer neural probes, surgical insertion, polyethylene glycol, microelectrode arrays, chronic implantation
Training Rats to Voluntarily Dive Underwater: Investigations of the Mammalian Diving Response
Institutions: Midwestern University.
Underwater submergence produces autonomic changes that are observed in virtually all diving animals. This reflexly-induced response consists of apnea, a parasympathetically-induced bradycardia and a sympathetically-induced alteration of vascular resistance that maintains blood flow to the heart, brain and exercising muscles. While many of the metabolic and cardiorespiratory aspects of the diving response have been studied in marine animals, investigations of the central integrative aspects of this brainstem reflex have been relatively lacking. Because the physiology and neuroanatomy of the rat are well characterized, the rat can be used to help ascertain the central pathways of the mammalian diving response. Detailed instructions are provided on how to train rats to swim and voluntarily dive underwater through a 5 m long Plexiglas maze. Considerations regarding tank design and procedure room requirements are also given. The behavioral training is conducted in such a way as to reduce the stressfulness that could otherwise be associated with forced underwater submergence, thus minimizing activation of central stress pathways. The training procedures are not technically difficult, but they can be time-consuming. Since behavioral training of animals can only provide a model to be used with other experimental techniques, examples of how voluntarily diving rats have been used in conjunction with other physiological and neuroanatomical research techniques, and how the basic training procedures may need to be modified to accommodate these techniques, are also provided. These experiments show that voluntarily diving rats exhibit the same cardiorespiratory changes typically seen in other diving animals. The ease with which rats can be trained to voluntarily dive underwater, and the already available data from rats collected in other neurophysiological studies, makes voluntarily diving rats a good behavioral model to be used in studies investigating the central aspects of the mammalian diving response.
Behavior, Issue 93, Rat, Rattus norvegicus, voluntary diving, diving response, diving reflex, autonomic reflex, central integration
Recording Human Electrocorticographic (ECoG) Signals for Neuroscientific Research and Real-time Functional Cortical Mapping
Institutions: New York State Department of Health, Albany Medical College, Albany Medical College, Washington University, Rensselaer Polytechnic Institute, State University of New York at Albany, University of Texas at El Paso .
Neuroimaging studies of human cognitive, sensory, and motor processes are usually based on noninvasive techniques such as electroencephalography (EEG), magnetoencephalography or functional magnetic-resonance imaging. These techniques have either inherently low temporal or low spatial resolution, and suffer from low signal-to-noise ratio and/or poor high-frequency sensitivity. Thus, they are suboptimal for exploring the short-lived spatio-temporal dynamics of many of the underlying brain processes. In contrast, the invasive technique of electrocorticography (ECoG) provides brain signals that have an exceptionally high signal-to-noise ratio, less susceptibility to artifacts than EEG, and a high spatial and temporal resolution (i.e., <1 cm/<1 millisecond, respectively). ECoG involves measurement of electrical brain signals using electrodes that are implanted subdurally on the surface of the brain. Recent studies have shown that ECoG amplitudes in certain frequency bands carry substantial information about task-related activity, such as motor execution and planning1
, auditory processing2
and visual-spatial attention3
. Most of this information is captured in the high gamma range (around 70-110 Hz). Thus, gamma activity has been proposed as a robust and general indicator of local cortical function1-5
. ECoG can also reveal functional connectivity and resolve finer task-related spatial-temporal dynamics, thereby advancing our understanding of large-scale cortical processes. It has especially proven useful for advancing brain-computer interfacing (BCI) technology for decoding a user's intentions to enhance or improve communication6
. Nevertheless, human ECoG data are often hard to obtain because of the risks and limitations of the invasive procedures involved, and the need to record within the constraints of clinical settings. Still, clinical monitoring to localize epileptic foci offers a unique and valuable opportunity to collect human ECoG data. We describe our methods for collecting recording ECoG, and demonstrate how to use these signals for important real-time applications such as clinical mapping and brain-computer interfacing. Our example uses the BCI2000 software platform8,9
and the SIGFRIED10
method, an application for real-time mapping of brain functions. This procedure yields information that clinicians can subsequently use to guide the complex and laborious process of functional mapping by electrical stimulation.
Prerequisites and Planning:
Patients with drug-resistant partial epilepsy may be candidates for resective surgery of an epileptic focus to minimize the frequency of seizures. Prior to resection, the patients undergo monitoring using subdural electrodes for two purposes: first, to localize the epileptic focus, and second, to identify nearby critical brain areas (i.e., eloquent cortex) where resection could result in long-term functional deficits. To implant electrodes, a craniotomy is performed to open the skull. Then, electrode grids and/or strips are placed on the cortex, usually beneath the dura. A typical grid has a set of 8 x 8 platinum-iridium electrodes of 4 mm diameter (2.3 mm exposed surface) embedded in silicon with an inter-electrode distance of 1cm. A strip typically contains 4 or 6 such electrodes in a single line. The locations for these grids/strips are planned by a team of neurologists and neurosurgeons, and are based on previous EEG monitoring, on a structural MRI of the patient's brain, and on relevant factors of the patient's history. Continuous recording over a period of 5-12 days serves to localize epileptic foci, and electrical stimulation via the implanted electrodes allows clinicians to map eloquent cortex. At the end of the monitoring period, explantation of the electrodes and therapeutic resection are performed together in one procedure.
In addition to its primary clinical purpose, invasive monitoring also provides a unique opportunity to acquire human ECoG data for neuroscientific research. The decision to include a prospective patient in the research is based on the planned location of their electrodes, on the patient's performance scores on neuropsychological assessments, and on their informed consent, which is predicated on their understanding that participation in research is optional and is not related to their treatment. As with all research involving human subjects, the research protocol must be approved by the hospital's institutional review board. The decision to perform individual experimental tasks is made day-by-day, and is contingent on the patient's endurance and willingness to participate. Some or all of the experiments may be prevented by problems with the clinical state of the patient, such as post-operative facial swelling, temporary aphasia, frequent seizures, post-ictal fatigue and confusion, and more general pain or discomfort.
At the Epilepsy Monitoring Unit at Albany Medical Center in Albany, New York, clinical monitoring is implemented around the clock using a 192-channel Nihon-Kohden Neurofax monitoring system. Research recordings are made in collaboration with the Wadsworth Center of the New York State Department of Health in Albany. Signals from the ECoG electrodes are fed simultaneously to the research and the clinical systems via splitter connectors. To ensure that the clinical and research systems do not interfere with each other, the two systems typically use separate grounds. In fact, an epidural strip of electrodes is sometimes implanted to provide a ground for the clinical system. Whether research or clinical recording system, the grounding electrode is chosen to be distant from the predicted epileptic focus and from cortical areas of interest for the research. Our research system consists of eight synchronized 16-channel g.USBamp amplifier/digitizer units (g.tec, Graz, Austria). These were chosen because they are safety-rated and FDA-approved for invasive recordings, they have a very low noise-floor in the high-frequency range in which the signals of interest are found, and they come with an SDK that allows them to be integrated with custom-written research software. In order to capture the high-gamma signal accurately, we acquire signals at 1200Hz sampling rate-considerably higher than that of the typical EEG experiment or that of many clinical monitoring systems. A built-in low-pass filter automatically prevents aliasing of signals higher than the digitizer can capture. The patient's eye gaze is tracked using a monitor with a built-in Tobii T-60 eye-tracking system (Tobii Tech., Stockholm, Sweden). Additional accessories such as joystick, bluetooth Wiimote (Nintendo Co.), data-glove (5th
Dimension Technologies), keyboard, microphone, headphones, or video camera are connected depending on the requirements of the particular experiment.
Data collection, stimulus presentation, synchronization with the different input/output accessories, and real-time analysis and visualization are accomplished using our BCI2000 software8,9
. BCI2000 is a freely available general-purpose software system for real-time biosignal data acquisition, processing and feedback. It includes an array of pre-built modules that can be flexibly configured for many different purposes, and that can be extended by researchers' own code in C++, MATLAB or Python. BCI2000 consists of four modules that communicate with each other via a network-capable protocol: a Source module that handles the acquisition of brain signals from one of 19 different hardware systems from different manufacturers; a Signal Processing module that extracts relevant ECoG features and translates them into output signals; an Application module that delivers stimuli and feedback to the subject; and the Operator module that provides a graphical interface to the investigator.
A number of different experiments may be conducted with any given patient. The priority of experiments will be determined by the location of the particular patient's electrodes. However, we usually begin our experimentation using the SIGFRIED (SIGnal modeling For Realtime Identification and Event Detection) mapping method, which detects and displays significant task-related activity in real time. The resulting functional map allows us to further tailor subsequent experimental protocols and may also prove as a useful starting point for traditional mapping by electrocortical stimulation (ECS).
Although ECS mapping remains the gold standard for predicting the clinical outcome of resection, the process of ECS mapping is time consuming and also has other problems, such as after-discharges or seizures. Thus, a passive functional mapping technique may prove valuable in providing an initial estimate of the locus of eloquent cortex, which may then be confirmed and refined by ECS. The results from our passive SIGFRIED mapping technique have been shown to exhibit substantial concurrence with the results derived using ECS mapping10
The protocol described in this paper establishes a general methodology for gathering human ECoG data, before proceeding to illustrate how experiments can be initiated using the BCI2000 software platform. Finally, as a specific example, we describe how to perform passive functional mapping using the BCI2000-based SIGFRIED system.
Neuroscience, Issue 64, electrocorticography, brain-computer interfacing, functional brain mapping, SIGFRIED, BCI2000, epilepsy monitoring, magnetic resonance imaging, MRI
Designing Silk-silk Protein Alloy Materials for Biomedical Applications
Institutions: Rowan University, Rowan University, Cooper Medical School of Rowan University, Rowan University.
Fibrous proteins display different sequences and structures that have been used for various applications in biomedical fields such as biosensors, nanomedicine, tissue regeneration, and drug delivery. Designing materials based on the molecular-scale interactions between these proteins will help generate new multifunctional protein alloy biomaterials with tunable properties. Such alloy material systems also provide advantages in comparison to traditional synthetic polymers due to the materials biodegradability, biocompatibility, and tenability in the body. This article used the protein blends of wild tussah silk (Antheraea pernyi
) and domestic mulberry silk (Bombyx mori
) as an example to provide useful protocols regarding these topics, including how to predict protein-protein interactions by computational methods, how to produce protein alloy solutions, how to verify alloy systems by thermal analysis, and how to fabricate variable alloy materials including optical materials with diffraction gratings, electric materials with circuits coatings, and pharmaceutical materials for drug release and delivery. These methods can provide important information for designing the next generation multifunctional biomaterials based on different protein alloys.
Bioengineering, Issue 90, protein alloys, biomaterials, biomedical, silk blends, computational simulation, implantable electronic devices
Environmentally-controlled Microtensile Testing of Mechanically-adaptive Polymer Nanocomposites for ex vivo Characterization
Institutions: Louis Stokes Cleveland Department of Veterans Affairs Medical Center, Case Western Reserve University, Case Western Reserve University.
Implantable microdevices are gaining significant attention for several biomedical applications1-4
. Such devices have been made from a range of materials, each offering its own advantages and shortcomings5,6
. Most prominently, due to the microscale device dimensions, a high modulus is required to facilitate implantation into living tissue. Conversely, the stiffness of the device should match the surrounding tissue to minimize induced local strain7-9
. Therefore, we recently developed a new class of bio-inspired materials to meet these requirements by responding to environmental stimuli with a change in mechanical properties10-14
. Specifically, our poly(vinyl acetate)-based nanocomposite (PVAc-NC) displays a reduction in stiffness when exposed to water and elevated temperatures (e.g.
body temperature). Unfortunately, few methods exist to quantify the stiffness of materials in vivo15
, and mechanical testing outside of the physiological environment often requires large samples inappropriate for implantation. Further, stimuli-responsive materials may quickly recover their initial stiffness after explantation. Therefore, we have developed a method by which the mechanical properties of implanted microsamples can be measured ex vivo
, with simulated physiological conditions maintained using moisture and temperature control13,16,17
To this end, a custom microtensile tester was designed to accommodate microscale samples13,17
with widely-varying Young's moduli (range of 10 MPa to 5 GPa). As our interests are in the application of PVAc-NC as a biologically-adaptable neural probe substrate, a tool capable of mechanical characterization of samples at the microscale was necessary. This tool was adapted to provide humidity and temperature control, which minimized sample drying and cooling17
. As a result, the mechanical characteristics of the explanted sample closely reflect those of the sample just prior to explantation.
The overall goal of this method is to quantitatively assess the in vivo
mechanical properties, specifically the Young's modulus, of stimuli-responsive, mechanically-adaptive polymer-based materials. This is accomplished by first establishing the environmental conditions that will minimize a change in sample mechanical properties after explantation without contributing to a reduction in stiffness independent of that resulting from implantation. Samples are then prepared for implantation, handling, and testing (Figure 1A
). Each sample is implanted into the cerebral cortex of rats, which is represented here as an explanted rat brain, for a specified duration (Figure 1B
). At this point, the sample is explanted and immediately loaded into the microtensile tester, and then subjected to tensile testing (Figure 1C
). Subsequent data analysis provides insight into the mechanical behavior of these innovative materials in the environment of the cerebral cortex.
Bioengineering, Issue 78, Biophysics, Biomedical Engineering, Molecular Biology, Cellular Biology, Electrical Engineering, Materials Science, Nanotechnology, Nanocomposites, Electrodes, Implanted, Neural Prostheses, Micro-Electrical-Mechanical Systems, Implants, Experimental, mechanical properties (composite materials), Dynamic materials, polymer nanocomposite, Young's modulus, modulus of elasticity, intracortical microelectrode, polymers, biomaterials
A Method for Systematic Electrochemical and Electrophysiological Evaluation of Neural Recording Electrodes
Institutions: La Trobe University, University of Wollongong, ARC Centre of Excellence for Electromaterials Science, RMIT University.
New materials and designs for neural implants are typically tested separately, with a demonstration of performance but without reference to other implant characteristics. This precludes a rational selection of a particular implant as optimal for a particular application and the development of new materials based on the most critical performance parameters. This article develops a protocol for in vitro
and in vivo
testing of neural recording electrodes. Recommended parameters for electrochemical and electrophysiological testing are documented with the key steps and potential issues discussed. This method eliminates or reduces the impact of many systematic errors present in simpler in vivo
testing paradigms, especially variations in electrode/neuron distance and between animal models. The result is a strong correlation between the critical in vitro
and in vivo
responses, such as impedance and signal-to-noise ratio. This protocol can easily be adapted to test other electrode materials and designs. The in vitro
techniques can be expanded to any other nondestructive method to determine further important performance indicators. The principles used for the surgical approach in the auditory pathway can also be modified to other neural regions or tissue.
Neuroscience, Issue 85, Electrochemistry, Electrophysiology, Neural Recording, Neural Implant, Electrode Coating, Bionics
Performing Behavioral Tasks in Subjects with Intracranial Electrodes
Institutions: Cleveland Clinic Foundation, Cleveland Clinic Foundation, Cleveland Clinic Foundation, Johns Hopkins University.
Patients having stereo-electroencephalography (SEEG) electrode, subdural grid or depth electrode implants have a multitude of electrodes implanted in different areas of their brain for the localization of their seizure focus and eloquent areas. After implantation, the patient must remain in the hospital until the pathological area of brain is found and possibly resected. During this time, these patients offer a unique opportunity to the research community because any number of behavioral paradigms can be performed to uncover the neural correlates that guide behavior. Here we present a method for recording brain activity from intracranial implants as subjects perform a behavioral task designed to assess decision-making and reward encoding. All electrophysiological data from the intracranial electrodes are recorded during the behavioral task, allowing for the examination of the many brain areas involved in a single function at time scales relevant to behavior. Moreover, and unlike animal studies, human patients can learn a wide variety of behavioral tasks quickly, allowing for the ability to perform more than one task in the same subject or for performing controls. Despite the many advantages of this technique for understanding human brain function, there are also methodological limitations that we discuss, including environmental factors, analgesic effects, time constraints and recordings from diseased tissue. This method may be easily implemented by any institution that performs intracranial assessments; providing the opportunity to directly examine human brain function during behavior.
Behavior, Issue 92, Cognitive neuroscience, Epilepsy, Stereo-electroencephalography, Subdural grids, Behavioral method, Electrophysiology
Simultaneous fMRI and Electrophysiology in the Rodent Brain
Institutions: Emory University, Georgia Institute of Technology, Emory University.
To examine the neural basis of the blood oxygenation level dependent (BOLD) magnetic resonance imaging (MRI) signal, we have developed a rodent model in which functional MRI data and in vivo
intracortical recording can be performed simultaneously. The combination of MRI and electrical recording is technically challenging because the electrodes used for recording distort the MRI images and the MRI acquisition induces noise in the electrical recording. To minimize the mutual interference of the two modalities, glass microelectrodes were used rather than metal and a noise removal algorithm was implemented for the electrophysiology data. In our studies, two microelectrodes were separately implanted in bilateral primary somatosensory cortices (SI) of the rat and fixed in place. One coronal slice covering the electrode tips was selected for functional MRI. Electrode shafts and fixation positions were not included in the image slice to avoid imaging artifacts. The removed scalp was replaced with toothpaste to reduce susceptibility mismatch and prevent Gibbs ringing artifacts in the images. The artifact structure induced in the electrical recordings by the rapidly-switching magnetic fields during image acquisition was characterized by averaging all cycles of scans for each run. The noise structure during imaging was then subtracted from original recordings. The denoised time courses were then used for further analysis in combination with the fMRI data. As an example, the simultaneous acquisition was used to determine the relationship between spontaneous fMRI BOLD signals and band-limited intracortical electrical activity. Simultaneous fMRI and electrophysiological recording in the rodent will provide a platform for many exciting applications in neuroscience in addition to elucidating the relationship between the fMRI BOLD signal and neuronal activity.
Neuroscience, Issue 42, fMRI, electrophysiology, rat, BOLD, brain, resting state
Functional Imaging with Reinforcement, Eyetracking, and Physiological Monitoring
Institutions: Columbia University, Columbia University, Columbia University.
We use functional brain imaging (fMRI) to study neural circuits that underlie decision-making. To understand how outcomes affect decision processes, simple perceptual tasks are combined with appetitive and aversive reinforcement. However, the use of reinforcers such as juice and airpuffs can create challenges for fMRI. Reinforcer delivery can cause head movement, which creates artifacts in the fMRI signal. Reinforcement can also lead to changes in heart rate and respiration that are mediated by autonomic pathways. Changes in heart rate and respiration can directly affect the fMRI (BOLD) signal in the brain and can be confounded with signal changes that are due to neural activity. In this presentation, we demonstrate methods for administering reinforcers in a controlled manner, for stabilizing the head, and for measuring pulse and respiration.
Medicine, Issue 21, Neuroscience, Psychiatry, fMRI, Decision Making, Reward, Punishment, Pulse, Respiration, Eye Tracking, Psychology