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Pubmed Article
Describing temperament in an ungulate: a multidimensional approach.
PUBLISHED: 01-01-2013
Studies on animal temperament have often described temperament using a one-dimensional scale, whereas theoretical framework has recently suggested two or more dimensions using terms like "valence" or "arousal" to describe these dimensions. Yet, the valence or assessment of a situation is highly individual. The aim of this study was to provide support for the multidimensional framework with experimental data originating from an economically important species (Bos taurus). We tested 361 calves at 90 days post natum (dpn) in a novel-object test. Using a principal component analysis (PCA), we condensed numerous behaviours into fewer variables to describe temperament and correlated these variables with simultaneously measured heart rate variability (HRV) data. The PCA resulted in two behavioural dimensions (principal components, PC): novel-object-related (PC 1) and exploration-activity-related (PC 2). These PCs explained 58% of the variability in our data. The animals were distributed evenly within the two behavioural dimensions independent of their sex. Calves with different scores in these PCs differed significantly in HRV, and thus in the autonomous nervous systems activity. Based on these combined behavioural and physiological data we described four distinct temperament types resulting from two behavioural dimensions: "neophobic/fearful--alert", "interested--stressed", "subdued/uninterested--calm", and "neoophilic/outgoing--alert". Additionally, 38 calves were tested at 90 and 197 dpn. Using the same PCA-model, they correlated significantly in PC 1 and tended to correlate in PC 2 between the two test ages. Of these calves, 42% expressed a similar behaviour pattern in both dimensions and 47% in one. No differences in temperament scores were found between sexes or breeds. In conclusion, we described distinct temperament types in calves based on behavioural and physiological measures emphasising the benefits of a multidimensional approach.
Authors: Phoebe Spetsieris, Yilong Ma, Shichun Peng, Ji Hyun Ko, Vijay Dhawan, Chris C. Tang, David Eidelberg.
Published: 06-26-2013
The scaled subprofile model (SSM)1-4 is a multivariate PCA-based algorithm that identifies major sources of variation in patient and control group brain image data while rejecting lesser components (Figure 1). Applied directly to voxel-by-voxel covariance data of steady-state multimodality images, an entire group image set can be reduced to a few significant linearly independent covariance patterns and corresponding subject scores. Each pattern, termed a group invariant subprofile (GIS), is an orthogonal principal component that represents a spatially distributed network of functionally interrelated brain regions. Large global mean scalar effects that can obscure smaller network-specific contributions are removed by the inherent logarithmic conversion and mean centering of the data2,5,6. Subjects express each of these patterns to a variable degree represented by a simple scalar score that can correlate with independent clinical or psychometric descriptors7,8. Using logistic regression analysis of subject scores (i.e. pattern expression values), linear coefficients can be derived to combine multiple principal components into single disease-related spatial covariance patterns, i.e. composite networks with improved discrimination of patients from healthy control subjects5,6. Cross-validation within the derivation set can be performed using bootstrap resampling techniques9. Forward validation is easily confirmed by direct score evaluation of the derived patterns in prospective datasets10. Once validated, disease-related patterns can be used to score individual patients with respect to a fixed reference sample, often the set of healthy subjects that was used (with the disease group) in the original pattern derivation11. These standardized values can in turn be used to assist in differential diagnosis12,13 and to assess disease progression and treatment effects at the network level7,14-16. We present an example of the application of this methodology to FDG PET data of Parkinson's Disease patients and normal controls using our in-house software to derive a characteristic covariance pattern biomarker of disease.
27 Related JoVE Articles!
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The Tail Suspension Test
Authors: Adem Can, David T. Dao, Chantelle E. Terrillion, Sean C. Piantadosi, Shambhu Bhat, Todd D. Gould.
Institutions: University of Maryland School of Medicine, Tulane University School of Medicine, University of Maryland , University of Maryland School of Medicine.
The tail-suspension test is a mouse behavioral test useful in the screening of potential antidepressant drugs, and assessing of other manipulations that are expected to affect depression related behaviors. Mice are suspended by their tails with tape, in such a position that it cannot escape or hold on to nearby surfaces. During this test, typically six minutes in duration, the resulting escape oriented behaviors are quantified. The tail-suspension test is a valuable tool in drug discovery for high-throughput screening of prospective antidepressant compounds. Here, we describe the details required for implementation of this test with additional emphasis on potential problems that may occur and how to avoid them. We also offer a solution to the tail climbing behavior, a common problem that renders this test useless in some mouse strains, such as the widely used C57BL/6. Specifically, we prevent tail climbing behaviors by passing mouse tails through a small plastic cylinder prior to suspension. Finally, we detail how to manually score the behaviors that are manifested in this test.
Neuroscience, Issue 59, animal models, behavioral analysis, neuroscience, neurobiology, mood disorder, depression, mood stabilizer, antidepressant
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A Novel Method of Drug Administration to Multiple Zebrafish (Danio rerio) and the Quantification of Withdrawal
Authors: Adam Holcombe, Melike Schalomon, Trevor James Hamilton.
Institutions: MacEwan University.
Anxiety testing in zebrafish is often studied in combination with the application of pharmacological substances. In these studies, fish are routinely netted and transported between home aquaria and dosing tanks. In order to enhance the ease of compound administration, a novel method for transferring fish between tanks for drug administration was developed. Inserts that are designed for spawning were used to transfer groups of fish into the drug solution, allowing accurate dosing of all fish in the group. This increases the precision and efficiency of dosing, which becomes very important in long schedules of repeated drug administration. We implemented this procedure for use in a study examining the behavior of zebrafish in the light/dark test after administering ethanol with differing 21 day schedules. In fish exposed to daily-moderate amounts of alcohol there was a significant difference in location preference after 2 days of withdrawal when compared to the control group. However, a significant difference in location preference in a group exposed to weekly-binge administration was not observed. This protocol can be generalized for use with all types of compounds that are water-soluble and may be used in any situation when the behavior of fish during or after long schedules of drug administration is being examined. The light/dark test is also a valuable method of assessing withdrawal-induced changes in anxiety.
Neuroscience, Issue 93, Zebrafish, Ethanol, Behavior, Anxiety, Pharmacology, Fish, Neuroscience, Drug administration, Scototaxis
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An Investigation of the Effects of Sports-related Concussion in Youth Using Functional Magnetic Resonance Imaging and the Head Impact Telemetry System
Authors: Michelle Keightley, Stephanie Green, Nick Reed, Sabrina Agnihotri, Amy Wilkinson, Nancy Lobaugh.
Institutions: University of Toronto, University of Toronto, University of Toronto, Bloorview Kids Rehab, Toronto Rehab, Sunnybrook Health Sciences Centre, University of Toronto.
One of the most commonly reported injuries in children who participate in sports is concussion or mild traumatic brain injury (mTBI)1. Children and youth involved in organized sports such as competitive hockey are nearly six times more likely to suffer a severe concussion compared to children involved in other leisure physical activities2. While the most common cognitive sequelae of mTBI appear similar for children and adults, the recovery profile and breadth of consequences in children remains largely unknown2, as does the influence of pre-injury characteristics (e.g. gender) and injury details (e.g. magnitude and direction of impact) on long-term outcomes. Competitive sports, such as hockey, allow the rare opportunity to utilize a pre-post design to obtain pre-injury data before concussion occurs on youth characteristics and functioning and to relate this to outcome following injury. Our primary goals are to refine pediatric concussion diagnosis and management based on research evidence that is specific to children and youth. To do this we use new, multi-modal and integrative approaches that will: 1.Evaluate the immediate effects of head trauma in youth 2.Monitor the resolution of post-concussion symptoms (PCS) and cognitive performance during recovery 3.Utilize new methods to verify brain injury and recovery To achieve our goals, we have implemented the Head Impact Telemetry (HIT) System. (Simbex; Lebanon, NH, USA). This system equips commercially available Easton S9 hockey helmets (Easton-Bell Sports; Van Nuys, CA, USA) with single-axis accelerometers designed to measure real-time head accelerations during contact sport participation 3 - 5. By using telemetric technology, the magnitude of acceleration and location of all head impacts during sport participation can be objectively detected and recorded. We also use functional magnetic resonance imaging (fMRI) to localize and assess changes in neural activity specifically in the medial temporal and frontal lobes during the performance of cognitive tasks, since those are the cerebral regions most sensitive to concussive head injury 6. Finally, we are acquiring structural imaging data sensitive to damage in brain white matter.
Medicine, Issue 47, Mild traumatic brain injury, concussion, fMRI, youth, Head Impact Telemetry System
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Bottom-up and Shotgun Proteomics to Identify a Comprehensive Cochlear Proteome
Authors: Lancia N.F. Darville, Bernd H.A. Sokolowski.
Institutions: University of South Florida.
Proteomics is a commonly used approach that can provide insights into complex biological systems. The cochlear sensory epithelium contains receptors that transduce the mechanical energy of sound into an electro-chemical energy processed by the peripheral and central nervous systems. Several proteomic techniques have been developed to study the cochlear inner ear, such as two-dimensional difference gel electrophoresis (2D-DIGE), antibody microarray, and mass spectrometry (MS). MS is the most comprehensive and versatile tool in proteomics and in conjunction with separation methods can provide an in-depth proteome of biological samples. Separation methods combined with MS has the ability to enrich protein samples, detect low molecular weight and hydrophobic proteins, and identify low abundant proteins by reducing the proteome dynamic range. Different digestion strategies can be applied to whole lysate or to fractionated protein lysate to enhance peptide and protein sequence coverage. Utilization of different separation techniques, including strong cation exchange (SCX), reversed-phase (RP), and gel-eluted liquid fraction entrapment electrophoresis (GELFrEE) can be applied to reduce sample complexity prior to MS analysis for protein identification.
Biochemistry, Issue 85, Cochlear, chromatography, LC-MS/MS, mass spectrometry, Proteomics, sensory epithelium
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Linking Predation Risk, Herbivore Physiological Stress and Microbial Decomposition of Plant Litter
Authors: Oswald J. Schmitz, Mark A. Bradford, Michael S. Strickland, Dror Hawlena.
Institutions: Yale University, Virginia Tech, The Hebrew University of Jerusalem.
The quantity and quality of detritus entering the soil determines the rate of decomposition by microbial communities as well as recycle rates of nitrogen (N) and carbon (C) sequestration1,2. Plant litter comprises the majority of detritus3, and so it is assumed that decomposition is only marginally influenced by biomass inputs from animals such as herbivores and carnivores4,5. However, carnivores may influence microbial decomposition of plant litter via a chain of interactions in which predation risk alters the physiology of their herbivore prey that in turn alters soil microbial functioning when the herbivore carcasses are decomposed6. A physiological stress response by herbivores to the risk of predation can change the C:N elemental composition of herbivore biomass7,8,9 because stress from predation risk increases herbivore basal energy demands that in nutrient-limited systems forces herbivores to shift their consumption from N-rich resources to support growth and reproduction to C-rich carbohydrate resources to support heightened metabolism6. Herbivores have limited ability to store excess nutrients, so stressed herbivores excrete N as they increase carbohydrate-C consumption7. Ultimately, prey stressed by predation risk increase their body C:N ratio7,10, making them poorer quality resources for the soil microbial pool likely due to lower availability of labile N for microbial enzyme production6. Thus, decomposition of carcasses of stressed herbivores has a priming effect on the functioning of microbial communities that decreases subsequent ability to of microbes to decompose plant litter6,10,11. We present the methodology to evaluate linkages between predation risk and litter decomposition by soil microbes. We describe how to: induce stress in herbivores from predation risk; measure those stress responses, and measure the consequences on microbial decomposition. We use insights from a model grassland ecosystem comprising the hunting spider predator (Pisuarina mira), a dominant grasshopper herbivore (Melanoplus femurrubrum),and a variety of grass and forb plants9.
Environmental Sciences, Issue 73, Microbiology, Plant Biology, Entomology, Organisms, Investigative Techniques, Biological Phenomena, Chemical Phenomena, Metabolic Phenomena, Microbiological Phenomena, Earth Resources and Remote Sensing, Life Sciences (General), Litter Decomposition, Ecological Stoichiometry, Physiological Stress and Ecosystem Function, Predation Risk, Soil Respiration, Carbon Sequestration, Soil Science, respiration, spider, grasshoper, model system
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Planar and Three-Dimensional Printing of Conductive Inks
Authors: Bok Yeop Ahn, Steven B. Walker, Scott C. Slimmer, Analisa Russo, Ashley Gupta, Steve Kranz, Eric B. Duoss, Thomas F. Malkowski, Jennifer A. Lewis.
Institutions: University of Illinois at Urbana-Champaign, Lawrence Livermore National Laboratory, University Of California Santa Barbara .
Printed electronics rely on low-cost, large-area fabrication routes to create flexible or multidimensional electronic, optoelectronic, and biomedical devices1-3. In this paper, we focus on one- (1D), two- (2D), and three-dimensional (3D) printing of conductive metallic inks in the form of flexible, stretchable, and spanning microelectrodes. Direct-write assembly4,5 is a 1-to-3D printing technique that enables the fabrication of features ranging from simple lines to complex structures by the deposition of concentrated inks through fine nozzles (~0.1 - 250 μm). This printing method consists of a computer-controlled 3-axis translation stage, an ink reservoir and nozzle, and 10x telescopic lens for visualization. Unlike inkjet printing, a droplet-based process, direct-write assembly involves the extrusion of ink filaments either in- or out-of-plane. The printed filaments typically conform to the nozzle size. Hence, microscale features (< 1 μm) can be patterned and assembled into larger arrays and multidimensional architectures. In this paper, we first synthesize a highly concentrated silver nanoparticle ink for planar and 3D printing via direct-write assembly. Next, a standard protocol for printing microelectrodes in multidimensional motifs is demonstrated. Finally, applications of printed microelectrodes for electrically small antennas, solar cells, and light-emitting diodes are highlighted.
Bioengineering, Issue 58, Direct-write assembly, silver ink, 3D printing, planar, three-dimensional, microelectrodes, flexible electronics, printed electronics
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Using a Comparative Species Approach to Investigate the Neurobiology of Paternal Responses
Authors: Catherine L. Franssen, Massimo Bardi, Kelly G. Lambert.
Institutions: Randolph-Macon College, Marshall University.
A goal of behavioral neuroscience is to identify underlying neurobiological factors that regulate specific behaviors. Using animal models to accomplish this goal, many methodological strategies require invasive techniques to manipulate the intensity of the behavior of interest (e.g., lesion methods, pharmacological manipulations, microdialysis techniques, genetically-engineered animal models). The utilization of a comparative species approach allows researchers to take advantage of naturally occurring differences in response strategies existing in closely related species. In our lab, we use two species of the Peromyscus genus that differ in paternal responses. The male California deer mouse (Peromyscus californicus) exhibits the same parental responses as the female whereas its cousin, the common deer mouse (Peromyscus maniculatus) exhibits virtually no nurturing/parental responses in the presence of pups. Of specific interest in this article is an exploration of the neurobiological factors associated with the affiliative social responses exhibited by the paternal California deer mouse. Because the behavioral neuroscience approach is multifaceted, the following key components of the study will be briefly addressed: the identification of appropriate species for this type of research; data collection for behavioral analysis; preparation and sectioning of the brains; basic steps involved in immunocytochemistry for the quantification of vasopressin-immunoreactivity; the use of neuroimaging software to quantify the brain tissue; the use of a microsequencing video analysis to score behavior and, finally, the appropriate statistical analyses to provide the most informed interpretations of the research findings.
Neuroscience, Issue 55, Peromyscus, mouse, paternal behavior, vasopressin, immunocytochemistry, microsequencing behavioral analysis
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Improved Visualization and Quantitative Analysis of Drug Effects Using Micropatterned Cells
Authors: Sébastien Degot, Muriel Auzan, Violaine Chapuis, Anne Béghin, Amélie Chadeyras, Constantin Nelep, Maria Luisa Calvo-Muñoz, Joanne Young, François Chatelain, Alexandra Fuchs.
Institutions: Grenoble, France, Faculté de Médecine Rockefeller, Lyon, France.
To date, most HCA (High Content Analysis) studies are carried out with adherent cell lines grown on a homogenous substrate in tissue-culture treated micro-plates. Under these conditions, cells spread and divide in all directions resulting in an inherent variability in cell shape, morphology and behavior. The high cell-to-cell variance of the overall population impedes the success of HCA, especially for drug development. The ability of micropatterns to normalize the shape and internal polarity of every individual cell provides a tremendous opportunity for solving this critical bottleneck 1-2. To facilitate access and use of the micropatterning technology, CYTOO has developed a range of ready to use micropatterns, available in coverslip and microwell formats. In this video article, we provide detailed protocols of all the procedures from cell seeding on CYTOOchip micropatterns, drug treatment, fixation and staining to automated acquisition, automated image processing and final data analysis. With this example, we illustrate how micropatterns can facilitate cell-based assays. Alterations of the cell cytoskeleton are difficult to quantify in cells cultured on homogenous substrates, but culturing cells on micropatterns results in a reproducible organization of the actin meshwork due to systematic positioning of the cell adhesion contacts in every cell. Such normalization of the intracellular architecture allows quantification of even small effects on the actin cytoskeleton as demonstrated in these set of protocols using blebbistatin, an inhibitor of the actin-myosin interaction.
Cellular Biology, Issue 46, Adhesive micropatterns, cell normalization, High Content Analysis, actin, image analysis, cytoskeleton, blebbistatin, cell-based assay, cell imaging, drug screening
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Transabdominal Ultrasound for Pregnancy Diagnosis in Reeves' Muntjac Deer
Authors: Kelly D. Walton, Erin McNulty, Amy V. Nalls, Candace K. Mathiason.
Institutions: Colorado State University.
Reeves' muntjac deer (Muntiacus reevesi) are a small cervid species native to southeast Asia, and are currently being investigated as a potential model of prion disease transmission and pathogenesis. Vertical transmission is an area of interest among researchers studying infectious diseases, including prion disease, and these investigations require efficient methods for evaluating the effects of maternal infection on reproductive performance. Ultrasonographic examination is a well-established tool for diagnosing pregnancy and assessing fetal health in many animal species1-7, including several species of farmed cervids8-19, however this technique has not been described in Reeves' muntjac deer. Here we describe the application of transabdominal ultrasound to detect pregnancy in muntjac does and to evaluate fetal growth and development throughout the gestational period. Using this procedure, pregnant animals were identified as early as 35 days following doe-buck pairing and this was an effective means to safely monitor the pregnancy at regular intervals. Future goals of this work will include establishing normal fetal measurement references for estimation of gestational age, determining sensitivity and specificity of the technique for diagnosing pregnancy at various stages of gestation, and identifying variations in fetal growth and development under different experimental conditions.
Medicine, Issue 83, Ultrasound, Reeves' muntjac deer, Muntiacus reevesi, fetal development, fetal growth, captive cervids
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ScanLag: High-throughput Quantification of Colony Growth and Lag Time
Authors: Irit Levin-Reisman, Ofer Fridman, Nathalie Q. Balaban.
Institutions: The Hebrew University of Jerusalem.
Growth dynamics are fundamental characteristics of microorganisms. Quantifying growth precisely is an important goal in microbiology. Growth dynamics are affected both by the doubling time of the microorganism and by any delay in growth upon transfer from one condition to another, the lag. The ScanLag method enables the characterization of these two independent properties at the level of colonies originating each from a single cell, generating a two-dimensional distribution of the lag time and of the growth time. In ScanLag, measurement of the time it takes for colonies on conventional nutrient agar plates to be detected is automated on an array of commercial scanners controlled by an in house application. Petri dishes are placed on the scanners, and the application acquires images periodically. Automated analysis of colony growth is then done by an application that returns the appearance time and growth rate of each colony. Other parameters, such as the shape, texture and color of the colony, can be extracted for multidimensional mapping of sub-populations of cells. Finally, the method enables the retrieval of rare variants with specific growth phenotypes for further characterization. The technique could be applied in bacteriology for the identification of long lag that can cause persistence to antibiotics, as well as a general low cost technique for phenotypic screens.
Immunology, Issue 89, lag, growth rate, growth delay, single cell, scanners, image analysis, persistence, resistance, rare mutants, phenotypic screens, phenomics
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Use of a High-throughput In Vitro Microfluidic System to Develop Oral Multi-species Biofilms
Authors: Derek S. Samarian, Nicholas S. Jakubovics, Ting L. Luo, Alexander H. Rickard.
Institutions: The University of Michigan, Newcastle University.
There are few high-throughput in vitro systems which facilitate the development of multi-species biofilms that contain numerous species commonly detected within in vivo oral biofilms. Furthermore, a system that uses natural human saliva as the nutrient source, instead of artificial media, is particularly desirable in order to support the expression of cellular and biofilm-specific properties that mimic the in vivo communities. We describe a method for the development of multi-species oral biofilms that are comparable, with respect to species composition, to supragingival dental plaque, under conditions similar to the human oral cavity. Specifically, this methods article will describe how a commercially available microfluidic system can be adapted to facilitate the development of multi-species oral biofilms derived from and grown within pooled saliva. Furthermore, a description of how the system can be used in conjunction with a confocal laser scanning microscope to generate 3-D biofilm reconstructions for architectural and viability analyses will be presented. Given the broad diversity of microorganisms that grow within biofilms in the microfluidic system (including Streptococcus, Neisseria, Veillonella, Gemella, and Porphyromonas), a protocol will also be presented describing how to harvest the biofilm cells for further subculture or DNA extraction and analysis. The limits of both the microfluidic biofilm system and the current state-of-the-art data analyses will be addressed. Ultimately, it is envisioned that this article will provide a baseline technique that will improve the study of oral biofilms and aid in the development of additional technologies that can be integrated with the microfluidic platform.
Bioengineering, Issue 94, Dental plaque, biofilm, confocal laser scanning microscopy, three-dimensional structure, pyrosequencing, image analysis, image reconstruction, saliva, modeling, COMSTAT, IMARIS, IMAGEJ, multi-species biofilm communities.
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Non-contact, Label-free Monitoring of Cells and Extracellular Matrix using Raman Spectroscopy
Authors: Miriam Votteler, Daniel A. Carvajal Berrio, Marieke Pudlas, Heike Walles, Katja Schenke-Layland.
Institutions: Eberhard Karls University, Tübingen, Fraunhofer Institute of Interfacial Engineering and Biotechnology (IGB) Stuttgart, Germany, University of Stuttgart, Germany, Julius-Maximillians University, Würzburg, Germany.
Non-destructive, non-contact and label-free technologies to monitor cell and tissue cultures are needed in the field of biomedical research.1-5 However, currently available routine methods require processing steps and alter sample integrity. Raman spectroscopy is a fast method that enables the measurement of biological samples without the need for further processing steps. This laser-based technology detects the inelastic scattering of monochromatic light.6 As every chemical vibration is assigned to a specific Raman band (wavenumber in cm-1), each biological sample features a typical spectral pattern due to their inherent biochemical composition.7-9 Within Raman spectra, the peak intensities correlate with the amount of the present molecular bonds.1 Similarities and differences of the spectral data sets can be detected by employing a multivariate analysis (e.g. principal component analysis (PCA)).10 Here, we perform Raman spectroscopy of living cells and native tissues. Cells are either seeded on glass bottom dishes or kept in suspension under normal cell culture conditions (37 °C, 5% CO2) before measurement. Native tissues are dissected and stored in phosphate buffered saline (PBS) at 4 °C prior measurements. Depending on our experimental set up, we then either focused on the cell nucleus or extracellular matrix (ECM) proteins such as elastin and collagen. For all studies, a minimum of 30 cells or 30 random points of interest within the ECM are measured. Data processing steps included background subtraction and normalization.
Bioengineering, Issue 63, Raman spectroscopy, label-free analysis, living cells, extracellular matrix, tissue engineering
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Improved In-gel Reductive β-Elimination for Comprehensive O-linked and Sulfo-glycomics by Mass Spectrometry
Authors: David B. Nix, Tadahiro Kumagai, Toshihiko Katoh, Michael Tiemeyer, Kazuhiro Aoki.
Institutions: University of Georgia, University of Georgia, Ishikawa Prefectural University.
Separation of proteins by SDS-PAGE followed by in-gel proteolytic digestion of resolved protein bands has produced high-resolution proteomic analysis of biological samples. Similar approaches, that would allow in-depth analysis of the glycans carried by glycoproteins resolved by SDS-PAGE, require special considerations in order to maximize recovery and sensitivity when using mass spectrometry (MS) as the detection method. A major hurdle to be overcome in achieving high-quality data is the removal of gel-derived contaminants that interfere with MS analysis. The sample workflow presented here is robust, efficient, and eliminates the need for in-line HPLC clean-up prior to MS. Gel pieces containing target proteins are washed in acetonitrile, water, and ethyl acetate to remove contaminants, including polymeric acrylamide fragments. O-linked glycans are released from target proteins by in-gel reductive β-elimination and recovered through robust, simple clean-up procedures. An advantage of this workflow is that it improves sensitivity for detecting and characterizing sulfated glycans. These procedures produce an efficient separation of sulfated permethylated glycans from non-sulfated (sialylated and neutral) permethylated glycans by a rapid phase-partition prior to MS analysis, and thereby enhance glycomic and sulfoglycomic analyses of glycoproteins resolved by SDS-PAGE.
Chemistry, Issue 93, glycoprotein, glycosylation, in-gel reductive β-elimination, O-linked glycan, sulfated glycan, mass spectrometry, protein ID, SDS-PAGE, glycomics, sulfoglycomics
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Assessment and Evaluation of the High Risk Neonate: The NICU Network Neurobehavioral Scale
Authors: Barry M. Lester, Lynne Andreozzi-Fontaine, Edward Tronick, Rosemarie Bigsby.
Institutions: Brown University, Women & Infants Hospital of Rhode Island, University of Massachusetts, Boston.
There has been a long-standing interest in the assessment of the neurobehavioral integrity of the newborn infant. The NICU Network Neurobehavioral Scale (NNNS) was developed as an assessment for the at-risk infant. These are infants who are at increased risk for poor developmental outcome because of insults during prenatal development, such as substance exposure or prematurity or factors such as poverty, poor nutrition or lack of prenatal care that can have adverse effects on the intrauterine environment and affect the developing fetus. The NNNS assesses the full range of infant neurobehavioral performance including neurological integrity, behavioral functioning, and signs of stress/abstinence. The NNNS is a noninvasive neonatal assessment tool with demonstrated validity as a predictor, not only of medical outcomes such as cerebral palsy diagnosis, neurological abnormalities, and diseases with risks to the brain, but also of developmental outcomes such as mental and motor functioning, behavior problems, school readiness, and IQ. The NNNS can identify infants at high risk for abnormal developmental outcome and is an important clinical tool that enables medical researchers and health practitioners to identify these infants and develop intervention programs to optimize the development of these infants as early as possible. The video shows the NNNS procedures, shows examples of normal and abnormal performance and the various clinical populations in which the exam can be used.
Behavior, Issue 90, NICU Network Neurobehavioral Scale, NNNS, High risk infant, Assessment, Evaluation, Prediction, Long term outcome
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Perceptual and Category Processing of the Uncanny Valley Hypothesis' Dimension of Human Likeness: Some Methodological Issues
Authors: Marcus Cheetham, Lutz Jancke.
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 (DHL) (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
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Quantification of Orofacial Phenotypes in Xenopus
Authors: Allyson E. Kennedy, Amanda J. Dickinson.
Institutions: Virginia Commonwealth University.
Xenopus has become an important tool for dissecting the mechanisms governing craniofacial development and defects. A method to quantify orofacial development will allow for more rigorous analysis of orofacial phenotypes upon abrogation with substances that can genetically or molecularly manipulate gene expression or protein function. Using two dimensional images of the embryonic heads, traditional size dimensions-such as orofacial width, height and area- are measured. In addition, a roundness measure of the embryonic mouth opening is used to describe the shape of the mouth. Geometric morphometrics of these two dimensional images is also performed to provide a more sophisticated view of changes in the shape of the orofacial region. Landmarks are assigned to specific points in the orofacial region and coordinates are created. A principle component analysis is used to reduce landmark coordinates to principle components that then discriminate the treatment groups. These results are displayed as a scatter plot in which individuals with similar orofacial shapes cluster together. It is also useful to perform a discriminant function analysis, which statistically compares the positions of the landmarks between two treatment groups. This analysis is displayed on a transformation grid where changes in landmark position are viewed as vectors. A grid is superimposed on these vectors so that a warping pattern is displayed to show where significant landmark positions have changed. Shape changes in the discriminant function analysis are based on a statistical measure, and therefore can be evaluated by a p-value. This analysis is simple and accessible, requiring only a stereoscope and freeware software, and thus will be a valuable research and teaching resource.
Developmental Biology, Issue 93, Orofacial quantification, geometric morphometrics, Xenopus, orofacial development, orofacial defects, shape changes, facial dimensions
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Oscillation and Reaction Board Techniques for Estimating Inertial Properties of a Below-knee Prosthesis
Authors: Jeremy D. Smith, Abbie E. Ferris, Gary D. Heise, Richard N. Hinrichs, Philip E. Martin.
Institutions: University of Northern Colorado, Arizona State University, Iowa State University.
The purpose of this study was two-fold: 1) demonstrate a technique that can be used to directly estimate the inertial properties of a below-knee prosthesis, and 2) contrast the effects of the proposed technique and that of using intact limb inertial properties on joint kinetic estimates during walking in unilateral, transtibial amputees. An oscillation and reaction board system was validated and shown to be reliable when measuring inertial properties of known geometrical solids. When direct measurements of inertial properties of the prosthesis were used in inverse dynamics modeling of the lower extremity compared with inertial estimates based on an intact shank and foot, joint kinetics at the hip and knee were significantly lower during the swing phase of walking. Differences in joint kinetics during stance, however, were smaller than those observed during swing. Therefore, researchers focusing on the swing phase of walking should consider the impact of prosthesis inertia property estimates on study outcomes. For stance, either one of the two inertial models investigated in our study would likely lead to similar outcomes with an inverse dynamics assessment.
Bioengineering, Issue 87, prosthesis inertia, amputee locomotion, below-knee prosthesis, transtibial amputee
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Analysis of Tubular Membrane Networks in Cardiac Myocytes from Atria and Ventricles
Authors: Eva Wagner, Sören Brandenburg, Tobias Kohl, Stephan E. Lehnart.
Institutions: Heart Research Center Goettingen, University Medical Center Goettingen, German Center for Cardiovascular Research (DZHK) partner site Goettingen, University of Maryland School of Medicine.
In cardiac myocytes a complex network of membrane tubules - the transverse-axial tubule system (TATS) - controls deep intracellular signaling functions. While the outer surface membrane and associated TATS membrane components appear to be continuous, there are substantial differences in lipid and protein content. In ventricular myocytes (VMs), certain TATS components are highly abundant contributing to rectilinear tubule networks and regular branching 3D architectures. It is thought that peripheral TATS components propagate action potentials from the cell surface to thousands of remote intracellular sarcoendoplasmic reticulum (SER) membrane contact domains, thereby activating intracellular Ca2+ release units (CRUs). In contrast to VMs, the organization and functional role of TATS membranes in atrial myocytes (AMs) is significantly different and much less understood. Taken together, quantitative structural characterization of TATS membrane networks in healthy and diseased myocytes is an essential prerequisite towards better understanding of functional plasticity and pathophysiological reorganization. Here, we present a strategic combination of protocols for direct quantitative analysis of TATS membrane networks in living VMs and AMs. For this, we accompany primary cell isolations of mouse VMs and/or AMs with critical quality control steps and direct membrane staining protocols for fluorescence imaging of TATS membranes. Using an optimized workflow for confocal or superresolution TATS image processing, binarized and skeletonized data are generated for quantitative analysis of the TATS network and its components. Unlike previously published indirect regional aggregate image analysis strategies, our protocols enable direct characterization of specific components and derive complex physiological properties of TATS membrane networks in living myocytes with high throughput and open access software tools. In summary, the combined protocol strategy can be readily applied for quantitative TATS network studies during physiological myocyte adaptation or disease changes, comparison of different cardiac or skeletal muscle cell types, phenotyping of transgenic models, and pharmacological or therapeutic interventions.
Bioengineering, Issue 92, cardiac myocyte, atria, ventricle, heart, primary cell isolation, fluorescence microscopy, membrane tubule, transverse-axial tubule system, image analysis, image processing, T-tubule, collagenase
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Coordinate Mapping of Hyolaryngeal Mechanics in Swallowing
Authors: Thomas Z. Thompson, Farres Obeidin, Alisa A. Davidoff, Cody L. Hightower, Christohper Z. Johnson, Sonya L. Rice, Rebecca-Lyn Sokolove, Brandon K. Taylor, John M. Tuck, William G. Pearson, Jr..
Institutions: Georgia Regents University, New York University, Georgia Regents University, Georgia Regents University.
Characterizing hyolaryngeal movement is important to dysphagia research. Prior methods require multiple measurements to obtain one kinematic measurement whereas coordinate mapping of hyolaryngeal mechanics using Modified Barium Swallow (MBS) uses one set of coordinates to calculate multiple variables of interest. For demonstration purposes, ten kinematic measurements were generated from one set of coordinates to determine differences in swallowing two different bolus types. Calculations of hyoid excursion against the vertebrae and mandible are correlated to determine the importance of axes of reference. To demonstrate coordinate mapping methodology, 40 MBS studies were randomly selected from a dataset of healthy normal subjects with no known swallowing impairment. A 5 ml thin-liquid bolus and a 5 ml pudding swallows were measured from each subject. Nine coordinates, mapping the cranial base, mandible, vertebrae and elements of the hyolaryngeal complex, were recorded at the frames of minimum and maximum hyolaryngeal excursion. Coordinates were mathematically converted into ten variables of hyolaryngeal mechanics. Inter-rater reliability was evaluated by Intraclass correlation coefficients (ICC). Two-tailed t-tests were used to evaluate differences in kinematics by bolus viscosity. Hyoid excursion measurements against different axes of reference were correlated. Inter-rater reliability among six raters for the 18 coordinates ranged from ICC = 0.90 - 0.97. A slate of ten kinematic measurements was compared by subject between the six raters. One outlier was rejected, and the mean of the remaining reliability scores was ICC = 0.91, 0.84 - 0.96, 95% CI. Two-tailed t-tests with Bonferroni corrections comparing ten kinematic variables (5 ml thin-liquid vs. 5 ml pudding swallows) showed statistically significant differences in hyoid excursion, superior laryngeal movement, and pharyngeal shortening (p < 0.005). Pearson correlations of hyoid excursion measurements from two different axes of reference were: r = 0.62, r2 = 0.38, (thin-liquid); r = 0.52, r2 = 0.27, (pudding). Obtaining landmark coordinates is a reliable method to generate multiple kinematic variables from video fluoroscopic images useful in dysphagia research.
Medicine, Issue 87, videofluoroscopy, modified barium swallow studies, hyolaryngeal kinematics, deglutition, dysphagia, dysphagia research, hyolaryngeal complex
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A Proboscis Extension Response Protocol for Investigating Behavioral Plasticity in Insects: Application to Basic, Biomedical, and Agricultural Research
Authors: Brian H. Smith, Christina M. Burden.
Institutions: Arizona State University.
Insects modify their responses to stimuli through experience of associating those stimuli with events important for survival (e.g., food, mates, threats). There are several behavioral mechanisms through which an insect learns salient associations and relates them to these events. It is important to understand this behavioral plasticity for programs aimed toward assisting insects that are beneficial for agriculture. This understanding can also be used for discovering solutions to biomedical and agricultural problems created by insects that act as disease vectors and pests. The Proboscis Extension Response (PER) conditioning protocol was developed for honey bees (Apis mellifera) over 50 years ago to study how they perceive and learn about floral odors, which signal the nectar and pollen resources a colony needs for survival. The PER procedure provides a robust and easy-to-employ framework for studying several different ecologically relevant mechanisms of behavioral plasticity. It is easily adaptable for use with several other insect species and other behavioral reflexes. These protocols can be readily employed in conjunction with various means for monitoring neural activity in the CNS via electrophysiology or bioimaging, or for manipulating targeted neuromodulatory pathways. It is a robust assay for rapidly detecting sub-lethal effects on behavior caused by environmental stressors, toxins or pesticides. We show how the PER protocol is straightforward to implement using two procedures. One is suitable as a laboratory exercise for students or for quick assays of the effect of an experimental treatment. The other provides more thorough control of variables, which is important for studies of behavioral conditioning. We show how several measures for the behavioral response ranging from binary yes/no to more continuous variable like latency and duration of proboscis extension can be used to test hypotheses. And, we discuss some pitfalls that researchers commonly encounter when they use the procedure for the first time.
Neuroscience, Issue 91, PER, conditioning, honey bee, olfaction, olfactory processing, learning, memory, toxin assay
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Using an Automated 3D-tracking System to Record Individual and Shoals of Adult Zebrafish
Authors: Hans Maaswinkel, Liqun Zhu, Wei Weng.
Institutions: xyZfish.
Like many aquatic animals, zebrafish (Danio rerio) moves in a 3D space. It is thus preferable to use a 3D recording system to study its behavior. The presented automatic video tracking system accomplishes this by using a mirror system and a calibration procedure that corrects for the considerable error introduced by the transition of light from water to air. With this system it is possible to record both single and groups of adult zebrafish. Before use, the system has to be calibrated. The system consists of three modules: Recording, Path Reconstruction, and Data Processing. The step-by-step protocols for calibration and using the three modules are presented. Depending on the experimental setup, the system can be used for testing neophobia, white aversion, social cohesion, motor impairments, novel object exploration etc. It is especially promising as a first-step tool to study the effects of drugs or mutations on basic behavioral patterns. The system provides information about vertical and horizontal distribution of the zebrafish, about the xyz-components of kinematic parameters (such as locomotion, velocity, acceleration, and turning angle) and it provides the data necessary to calculate parameters for social cohesions when testing shoals.
Behavior, Issue 82, neuroscience, Zebrafish, Danio rerio, anxiety, Shoaling, Pharmacology, 3D-tracking, MK801
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Getting to Compliance in Forced Exercise in Rodents: A Critical Standard to Evaluate Exercise Impact in Aging-related Disorders and Disease
Authors: Jennifer C. Arnold, Michael F. Salvatore.
Institutions: Louisiana State University Health Sciences Center.
There is a major increase in the awareness of the positive impact of exercise on improving several disease states with neurobiological basis; these include improving cognitive function and physical performance. As a result, there is an increase in the number of animal studies employing exercise. It is argued that one intrinsic value of forced exercise is that the investigator has control over the factors that can influence the impact of exercise on behavioral outcomes, notably exercise frequency, duration, and intensity of the exercise regimen. However, compliance in forced exercise regimens may be an issue, particularly if potential confounds of employing foot-shock are to be avoided. It is also important to consider that since most cognitive and locomotor impairments strike in the aged individual, determining impact of exercise on these impairments should consider using aged rodents with a highest possible level of compliance to ensure minimal need for test subjects. Here, the pertinent steps and considerations necessary to achieve nearly 100% compliance to treadmill exercise in an aged rodent model will be presented and discussed. Notwithstanding the particular exercise regimen being employed by the investigator, our protocol should be of use to investigators that are particularly interested in the potential impact of forced exercise on aging-related impairments, including aging-related Parkinsonism and Parkinson’s disease.
Behavior, Issue 90, Exercise, locomotor, Parkinson’s disease, aging, treadmill, bradykinesia, Parkinsonism
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Electrophysiological Measurements and Analysis of Nociception in Human Infants
Authors: L. Fabrizi, A. Worley, D. Patten, S. Holdridge, L. Cornelissen, J. Meek, S. Boyd, R. Slater.
Institutions: University College London, Great Ormond Street Hospital, University College Hospital, University of Oxford.
Pain is an unpleasant sensory and emotional experience. Since infants cannot verbally report their experiences, current methods of pain assessment are based on behavioural and physiological body reactions, such as crying, body movements or changes in facial expression. While these measures demonstrate that infants mount a response following noxious stimulation, they are limited: they are based on activation of subcortical somatic and autonomic motor pathways that may not be reliably linked to central sensory processing in the brain. Knowledge of how the central nervous system responds to noxious events could provide an insight to how nociceptive information and pain is processed in newborns. The heel lancing procedure used to extract blood from hospitalised infants offers a unique opportunity to study pain in infancy. In this video we describe how electroencephalography (EEG) and electromyography (EMG) time-locked to this procedure can be used to investigate nociceptive activity in the brain and spinal cord. This integrative approach to the measurement of infant pain has the potential to pave the way for an effective and sensitive clinical measurement tool.
Neuroscience, Issue 58, pain, infant, electrophysiology, human development
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Brain Imaging Investigation of the Neural Correlates of Observing Virtual Social Interactions
Authors: Keen Sung, Sanda Dolcos, Sophie Flor-Henry, Crystal Zhou, Claudia Gasior, Jennifer Argo, Florin Dolcos.
Institutions: University of Alberta, University of Illinois, University of Alberta, University of Alberta, University of Alberta, University of Illinois at Urbana-Champaign, University of Illinois at Urbana-Champaign.
The ability to gauge social interactions is crucial in the assessment of others’ intentions. Factors such as facial expressions and body language affect our decisions in personal and professional life alike 1. These "friend or foe" judgements are often based on first impressions, which in turn may affect our decisions to "approach or avoid". Previous studies investigating the neural correlates of social cognition tended to use static facial stimuli 2. Here, we illustrate an experimental design in which whole-body animated characters were used in conjunction with functional magnetic resonance imaging (fMRI) recordings. Fifteen participants were presented with short movie-clips of guest-host interactions in a business setting, while fMRI data were recorded; at the end of each movie, participants also provided ratings of the host behaviour. This design mimics more closely real-life situations, and hence may contribute to better understanding of the neural mechanisms of social interactions in healthy behaviour, and to gaining insight into possible causes of deficits in social behaviour in such clinical conditions as social anxiety and autism 3.
Neuroscience, Issue 53, Social Perception, Social Knowledge, Social Cognition Network, Non-Verbal Communication, Decision-Making, Event-Related fMRI
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Brain Imaging Investigation of the Neural Correlates of Emotion Regulation
Authors: Sanda Dolcos, Keen Sung, Ekaterina Denkova, Roger A. Dixon, Florin Dolcos.
Institutions: University of Illinois, Urbana-Champaign, University of Alberta, Edmonton, University of Alberta, Edmonton, University of Alberta, Edmonton, University of Alberta, Edmonton, University of Illinois, Urbana-Champaign, University of Illinois, Urbana-Champaign.
The ability to control/regulate emotions is an important coping mechanism in the face of emotionally stressful situations. Although significant progress has been made in understanding conscious/deliberate emotion regulation (ER), less is known about non-conscious/automatic ER and the associated neural correlates. This is in part due to the problems inherent in the unitary concepts of automatic and conscious processing1. Here, we present a protocol that allows investigation of the neural correlates of both deliberate and automatic ER using functional magnetic resonance imaging (fMRI). This protocol allows new avenues of inquiry into various aspects of ER. For instance, the experimental design allows manipulation of the goal to regulate emotion (conscious vs. non-conscious), as well as the intensity of the emotional challenge (high vs. low). Moreover, it allows investigation of both immediate (emotion perception) and long-term effects (emotional memory) of ER strategies on emotion processing. Therefore, this protocol may contribute to better understanding of the neural mechanisms of emotion regulation in healthy behaviour, and to gaining insight into possible causes of deficits in depression and anxiety disorders in which emotion dysregulation is often among the core debilitating features.
Neuroscience, Issue 54, Emotion Suppression, Automatic Emotion Control, Deliberate Emotion Control, Goal Induction, Neuroimaging
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Basics of Multivariate Analysis in Neuroimaging Data
Authors: Christian Georg Habeck.
Institutions: Columbia University.
Multivariate analysis techniques for neuroimaging data have recently received increasing attention as they have many attractive features that cannot be easily realized by the more commonly used univariate, voxel-wise, techniques1,5,6,7,8,9. Multivariate approaches evaluate correlation/covariance of activation across brain regions, rather than proceeding on a voxel-by-voxel basis. Thus, their results can be more easily interpreted as a signature of neural networks. Univariate approaches, on the other hand, cannot directly address interregional correlation in the brain. Multivariate approaches can also result in greater statistical power when compared with univariate techniques, which are forced to employ very stringent corrections for voxel-wise multiple comparisons. Further, multivariate techniques also lend themselves much better to prospective application of results from the analysis of one dataset to entirely new datasets. Multivariate techniques are thus well placed to provide information about mean differences and correlations with behavior, similarly to univariate approaches, with potentially greater statistical power and better reproducibility checks. In contrast to these advantages is the high barrier of entry to the use of multivariate approaches, preventing more widespread application in the community. To the neuroscientist becoming familiar with multivariate analysis techniques, an initial survey of the field might present a bewildering variety of approaches that, although algorithmically similar, are presented with different emphases, typically by people with mathematics backgrounds. We believe that multivariate analysis techniques have sufficient potential to warrant better dissemination. Researchers should be able to employ them in an informed and accessible manner. The current article is an attempt at a didactic introduction of multivariate techniques for the novice. A conceptual introduction is followed with a very simple application to a diagnostic data set from the Alzheimer s Disease Neuroimaging Initiative (ADNI), clearly demonstrating the superior performance of the multivariate approach.
JoVE Neuroscience, Issue 41, fMRI, PET, multivariate analysis, cognitive neuroscience, clinical neuroscience
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The Successive Alleys Test of Anxiety in Mice and Rats
Authors: Robert M.J. Deacon.
Institutions: University of Oxford.
The plus-maze was derived from the early work of Montgomery. He observed that rats tended to avoid the open arms of a maze, preferring the enclosed ones. Handley, Mithani and File et al. performed the first studies on the plus-maze design we use today, and in 1987 Lister published a design for use with mice. Time spent on, and entries into, the open arms are an index of anxiety; the lower these indices, the more anxious the mouse is. Alternatively, a mouse that spends most of its time in the closed arms is classed as anxious. One of the problems of the plus-maze is that, while time spent on, and entries into, the open arms is a fairly unambiguous measure of anxiety, time in the central area is more difficult to interpret, although time spent here has been classified as “decision making”. In many tests central area time is a considerable part of the total test time. Shepherd et al. produced an ingenious design to eliminate the central area, which they called the “zero maze”. However, although used by several groups, it has never been as widely adopted as the plus-maze. In the present article I describe a modification of the plus-maze design that not only eliminates the central area but also incorporates elements from other anxiety tests, such as the light-dark box and emergence tests. It is a linear series of four alleys, each having increasing anxiogenic properties. It has given similar results to the plus-maze in general. Although it may not be more sensitive than the plus-maze (more data is needed before a firm conclusion can be reached on this point), it provides a useful confirmation of plus-maze results which would be useful when, for example, only a single example of a mutant mouse was available, as, for example, in ENU-based mutagenesis programs.
Behavior, Issue 76, Neuroscience, Neurobiology, Medicine, Psychology, Mice, rats, anxiety-like behaviour, plus-maze, behaviour, prefrontal cortex, hippocampus, medial septum, successive alleys, animal model
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What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.