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Current Protocols in Neuroscience

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https://www.readbyqxmd.com/read/28398646/fluorescein-isothiocyanate-fitc-dextran-extravasation-as-a-measure-of-blood-brain-barrier-permeability
#1
Reka Natarajan, Nicole Northrop, Bryan Yamamoto
The blood-brain barrier (BBB) is formed in part by vascular endothelial cells that constitute the capillaries and microvessels of the brain. The function of this barrier is to maintain homeostasis within the brain microenvironment and buffer the brain from changes in the periphery. A dysfunction of the BBB would permit circulating molecules and pathogens typically restricted to the periphery to enter the brain and interfere with normal brain function. As increased permeability of the BBB is associated with several neuropathologies, it is important to have a reliable and sensitive method that determines BBB permeability and the degree of BBB disruption...
April 10, 2017: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/28398645/crispr-cas9-mediated-gene-knockout-in-the-mouse-brain-using-in-utero-electroporation
#2
Yohei Shinmyo, Hiroshi Kawasaki
This unit describes a highly efficient and rapid procedure for brain-specific disruption of genes in the developing mouse brain using pX330 plasmids expressing humanized Cas9 and single-guide RNAs (sgRNAs) against target genes. The pX330 plasmids are delivered into the rodent brain using in utero electroporation. Focusing on the Satb2 gene, which encodes an AT-rich DNA-binding transcription factor, we found that the introduction of pX330-Satb2 induced insertion/deletion (indel) mutations near the predicted cleavage site in the Satb2 gene, resulting in a dramatic reduction of Satb2 expression in post-mitotic neurons...
April 10, 2017: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/28398644/quantitative-high-throughput-screening-using-a-coincidence-reporter-biocircuit
#3
Brittany W Schuck, Ryan MacArthur, James Inglese
Reporter-biased artifacts-i.e., compounds that interact directly with the reporter enzyme used in a high-throughput screening (HTS) assay and not the biological process or pharmacology being interrogated-are now widely recognized to reduce the efficiency and quality of HTS used for chemical probe and therapeutic development. Furthermore, narrow or single-concentration HTS perpetuates false negatives during primary screening campaigns. Titration-based HTS, or quantitative HTS (qHTS), and coincidence reporter technology can be employed to reduce false negatives and false positives, respectively, thereby increasing the quality and efficiency of primary screening efforts, where the number of compounds investigated can range from tens of thousands to millions...
April 10, 2017: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/28398643/systems-genetic-analysis-in-genenetwork-org
#4
Clarissa C Parker, Price E Dickson, Vivek M Philip, Mary Thomas, Elissa J Chesler
Genome-wide association studies (GWAS) have emerged as a powerful tool to identify alleles and molecular pathways that influence susceptibility to psychiatric disorders and other diseases. Forward genetics using mouse mapping populations allows for a complementary approach that provides rigorous genetic and environmental control. In this unit, we describe techniques and tools that reduce the technical burden traditionally associated with genetic mapping in mice and enhance their translational utility to human psychiatric disorders...
April 10, 2017: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/28398642/wga-alexa-conjugates-for-axonal-tracing
#5
Sabrina L Levy, Joshua J White, Elizabeth P Lackey, Lindsey Schwartz, Roy V Sillitoe
Anatomical labeling approaches are essential for understanding brain organization. Among these approaches are various methods of performing tract tracing. However, a major hurdle to overcome when marking neurons in vivo is visibility. Poor visibility makes it challenging to image a desired neuronal pathway so that it can be easily differentiated from a closely neighboring pathway. As a result, it becomes impossible to analyze individual projections or their connections. The tracer that is chosen for a given purpose has a major influence on the quality of the tracing...
April 10, 2017: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/28398641/time-lapse-imaging-and-cell-tracking-of-migrating-cells-in-slices-and-flattened-telencephalic-vesicles
#6
Verónica Murcia-Belmonte, Giovanna Expósito, Eloísa Herrera
Neuronal migration is a vital process needed for subsequent assembly and function of neural circuitry during embryonic development. The vast majority of neural progenitors are generated far from their final destination and need to migrate considerable distances to reach their specific cortical layer. Innovations in cell culture techniques and fluorescence microscopy now facilitate the direct visualization of cell movements during cortical development. Here, a detailed protocol to record and analyze a particular type of early migrating neurons, the Cajal Retzius Cells, during the development of the telencephalic vesicles in mammals is described...
April 10, 2017: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/28398640/fluorescence-microscopy-a-concise-guide-to-current-imaging-methods
#7
Christian A Combs, Hari Shroff
The field of fluorescence microscopy is rapidly growing and offers ever more imaging capabilities for biologists. Over the past decade, many new technologies and techniques have been developed that allow for combinations of deeper, faster, and higher resolution imaging. These have included the commercialization of many super-resolution and light sheet fluorescence microscopy techniques. For the non-expert, it can be difficult to match the best imaging techniques to biological questions. Picking the most appropriate imaging modality requires a basic understanding of the underlying physics governing each of them, as well as information comparing potentially competing imaging properties in the context of the sample to be imaged...
April 10, 2017: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/28046204/electroporation-in-the-rodent-embryonic-brain-using-whole-embryo-culture-system
#8
Takako Kikkawa, Masanori Takahashi, Noriko Osumi
This unit describes basic methods for mammalian whole embryo culture (WEC) using embryonic day 10.5 mouse embryos, including the preparation of high-quality immediately centrifuged (IC) rat serum that is commonly used for WEC and is essential for normal growth and development of cultured mouse and rat embryos in vitro. An alternative protocol for different stages of rodent embryos is also introduced. Since embryos for WEC are dissected out of the uterus and manipulated under the microscope, one can overcome many of the difficulties of gene delivery encountered using in utero electroporation...
January 3, 2017: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/28046203/visualizing-changes-in-neuronal-dendritic-morphology-in-response-to-stress-and-pharmacological-challenge
#9
Cara L Wellman
This unit outlines a protocol for Golgi staining, which has been used extensively to reliably and quantitatively assess alterations in dendritic arborization and spine density as a result of a variety of factors, including chronic administration of glucocorticoids, chronic stress, and pharmacological manipulations. The method stains neurons in their entirety, allowing for sophisticated analyses of branch lengths and numbers as well as patterns of dendritic branching. Advantages of the technique include its usefulness in multisite collaborations and its utility in visualizing neurons in multiple regions within the same brain...
January 3, 2017: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/28046202/intracerebral-injections-and-ultrastructural-analysis-of-high-pressure-frozen-brain-tissue
#10
Marie-Theres Weil, Torben Ruhwedel, Wiebke Möbius, Mikael Simons
Intracerebral injections are an invasive method to bypass the blood brain barrier and are widely used to study molecular and cellular mechanisms of the central nervous system. The administered substances are injected directly at the site of interest, executing their effect locally. By combining injections in the rat brain with state-of-the-art electron microscopy, subtle changes in ultrastructure of the nervous tissue can be detected prior to overt damage or disease. The protocol presented here involves stereotactic injection into the corpus callosum of Lewis rats and the cryopreparation of freshly dissected tissue for electron microscopy...
January 3, 2017: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/28046201/measuring-cognitive-judgement-bias-in-rats-using-the-ambiguous-cue-interpretation-test
#11
Justyna Papciak, Rafal Rygula
An active-choice, operant, ambiguous-cue interpretation (ACI) paradigm is described that can be used for measuring cognitive judgement bias in rats. In this behavioral test, animals in an operant conditioning chamber are trained to press a lever to receive a food reward when a specific tone is presented, and to press another lever in response to a different tone to avoid punishment by an electric foot-shock. The tones, which serve as discriminative stimuli, acquire a positive or negative valence, and the training continues until the rats demonstrate a stable, correct discrimination between these two stimuli...
January 3, 2017: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/28046200/the-five-choice-continuous-performance-task-5c-cpt-a-cross-species-relevant-paradigm-for-assessment-of-vigilance-and-response-inhibition-in-rodents
#12
Zackary A Cope, Jared W Young
Deficits in the domains of attention and response inhibition are central to many psychiatric disorders. As such, animal models of disorders purporting to replicate these behavioral deficits first require tests that can accurately assess the behaviors with high fidelity. The gold-standard clinical test of attention and response inhibition is the continuous performance test (CPT). Although there are a number of CPTs, all share the premise of responding to target stimuli and inhibiting from responding to non-target stimuli...
January 3, 2017: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/27696363/cerebral-cortex-electroporation-to-study-projection-neuron-migration
#13
Emilie Pacary, François Guillemot
Brain electroporation is a rapid and powerful approach to study neuronal development. In particular, this technique has become a method of choice for studying the process of radial migration of projection neurons in the embryonic cerebral cortex. This method has considerably helped to describe in detail the different steps of radial migration and to characterize the molecular mechanisms controlling this process. Delineating the complexities of neuronal migration is critical to our understanding not only of normal cerebral cortex formation but also of neurodevelopmental disorders resulting from neuronal migration defects...
October 3, 2016: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/27696362/cigarette-smoke-extract-a-preclinical-model-of-tobacco-dependence
#14
Candice A Gellner, Daisy D Reynaga, Frances M Leslie
Animal models are used to study many human diseases, one of which is tobacco addiction. Most preclinical models use nicotine alone, although there are >7000 constituents present in tobacco smoke. The clinical literature suggests that cigarettes have a strong addictive potential, which is not paralleled in preclinical studies using nicotine alone. In order to address the gap between clinical and preclinical literature on tobacco dependence, cigarette smoke extracts containing tobacco constituents have been developed...
October 3, 2016: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/27696361/the-neonatal-ventral-hippocampal-lesion-nvhl-rodent-model-of-schizophrenia
#15
Anne Marie Brady
Animal models are crucial to the study of the neurobiological bases of psychiatric disorders, but schizophrenia is a particularly challenging disorder to model given the complexity and heavily verbal nature of its symptoms. This unit describes a developmental surgical rodent model of schizophrenia, the neonatal ventral hippocampal lesion (NVHL) model. This widely used model produces reliable behavioral abnormalities that are comparable to those observed in patients, as well as anatomical and neurophysiological disruptions in forebrain areas that are also implicated in schizophrenia...
October 3, 2016: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/27696360/automatic-dendritic-spine-quantification-from-confocal-data-with-neurolucida-360
#16
Dara L Dickstein, Daniel R Dickstein, William G M Janssen, Patrick R Hof, Jacob R Glaser, Alfredo Rodriguez, Nate O'Connor, Paul Angstman, Susan J Tappan
Determining the density and morphology of dendritic spines is of high biological significance given the role of spines in synaptic plasticity and in neurodegenerative and neuropsychiatric disorders. Precise quantification of spines in three dimensions (3D) is essential for understanding the structural determinants of normal and pathological neuronal function. However, this quantification has been restricted to time- and labor-intensive methods such as electron microscopy and manual counting, which have limited throughput and are impractical for studies of large samples...
October 3, 2016: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/27696359/pica-in-rats-as-a-preclinical-model-of-emesis
#17
T Gregg Davis
The ability to assess the potential for gastrointestinal adverse events in a preclinical setting is a challenge in the development of new drugs, as the vast majority of in vivo research is conducted in rodent species lacking a vomiting reflex. The use of higher species capable of emesis is often limited by cost, technical experience, and relevant efficacy models to define a therapeutic index. Additionally, investigators should be mindful of ethical considerations when using more sentient species when an alternative in lower species is available...
October 3, 2016: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/27696358/the-movement-tracker-a-flexible-system-for-automated-movement-analysis-in-invertebrate-model-organisms
#18
Laurent Mouchiroud, Vincenzo Sorrentino, Evan G Williams, Matteo Cornaglia, Michael V Frochaux, Tao Lin, Amandine A Nicolet-Dit-Félix, Gopal Krishnamani, Tarik Ouhmad, Martin A M Gijs, Bart Deplancke, Johan Auwerx
Phenotyping strategies in simple model organisms such as D. melanogaster and C. elegans are often broadly limited to growth, aging, and fitness. Recently, a number of physical setups and video tracking software suites have been developed to allow for accurate, quantitative, and high-throughput analysis of movement in flies and worms. However, many of these systems require precise experimental setups and/or fixed recording formats. We report here an update to the Parallel Worm Tracker software, which we termed the Movement Tracker...
October 3, 2016: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/27063788/the-revised-neurobehavioral-severity-scale-nss-r-for-rodents
#19
Angela M Yarnell, Erin S Barry, Andrea Mountney, Deborah Shear, Frank Tortella, Neil E Grunberg
Motor and sensory deficits are common following traumatic brain injury (TBI). Although rodent models provide valuable insight into the biological and functional outcomes of TBI, the success of translational research is critically dependent upon proper selection of sensitive, reliable, and reproducible assessments. Published literature includes various observational scales designed to evaluate post-injury functionality; however, the heterogeneity in TBI location, severity, and symptomology can complicate behavioral assessments...
April 8, 2016: Current Protocols in Neuroscience
https://www.readbyqxmd.com/read/27063787/studying-socio-affective-communication-in-rats-through-playback-of-ultrasonic-vocalizations
#20
Markus Wöhr, Dominik Seffer, Rainer K W Schwarting
Rats are able to produce ultrasonic vocalizations (USVs). Such USVs are an important component of the rat social behavior repertoire and serve distinct communicative functions as socio-affective signals. Depending on the emotional valence of the situation, juvenile and adult rats utter (1) aversive 22-kHz USVs conveying an appeasing and/or alarming function; or (2) appetitive 50-kHz USVs, which act as social contact calls, amongst others. A 50-kHz USV radial maze playback paradigm that allows assessment of the behavioral responses displayed by the recipients in a highly standardized manner has been developed...
April 8, 2016: Current Protocols in Neuroscience
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