Name: revealing neural circuit topography in multi-color
Uploaded: 2011-11-14T12:00:00
Description: Watch this Scientific Journal Video about Revealing Neural Circuit Topography in Multi-Color at JoVE.com

Dette arbeidet ble støttet av ny etterforsker oppstart midler fra Albert Einstein College of Medicine ved Yeshiva University for å RVS.

1. Levere sporstoffer in vivo ved hjelp av sterile kirurgi (§ § 1.1 til 1.16) Gjennom hele prosedyren anbefaler vi at standard sterile kirurgiske teknikker brukes. Dette inkluderer bruk av sterile hansker, kappe, og ansiktsmaske. Verktøy skal enten autoklaveres før bruk eller rengjøres grundig med vann og etanol. Under operasjonen, og spesielt mellom dyr, rene vev fragmenter av verktøy og tørre sterilisere instrumenter med en varm perle sterilisator (Steri 250 Cat. # 18000-45, ...

<ol>
	<li>Apps, R., Hawkes, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cerebellar+cortical+organization:+a+one-map+hypothesis.">Cerebellar cortical organization: a one-map hypothesis.</a> Nat. Rev. Neurosci. 10, 670-681 (2009).</li><li>Grishkat, H. L., Eisenman, L. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+the+spinocerebellar+projection+in+the+prenatal+mouse.">Development of the spinocerebellar projection in the prenatal mouse.</a> J. Comp. Neurol. 363, 93-108 (1995).</li><li>Mesulam, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Tracing neural connections with horseradish peroxidase. , (1982).</li><li>Reeber, S. L., Sillitoe, R. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Patterned+expression+of+a+cocaine-+and+amphetamine+regulated+transcript+(CART)+peptide+reveals+complex+circuit+topography+in+the+rodent+cerebellar+cortex.">Patterned expression of a cocaine- and amphetamine regulated transcript (CART) peptide reveals complex circuit topography in the rodent cerebellar cortex.</a> Journal of Comparative Neurology. , (2011).</li><li>Reeber, S. L., Gebre, S. A., Sillitoe, R. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fluorescence+mapping+of+afferent+topography+in+three+dimensions.">Fluorescence mapping of afferent topography in three dimensions.</a> Brain. Struct. Funct. , (2011).</li><li>Sillitoe, R. V., Stephen, D., Lao, Z., Joyner, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Engrailed+homeobox+genes+determine+the+organization+of+Purkinje+cell+sagittal+stripe+gene+expression+in+the+adult+cerebellum.">Engrailed homeobox genes determine the organization of Purkinje cell sagittal stripe gene expression in the adult cerebellum.</a> J. Neurosci. 28, 12150-12162 (2008).</li><li>Sillitoe, R. V., Vogel, M. W., Joyner, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Engrailed+homeobox+genes+regulate+establishment+of+the+cerebellar+afferent+circuit+map.">Engrailed homeobox genes regulate establishment of the cerebellar afferent circuit map.</a> J. Neurosci. 30, 10015-10024 (2010).</li><li>Vig, J., Goldowitz, D., Steindler, D. A., Eisenman, L. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Compartmentation+of+the+reeler+cerebellum:+segregation+and+overlap+of+spinocerebellar+and+secondary+vestibulocerebellar+fibers+and+their+target+cells.">Compartmentation of the reeler cerebellum: segregation and overlap of spinocerebellar and secondary vestibulocerebellar fibers and their target cells.</a> Neuroscience. 130, 735-744 (2005).</li><li>Vogel, M. W., Prittie, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Topographic+spinocerebellar+mossy+fiber+projections+are+maintained+in+the+lurcher+mutant.">Topographic spinocerebellar mossy fiber projections are maintained in the lurcher mutant.</a> J. Comp. Neurol. 343, 341-351 (1994).</li><li>Voogd, J., Broere, G., van Rossum, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+medio-lateral+distribution+of+the+spinocerebellar+projection+in+the+anterior+lobe+and+the+simple+lobule+in+the+cat+and+a+comparison+with+some+other+afferent+fibre+systems.">The medio-lateral distribution of the spinocerebellar projection in the anterior lobe and the simple lobule in the cat and a comparison with some other afferent fibre systems.</a> Psychiatr. Neurol. Neurochir. 72, 137-151 (1969).</li><li>Wu, H. S., Sugihara, I., Shinoda, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Projection+patterns+of+single+mossy+fibers+originating+from+the+lateral+reticular+nucleus+in+the+rat+cerebellar+cortex+and+nuclei.">Projection patterns of single mossy fibers originating from the lateral reticular nucleus in the rat cerebellar cortex and nuclei.</a> J. Comp. Neurol. 411, 97-118 (1999).</li></ol>

Vi har beskrevet det kirurgiske og tekniske detaljer som kreves for vellykket aksonal og dendrite sporing ved hjelp av en roman fluorescerende tilnærming for rask merking nevrale anslagene i utvikling og voksne mus. Ved hjelp av WGA-Alexa viser vi hvordan sporstoffer og markører kan brukes for å analysere mønstrede krets topografien i høy oppløsning og i tre dimensjoner. Mange sporstoff tilgjengelig for sporing nevrale kretser inkludert koleratoksin subenhet B, Biocytin, Neurobiotin, P...

<table border="1"><tbody><tr><td> Utstyr / reagenser </td><td> Model / Katalognummer </td><td> Firma </td></tr><tr><td> Bead sterilizer </td><td> Model Steri 250 Kat. # 18000-45 </td><td> Fin Science Verktøy </td></tr><tr><td> Cauterizer </td><td> Cat. # 18000-00 </td><td> Fin Science Verktøy </td></tr><tr><td> Borsilikatglass kapillærer </td><td> Cat. # 300056 </td><td> Harvard Apparatus </td></tr><tr><td> Totrinns Glass mikropipette Puller </td><td> Modell 001-PC-10 </td><td> Narishige </td></tr><tr><td> Mikrometer sprøyte </td><td> Cat. # GS-1100 </td><td> Gilmont Instrumenter </td></tr><tr><td> Small Animal Stereotaxic Instrument </td><td> Modell 940-A </td><td> Kopf Instrumentation </td></tr><tr><td> Elektrode manipulator </td><td> Model 960 </td><td> Kopf Instrumentation </td></tr><tr><td> Vetcare kammer </td><td> Cat. #340508 </td><td> Harvard Apparatus </td></tr><tr><td> Oppvarming Pad </td><td> Cat. # 341241 </td><td> Harvard Apparatus </td></tr><tr><td> Leica DFC360 FX-kamera </td><td> DFC360 FX </td><td> Leica </td></tr><tr><td> Leica DFC490 kamera </td><td> DFC490 </td><td> Leica </td></tr><tr><td> Leica DM5500 mikroskop </td><td> DM5500 </td><td> Leica </td></tr><tr><td> Leica DFC3000 FX-kamera </td><td> DFC3000 FX </td><td> Leica </td></tr><tr><td> Leica MZ16 FA mikroskop </td><td> MZ16 FA </td><td> Leica </td></tr><tr><td> CY3 Filter </td><td> Model # 11600231 </td><td> Leica </td></tr><tr><td> FITC Filter </td><td> Model # 11513880 </td><td> Leica </td></tr><tr><td> A4 DAPI / UV filter </td><td> Model # 11504162 </td><td> Leica </td></tr><tr><td> Hvetespirer agglutinin, Alexa Fluor 488 konjugat </td><td> Cat. # W11261 </td><td> Invitrogen </td></tr><tr><td> Hvete germ agglutinin, Alexa Fluor 555 konjugat </td><td> Cat. # W32464 </td><td> Invitrogen </td></tr></tbody></table>

revealing neural circuit topography in multi-color

Nevrale kretsene er organisert i funksjonelle topografiske kart. For å visualisere komplekse krets arkitekturen vi utviklet en tilnærming til pålitelig merke den globale fordelingen av flere topografiske projeksjoner. Lillehjernen er en ideell modell for å studere ordnet arrangement av nevrale kretser. For eksempel har compartment organiseringen av spinocerebellar mosegrodde fibre vist seg å være en uunnværlig system for å studere mosegrodde fiber mønster. Vi har nylig viste at hvetespirer agglutinin (WGA) konjugert til Alexa 555 og 488 kan brukes til å spore spinocerebellar mosegrodde fiber anslagene i å utvikle og voksne mus (Reeber et al. 2011). Vi fant tre store egenskaper som gjør at WGA-Alexa tracere ønskelig verktøy for merking nevrale projeksjoner. Først Alexa fluorophores er intense og deres lysstyrke tillater wholemount bildebehandling rett etter tracing. Sekund, label WGA-Alexa tracers hele banen for å utvikle og voksen nevrale projections. Tredje, WGA-Alexa sporstoff raskt transporteres i både retrograd og anterograd retninger. Her beskriver vi i detalj hvordan man skal forberede sporingsstoffene og andre nødvendige verktøy, hvordan å utføre kirurgi for spinocerebellar sporing og hvordan man best bilde spores anslagene i tre dimensjoner. Oppsummert kan vi tilby en steg-for-steg tracing-protokollen som vil være nyttig for å tyde organisering og tilkobling av funksjonelle kart ikke bare i lillehjernen, men også i cortex, hjernestammen og ryggmargen.

Watch this Scientific Journal Video about Revealing Neural Circuit Topography in Multi-Color at JoVE.com

Revealing Neural Circuit Topography in Multi-Color

Vi tilbyr en praktisk guide for å levere sporstoffer<em&gt; In vivo</em&gt; Og bruke spinocerebellar veien som et modellsystem for å demonstrere viktige skritt for å lykkes neuronal kretsanalyse hos mus. Vi beskriver i detalj vår allsidige tracing protokoll som utnytter hvetespirer agglutinin (WGA) konjugert til Alexa fluorophores.

Avslører Neural Circuit Topografi i Multi-Color

Neural circuits are organized into functional topographic maps. In order to visualize complex circuit architecture we developed an approach to reliably ...

Research

JoVE Journal

Neuroscience

Dissection and Culture of Commissural Neurons from Embryonic Spinal Cord

Commissural neurons have been widely used to investigate the mechanisms underlying axon guidance during embryonic spinal cord development. The cell bodies ...

Combining Behavioral Endocrinology and Experimental Economics: Testosterone and Social Decision Making

Behavioral endocrinological research in humans as well as in animals suggests that testosterone plays a key role in social interactions. Studies in ...

Time-lapse Live Imaging of Clonally Related Neural Progenitor Cells in the Developing Zebrafish Forebrain

Precise patterns of division, migration and differentiation of neural progenitor cells are crucial for proper brain development and function1,2. To ...

Detection of Microregional Hypoxia in Mouse Cerebral Cortex by Two-photon Imaging of Endogenous NADH Fluorescence

Påvisning av Microregional Hypoksi i Mus Cerebral Cortex av to-foton Imaging av endogen NADH Fluorescence

The brain's ability to function at high levels of metabolic demand depends on continuous oxygen supply through blood flow and tissue oxygen diffusion. ...

Wholemount Immunohistochemistry for Revealing Complex Brain Topography

Wholemount Immunhistokjemi for å avdekke Complex Brain Topografi

The repeated and well-understood cellular architecture of the cerebellum make it an ideal model system for exploring brain topography. Underlying its ...

Design and Assembly of an Ultra-light Motorized Microdrive for Chronic Neural Recordings in Small Animals

The ability to chronically record from populations of neurons in freely behaving animals has proven an invaluable tool for dissecting the function of ...

Multi-unit Recording Methods to Characterize Neural Activity in the Locust (Schistocerca Americana) Olfactory Circuits

Multi-unit Recording Methods to Characterize Neural Activity in the Locust Schistocerca Americana Olfactory Circuits

Multi-enheten opptaket Metoder for å karakterisere nevrale aktiviteten i Locust (<em&gt; Schistocerca Americana</em&gt;) Olfactory kretser

Detection and interpretation of olfactory cues are critical for the survival  of many organisms. Remarkably, species across phyla have strikingly similar ...

Neural Circuit Recording from an Intact Cockroach Nervous System

Neural Circuit Opptak fra et intakt Cockroach Nervous System

The cockroach ventral nerve cord preparation is a tractable system for neuroethology experiments, neural network modeling, and testing the physiological ...

Functional Analysis of the Larval Feeding Circuit in Drosophila

Functional Analysis of the Larval Feeding Circuit in Drosophila

Funksjonell analyse av larve Fôring Circuit i Drosophila</i

The serotonergic  feeding circuit in Drosophila  melanogaster larvae can be used to investigate neuronal substrates of  critical importance during the ...

Time-dependent Increase in the Network Response to the Stimulation of Neuronal Cell Cultures on Micro-electrode Arrays

Tidsavhengig økning i nettverksresponsen til stimulering av neuronale cellekulturer på mikroelektrodearrayer

Micro-electrode arrays (MEAs) can be used to investigate drug toxicity, design paradigms for next-generation personalized medicine, and study network ...