Name: neonatal subventricular zone electroporation
Uploaded: 2013-02-11T13:00:00
Description: Watch this Scientific Journal Video about Neonatal Subventricular Zone Electroporation at JoVE.com

Dette arbeidet ble støttet med tilskudd fra Department of Defense (Idéutvikling award, W81XWH-10-1-0041, AB), CT Stamcelleforskningen stipend (AB), og en National Institute of Health NRSA 10668225 (DMF). Det foreliggende materiale er basert på arbeid delvis støttet av staten Connecticut under Connecticut Stem Cell Research Grants Program. Innholdet er utelukkende ansvaret av forfatterne og ikke nødvendigvis representerer den offisielle syn i staten Connecticut, Department of Public Health i delstaten Connecticut eller CT Innovations, hadde Inc. organer ingen rolle i studiedesign, datainnsamling og analyse, beslutning om å publisere, eller utarbeidelse av manuskriptet.

Denne fremgangsmåten er i samsvar med Yale IACUC krav. Forskere bør sørge for at IACUC retningslinjer er godkjent og fulgt i henhold til deres institusjonelle krav. 1. § 1. Utarbeidelse av DNA, Solutions, og Glass Pipettes <ol><li> Generere høy renhet (OD 260/280&gt; 1,80) høy konsentrasjon (ug / pl&gt; 2,5) endotoksin-fri DNA. </li><li> Forbered 0,9% saltløsning og sterilt filter gjennom et 22 um filter. </li><li> Sted 10 cm brann polert borosilicate glass kapillarrør (OD: 1,5 mm,...

<ol>
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Her har vi detalj teknikken neonatal SVZ electroporation, en teknikk for å raskt og robust merke og manipulere SVZ stamceller og deres avkom. Det er flere fordeler som elektroporering har i forhold til andre teknikker. Først, gitt fokal merking av celler, er man i stand til å skjelne celle selvstyrte og ikke-celle autonome effekter. Sekund, genetisk manipulasjon hjelp induserbare systemer tillater en å sammenligne effekter før eller etter synaptiske integrasjon. Videre kan man omgå bruken av flere plasmider ved el...

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neonatal subventricular zone electroporation

Nevrale stamceller (NSCs) linje postnatal sideventriklene og gi opphav til flere celletyper som inkluderer nevroner, astrocytes og ependymal celler en. Forstå de molekylære veier som er ansvarlige for NSC selvfornyelse, engasjement og differensiering er kritisk for å utnytte deres unike potensial til å reparere hjernen og bedre forstå sentrale nervesystemet lidelser. Tidligere metoder for manipulering av pattedyr systemer kreves det tidkrevende og dyrt bestrebelse av genteknologi ved hele dyret nivå 2. Dermed har det store flertallet av studier utforsket funksjonene NSC molekyler in vitro eller in virvelløse dyr. Her viser vi den enkle og raske teknikk for å manipulere neonatal NPCer som er referert til som neonatal subventricular sone (SVZ) electroporation. Lignende teknikker ble utviklet for et tiår siden for å studere embryonale NSCs og har hjulpet studier på cortical utvikling 3,4. Mer nylig denne ble brukt til å studere postnatal gnager forebrain 5-7. Denne teknikken resulterer i robust merking av SVZ NSCs og deres avkom. Dermed gir postnatal SVZ electroporation en kostnadseffektiv og tidsbesparende alternativ for pattedyr NSC genteknologi.

Neonatal subventricular sone electroporation resultater i merking av nesten alle radial gliaceller sammenhengende med dorsal, dorsal lateral, og lateral subventricular sone etter &quot;feiing&quot; bevegelse av pinsetten elektroden (Figur 1E). Imidlertid kan elektroporering tilpasses kravet av den respektive eksperimentet, men ikke feiende elektroden og med bestemte plassering og orientering som beskrevet i videoen. For eksempel, siden dorsally lokalisert radial gliaceller forskjellige fra de fôr den l...

Watch this Scientific Journal Video about Neonatal Subventricular Zone Electroporation at JoVE.com

Neonatal Subventricular Zone Electroporation

Vi viser en minimalt invasiv teknikk kalt neonatal subventricular sone electroporation. Teknikken består i innsprøyting plasmid DNA inn i laterale ventrikkel av neonatale valper og tilføre elektrisk strøm for å levere og genetisk manipulere nevrale stamceller

Neonatal subventricular Zone Electroporation

Neural stem cells (NSCs) line the postnatal lateral ventricles and give rise to multiple cell types which include neurons, astrocytes, and ependymal ...

Research

JoVE Journal

Neuroscience

The Subventricular Zone En-face: Wholemount Staining and Ependymal Flow

The walls of the lateral ventricles contain the largest germinal region in the adult mammalian brain. The subventricular zone (SVZ) in these walls is an ...

In vivo Electroporation of Developing Mouse Retina

In vivo Electroporation of Developing Mouse Retina

The functional characterization of genes expressed during mammalian retinal development remains a significant challenge.  Gene targeting to generate ...

Mouse in Utero Electroporation: Controlled Spatiotemporal Gene Transfection

Mouse in Utero Electroporation: Controlled Spatiotemporal Gene Transfection

In order to understand the function of genes expressed in specific region of the developing brain, including signaling molecules and axon guidance ...

Preparation of Acute Subventricular Zone Slices for Calcium Imaging

Utarbeidelse av Akutt subventricular sone Slices for kalsium Imaging

The subventricular zone (SVZ) is one of the two neurogenic zones in the postnatal brain. The SVZ contains densely packed cells, including neural ...

Neonatal Pial Surface Electroporation

Neonatal PIAL Surface Electroporation

Over the past several years the pial surface has been identified as a germinal niche of importance during embryonic, perinatal and adult neuro- and ...

Subtype-selective Electroporation of Cortical Interneurons

Subtype-selektiv Electroporation av Cortical interneuroner

The study of central nervous system (CNS) maturation relies on genetic targeting of neuronal populations. However, the task of restricting the expression ...

Cell Sorting of Neural Stem and Progenitor Cells from the Adult Mouse Subventricular Zone and Live-imaging of their Cell Cycle Dynamics

Cell Sortering av Neural stilk og stamceller fra Adult Mouse subventricular Zone og Live-avbildning av deres Cell Cycle Dynamics

Neural stem cells (NSCs) in the subventricular zone of the lateral ventricles (SVZ) sustain olfactory neurogenesis throughout life in the mammalian brain. ...

Isolation and Culture of Adult Neural Stem Cells from the Mouse Subcallosal Zone

Isolasjon og Culture of Adult Neural stamceller fra mus Subcallosal Zone

Adult neural stem cells (aNSCs) can be used for the regeneration of damaged brain tissue. NSCs have the potential for differentiation and proliferation ...

Spinal Cord Neurons Isolation and Culture from Neonatal Mice

Spinal Cord Neurons Isolasjon og kultur fra neonatale mus

We present a protocol for the isolation and culture of spinal cord neurons. The neurons are obtained from neonatal C57BL/6 mice and are isolated on ...

Continuous Video Electroencephalogram during Hypoxia-Ischemia in Neonatal Mice

Kontinuerlig videoelektroencefalogram under Hypoksi-iskemi hos neonatale mus

Hypoxia ischemia is the most common cause of neonatal seizures. Animal models are crucial for understanding the mechanisms and physiology underlying ...