The Use of Herpes Simplex Virus in Ex Vivo Slice Culture

Allyson K. Friedman1, Ming‐Hu Han1

1 Department of Pharmacology and Systems Therapeutics, Friedman Brain Institute, Icahn School of Medicine at Mount Sinai, New York, New York
Publication Name:  Current Protocols in Neuroscience
Unit Number:  Unit 4.36
DOI:  10.1002/0471142301.ns0436s72
Online Posting Date:  July, 2015
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Abstract

Herpes simplex virus (HSV) can be used for a wide range of genetic manipulations in ex vivo slices of central nervous system tissue from both young and adult rodents. The fast expression of the HSV viral‐mediated gene transfer, which can be engineered to produce cell‐type specificity, can be utilized in slice cultures for a variety of purposes over a 1‐ to 4‐day period with spatial and temporal specificity. This protocol exploits the rapid expression of HSV viral vectors by utilizing slice culture for electrophysiological recordings, avoiding the need to do intracranial viral injections. Brain slice cultures maintain many aspects of in vivo biology, including functional local synaptic circuitry with preserved brain architecture, while allowing good experimental access and precise control of the extracellular environment, making them ideal platforms for quick access to evaluate expression effects of HSV viral‐mediated gene transfer on the molecular and cellular properties of specific neurons. This protocol provides an easy way to study neuronal function following viral expression of a gene of interest. © 2015 by John Wiley & Sons, Inc.

Keywords: slice culture; HSV; gene transfer; neuronal property

     
 
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Table of Contents

  • Reagents and Solutions
  • Commentary
  • Literature Cited
     
 
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Materials

Basic Protocol 1:

  Materials
  • Brain slice culture medium (see recipe)
  • Artificial cerebrospinal fluid (ACSF; see recipe)
  • Carbogen tank (95% oxygen and 5% carbon dioxide)
  • 70% ethanol solution
  • Mice or rats for testing
  • Super glue (Loctite gel control)
  • Herpes simplex virus (HSV) viral vector containing gene construct of interest [see, e.g., unit 4.12 (Neve, 2012) and other relevant units in Chapter 4] and control virus
  • 0.22‐μm filter (Corning, cat. no. 430758)
  • Airstone
  • Slice recovery chamber (250‐ml beaker with a mesh membrane)
  • Razor blades
  • Iris scissors
  • Styrofoam brain slice platform (4‐in. × 4‐in. × 1‐in. platform for coarse cutting of brain area to be sliced)
  • 34°C water bath
  • 34°C incubator (5% CO 2, 100% humidity)
  • 6‐well plate (Corning, CellBIND 6‐Well Clear Multiple‐Well Plates, flat bottom, with lid, sterile; cat. no. 3335)
  • Millicell culture plate insert (0.4 μm; EMD Millipore, cat. no. PICM03050)
  • Surgical scissors
  • Paint brushes, size 1 and 2 (Dick Blick Wonder White, cat. no. 2026)
  • Spatula
  • Microslicer (Ted Pella, cat. no. DTK‐1000)
  • Dissection microscope (e.g., Leica, WILD M3C)
  • Standard electrophysiology setup
  • Additional reagents for isoflurane anesthesia of rodents ( appendix 4B; Davis, )
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Figures

Videos

Literature Cited

Literature Cited
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