Micro-dissection of Rat Brain for RNA or Protein Extraction from Specific Brain Region

Published 8/30/2007
9 Comments
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Biology

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Summary

Micro-dissection of rat brain into various regions is extremely important for the study of different neurodegenerative diseases. This video demonstrates micro-dissection of four major brain regions include olfactory bulb, frontal cortex, striatum and hippocampus in fresh rat brain tissue. Useful tips for quick removal of respective regions to avoid RNA and protein degradation of the tissue are given.

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Chiu, K., Lau, W. M., Lau, H. T., So, K., Chang, R. C. Micro-dissection of Rat Brain for RNA or Protein Extraction from Specific Brain Region. J. Vis. Exp. (7), e269, doi:10.3791/269 (2007).

Abstract

Micro-dissection of rat brain into various regions is extremely important for the study of different neurodegenerative diseases. This video demonstrates micro-dissection of four major brain regions include olfactory bulb, frontal cortex, striatum and hippocampus in fresh rat brain tissue. Useful tips for quick removal of respective regions to avoid RNA and protein degradation of the tissue are given.

Protocol

Step 1

Sacrifice the rat according to established protocols. Remove the brain from the skull and rinse it in ice cold DEPC treated Milli Q water to remove any surface blood.

Step 2

Place on cold metal plate. Cut the brain bi-half into right and left hemisphere.

269_Figure-01

Figure 1.  The cutting positions from the medial view of rat hemisphere
 

Step 3

Collect the olfactory bulb right after the first cut. Flash freeze the specimen in liquid nitrogen and store at -80°C.

269_Figure-02

Figure 2. Olfactory bulb dissection

Step 4

Collect the frontal cortex right after the second cut, using a blade and a Dumont No. 5 forceps. Flash freeze the specimen in liquid nitrogen and store at -80°C.

269_Figure-03

Figure 3. Frontal cortex dissection

Step 5

Collect the striatum (caudate nucleus) right after the third cut with the help of two Dumont No.5 forceps. Flash freeze the specimen in liquid nitrogen and store at -80°C.

269_Figure-04

Figure 4. Striatum (caudate nucleus) dissection

Step 6

To collect the hippocampus, put the ventral side of the brain up and then remove midbrain to expose the hippocampus. Dissect the hippocampus from the cortex using two Dumont No.5 forceps. Flash freeze the specimen in liquid nitrogen and store at -80°C.

269_Figure-05

Figure 5. Hippocampus dissection

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Discussion

Different brain regions associate with different neurodegenerative diseases. For example, frontal cortex and striatum relate with Parkinson s disease while hippocampus relates with Alzheimer's disease. Precise micro-dissection of the brain allows us to detect the mRNA and protein changes in diseased region more accurately.

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Disclosures

The animals were handled according to the protocol for the use of animal in research approved by Committee on the Use of Live Animals in Teaching and Research (CULATR) in the University of Hong Kong and according to the National Institutes of Health guidelines for animal use.

Materials

Name Company Catalog Number Comments
Microscope Olympus Corporation
Dissection tools Metal plate, forceps, blade, etc. Individually wrapped with aluminum foil and heated at 180oC for eight hours to eliminate RNase. Made ice cold before use.

DOWNLOAD MATERIALS LIST

References

  1. Palkovits, M. Maps and guide to micro-dissection of the rat brain. Elsevier. New York. (c.1988).
  2. Swanson, L. W. Brain maps: structure of the rat brain. 3rd revised edition. Elsevier Academic Press. (2004).

Comments

9 Comments

  1. thank u very much. good protocol

    Reply
    Posted by: Anonymous
    January 8, 2009 - 4:37 AM
  2. very nice and great

    Reply
    Posted by: Anonymous
    February 11, 2009 - 1:27 PM
  3. Approximately how much time/rat from sacrifice to remove the frontal cortex and the hippocampus?  Thank you.

    Reply
    Posted by: Anonymous
    February 16, 2009 - 10:59 AM
  4. Hi, can be done within 3 minutes. When you practice more, ² minutes will be enough.

    Reply
    Posted by: kin c.
    March 28, 2012 - 9:28 PM
  5. Great, Thanks for your sharing̀²; maybe it's better to have more brain regions to dissect  

    Reply
    Posted by: Anonymous
    April 1, 2009 - 8:47 AM
  6. How dŒs this protocol change if we were to attempting to dissect the striatum of embryonic rats or mice for culture?

    Reply
    Posted by: Anonymous
    July 14, 2010 - 1:59 PM
  7. Yeah nicely done. Although I am using mice this seems to work just fine for me as well.

    Reply
    Posted by: Anonymous
    March 28, 2012 - 6:23 AM
  8. Thanks. I think this is just a demostration that we can dissect the brain into regions. When we deal with different parts, normally we go back to the atlas first (you can use the atlas listed in the reference). Then we try to identify the region in the brain samples.

    Reply
    Posted by: kin c.
    March 28, 2012 - 9:26 PM
  9. Thanks. If we want to isolate some specific nuclei from brain to study them under microscope how we must do that?

    Reply
    Posted by: Jamal E.
    May 8, 2012 - 2:10 PM

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