Name: knowing what counts: unbiased stereology in the non-human primate brain
Uploaded: 2009-05-14T00:00:00
Description: Watch this Scientific Journal Video about Knowing What Counts: Unbiased Stereology in the Non-human Primate Brain at JoVE.com

The authors would like to thank Ikiel Ptito for his continued technical support. NSERC grant to MP.

Part 1: Pre-processing of tissue should be done according to Burke et al. (2009)5.
<ol>
<li>Briefly, tissue should be well 	perfused with paraformaldehyde, glutaraldehyde, or formalin. This 	can be achieved through standard transcardial perfusion typically 	used to harvest other organs. In the present study the subject was 	deeply sedated with ketamine hydrochloride (10 mg/kg, i.m.), 	euthanized with an overdose of sodium pentobarbital (25 mg/kg, i.v.) 	and perfused transcardially with 0.1 M PBS until comp...

<ol>
	<li>West, M., Slomianka, L., Gundersen, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Unbiased+stereological+estimation+of+the+total+number+of+neurons+in+the+subdivisions+of+the+rat+hippocampus+using+the+optical+fractionator.">Unbiased stereological estimation of the total number of neurons in the subdivisions of the rat hippocampus using the optical fractionator.</a> Aat Rec. 231, 482-497 (1991).</li><li>Saper, C. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Any+way+you+cut+it:+a+new+journal+policy+for+the+use+of+unbiased+counting+methods.">Any way you cut it: a new journal policy for the use of unbiased counting methods.</a> J Comp Neurol. 364, 5-5 (1996).</li><li>Sterio, D. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+unbiased+estimation+of+number+and+sizes+of+arbitrary+particles+using+the+disector.">The unbiased estimation of number and sizes of arbitrary particles using the disector.</a> J Microsc. 134, 127-136 (1984).</li><li>Mouton, P. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Principles and Practices of Unbiased Stereology. , (2002).</li><li>Burke, M. W., Zangenehpour, S., Boire, D., Ptito, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dissecting+the+non-human+primate+brain+in+stereotaxic+space.">Dissecting the non-human primate brain in stereotaxic space.</a> J Vis Exp. 29, (2009).</li><li>Burke, M. W., Zangenehpour, S., Boire, D., Ptito, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Brain+banking:+Making+the+most+of+your+research.">Brain banking: Making the most of your research.</a> J Vis Exp. 29, (2009).</li><li>Manaye, K. F., Wang, P., O'Neil, J., Huafu, S., Tizabi, Y., Thompson, N., Ottinger, M. A., Ingram, D. K., Mouton, P. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuropathological+Quantification+Of+Dtg+APP/PS1:+Neuroimaging,+Stereology,+And+Biochemistry.">Neuropathological Quantification Of Dtg APP/PS1: Neuroimaging, Stereology, And Biochemistry.</a> AGE. 29, 87-96 (2009).</li><li>Burke, M. W., Palmour, R. M., Ervin, F. R., Ptito, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuronal+reduction+in+frontal+cortex+of+primates+after+prenatal+alcohol+exposure.">Neuronal reduction in frontal cortex of primates after prenatal alcohol exposure.</a> Neuroreport. 20, 13-17 (2009).</li><li>Zangenehpour, S., Burke, M. W., Chaudhuri, A., Ptito, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Batch+immunostaining+for+large-scale+protein+detection+in+the+whole+monkey+brain.">Batch immunostaining for large-scale protein detection in the whole monkey brain.</a> J Vis Exp. 29, (2009).</li></ol>

Unbiased stereology in a primate brain is similar to that of other biological specimens. However, given the size of the primate brain, a careful processing plan is recommended since a single hemisphere of the vervet monkey yields more than 1200 sections when sliced at 50&micro;m. First we recommend stereotaxically blocking the brain into 1cm blocks, which provides a standard plane of section between blocks and subjects, minimizes partial sections between blocks, and provides manageable sized blocks5. A crucial...

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		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
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<tr>
<td>Stereologer</td>
<td>&nbsp;</td>
<td>Stereology Resource Center (Chester, MD)</td>
<td>www.disector.com</td>
<td>&nbsp;</td>
<tr>
<td>Stereology Tool-Kit Stereologer system</td>
<td>&nbsp;</td>
<td>BioQuant Life Sciences (Nashville, TN)</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>StereoInvestigator</td>
<td>&nbsp;</td>
<td>MicroBright Field (Willston, VT)</td>
<td>&nbsp;</td>
<td>&nbsp;</td>		</tbody>
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knowing what counts: unbiased stereology in the non-human primate brain

The non-human primate is an important translational species for understanding the normal function and disease processes of the human brain. Unbiased stereology, the method accepted as state-of-the-art for quantification of biological objects in tissue sections2, generates reliable structural data for biological features in the mammalian brain3. The key components of the approach are unbiased (systematic-random) sampling of anatomically defined structures (reference spaces), combined with quantification of cell numbers and size, fiber and capillary lengths, surface areas, regional volumes and spatial distributions of biological objects within the reference space4. Among the advantages of these stereological approaches over previous methods is the avoidance of all known sources of systematic (non-random) error arising from faulty assumptions and non-verifiable models. This study documents a biological application of computerized stereology to estimate the total neuronal population in the frontal cortex of the vervet monkey brain (Chlorocebus aethiops sabeus), with assistance from two commercially available stereology programs, BioQuant Life Sciences and Stereologer (Figure 1). In addition to contrast and comparison of results from both the BioQuant and Stereologer systems, this study provides a detailed protocol for the Stereologer system.

Watch this Scientific Journal Video about Knowing What Counts: Unbiased Stereology in the Non-human Primate Brain at JoVE.com

Knowing What Counts: Unbiased Stereology in the Non-human Primate Brain

The anatomical organization of the primate brain can provide important insights into normal and pathological conditions in humans. Unbiased stereology is a method for accurately and efficiently estimating the total neuron number (or other cell type) in a given reference space1.

The non-human primate is an important translational species for understanding the normal function and disease processes of the human brain. Unbiased ...

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