Introduction - Brain Sampling


The NNTC Neuropathology Subcommittee suggested that three samples are required. 1) Frontal cortex was selected as a sample of neocortex. 2) Frontal lobe white matter. 3) A sample of gray matter from the basal ganglia (head of caudate). Some type of inventory checking is going to be required to identify cases with available material in these proposed anatomical sites. It is understood that the three samplings are mostly independent for computational purposes, at least in the initial stages of the study.


Frontal cortex: The primary target of neocortex was the middle frontal gyrus. In order of preference, the sample would come from Brodmann area 9, area 8, or area 46. The area of interest corresponds generally to brain section number 1 of the NNTC fixed brain dissection protocol Area 6 is too close to premotor areas and probably should be avoided. The drawing above is from a protocol that was similar but not identical to this one. The slice shown above is anterior to area 8 and shows area 9 instead. Ideally, the sampling of cortex should be full thickness to include all neuronal lamina. An illustration of marked neural lamina is included here to show how the distribution of chemicals in the cortex varies (this is a stain for a kinase). Note how lamina II and V are enriched with this kinase. A rectangular piece of gyrus that has all six layers of neurons and no underlying white matter was sampled. The top of the gyrus was given preference, but the side of the gyrus was satisfactory. Perisulcal cortex was a last resort.

Frontal lobe white matter: The primary site that was sampled in the frontal lobe was a place that is clear of the cortex. An area close to the middle frontal gyrus was acceptable. Perisulcal white matter lying about 0.5 cm from the sulcus is easy to dissect. With more precision one can obtain gyral white matter as shown in the diagram on the previous page. Also acceptable was a sample from the deeper white matter of the frontal lobe. The rationale for trying to sample white matter adjacent to the cortical sample was that afferent and efferent axon connectivity between the two samplings is likely. Deeper areas of white matter were quite acceptable when more superficial white matter could not be dissected cleanly and confidently.

Caudate nucleus: Neostriatum is the part of basal ganglia in which it is most possible to obtain pure gray matter. The head of the caudate nucleus was a large focus that is easily dissected because it borders the lateral ventricle, providing a convenient lateral entry point for the dissecting tool. Other sectors of caudate were also acceptable. Putamen was the third area of preference even though some pencil fibers that contain white matter could have introduced some contamination with white matter. If none of the above were available, globus pallidus was considered next in line.

Physical dissection of frozen brain samples: Some prefer to avoid warming below –80C for RNA work; theoretically the RNA is vulnerable to degradation. A saw was used to excise the sample at -80C.