The overall goal of this procedure is to systematically collect and analyze a constant series of cerebral regions symmetrically dissected from both hemispheres of the human brain in order to assess severity, clinical pathologic staging and hemispheric distribution of various brain diseases. This method can help answer key questions in the neuropathology field providing a systematically organized collection of qualitative and quantitative neuropathologic data for detailed clinical pathologic and imaging pathologic correlations. The main advantage of this technique consists in yielding relevant data related to the pathologic processes per se as well as the histofunctional specialized aspect of each cerebral hemisphere.
After fixing post-mortem human brain tissue according to the text protocol, place the brain on a flat surface with the frontal poles directed opposite the investigator. Orient the brain to allow full and clear visualization of all cortical gyri and sulci of the entire cerebrum. Examine the tissue for meningeal anomalies, macroscopic hemispheric asymmetries, macroscopic tissue allegiance, congenital malformations, vessel abnormalities or any other abnormalities or unusual presentations of the cerebral surface.
With the frontal poles oriented away from the investigator and the superficial aspects of the hemispheres facing the investigator, take as many digital pictures as necessary to document possible macro anomalies and to account for possible clinic, neural, anatomical and post-cutting considerations. Take digital pictures perpendicularly to the brain to capture the entire cortical surface. Then using ink or colored needles, mark pre and post central cortical gyri.
Next, with the frontal poles still facing away, flip the brain 180 degrees then carefully inspect the base of the brain paying special attention to the cerebrovascular systems and the cranial nerves at the brain stem exit and entrance levels. Manage the olfactory bulbs and tracts with special care to avoid tissue laceration due to their extreme frailty. Take digital photographs perpendicularly to the brain to capture the entirety of the cortical and brain stem surfaces.
Facing the base of the brain and using a scalpel, cut the brain stem transversely at the upper portion of the pons as close as possible to the base of the cerebrum. Carefully inspect the substantia nigra for pallor and inspect other neighboring structures. Using an audio recording device, take note of any unusual appearance of the brain in comparison to a normal brain.
Here, the olfactory bulb and tract of the left and then right hemispheres are being dissected. Flip the brain back over and using a sharp knife, follow a fronto-occipital direction to separate the two hemispheres by cutting the corpus callosum centrally through the medial longitudinal fissure. Inspect each side of each hemisphere for possible anomalies.
After taking as many pictures as necessary, including perpendicularly to the brain to capture the entire cortical surface, place the two hemispheres flat laying on their medial aspects with the frontal lobes facing away from the investigator and with their centers touching. Using a sharp knife, manually cut through both cerebral hemispheres starting at the frontal poles and move towards the occipital poles through the entire length of the hemispheres to obtain two series of one centimeter thick slabs of brain tissue. Position the brain slabs in an anatomically organized and symmetric sequence on a flat white surface with a ruler printed on it for photographing.
Ensure that the slab's coronal surfaces are visible for direct eye inspection and digital photography. Use cutting surfaces with printed millimetric grids on both sides to localize brain structures, sizes, and possible abnormalities in a more accurate manner. With the coronal surfaces visible, take digital photographs perpendicularly to capture the entire cortical surface.
Take notes of any unusual aspect of the brain in comparison to a normal brain. Next, using a sharp scalpel, manually dissect small rectangular blocks of brain tissue for each established cerebral region. Follow the proposed cerebral region collection scheme described in this table.
Create the identifying codes to label the histo cassettes by generating random or semi-random numbers for each case where AD stands for Alzheimer's Disease Study, 16 stands for the year the autopsy was performed and 0001 is a progressive accession specimen number. Place each tissue block in separate histo cassettes. Take as many digital pictures as described earlier of the entire cut brain and the associated histo cassettes.
After taking a complete set of pictures, punch two to five millimeter pieces of tissue for DNA extraction and genetic analysis. Finally, immerse all histo cassettes containing brain tissue blocks in the same type of fixative solution as previously used and process the tissue according to the text protocol. This table describes some preliminary semi-quantitative data using a symmetric bihemispheric brain cutting procedure performed on a series of human brains collected in the brain bank.
Preliminary data show that most control brains from older subjects were actually positive for beta-A neuritic plaques and tau-neurofibrillary tangles and that levels of insoluble of beta-A neuritic plaques were higher in the left entorhinal cortex and hippocampus in comparison to the same brain regions on the right side. Once mastered, this technique can be completed in an hour for fixed brains and in two hours for fresh brains if it's properly performed. While attempting this procedure, it's important to remember following the proposed cutting scheme, collect each cerebral region from both hemispheres based upon precise human anatomical knowledge.
Following this procedure, other methods like DNA, RNA or protein extraction can be performed in order to answer additional questions such as what are the activation levels of a specific gene in a specific brain region in a specific hemisphere in a specific brain disease. With each diffusion, this technique will pave the way for researchers in the field of neuropathology, clinical pathology and imaging pathology correlation to explore the hemispheric related pathologic aspects of various neuropsychiatric diseases. After watching this video, you should have a good understanding of how to collect specific anatomical regions from both hemispheres of the human brain to assess severity of the disease, pathologic stage of the disease and hemispheric distribution of different neuropathologic processes.
Don't forget that working with the human brain is working with the history of a person with complex neuroanatomical circuits and possibly with different diseases and that blades, scalpels, formalin must be handled with extreme precaution.
