A Quantitative Histochemistry Technique for Measuring Regional Distribution of Acetylcholinesterase in the Brain Using Digital Scanning Densitometry

• Published in: Analytical biochemistry 2001-09, Vol.296 (1), p.18-28
• Main Authors: Ma, Tangeng, Cai, Zhengwei, Wellman, Susan E., Ho, Ing K.
• Format: Article
• Language: English
• Subjects: acetylcholinesterase; Acetylcholinesterase - analysis; Animals; Bio-Rad GS-700 imaging densitometer; Brain - anatomy & histology; Brain - enzymology; Brain Stem - enzymology; butyrylcholinesterase; Butyrylcholinesterase - analysis; Cerebellum - enzymology; Cerebral Cortex - enzymology; Cholinesterase Inhibitors; Densitometry - methods; digital scanning densitometry; Hippocampus - enzymology; Histocytological Preparation Techniques; Image Processing, Computer-Assisted; Male; Microscopy, Video; Microtomy; Molecular Dynamics personal densitometer; Paraoxon - analogs & derivatives; Putamen - enzymology; quantitative histochemistry; rat brain; Rats; Staining and Labeling; Thalamic Nuclei - enzymology; Urodela - metabolism
• ISSN: 0003-2697; 1096-0309
• DOI: 10.1006/abio.2001.5208

Studies of brain acetylcholinesterase (AChE) are traditionally based on biochemical assays, immunoreactivity, and histochemistry. Conventional histochemistry yields rich morphological data from tissue sections but yields quantitative results only with great difficulty. Several histochemical methods developed in recent years, including microdensitometry, microphotometry, and video-based histochemistry, are effective in quantitative and detailed study of AChE in tissue sections. However, they are usually time-consuming. As we report here, we adapted digital scanning densitometry to quantitate AChE histochemical staining in brain sections. The AChE and butyrylcholinesterase (BuChE), as measured by the method, were heterogeneously distributed throughout the brain, results that are consistent with those obtained by biochemical methods. The staining intensity is dependent on section thickness, substrate concentration, and reaction time. The cholinesterase inhibitor methyl paraoxon significantly decreased AChE staining intensity. Furthermore, data acquired from densitometry are similar to those obtained by video-based microscopy or by spectrophotometry. The advantage of the densitometric measurements compared to other quantitative histochemical methods is that it is very rapid while collecting data that are equivalent in quality. Because the digital scanning densitometers provide high quality and sensitive imaging, wide dynamic ranges, and convenient image analysis software, they are very useful tools in quantitative histochemistry.