A new method for quantitative measurement of brain acetylcholinesterase (AChE) activity in living human brain using positron emission tomography (PET) is described. We tested several radiolabeled lipophilic acetylcholine analogs, e.g., N-methylpiperidyl esters, which readily entered the brain via the blood-brain barrier, were hydrolyzed selectively by AChE, and were then trapped in the brain. Among them, and tested and N-[11C]methylpiperidin-4-yl acetate ([11C]MP4A) was chosen as the tracer for PET. Quantitative measurement of cortical AChE was accomplished by fitting the time course of cerebral radioactivity concentration measured by PET and the metabolite-corrected arterial plasma input function using a nonlinear least-squares fitting method. Normal control studies of subjects with a wide range in age (24-89 years) showed no decrease in AChE activity in the cerebral cortex with age. Studies on patients with Alzheimer's disease demonstrated a widespread reduction of AChE activity in the cerebral cortex (more profound in early-onset than in late-onset Alzheimer's disease). Parkinson's disease and progressive supranuclear palsy, clinically similar disorders, could be differentiated with [11C]MP4A/PET studies. Simple methods without using an arterial input function are also proposed. The method provides a quantitative measure of the cholinergic aspect of brain function and proved to be useful in diagnosis of neurodegenerative disorders including Alzheimer's disease.